ORCID Profile
0000-0002-9400-893X
Current Organisations
Perron Institute for Neurological and Translational Science
,
Murdoch University
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Proteomics and Intermolecular Interactions (excl. Medical Proteomics) | Biochemistry and Cell Biology | Medical Biochemistry and Metabolomics not elsewhere classified | Biologically Active Molecules
Expanding Knowledge in the Chemical Sciences | Expanding Knowledge in the Agricultural and Veterinary Sciences | Expanding Knowledge in the Biological Sciences |
Publisher: Springer Science and Business Media LLC
Date: 31-07-2018
Publisher: ClinMed International Library
Date: 31-12-2018
Publisher: Elsevier BV
Date: 08-2013
DOI: 10.1016/J.JNEUROIM.2013.04.022
Abstract: Non-Human Leukocyte Antigen (HLA) genes have concomitant, although modest, effects on multiple sclerosis (MS) susceptibility however findings have varied in different populations. Here we present the results of an association study of 16 single nucleotide polymorphisms (SNPs) in 10 non-HLA genes (IL7R, IL2RA, CLEC-16A, TYK2, CD58, IRF5, STAT3, CTLA-4, APOE, ICAM-1) in a Western Australian cohort of 350 MS patients and 498 population control subjects. Our results indicate that in this population, SNPs in IL7R, TYK2, IRF5 and APOE have modifying effects on MS susceptibility. We also found evidence of interactive protective effects between polymorphisms in the IL7R/CD58, CLEC-16A/CTLA-4, and TYK2/IRF5 genes, which in some instances are restricted within HLA- or gender-defined groups.
Publisher: MDPI AG
Date: 25-06-2020
DOI: 10.3390/IJMS21124511
Abstract: Duchenne muscular dystrophy (DMD) is a severe muscle wasting disease typically caused by protein-truncating mutations that preclude synthesis of a functional dystrophin. Exonic deletions are the most common type of DMD lesion, however, whole exon duplications account for between 10–15% of all reported mutations. Here, we describe in vitro evaluation of antisense oligonucleotide-induced splice switching strategies to re-frame the transcript disrupted by a multi-exon duplication within the DMD gene. Phosphorodiamidate morpholino oligomers and phosphorodiamidate morpholino oligomers coupled to a cell penetrating peptide were evaluated in a Duchenne muscular dystrophy patient cell strain carrying an exon 14–17 duplication. Two strategies were employed the conventional approach was to remove both copies of exon 17 in addition to exon 18, and the second strategy was to remove only the first copy of exon 17. Both approaches result in a larger than normal but in-frame DMD transcript, but surprisingly, the removal of only the first exon 17 appeared to be more efficient in restoring dystrophin, as determined using western blotting. The emergence of a normal sized DMD mRNA transcript that was not apparent in untreated s les may have arisen from back splicing and could also account for some of the dystrophin protein being produced.
Publisher: Wiley
Date: 23-08-2013
DOI: 10.1111/AGE.12082
Abstract: Obesity has reached epidemic proportions globally and has become the cause of several major health risks worldwide. Presently, more than 100 loci have been related to obesity and metabolic traits in humans by genome-wide association studies. The complex genetic architecture behind obesity has triggered a need for the development of better animal models than rodents. The pig has emerged as a very promising biomedical model to study human obesity traits. In this study, we have characterized the expression patterns of six obesity-related genes, leptin (LEP), leptin receptor (LEPR), melanocortin 4 receptor (MC4R), fat mass and obesity associated (FTO), neuronal growth regulator 1 (NEGR)1 and adiponectin (ADIPOQ), in seven obesity-relevant tissues (liver muscle pancreas hypothalamus and retroperitoneal, subcutaneous and mesenteric adipose tissues) in two pig breeds (production pigs and Göttingen minipigs) that deviate phenotypically and genetically from each other with respect to obesity traits. We observe significant differential expression for LEP, LEPR and ADIPOQ in muscle and in all three adipose tissues. Interestingly, in pancreas, LEP expression is only detected in the fat minipigs. FTO shows significant differential expression in all tissues analyzed, and NEGR1 shows significant differential expression in muscle, pancreas, hypothalamus and subcutaneous adipose tissue. The MC4R transcript can be detected only in hypothalamus. In general, the expression profiles of the investigated genes are in accordance with those observed in human studies. Our study shows that both the differences between the investigated breeds and the phenotypic state with respect to obesity/leanness play a large role for differential expression of the obesity-related genes.
Publisher: Elsevier BV
Date: 08-2009
DOI: 10.1038/MT.2009.49
Publisher: Frontiers Media SA
Date: 09-09-2014
Publisher: Hindawi Limited
Date: 1994
Abstract: Duchenne and Becker muscular dystrophies (DMD and BMD) are allelic X-linked disorders arising from mutations in the (2.4 Mb) dystrophin gene at Xp21. We have applied the reverse transcriptase-polymerase chain reaction (RT-PCR) to identify a larger than normal dystrophin mRNA from a male with Duchenne muscular dystrophy and his younger affected brother. The increased size of the dystrophin mRNA was due to a splice-site mutation at the exon 26:intron 26 junction where a T to G substitution prevented normal RNA processing. A cryptic splice-site, downstream of the mutation, was activated during processing, resulting in the inclusion of 117 bases of intron 26. This insertion introduced an in-frame stop codon into the mature dystrophin mRNA. An allele-specific test was developed to identify the mutation and was applied to this family. Interestingly, the mother of the two affected boys did not carry the mutation, as determined by allele-specific lification and direct DNA sequence analysis, indicating gonadal mosaicism. Her eldest daughter, designated as a carrier based upon conventional testing and haplotype analysis, also did not carry the family mutation. Initial haplotyping of the family appeared to be straightforward with gonadal mosaicism becoming evident only after allele-specific analysis. The application of linked markers to identify the disease allele for conventional genetic counselling would have been erroneous in this family and highlights the diagnostic power of precise identification of the disease-causing mutation.
Publisher: Research Square Platform LLC
Date: 18-01-2021
DOI: 10.21203/RS.3.RS-144809/V1
Abstract: Antisense oligomers (AOs) are increasingly being used for modulating RNA splicing in live cells, both for research and for therapeutic purposes. While the most common intended effect of these AOs is to induce skipping of whole exons, rare ex les are emerging of AOs that induce skipping of only part of an exon, through activation of an internal cryptic splice site. In this report, we examined seven such ex les of AO-induced cryptic splice site activation – five new ex les from our own experiments and three from reports published by others. We modelled the predicted effects that AO binding would have on the secondary structure of each of the RNA targets, and how these alterations would in turn affect the accessibility of the RNA to splice factors. We observed that a common predicted effect of AO binding was a disruption to the exon definition signal within the exon’s excluded segment.
Publisher: Wiley
Date: 06-2020
DOI: 10.1111/RESP.13778
Publisher: Cambridge University Press (CUP)
Date: 2017
DOI: 10.1017/S175275620000418X
Abstract: Selection for milk production alone would lead to a genetic decline in cow fertility (e.g. Kadarmideen et al., 2000) thus incorporation of fertility in selection seems desirable. The Dairy Information System (DAISY) is a comprehensive recording scheme for insemination and health events and hence the data quality is more reliable for genetic analysis of cow fertility. The objective of this study was to analyse DAISY data to estimate sire breeding values and genetic parameters for various fertility measures based on insemination and calving events on daughters.
Publisher: MDPI AG
Date: 04-2022
DOI: 10.3390/IJMS23073937
Abstract: Spinal muscular atrophy (SMA) is a severe, debilitating neuromuscular condition characterised by loss of motor neurons and progressive muscle wasting. SMA is caused by a loss of expression of SMN1 that encodes the survival motor neuron (SMN) protein necessary for the survival of motor neurons. Restoration of SMN expression through increased inclusion of SMN2 exon 7 is known to ameliorate symptoms in SMA patients. As a consequence, regulation of pre-mRNA splicing of SMN2 could provide a potential molecular therapy for SMA. In this study, we explored if splice switching antisense oligonucleotides could redirect the splicing repressor hnRNPA1 to the hnRNPA1b isoform and restore SMN expression in fibroblasts from a type I SMA patient. Antisense oligonucleotides (AOs) were designed to promote exon 7b retention in the mature mRNA and induce the hnRNPA1b isoform. RT-PCR and western blot analysis were used to assess and monitor the efficiency of different AO combinations. A combination of AOs targeting multiple silencing motifs in hnRNPA1 pre-mRNA led to robust hnRNPA1b induction, which, in turn, significantly increased expression of full-length SMN (FL-SMN) protein. A combination of PMOs targeting the same motifs also strongly induced hnRNPA1b isoform, but surprisingly SMN2 exon 5 skipping was detected, and the PMO cocktail did not lead to a significant increase in expression of FL-SMN protein. We further performed RNA sequencing to assess the genome-wide effects of hnRNPA1b induction. Some 3244 genes were differentially expressed between the hnRNPA1b-induced and untreated SMA fibroblasts, which are functionally enriched in cell cycle and chromosome segregation processes. RT-PCR analysis demonstrated that expression of the master regulator of these enrichment pathways, MYBL2 and FOXM1B, were reduced in response to PMO treatment. These findings suggested that induction of hnRNPA1b can promote SMN protein expression, but not at sufficient levels to be clinically relevant.
Publisher: Wiley
Date: 29-01-2009
Publisher: Bentham Science Publishers Ltd.
Date: 2009
DOI: 10.2174/1874467210902010110
Abstract: Duchenne muscular dystrophy (DMD) arises from protein-truncating mutations in the large dystrophin gene that preclude synthesis of a functional protein that primarily stabilizes muscle fibre membranes. The absence of dystrophin leads to this most common and serious form of childhood muscle-wasting. Since the identification of the dystrophin gene in 1987, cell and gene repair or replacement therapies have been evaluated for DMD treatment and one genetic intervention, exon skipping, is now in clinical trials. Antisense oligomers have been designed to redirect dystrophin splicing patterns so that targeted exons may be removed from a defective dystrophin pre-mRNA to either restore the reading frame of a deletion, or excise an in-frame exon corrupted by a nonsense mutation or micro-insertion/deletion. This review discusses the evolution of oligomer induced exon skipping, including in vitro applications, evaluation of different oligomer chemistries, the treatment of animal models and alternative exon skipping strategies involving viral expression cassettes and ex vivo manipulation of stem cells. The discussion culminates with the current clinical trials and the great challenges that lie ahead. The major obstacle to the implementation of personalised genetic treatments to address the many different mutations that can lead to DMD, are considered to be establishing effective treatments for the different patients and their mutations. Furthermore, the view of regulatory authorities in assessing preclinical data on potentially scores of different but class-specific compounds will be of paramount importance in expediting the clinical application of exon skipping therapy for this serious and relentlessly progressive muscle wasting disease.
Publisher: MDPI AG
Date: 31-10-2019
DOI: 10.3390/IJMS20215434
Abstract: Spinocerebellar ataxia type 3 (SCA3) is a devastating neurodegenerative disease for which there is currently no cure, nor effective treatment strategy. One of nine polyglutamine disorders known to date, SCA3 is clinically heterogeneous and the main feature is progressive ataxia, which in turn affects speech, balance and gait of the affected in idual. SCA3 is caused by an expanded polyglutamine tract in the ataxin-3 protein, resulting in conformational changes that lead to toxic gain of function. The expanded glutamine tract is located at the 5′ end of the penultimate exon (exon 10) of ATXN3 gene transcript. Other studies reported removal of the expanded glutamine tract using splice switching antisense oligonucleotides. Here, we describe improved efficiency in the removal of the toxic polyglutamine tract of ataxin-3 in vitro using phosphorodiamidate morpholino oligomers, when compared to antisense oligonucleotides composed of 2′-O-methyl modified bases on a phosphorothioate backbone. Significant downregulation of both the expanded and non-expanded protein was induced by the morpholino antisense oligomer, with a greater proportion of ataxin-3 protein missing the polyglutamine tract. With growing concerns over toxicity associated with long-term administration of phosphorothioate oligonucleotides, the use of a phosphorodiamidate morpholino oligomer may be preferable for clinical application. These results suggest that morpholino oligomers may provide greater therapeutic benefit for the treatment of spinocerebellar ataxia type 3, without toxic effects.
Publisher: Ovid Technologies (Wolters Kluwer Health)
Date: 13-10-2021
Publisher: Elsevier BV
Date: 10-2005
DOI: 10.1016/J.COPH.2005.06.001
Abstract: The manipulation of pre-mRNA to alter gene transcript splicing patterns offers considerable potential for many genetic disorders. In particular, the targeted removal of one or more exons from a gene transcript can skip over, or compensate for, disease-causing mutations. Duchenne muscular dystrophy (DMD), the most common and severe form of muscular dystrophy, is one such disorder that could benefit from this strategy. Splicing modulation can convert a DMD phenotype into the less severe allelic Becker-like phenotype. Recent studies using antisense oligonucleotide-targeted exon skipping to induce near normal dystrophin in vivo in animal models, and in vitro in DMD cell lines, highlight the promise of this approach. On the basis of these successes, human clinical trials could be realized in the near future.
Publisher: S. Karger AG
Date: 1995
DOI: 10.1159/000133928
Abstract: A sequence-tagged site (STS) was developed for the human skeletal muscle α-tropomyosin gene (TPM1) and used to isolate a genomic clone, λTPM1.1, containing part of the TPM1 gene. Fluorescence in situ hybridization of this clone to metaphase chromosome spreads localised TPM1 to chromosome band 15q22. This localisation in humans is consistent with that recently described for the mouse.
Publisher: Elsevier BV
Date: 11-2022
Publisher: Elsevier BV
Date: 10-2008
Publisher: Elsevier BV
Date: 07-2021
Publisher: American Society for Clinical Investigation
Date: 21-03-2019
Publisher: Public Library of Science (PLoS)
Date: 18-10-2017
Publisher: Springer Science and Business Media LLC
Date: 14-01-2020
DOI: 10.1038/S41598-019-57182-4
Abstract: Improving feed efficiency (FE) is a major goal of pig breeding, reducing production costs and providing sustainability to the pig industry. Reliable predictors for FE could assist pig producers. We carried out untargeted blood metabolite profiling in uncastrated males from Danbred Duroc (n = 59) and Danbred Landrace (n = 50) pigs at the beginning and end of a FE testing phase to identify biomarkers and biological processes underlying FE and related traits. By applying linear modeling and clustering analyses coupled with WGCNA framework, we identified 102 and 73 relevant metabolites in Duroc and Landrace based on two s ling time points. Among them, choline and pyridoxamine were hub metabolites in Duroc in early testing phase, while, acetoacetate, cholesterol sulfate, xanthine, and deoxyuridine were identified in the end of testing. In Landrace, cholesterol sulfate, thiamine, L-methionine, chenodeoxycholate were identified at early testing phase, while, D-glutamate, pyridoxamine, deoxycytidine, and L-2-aminoadipate were found at the end of testing. Validation of these results in larger populations could establish FE prediction using metabolomics biomarkers. We conclude that it is possible to identify a link between blood metabolite profiles and FE. These results could lead to improved nutrient utilization, reduced production costs, and increased FE.
Publisher: Elsevier BV
Date: 04-2001
DOI: 10.1016/S0960-8966(00)00187-5
Abstract: Golden retriever muscular dystrophy arises from a mutation in the acceptor splice site of intron 6 of the dystrophin gene. Skipping of exon 7 disrupts the mRNA reading frame and results in premature termination of translation. We are using this animal model to evaluate treatments for Duchenne muscular dystrophy, including gene repair induced by chimeric oligonucleotides. After injection of golden retriever muscular dystrophy (GRMD) muscle with a chimeric oligonucleotide to repair the lesion, immunostaining revealed a modest increase in the number of dystrophin-positive fibres at the injection sites. Dystrophin gene transcripts containing exon 7 were detected by reverse transcription-polymerase chain reaction, suggesting that low levels of splice site correction may have occurred. However, DNA sequencing of these apparently normal dystrophin gene transcripts revealed that the first five bases of exon 7 were missing. It will be important to be aware of this phenomenon with respect to further gene correction studies in the canine model.
Publisher: SAGE Publications
Date: 03-2023
DOI: 10.1177/03000605231159335
Abstract: The use of artificial intelligence (AI) to generate automated early warnings in epidemic surveillance by harnessing vast open-source data with minimal human intervention has the potential to be both revolutionary and highly sustainable. AI can overcome the challenges faced by weak health systems by detecting epidemic signals much earlier than traditional surveillance. AI-based digital surveillance is an adjunct to—not a replacement of—traditional surveillance and can trigger early investigation, diagnostics and responses at the regional level. This narrative review focuses on the role of AI in epidemic surveillance and summarises several current epidemic intelligence systems including ProMED-mail, HealthMap, Epidemic Intelligence from Open Sources, BlueDot, Metabiota, the Global Biosurveillance Portal, Epitweetr and EPIWATCH. Not all of these systems are AI-based, and some are only accessible to paid users. Most systems have large volumes of unfiltered data only a few can sort and filter data to provide users with curated intelligence. However, uptake of these systems by public health authorities, who have been slower to embrace AI than their clinical counterparts, is low. The widespread adoption of digital open-source surveillance and AI technology is needed for the prevention of serious epidemics.
Publisher: Springer Science and Business Media LLC
Date: 28-09-2016
DOI: 10.1038/NATURE19806
Publisher: Springer Science and Business Media LLC
Date: 21-03-2016
DOI: 10.1007/S13353-016-0344-7
Abstract: The use of genome-wide association results combined with other genomic approaches may uncover genes and metabolic pathways related to complex traits. In this study, the phenotypic and genotypic data of 1475 Nellore (Bos indicus) cattle and 941,033 single nucleotide polymorphisms (SNPs) were used for genome-wide association study (GWAS) and copy number variations (CNVs) analysis in order to identify candidate genes and putative pathways involved with the feed conversion ratio (FCR). The GWAS was based on the Bayes B approach analyzing genomic windows with multiple regression models to estimate the proportion of genetic variance explained by each window. The CNVs were detected with PennCNV software using the log R ratio and B allele frequency data. CNV regions (CNVRs) were identified with CNVRuler and a linear regression was used to associate CNVRs and the FCR. Functional annotation of associated genomic regions was performed with the Database for Annotation, Visualization and Integrated Discovery (DAVID) and the metabolic pathways were obtained from the Kyoto Encyclopedia of Genes and Genomes (KEGG). We showed five genomic windows distributed over chromosomes 4, 6, 7, 8, and 24 that explain 12 % of the total genetic variance for FCR, and detected 12 CNVRs (chromosomes 1, 5, 7, 10, and 12) significantly associated [false discovery rate (FDR) < 0.05] with the FCR. Significant genomic regions (GWAS and CNV) harbor candidate genes involved in pathways related to energetic, lipid, and protein metabolism. The metabolic pathways found in this study are related to processes directly connected to feed efficiency in beef cattle. It was observed that, even though different genomic regions and genes were found between the two approaches (GWAS and CNV), the metabolic processes covered were related to each other. Therefore, a combination of the approaches complement each other and lead to a better understanding of the FCR.
Publisher: Cold Spring Harbor Laboratory
Date: 17-10-2018
DOI: 10.1101/442756
Abstract: Birth weight (BW) variation is influenced by fetal and maternal genetic and non-genetic factors, and has been reproducibly associated with future cardio-metabolic health outcomes. These associations have been proposed to reflect the lifelong consequences of an adverse intrauterine environment. In earlier work, we demonstrated that much of the negative correlation between BW and adult cardio-metabolic traits could instead be attributable to shared genetic effects. However, that work and other previous studies did not systematically distinguish the direct effects of an in idual’s own genotype on BW and subsequent disease risk from indirect effects of their mother’s correlated genotype, mediated by the intrauterine environment. Here, we describe expanded genome-wide association analyses of own BW (n=321,223) and offspring BW (n=230,069 mothers), which identified 278 independent association signals influencing BW (214 novel). We used structural equation modelling to decompose the contributions of direct fetal and indirect maternal genetic influences on BW, implicating fetal- and maternal-specific mechanisms. We used Mendelian randomization to explore the causal relationships between factors influencing BW through fetal or maternal routes, for ex le, glycemic traits and blood pressure. Direct fetal genotype effects dominate the shared genetic contribution to the association between lower BW and higher type 2 diabetes risk, whereas the relationship between lower BW and higher later blood pressure (BP) is driven by a combination of indirect maternal and direct fetal genetic effects: indirect effects of maternal BP-raising genotypes act to reduce offspring BW, but only direct fetal genotype effects (once inherited) increase the offspring’s later BP. Instrumental variable analysis using maternal BW-lowering genotypes to proxy for an adverse intrauterine environment provided no evidence that it causally raises offspring BP. In successfully separating fetal from maternal genetic effects, this work represents an important advance in genetic studies of perinatal outcomes, and shows that the association between lower BW and higher adult BP is attributable to genetic effects, and not to intrauterine programming.
Publisher: IEEE
Date: 2005
Publisher: Springer Science and Business Media LLC
Date: 30-09-2014
Publisher: Wiley
Date: 08-1997
DOI: 10.1111/J.1365-2052.1997.00138.X
Abstract: The cDNA sequences for Cu,Zn superoxide dismutase from two Cervus elaphus subspecies, North American wapiti and European red deer, were determined. The derived amino acid sequences showed two differences: residue 8 was Leu in wapiti and Met in red deer and residue 25 was His in wapiti and Asn in red deer. The extra positive charge at position 25 in the wapiti isoform accounted for its greater mobility towards the cathode during non-denaturing electrophoresis, a procedure widely used in the genetic analysis of deer. There was no difference in specific activity between the two Cu,Zn superoxide dismutase isoforms, but the wapiti isoform was slightly more susceptible to heat denaturation.
Publisher: AMPCo
Date: 07-1996
DOI: 10.5694/J.1326-5377.1996.TB124852.X
Abstract: To determine the relationship between the apolipoprotein E epsilon 4 allele and autopsy-verified Alzheimer's disease (AD) in an Australian population. Retrospective case-control study. Royal Perth Hospital, Perth, Western Australia (a tertiary referral hospital). 50 subjects with "definite" AD (according to the histological and clinical criteria of the Consortium to Establish a Registry for Alzheimer's Disease [CERAD]) and 30 control subjects who had died from a non-neurological disease were randomly selected from the hospital's neuropathology register. Histological grading of brain sections stained with the modified Bielschowsky stain according to the criteria of CERAD number (burden) of neuritic plaques apolipoprotein E genotype (APOE). Frequency of the epsilon 4 allele was significantly higher in the AD group (37%) than in the control group (2%) (chi 2 = 25.8 P < 0.00001). In the AD group, 50% of subjects were heterozygous for the epsilon 4 allele and 12% were homozygous, while in the control group one subject was heterozygous for the allele and none were homozygous. No association was seen between the epsilon 4 allele and neuritic plaque burden in the hippoc us, entorhinal cortex, middle frontal gyrus or inferior parietal lobule in subjects with AD. Our findings confirm an association between the epsilon 4 allele and autopsy-verified AD. The epsilon 4 allele may be an important risk factor for susceptibility to AD in the general Australian population.
Publisher: Elsevier BV
Date: 2014
DOI: 10.1038/MTNA.2014.8
Publisher: MDPI AG
Date: 09-2017
Publisher: Wiley
Date: 2003
DOI: 10.1002/JGM.361
Abstract: Duchenne muscular dystrophy (DMD) is an X-linked recessive muscle wasting disorder characterised by the absence of the protein dystrophin. Antisense oligonucleotides have been used to re-direct dystrophin pre-mRNA processing by blocking sequences crucial to pre-mRNA splicing, thereby inducing skipping of specific exons. We wished to determine which splicing motifs are most amenable as targets for antisense oligonucleotide induction of efficient and specific skipping of selected exons. Antisense oligonucleotides were directed at regions of dystrophin exon 19 involved in pre-mRNA splicing, including the donor and acceptor splice sites and the exon splicing enhancer (ESE). Cultured myotubes were transfected with antisense oligonucleotides at various concentrations and studies undertaken to determine both specificity and efficiency of induced exon 19 skipping. Antisense oligonucleotides as small as 12 nucleotides targeting the ESE induced consistent and specific skipping of only exon 19 in both human and normal and mdx mouse myotubes. Antisense oligonucleotides directed at the donor and acceptor splice sites also induced specific exon 19 skipping while mismatched antisense oligonucleotides could only induce skipping when delivered at higher concentrations. No other dystrophin exons were removed from the mature mRNA as a consequence of these antisense oligonucleotides treatments. Antisense oligonucleotides directed at the ESE tended to be marginally more efficient than those which targeted the donor or acceptor splice sites, based on their ability to induce specific skipping at lower concentrations. The specificity of exon removal does not appear to be a function of target selection, but may reflect the combination of the splicing motifs and position of that exon in the pre-mRNA.
Publisher: CSIRO Publishing
Date: 2019
DOI: 10.1071/RD18338
Abstract: In this paper we first provide a brief review of main results from our previously published studies on genome-wide gene expression (transcriptomics) in donor and recipient cattle used in invitro production (IVP) of embryos and embryo transfer (ET). Then, we present novel results from applying integrative systems genomics and biological analyses where transcriptomics data are combined with genomic data in both donor and recipient cattle to map expression quantitative trait loci (eQTLs). The eQTLs are genetic markers that can regulate or control the expression of genes in the entire genome, via complex molecular mechanisms, and thus can act as a powerful tool for genomic and gene-assisted selection. We identified significant eQTLs potentially controlling the expression of 13 candidate genes for donor cow quality (IVP parameters e.g. cyclin B1 (CCNB1), outer dense fiber of sperm tails 2 like (ODF2L)) and 19 candidate genes for recipient cows quality (endometrial receptivity e.g. ER membrane protein complex subunit 9 (EMC9), mannosidase beta (MANBA), peptidase inhibitor 16 (PI16)). Annotation and colocation of detected eQTLs show that some of the eQTLs are in the same genomic regions previously reported as QTLs for reproduction-related traits. However, eQTLs and the candidate genes identified should be further validated in larger populations before implementation as genetic markers or used in genomic selection for improving IVP and ET performance.
Publisher: Genetics and Molecular Research
Date: 2016
DOI: 10.4238/GMR15048930
Abstract: Feed intake, feed efficiency, and weight gain are important economic traits of beef cattle in feedlots. In the present study, we investigated the physiological processes underlying such traits from the point of view of systems genetics. Firstly, using data from 1334 Nellore (Bos indicus) cattle and 943,577 single nucleotide polymorphisms (SNPs), a genome-wide association analysis was performed for dry matter intake, average daily weight gain, feed conversion ratio, and residual feed intake with a Bayesian Lasso procedure. Genes within 50-kb SNPs, most relevant for explaining the genomic variance, were annotated and the biological processes underlying the traits were inferred from Database for Annotation, Visualization and Integrated Discovery (DAVID) and Kyoto Encyclopedia of Genes and Genomes (KEGG) databases. Our results indicated several putative genomic regions associated with the target phenotypes and showed that almost all genomic variances were in the SNPs located in the intergenic and intronic regions. We further identified five main metabolic pathways related to ion transport, body composition, and feed intake control, which influenced the four phenotypes simultaneously. The systems genetics approach used in this study revealed novel pathways related to feed efficiency traits in beef cattle.
Publisher: Elsevier BV
Date: 10-2017
Publisher: Public Library of Science (PLoS)
Date: 12-04-2017
Publisher: Elsevier BV
Date: 2013
DOI: 10.1016/J.JNEUROIM.2012.09.003
Abstract: We compared the carriage frequencies of HLA-DRB3 and its major alleles and of HLA-DRB4 and HLA-DRB5 in an Australian sIBM cohort and a population control group who had previously been genotyped for the HLA-DRB1*03:01 risk allele. There was a strong disease association with the carriage of the DRB3*01:01 allele which was accounted for by its linkage disequilibrium with DRB1*03:01. The carriage of HLA-DRB4 was found to be strongly protective and abrogated the risk effect of HLA-DRB1*03:01. The findings indicate that haplotypic combinations of alleles at the HLA-DRB1 and secondary HLA-DRB loci have important risk modifying effects in sIBM.
Publisher: MDPI AG
Date: 24-06-2019
DOI: 10.3390/F10060524
Abstract: In the inland riverine environment of Australia, wildfires not only threaten human life and cause economic loss but also make distinctive impacts on the ecosystem (e.g., injuring or killing fire-sensitive wetland species such as the river red gum). Understanding the drivers of wildfire occurrence patterns in this particular environment is vital for fire-risk reduction and ecologically sustainable management. This study investigated patterns and driving factors of wildfire occurrence over the years from 2001 to 2016 and across the New South Wales side of the Riverina bioregion. Descriptive analyses were conducted for fires of different causes and that burned different vegetation types. Logistic regression models were developed by incorporating factors that provide information on weather, climate, fuel, topography and ignition sources. Analyses revealed that most fires occurred in summer, with human-caused fires primarily in spring and summer, and natural fires in summer. Summer was the most fire-prone season in forested wetlands, whereas fires in drylands mostly occurred during spring and summer. Fire probabilities were higher under severe weather conditions, in areas with higher annual rainfall, in forested wetlands and in areas with intermediate inundation frequencies. Special attention needs to be paid to the effects of vegetation type and inundation frequency on fire occurrence. Weather, climate& fuel and ignition sources were comparably important in explaining human-caused fire occurrence, whereas weather was more important than climate& fuel in explaining natural fire occurrence. Understandings obtained from this study can potentially support the planning of fire and forest management, as well as to supplement the relatively scarce knowledge on riverine wildfire occurrence.
Publisher: F1000 Research Ltd
Date: 22-05-2019
DOI: 10.12688/F1000RESEARCH.18466.1
Abstract: Recent approvals of oligonucleotide analogue drugs to alter gene expression have been welcomed by patient communities but not universally supported. These compounds represent a class of drugs that are designed to target a specific gene transcript, and they include a number of chemical entities to evoke different antisense mechanisms, depending upon the disease aetiology. To date, oligonucleotide therapeutics that are in the clinic or at advanced stages of translation target rare diseases, posing challenges to clinical trial design, recruitment and evaluation and requiring new evaluation paradigms. This review discusses the currently available and emerging therapeutics that alter exon selection through an effect on pre-mRNA splicing and explores emerging concerns over safety and efficacy. Although modification of synthetic nucleic acids destined for therapeutic application is common practice to protect against nuclease degradation and to influence drug function, such modifications may also confer unexpected physicochemical and biological properties. Negatively charged oligonucleotides have a strong propensity to bind extra- and intra-cellular proteins, whereas those analogues with a neutral backbone show inefficient cellular uptake but excellent safety profiles. In addition, the potential for incorporation of chemically modified nucleic acid monomers, yielded by nuclease degradation of exogenous oligonucleotides, into biomolecules has been raised and the possibility not entirely discounted. We conclude with a commentary on the ongoing efforts to develop novel antisense compounds and enhance oligonucleotide delivery in order to further improve efficacy and accelerate implementation of antisense therapeutics for human disease.
Publisher: Informa UK Limited
Date: 06-2015
Publisher: Elsevier BV
Date: 07-2021
DOI: 10.1016/J.SCR.2021.102439
Abstract: Mutations in ABCA4 gene are causative for autosomal recessive Stargardt disease (STGD1), the most common inherited retinal dystrophy. Here, we report the generation of an induced pluripotent stem cell (iPSC) line from a STGD1 patient carrying biallelic c.[5461-10T>C A>T] [6077T>C] mutations in the ABCA4 gene. Episomes carrying OCT4, SOX2, KLF4, L-MYC, LIN28 and mp53DD were employed for the reprogramming of patient-derived fibroblasts. This iPSC line expressed comparable pluripotency markers as in a commercially available human iPSC line, displayed normal karyotype and potential for trilineage differentiation, and were negative for both reprogramming episomes and mycoplasma test.
Publisher: Elsevier BV
Date: 10-2008
Publisher: Ovid Technologies (Wolters Kluwer Health)
Date: 07-2022
DOI: 10.1097/APO.0000000000000546
Abstract: Usher syndrome (USH) is the most common form of deaf-blindness, with an estimated prevalence of 4.4 to 16.6 per 100,000 people worldwide. The most common form of USH is type IIA (USH2A), which is caused by homozygous or compound heterozygous mutations in the USH2A gene and accounts for around half of all USH cases. USH2A patients show moderate to severe hearing loss from birth, with diagnosis of retinitis pigmentosa in the second decade of life and variable vestibular involvement. Although hearing aids or cochlear implants can provide some mitigation of hearing deficits, there are currently no treatments aimed at preventing or restoring vision loss in USH2A patients. In this review, we first provide an overview of the molecular biology of the USH2A gene and its protein isoforms, which include a transmembrane protein (TM usherin) and an extracellular protein (EC usherin). The role of these proteins in the inner ear and retina and their impact on the pathogenesis of USH2A is discussed. We review animal cell-derived and patient cell-derived models currently used in USH2A research and conclude with an overview of potential treatment strategies currently in preclinical development and clinical trials.
Publisher: Elsevier BV
Date: 07-1997
DOI: 10.1016/S0960-8966(97)00058-8
Abstract: The mdx mouse, an animal model used to study Duchenne muscular dystrophy (DMD), has a nonsense mutation in exon 23 of the dystrophin gene which should result in a truncated protein that cannot be correctly localized at the sarcolemma of the muscle fibres. Immunohistochemical staining with anti-dystrophin antibodies had shown that while most of the muscle tissue was dystrophin-negative, a small percentage of muscle fibres were clearly dystrophin-positive and had somehow by-passed the primary nonsense mutation. A nested PCR-based examination of dystrophin gene transcripts around the mdx mutation revealed several alternatively processed transcripts, of which four mRNA species skipped the mutation in exon 23, were in-frame and could be translated into a shorter, but still functional dystrophin protein. Specific tests for these transcripts demonstrated these were also present in normal adult and embryonic mouse muscle tissue.
Publisher: IGI Global
Date: 2009
DOI: 10.4018/978-1-59140-995-3.CH037
Abstract: Over the past few decades, the technologies of mobile communication, positioning, and computing have gradually converged. The automobile has been a natural platform for this convergence where satellitebased positioning, wireless communication and on-board computing work in tandem offering various services to motorists. While there are many opportunities with these novel services, significant risks to the location privacy of motorists also exist as a result of the fast-paced technological evolution. These risks must be confronted if trust and confidence are to prevail between motorists and service providers. This chapter provides an overview of the current situation of location privacy in automotive telematics by exploring possible abuses and existing approaches to curb these abuses followed by a discussion of possible privacy-strengthening measures.
Publisher: Springer International Publishing
Date: 2017
Publisher: Elsevier BV
Date: 07-1997
DOI: 10.1016/S0960-8966(97)00062-X
Abstract: McArdle's disease is an autosomal recessive myopathy with symptoms of exercise intolerance caused by deficiency of the enzyme muscle glycogen phosphorylase which releases glucose for contraction during exercise. The human cDNA has been sequenced and disease-causing mutations identified. An ovine equivalent of McArdle's disease has been diagnosed and the mutation responsible identified by PCR- lification of the ovine glycogen myophosphorylase cDNA in six overlapping fragments followed by single strand conformation polymorphism (SSCP) analysis. Two fragments showed SSCPs in the glycogen myophosphorylase cDNA from affected sheep. The SSCP in fragment one was a silent polymorphism, while that in fragment six, was an eight base deletion at the 5' end of exon 20. This deletion will cause a frame-shift, a premature stop codon and remove the last 31 amino-acid residues from the protein. The cDNA deletion suggested that the genomic mutation most likely involved a splice-site. Sequencing intron 19 identified the mutation as an adenine for guanine substitution at the intron 19 3' splice-site. This eliminated an XbaI site present in normal sheep allowing diagnosis of normal, affected and carrier sheep. This ovine model of McArdle's disease is now available for therapeutic trials.
Publisher: Elsevier BV
Date: 2012
DOI: 10.1038/MTNA.2012.40
Publisher: Elsevier BV
Date: 09-2012
Publisher: Elsevier BV
Date: 07-1999
DOI: 10.1016/S0960-8966(99)00010-3
Abstract: The mdx mouse, which carries a nonsense mutation in exon 23 of the dystrophin gene, has been used as an animal model of Duchenne muscular dystrophy to evaluate cell or gene replacement therapies. Despite the mdx mutation, which should preclude the synthesis of a functional dystrophin protein, rare, naturally occurring dystrophin-positive fibres have been observed in mdx muscle tissue. These dystrophin-positive fibres are thought to have arisen from an exon-skipping mechanism, either somatic mutations or alternative splicing. Increasing the frequency of these fibres may offer another therapeutic approach to reduce the severity of Duchenne muscular dystrophy. Antisense oligonucleotides have been shown to block aberrant splicing in the human beta-globin gene. We wished to use a similar approach to re-direct normal processing of the dystrophin pre-mRNA and induce specific exon skipping. Antisense 2'-O-methyl-oligoribonucleotides, directed to the 3' and 5' splice sites of introns 22 and 23, respectively in the mdx pre-mRNA, were used to transfect myoblast cultures. The 5' antisense oligonucleotide appeared to efficiently displace factors normally involved in the removal of intron 23 so that exon 23 was also removed during the splicing of the dystrophin pre-mRNA. Approximately 50% of the dystrophin gene mRNAs were missing this exon 6 h after transfection of primary mdx myotubes, with all transcripts showing skipping of exon 23 after 24 h. Deletion of exon 23 does not disrupt the reading frame and should allow the synthesis of a shorter but presumably functional Becker-like dystrophin. Molecular intervention at dystrophin pre-mRNA splicing has the potential to reduce the severity of a Duchenne mutation to the milder Becker phenotype.
Publisher: Cambridge University Press (CUP)
Date: 2017
DOI: 10.1017/S1752756200001149
Abstract: Differences in banding scales for milk quality penalties, as determined by bulk tank somatic cell count (SCC), prevent the use of a single economic value for SCC in an overall economic-genetic selection index (Veerk et al., 1998) such as, Profitable Lifetime Index or £PLI. But SCC could be used as a predictor of mastitis as genetic correlation estimates between mastitis and SCC are medium to high (review of Mrode and Swanson, 1996). This suggests that, although deriving a direct single economic value (EV) for SCC based on bulk tank SCC is difficult, a single financial value could still be assigned to SCC based on its relationship with mastitis. Here we use a genetic regression method to calculate the EV of SCC (EV SCC ) as a predictor of mastitis. However, the dependency of regression coefficients on mastitis incidence (p) could make such EV SCC variable. The main objective of this study was to evaluate the impact of such relationship on EVSCC and genetic selection in dairy cattle using predicted transmitting abilities of SCC (PTA SCC ).
Publisher: Elsevier BV
Date: 07-2015
DOI: 10.1016/J.MOLMED.2015.04.006
Abstract: Targeted dystrophin exon removal is a promising therapy for Duchenne muscular dystrophy (DMD) however, dystrophin expression in some reports is not supported by the associated data. As in the account of 'The Emperor's New Clothes', the validity of such claims must be questioned, with critical re-evaluation of available data. Is it appropriate to report clinical benefit and induction of dystrophin as dose dependent when the baseline is unclear? The inability to induce meaningful levels of dystrophin does not mean that dystrophin expression as an end point is irrelevant, nor that induced exon skipping as a strategy is flawed, but demands that drug safety and efficacy, and study parameters be addressed, rather than questioning the strategy or the validity of dystrophin as a biomarker.
Publisher: Cold Spring Harbor Laboratory
Date: 05-03-2020
DOI: 10.1101/2020.03.04.977082
Abstract: Modifications of the endometrial transcriptome at day 7 of the estrus cycle are crucial to maintain gestation after transfer of in-vitro produced (IVP) embryos. The aim of this study was to identify genes, and their related biological mechanisms, in the endometria of recipient lactating dairy cows that become pregnant in the subsequent estrus cycle upon transfer of IVP embryos. Endometrial biopsies were taken from lactating Holstein Friesian cows on day 6-8 of the estrus cycle followed by embryo transfer in the following cycle. Animals were classified retrospectively as pregnant (PR, n=8) or non-pregnant (non-PR, n=11) cows, according to pregnancy status at 26-47 days. Extracted mRNAs from endometrial s les were sequenced with an Illumina platform to determine differentially expressed genes (DEG) between the endometrial transcriptome from PR and non-PR cows. There were 111 DEG (FDR .05), which were mainly related to extracellular matrix interaction, histotroph metabolic composition, prostaglandin synthesis, TGF-β signaling as well as inflammation and leukocyte activation. Comparison of these DEG with DEG identified in two public external datasets confirmed the more fertile endometrial molecular profile of PR cows. In conclusion, this study provides insights into the key early endometrial mechanisms for pregnancy establishment, after IVP embryo transfer in dairy cows.
Publisher: Elsevier BV
Date: 10-2007
Publisher: Informa UK Limited
Date: 1996
DOI: 10.3109/10425179609047575
Abstract: A PCR product was generated from embryonic chicken spinal cord cDNA using primers designed to conserved regions of the human and bovine amino and carboxyl-terminal coding sequences of the Cu,Zn superoxide dismutase (SOD1, EC 1.15.1.1) gene. DNA sequencing confirmed this product to be the chicken homologue of the SOD1 gene. This sequence was compared to SOD1 from bovine, human and Xenopus laevis. Important structural features of SOD1 are shown to be conserved in the chicken gene.
Publisher: Public Library of Science (PLoS)
Date: 12-2016
Publisher: MDPI AG
Date: 20-10-2021
DOI: 10.3390/BIOMEDICINES9111499
Abstract: Polyglutamine (polyQ) ataxias are a heterogenous group of neurological disorders all caused by an expanded CAG trinucleotide repeat located in the coding region of each unique causative gene. To date, polyQ ataxias encompass six disorders: spinocerebellar ataxia types 1, 2, 3, 6, 7, and 17 and account for a larger group of disorders simply known as polyglutamine disorders, which also includes Huntington’s disease. These diseases are typically characterised by progressive ataxia, speech and swallowing difficulties, lack of coordination and gait, and are unfortunately fatal in nature, with the exception of SCA6. All the polyQ spinocerebellar ataxias have a hallmark feature of neuronal aggregations and share many common pathogenic mechanisms, such as mitochondrial dysfunction, impaired proteasomal function, and autophagy impairment. Currently, therapeutic options are limited, with no available treatments that slow or halt disease progression. Here, we discuss the common molecular and clinical presentations of polyQ spinocerebellar ataxias. We will also discuss the promising antisense oligonucleotide therapeutics being developed as treatments for these devastating diseases. With recent advancements and therapeutic approvals of various antisense therapies, it is envisioned that some of the studies reviewed may progress into clinical trials and beyond.
Publisher: Elsevier BV
Date: 09-2007
Abstract: Duchenne and Becker muscular dystrophies are allelic disorders arising from mutations in the dystrophin gene. Duchenne muscular dystrophy is characterized by an absence of functional protein, whereas Becker muscular dystrophy, commonly caused by in-frame deletions, shows synthesis of partially functional protein. Anti-sense oligonucleotides can induce specific exon removal during processing of the dystrophin primary transcript, while maintaining or restoring the reading frame, and thereby overcome protein-truncating mutations. The mdx mouse has a non-sense mutation in exon 23 of the dystrophin gene that precludes functional dystrophin production, and this model has been used in the development of treatment strategies for dystrophinopathies. A phosphorodiamidate morpholino oligomer (PMO) has previously been shown to exclude exon 23 from the dystrophin gene transcript and induce dystrophin expression in the mdxmouse, in vivo and in vitro. In this report, a cell-penetrating peptide (CPP)-conjugated oligomer targeted to the mouse dystrophin exon 23 donor splice site was administered to mdxmice by intraperitoneal injection. We demonstrate dystrophin expression and near-normal muscle architecture in all muscles examined, except for cardiac muscle. The CPP greatly enhanced uptake of the PMO, resulting in widespread dystrophin expression.
Publisher: Springer Science and Business Media LLC
Date: 2007
Publisher: Springer Science and Business Media LLC
Date: 06-07-2003
DOI: 10.1038/NM897
Publisher: Springer Science and Business Media LLC
Date: 28-04-2016
Publisher: Informa UK Limited
Date: 28-11-2016
Publisher: Elsevier BV
Date: 2010
DOI: 10.1038/MT.2009.248
Publisher: MDPI AG
Date: 14-05-2021
DOI: 10.3390/BIOMEDICINES9050552
Abstract: The literature surrounding the use of antisense oligonucleotides continues to grow, with new disease and mechanistic applications constantly evolving. Furthermore, the discovery and advancement of novel chemistries continues to improve antisense delivery, stability and effectiveness. For each new application, a rational sequence design is recommended for each oligomer, as is chemistry and delivery optimization. To confirm oligomer delivery and antisense activity, a positive control AO sequence with well characterized target-specific effects is recommended. Here, we describe splice-switching antisense oligomer sequences targeting the ubiquitously expressed human and mouse SMN and Smn genes for use as control AOs for this purpose. We report two AO sequences that induce targeted skipping of SMN1/SMN2 exon 7 and two sequences targeting the Smn gene, that induce skipping of exon 5 and exon 7. These antisense sequences proved effective in inducing alternative splicing in both in vitro and in vivo models and are therefore broadly applicable as controls. Not surprisingly, we discovered a number of differences in efficiency of exon removal between the two species, further highlighting the differences in splice regulation between species.
Publisher: Elsevier BV
Date: 2007
Abstract: Antisense oligonucleotides (AOs) can be used to redirect dystrophin pre-messenger RNA (mRNA) processing, to remove selected exons from the mature dystrophin mRNA, to overcome nonsense mutations, and/or restore the reading frame. Redundancy within the dystrophin protein allows some domains to be removed without seriously compromising function. One of the challenges for splicing blockade is to design AOs that efficiently remove targeted exons across the dystrophin pre-mRNA. AOs are initially designed to anneal to the more obvious motifs implicated in the splicing process, such as acceptor or donor splice sites and in silico predicted exonic splicing enhancers. The AOs are evaluated for their ability to induce targeted exon skipping after transfection into cultured myoblasts. Although no single motif has been implicated in the consistent induction of exon skipping, the length of the AO has emerged as an important parameter in designing compounds that redirect dystrophin pre-mRNA processing. We present data from in vitro studies in murine and human cells showing that appropriately designed AOs of 25-31 nucleotides are generally more effective at inducing exon skipping than shorter counterparts. However, there appears to be an upper limit in optimal length, which may have to be established on a case-by-case basis.
Publisher: Elsevier BV
Date: 06-2010
DOI: 10.1038/MT.2010.45
Publisher: Elsevier BV
Date: 10-2008
Publisher: Informa UK Limited
Date: 04-05-2022
Publisher: JMIR Publications Inc.
Date: 27-05-2022
Abstract: n February 25, 2022, Russian forces took control of the Chernobyl power plant after continuous fighting within the Chernobyl exclusion zone. Continual events occurred in the month of March, which raised the risk of potential contamination of previously uncontaminated areas and the potential for impacts on human and environmental health. The disruption of war has caused interruptions to normal preventive activities, and radiation monitoring sensors have been nonfunctional. Open-source intelligence can be informative when formal reporting and data are unavailable. his paper aimed to demonstrate the value of open-source intelligence in Ukraine to identify signals of potential radiological events of health significance during the Ukrainian conflict. ata were collected from search terminology for radiobiological events and acute radiation syndrome detection between February 1 and March 20, 2022, using 2 open-source intelligence (OSINT) systems, EPIWATCH and Epitweetr. oth EPIWATCH and Epitweetr identified signals of potential radiobiological events throughout Ukraine, particularly on March 4 in Kyiv, Bucha, and Chernobyl. pen-source data can provide valuable intelligence and early warning about potential radiation hazards in conditions of war, where formal reporting and mitigation may be lacking, to enable timely emergency and public health responses.
Publisher: Informa Healthcare
Date: 15-12-2015
Publisher: Elsevier BV
Date: 10-2008
Publisher: Elsevier BV
Date: 04-2012
Publisher: Wiley
Date: 15-04-2015
DOI: 10.1002/MGG3.144
Publisher: Wiley
Date: 12-11-2008
DOI: 10.1002/JGM.1265
Abstract: Duchenne muscular dystrophy (DMD), a severe neuromuscular disorder, is caused by protein-truncating mutations in the dystrophin gene. Absence of functional dystrophin renders muscle fibres more vulnerable to damage and necrosis. We report antisense oligomer (AO) induced exon skipping in the B6Ros.Cg-Dmd(mdx-4Cv)/J (4(CV)) mouse, a muscular dystrophy model arising from a nonsense mutation in dystrophin exon 53. Both exons 52 and 53 must be excised to remove the mutation and maintain the reading frame. A series of 2'-O-methyl modified oligomers on a phosphorothioate backbone (2OMeAOs) were designed and evaluated for the removal of each exon, and the most effective compounds were then combined to induce dual exon skipping in both myoblast cultures and in vivo. Exon skipping efficiency of 2OMeAOs and phosphorodiamidate morpholino oligomers (PMOs) was evaluated both in vitro and in vivo at the RNA and protein levels. Compared to the original mdx mouse studies, induction of exon skipping from the 4(CV) dystrophin mRNA was far more challenging. PMO cocktails could restore synthesis of near-full length dystrophin protein in cultured 4(CV) myogenic cells and in vivo, after a single intramuscular injection. By-passing the protein-truncating mutation in the 4(CV) mouse model of muscular dystrophy could not be achieved with single oligomers targeting both exons and was only achieved after the application of AO cocktails to remove exons 52 and 53. As in previous studies, the stability and efficiency of PMOs proved superior to 2OMeAOs for consistent and sustained protein induction in vivo.
Publisher: Cold Spring Harbor Laboratory
Date: 18-04-2020
DOI: 10.1101/2020.04.17.047027
Abstract: Feed efficiency (FE) is a key trait in pig production, as it has both high economic and environmental impact. FE is a challenging phenotype to study, as it is complex and affected by many factors, such as metabolism, growth and activity level. Furthermore, testing for FE is expensive, as it requires costly equipment to measure feed intake of in idual animals, making FE biomarkers valuable. Therefore, there has been a desire to find single nucleotide polymorphisms (SNPs) as biomarkers, to assist with improved selection and improve our biological understanding of FE. We have done a cis- and trans-eQTL (expressed quantitative trait loci) analysis, in a population of Danbred Durocs (N=11) and Danbred Landrace (N=27) using both a linear and Anova model. We used bootstrapping and enrichment analysis to validate and analyze our detected eQTLs. We identified 15 eQTLs with FDR 0.01, affecting several genes found in previous studies of commercial pig breeds. Ex les include IFI6, PRPF39, TMEM222, CSRNP1,PARK7 and MFF. The bootstrapping results showed statistically significant enrichment of eQTLs with p-value 0.01 (p-value 2.2×0 -16 ) in both cis and trans-eQTLs. Based on this, enrichment analysis of top trans-eQTLs revealed high enrichment for gene categories and gene ontologies associated with genomic context and expression regulation. This includes transcription factors (p-value=1.0×10 -13 ), DNA-binding (GO:0003677, p-value=8.9×10 -14 ), DNA-binding transcription factor activity (GO:0003700,) nucleus gene (GO:0005634, p-value .2×10 -16 ), positive regulation of expression (GO:0010628), negative regulation of expression (GO:0010629, p-value .2×10 -16 ). These results would be useful for future genome assisted breeding of pigs to improve FE, and in the improved understanding of the functional mechanism of trans-eQTLs.
Publisher: International Academy Publishing (IAP)
Date: 2015
Publisher: Elsevier BV
Date: 11-1996
DOI: 10.1016/0168-9525(96)99991-6
Abstract: We report the synthesis and practical application of a novel scavenger for precious metals. The scavenger was prepared from cellulose filter paper with grafted chains of poly(glycidyl methacrylate) modified with a novel ligand group of
Publisher: Frontiers Media SA
Date: 30-04-2019
Publisher: Elsevier BV
Date: 12-2022
DOI: 10.1016/J.EXER.2022.109276
Abstract: The ATP-binding cassette subfamily A member 4 gene (ABCA4)-associated retinopathy, Stargardt disease, is the most common monogenic inherited retinal disease. Given the pathogenicity of numerous ABCA4 variants is yet to be examined and a significant proportion (more than 15%) of ABCA4 variants are categorized as splice variants in silico, we therefore established a fibroblast-based splice assay to analyze ABCA4 variants in an Australian Stargardt disease cohort and characterize the pathogenic mechanisms of ABCA4 variants. A cohort of 67 patients clinically diagnosed with Stargardt disease was recruited. Genomic DNA was analysed using a commercial panel for ABCA4 variant detection and the consequences of ABCA4 variants were predicted in silico. Dermal fibroblasts were propagated from skin biopsies, total RNA was extracted and the ABCA4 transcript was lified by RT-PCR. Our analysis identified a total of 67 unique alleles carrying 74 unique variants. The most prevalent splice-affecting complex allele c.[5461-10T>C 5603A>T] was carried by 10% of patients in a compound heterozygous state. ABCA4 transcripts from exon 13 to exon 50 were readily detected in fibroblasts. In this region, aberrant splicing was evident in 10 out of 57 variant transcripts (18%), carried by 19 patients (28%). Patient-derived fibroblasts provide a feasible platform for identification of ABCA4 splice variants located within exons 13-50. Experimental evidence of aberrant splicing contributes to the pathogenic classification for ABCA4 variants. Moreover, identification of variants that affect splicing processes provides opportunities for intervention, in particular antisense oligonucleotide-mediated splice correction.
Publisher: Elsevier BV
Date: 11-2013
DOI: 10.1016/J.JNEUROIM.2013.08.008
Abstract: The mechanism of necrotizing myopathy associated with antibodies to signal recognition particle (SRP) remains unclear. We investigated the effect of anti-SRP+serum and complement on cell viability in myoblast cultures. Cell viability was only slightly reduced by incubation with anti-SRP+serum compared with control serum. However, the addition of fresh complement resulted in a marked reduction in cell survival. Surface immunostaining for SRP, C3c and C5b-9 was demonstrated in cultures pre-incubated with anti-SRP+serum and complement, and in muscle biopsies from patients with myopathy. These findings provide further support for a complement-dependent antibody-mediated mechanism in anti-SRP associated myopathy.
Publisher: Institute of Electrical and Electronics Engineers (IEEE)
Date: 2010
Publisher: Frontiers Media SA
Date: 09-06-2022
Publisher: Wiley
Date: 04-12-2007
DOI: 10.1002/ANA.21290
Abstract: The degenerative muscle diseases Duchenne (DMD) and Becker muscular dystrophy result from mutations in the DMD gene, which encodes the dystrophin protein. Recent improvements in mutational analysis techniques have resulted in the increasing identification of deep intronic point mutations, which alter splicing such that intronic sequences are included in the messenger RNA as "pseudoexons." We sought to test the hypothesis that the clinical phenotype correlates with splicing efficiency of these mutations, and to test the feasibility of antisense oligonucleotide (AON)-mediated pseudoexon skipping. We identified three pseudoexon insertion mutations in dystrophinopathy patients, two of whom had tissue available for further analysis. For these two out-of-frame pseudoexon mutations (one associated with Becker muscular dystrophy and one with DMD), mutation-induced splicing was tested by quantitative reverse transcription polymerase chain reaction pseudoexon skipping was tested using AONs composed of 2'-O-methyl-modified bases on a phosphorothioate backbone to treat cultured primary myoblasts. Variable amounts of pseudoexon inclusion correlates with the severity of the dystrophinopathy phenotype in these two patients. AON treatment directed at the pseudoexon results in the expression of full-length dystrophin in a DMD myoblast line. Both DMD and Becker muscular dystrophy can result from out-of-frame pseudoexons, with the difference in phenotype being due to variable efficiency of the newly generated splicing signal. AON-mediated pseudoexon skipping therapy is a viable approach to these patients and would be predicted to result in increased expression of wild-type dystrophin protein.
Publisher: Wiley
Date: 28-11-2011
Publisher: Wiley
Date: 27-08-2003
DOI: 10.1016/S0014-5793(03)00904-9
Abstract: The use of antisense oligonucleotides (AOs) to induce exon skipping leading to generation of an in-frame dystrophin protein product could be of benefit in around 70% of Duchenne muscular dystrophy patients. We describe the use of hyaluronidase enhanced electrotransfer to deliver uncomplexed 2'-O-methyl modified phosphorothioate AO to adult dystrophic mouse muscle, resulting in dystrophin expression in 20-30% of fibres in tibialis anterior muscle after a single injection. Although expression was transient, many of the corrected fibres initially showed levels of dystrophin expression well above the 20% of endogenous previously shown to be necessary for phenotypic correction of the dystrophic phenotype.
Publisher: Proceedings of the National Academy of Sciences
Date: 19-12-2001
DOI: 10.1073/PNAS.98.1.42
Abstract: Duchenne muscular dystrophy (DMD) is a severe muscle wasting disease arising from defects in the dystrophin gene, typically nonsense or frameshift mutations, that preclude the synthesis of a functional protein. A milder, allelic version of the disease, Becker muscular dystrophy, generally arises from in-frame deletions that allow synthesis of a shorter but still semifunctional protein. Therapies to introduce functional dystrophin into dystrophic tissue through either cell or gene replacement have not been successful to date. We report an alternative approach where 2′- O -methyl antisense oligoribonucleotides have been used to modify processing of the dystrophin pre-mRNA in the mdx mouse model of DMD. By targeting 2′- O -methyl antisense oligoribonucleotides to block motifs involved in normal dystrophin pre-mRNA splicing, we induced excision of exon 23, and the mdx nonsense mutation, without disrupting the reading frame. Exon 23 skipping was first optimized in vitro in transfected H-2K b -tsA58 mdx myoblasts and then induced in vivo . Immunohistochemical staining demonstrated the synthesis and correct subsarcolemmal localization of dystrophin and γ-sarcoglycan in the mdx mouse after intramuscular delivery of antisense oligoribonucleotide:liposome complexes. This approach should reduce the severity of DMD by allowing a dystrophic gene transcript to be modified, such that it can be translated into a Becker-dystrophin-like protein.
Publisher: Elsevier BV
Date: 09-1997
Publisher: Elsevier BV
Date: 10-1996
DOI: 10.1016/0960-8966(96)00353-7
Abstract: Autosomal dominant inheritance is exhibited by about 10% of cases of amyotrophic lateral sclerosis (ALS), a paralytic disorder characterized by the death of motor neurons in the brain and spinal cord. A subgroup of these familial cases are linked to mutations in the gene which codes for Cu/Zn superoxide dismutase (SOD1). We report three additional mutations occurring in the SOD1 gene in ALS patients and two single base pair variant changes. The single base pair change in an ALS family causes a glycine 93 to valine substitution, which is the fifth distinct amino acid change reported for the glycine 93 residue. One missense mutation in exon 5 would substitute neutral valine for the negatively-charged aspartate 124 (aspartate 124 to valine). An in idual with an apparently sporadic case of ALS carries a three base pair deletion in exon 5 of the SOD1 gene. These three mutations bring to 38 the total number of distinct SOD1 mutations associated with familial ALS.
Publisher: Hindawi Limited
Date: 1998
DOI: 10.1002/(SICI)1098-1004(1998)11:3<252::AID-HUMU11>3.0.CO;2-Y
Abstract: Early prognostic stratification of patients with sepsis is a difficult clinical challenge. Aim of this study was to evaluate novel molecules in association with clinical parameters as predictors of 90-days mortality in patients admitted with sepsis at Humanitas Research Hospital. Plasma s les were collected from 178 patients, diagnosed based on Sepsis-3 criteria, at admission to the Emergency Department and after 5 days of hospitalization. Levels of pentraxin 3 (PTX3), soluble IL-1 type 2 receptor (sIL-1R2), and of a panel of pro- and anti-inflammatory cytokines were measured by ELISA. Cox proportional-hazard models were used to evaluate predictors of 90-days mortality. Circulating levels of PTX3, sIL-1R2, IL-1β, IL-6, IL-8, IL-10, IL-18, IL-1ra, TNF-α increased significantly in sepsis patients on admission, with the highest levels measured in shock patients, and correlated with SOFA score (PTX3: r=0.44, p<0.0001 sIL-1R2: r=0.35, p<0.0001), as well as with 90-days mortality. After 5 days of hospitalization, PTX3 and cytokines, but not sIL-1R2 levels, decreased significantly, in parallel with a general improvement of clinical parameters. The combination of age, blood urea nitrogen, PTX3, IL-6 and IL-18, defined a prognostic index predicting 90-days mortality in Sepsis-3 patients and showing better apparent discrimination capacity than the SOFA score (AUC=0.863, 95% CI: 0.780-0.945 These data suggest that a prognostic index based on selected cytokines, PTX3 and clinical parameters, and hence easily adoptable in clinical practice, performs in predicting 90-days mortality better than SOFA. An independent validation is required.
Publisher: Wiley
Date: 12-04-2019
Publisher: MDPI AG
Date: 11-10-2019
DOI: 10.3390/IJMS20205030
Abstract: The process of pre-mRNA splicing is a common and fundamental step in the expression of most human genes. Alternative splicing, whereby different splice motifs and sites are recognised in a developmental and/or tissue-specific manner, contributes to genetic plasticity and ersity of gene expression. Redirecting pre-mRNA processing of various genes has now been validated as a viable clinical therapeutic strategy, providing treatments for Duchenne muscular dystrophy (inducing specific exon skipping) and spinal muscular atrophy (promoting exon retention). We have designed and evaluated over 5000 different antisense oligonucleotides to alter splicing of a variety of pre-mRNAs, from the longest known human pre-mRNA to shorter, exon-dense primary gene transcripts. Here, we present our guidelines for designing, evaluating and optimising splice switching antisense oligomers in vitro. These systematic approaches assess several critical factors such as the selection of target splicing motifs, choice of cells, various delivery reagents and crucial aspects of validating assays for the screening of antisense oligonucleotides composed of 2′-O-methyl modified bases on a phosphorothioate backbone.
Publisher: Elsevier BV
Date: 09-2012
DOI: 10.1016/J.NEUROSCIENCE.2012.06.042
Abstract: Spinal muscular atrophy (SMA), a fatal genetic motor disorder of infants, is caused by diminished full-length survival of motor neuron (SMN) protein levels. Normally involved in small nuclear ribonucleoprotein (snRNP) assembly and pre-mRNA splicing, recent studies suggest that SMN plays a critical role in regulating apoptosis. Interestingly, the anti-apoptotic Bcl-x isoform, Bcl-xL, is reduced in SMA. In a related finding, Sam68, an RNA-binding protein, was found to modulate splicing of SMN and Bcl-xL transcripts, promoting SMNΔ7 and pro-apoptotic Bcl-xS transcripts. Here we demonstrate that Bcl-xL expression increases SMN protein by ∼2-fold in SH-SY5Y cells. Conversely, SMN expression increases Bcl-xL protein levels by ∼6-fold in SH-SY5Y cells, and ∼2.5-fold in the brains of transgenic mice over-expressing SMN (PrP-SMN). Moreover, Sam68 protein levels were markedly reduced following SMN and Bcl-xL expression in SH-SY5Y cells, suggesting a feedback mechanism co-regulating levels of both proteins. We also found that exogenous SMN expression increased full-length SMN transcripts, possibly by promoting exon 7 inclusion. Finally, co-expression of SMN and Bcl-xL produced an additive anti-apoptotic effect following PI3-kinase inhibition in SH-SY5Y cells. Our findings implicate Bcl-xL as another potential target in SMA therapeutics, and indicate that therapeutic increases in SMN may arise from modest increases in total SMN.
Publisher: Wiley
Date: 08-1998
DOI: 10.1002/(SICI)1097-4598(199808)21:8<991::AID-MUS2>3.0.CO;2-0
Abstract: Golden retriever muscular dystrophy (GRMD), the canine model of Duchenne muscular dystrophy (DMD), is caused by a splice site mutation in the dystrophin gene. This mutation predicts a premature termination codon in exon 8 and a peptide that is 5% the size of normal dystrophin. Western blot analysis of skeletal muscle from GRMD dogs reveals a slightly truncated 390-kD protein that is approximately 91% the size of normal dystrophin. This 390-kD dystrophin suggests that GRMD dogs, like some DMD patients, employ a mechanism to overcome their predicted frameshift. Reverse-transcriptase polymerase chain reaction on GRMD muscle has revealed two in-frame dystrophin transcripts which lack either exons 3-9 or exons 5-12. Both transcripts could be translated into a dystrophin protein of approximately 390 kD. An understanding of how truncated dystrophin is produced in GRMD may allow this mechanism to be manipulated toward a potential therapy for DMD.
Publisher: Public Library of Science (PLoS)
Date: 24-02-2017
Publisher: Elsevier BV
Date: 03-1998
Publisher: MDPI AG
Date: 02-04-2018
DOI: 10.3390/IJGI7040139
Publisher: Public Library of Science (PLoS)
Date: 22-04-2013
Publisher: Springer Science and Business Media LLC
Date: 10-09-2019
DOI: 10.1038/S41598-019-49385-6
Abstract: With recent approvals of antisense oligonucleotides as therapeutics, there is an increasing interest in expanding the application of these compounds to many other diseases. Our laboratory focuses on developing therapeutic splice modulating antisense oligonucleotides to treat diseases potentially amendable to intervention during pre-mRNA processing, and here we report the use of oligomers to down-regulate integrin alpha 4 protein levels. Over one hundred antisense oligonucleotides were designed to induce skipping of in idual exons of the ITGA4 transcript and thereby reducing protein expression. Integrin alpha 4-mediated activities were evaluated in human dermal fibroblasts and Jurkat cells, an immortalised human T lymphocyte cell line. Peptide conjugated phosphorodiamidate morpholino antisense oligomers targeting ITGA4 were also assessed for their effect in delaying disease progression in the experimental autoimmune encephalomyelitis mouse model of multiple sclerosis. With the promising results in ameliorating disease progression, we are optimistic that the candidate oligomer may also be applicable to many other diseases associated with integrin alpha 4 mediated inflammation. This highly specific strategy to down-regulate protein expression through interfering with normal exon selection during pre-mRNA processing should be applicable to many other gene targets that undergo splicing during expression.
Publisher: Frontiers Media SA
Date: 06-12-2019
Publisher: Elsevier BV
Date: 07-1999
DOI: 10.1016/S0960-8966(99)00011-5
Abstract: We have determined the molecular basis for skeletal myopathy and dilated cardiomyopathy in two male German short-haired pointer (GSHP) littermates. Analysis of skeletal muscle demonstrated a complete absence of dystrophin on Western blot analysis. PCR analysis of genomic DNA revealed a deletion encompassing the entire dystrophin gene. Molecular cytogenetic analysis of lymphocytes from the dam and both dystrophic pups confirmed a visible deletion in the p21 region of the affected canine X chromosome. Utrophin is up-regulated in the skeletal muscle, but does not appear to ameliorate the dystrophic canine phenotype. This new canine model should further our understanding of the physiological and biochemical processes in Duchenne muscular dystrophy.
Publisher: Oxford University Press (OUP)
Date: 05-2015
Abstract: The study investigated genetic architecture and predictive ability using genomic annotation of residual feed intake (RFI) and its component traits (daily feed intake [DFI], ADG, and back fat [BF]). A total of 1,272 Duroc pigs had both genotypic and phenotypic records, and the records were split into a training (968 pigs) and a validation dataset (304 pigs) by assigning records as before and after January 1, 2012, respectively. SNP were annotated by 14 different classes using Ensembl variant effect prediction. Predictive accuracy and prediction bias were calculated using Bayesian Power LASSO, Bayesian A, B, and Cπ, and genomic BLUP (GBLUP) methods. Predictive accuracy ranged from 0.508 to 0.531, 0.506 to 0.532, 0.276 to 0.357, and 0.308 to 0.362 for DFI, RFI, ADG, and BF, respectively. BayesCπ100.1 increased accuracy slightly compared to the GBLUP model and other methods. The contribution per SNP to total genomic variance was similar among annotated classes across different traits. Predictive performance of SNP classes did not significantly differ from randomized SNP groups. Genomic prediction has accuracy comparable to observed phenotype, and use of genomic prediction can be cost effective by replacing feed intake measurement. Genomic annotation had less impact on predictive accuracy traits considered here but may be different for other traits. It is the first study to provide useful insights into biological classes of SNP driving the whole genomic prediction for complex traits in pigs.
Publisher: Cold Spring Harbor Laboratory
Date: 18-10-2018
DOI: 10.1101/446773
Abstract: Oligonucleotides and nucleic acid analogues that alter gene expression are showing therapeutic promise for selected human diseases. The modification of synthetic nucleic acids to protect against nuclease degradation and to influence drug function is common practice, however, such modifications may also confer unexpected physicochemical and biological properties. Here we report backbone-specific effects of modified oligonucleotides on subnuclear organelles, altered distribution of nuclear proteins, the appearance of novel structured nuclear inclusions, and modification of RNA processing in cultured cells transfected with antisense oligonucleotides on a phosphorothioate backbone. Phosphodiester and phosphorodiamidate morpholino oligomers elicited no such consequences. Disruption of subnuclear structures and proteins elicit severe phenotypic disturbances, revealed by transcriptomic analysis of fibroblasts exhibiting such disruption. These data suggest that the toxic effects and adverse events reported after clinical evaluation of phosphorothioate nucleic acid drugs may be mediated, at least in part, by non-specific interaction of nuclear components with the phosphorothioate backbone.
Publisher: Public Library of Science (PLoS)
Date: 08-01-2012
Publisher: Elsevier BV
Date: 05-2011
Publisher: Wiley
Date: 04-2021
DOI: 10.1111/RESP.14021
Publisher: Mary Ann Liebert Inc
Date: 09-2007
DOI: 10.1089/HUM.2006.061
Abstract: Duchenne muscular dystrophy (DMD) is caused by mutations in the dystrophin gene that result in the absence of functional protein. In the majority of cases these are out-of-frame deletions that disrupt the reading frame. Several attempts have been made to restore the dystrophin mRNA reading frame by modulation of pre-mRNA splicing with antisense oligonucleotides (AOs), demonstrating success in cultured cells, muscle explants, and animal models. We are preparing for a phase I/IIa clinical trial aimed at assessing the safety and effect of locally administered AOs designed to inhibit inclusion of exon 51 into the mature mRNA by the splicing machinery, a process known as exon skipping. Here, we describe a series of systematic experiments to validate the sequence and chemistry of the exon 51 AO reagent selected to go forward into the clinical trial planned in the United Kingdom. Eight specific AO sequences targeting exon 51 were tested in two different chemical forms and in three different preclinical models: cultured human muscle cells and explants (wild type and DMD), and local in vivo administration in transgenic mice harboring the entire human DMD locus. Data have been validated independently in the different model systems used, and the studies describe a rational collaborative path for the preclinical selection of AOs for evaluation in future clinical trials.
Publisher: Elsevier BV
Date: 10-2006
DOI: 10.1016/J.NMD.2006.05.017
Abstract: Antisense oligonucleotide (AO) manipulation of pre-mRNA splicing of the dystrophin gene is showing promise in overcoming Duchenne muscular dystrophy (DMD)-causing mutations. To date, this approach has been limited to studies using animal models or cultured human muscle cells, and evidence that AOs can induce exon skipping in human muscle has yet to be shown. In this study, we used different AO analogues to induce exon skipping in muscle explants derived from normal and DMD human tissue. We propose that inducing exon skipping in human muscle explants is closer to in vivo conditions than cells in monolayer cultures, and may minimize the numbers of participants in Phase I clinical studies to demonstrate proof of principle of exon skipping in human muscle.
Publisher: Springer Science and Business Media LLC
Date: 06-02-2018
Publisher: Elsevier BV
Date: 05-2008
Publisher: Springer Science and Business Media LLC
Date: 20-05-2021
DOI: 10.1186/S40035-021-00240-7
Abstract: Precursor messenger RNA (pre-mRNA) splicing is a fundamental step in eukaryotic gene expression that systematically removes non-coding regions (introns) and ligates coding regions (exons) into a continuous message (mature mRNA). This process is highly regulated and can be highly flexible through a process known as alternative splicing, which allows for several transcripts to arise from a single gene, thereby greatly increasing genetic plasticity and the ersity of proteome. Alternative splicing is particularly prevalent in neuronal cells, where the splicing patterns are continuously changing to maintain cellular homeostasis and promote neurogenesis, migration and synaptic function. The continuous changes in splicing patterns and a high demand on many cis- and trans- splicing factors contribute to the susceptibility of neuronal tissues to splicing defects. The resultant neurodegenerative diseases are a large group of disorders defined by a gradual loss of neurons and a progressive impairment in neuronal function. Several of the most common neurodegenerative diseases involve some form of splicing defect(s), such as Alzheimer’s disease, Parkinson’s disease and spinal muscular atrophy. Our growing understanding of RNA splicing has led to the explosion of research in the field of splice-switching antisense oligonucleotide therapeutics. Here we review our current understanding of the effects alternative splicing has on neuronal differentiation, neuronal migration, synaptic maturation and regulation, as well as the impact on neurodegenerative diseases. We will also review the current landscape of splice-switching antisense oligonucleotides as a therapeutic strategy for a number of common neurodegenerative disorders.
Publisher: Elsevier BV
Date: 06-2005
Publisher: Bentham Science Publishers Ltd.
Date: 08-2011
DOI: 10.2174/156652311796150381
Abstract: Antisense oligomers initially showed promise as compounds to modify gene expression, primarily through RNaseH induced degradation of the target transcript. Expansion of the field has led to new chemistries capable of invoking different mechanisms, including suppression of protein synthesis by translational blockade and gene silencing using short interfering RNAs. It is now apparent that the majority of the eukaryotic genome is transcribed and non-protein coding RNAs have been implicated in the regulation of gene expression at many levels. This review considers potential therapeutic applications of antisense oligomers to modify gene expression, primarily by interfering with the process of exon recognition and intron removal during gene transcript splicing. While suppression of gene expression will be necessary to address some conditions, it is likely that antisense oligomer splice modification will have extensive clinical application. Pre-mRNA splicing is a tightly co-ordinated, multifactorial process that can be disrupted by antisense oligomers in a highly specific manner to suppress aberrant splicing, remove exons to by-pass nonsense or frame-shifting mutations or influence exon selection to alter spliceoform ratios. Manipulation of splicing patterns has been applied to a erse range of conditions, including b-thalassemia, Duchenne muscular dystrophy, spinal muscular atrophy and certain cancers. Alternative exon usage has been identified as a major mechanism for generating ersity from a limited repertoire of genes in higher eukaryotes. Considering that the majority of all human primary gene transcripts are reportedly alternatively spliced, intervention at the level of pre-mRNA processing is likely to become increasingly significant in the fight against genetic and acquired disorders.
Publisher: American Society of Civil Engineers (ASCE)
Date: 09-2021
Publisher: Wiley
Date: 11-1995
Publisher: Elsevier BV
Date: 08-2010
Publisher: Frontiers Media SA
Date: 14-09-2017
Publisher: Public Library of Science (PLoS)
Date: 28-08-2013
Publisher: Elsevier BV
Date: 11-2018
DOI: 10.1016/J.TIPS.2018.09.001
Abstract: Clinical implementation of two recently approved antisense RNA therapeutics - Exondys51
Publisher: American Veterinary Medical Association (AVMA)
Date: 12-2001
DOI: 10.2460/AJVR.2001.62.1964
Abstract: Objectives —To determine the distribution of a 231- base pair (bp) element in the dystrophin gene 3' untranslated region (UTR) in a colony of Golden Retrievers with muscular dystrophy and other unrelated dogs and to estimate the frequency of recombination for the canine dystrophin gene. Animals —77 dogs from the Golden Retriever Muscular Dystrophy (GRMD) colony at the Murdoch Veterinary School and 30 unrelated dogs from the Murdoch University Veterinary Clinic. Procedure —S les of blood or hair from dogs were used for lification of DNA, using primers to the canine dystrophin 3' UTR. Results —The DNA from affected dogs generated a larger PCR product than that obtained from clinically normal dogs. Products were cloned and sequenced, and the difference in size was found to be attributable to a 231-bp short interspersed nucleotide element (SINE). The SINE was found in all affected dogs in the colony but not in most unaffected puppies in the colony. Eighteen of 19 dogs in the colony were heterozygous for the GRMD mutation, and 7 of 30 unrelated dogs also were heterozygous for the SINE. Conclusion and Clinical Relevance —Evidence of recombination between the GRMD mutation and the SINE was observed in only 4 dogs (2 sets of littermates) in the GRMD colony. Incidence of this SINE in a few unrelated dogs suggests that this particular insertion into the dystrophin gene may have been a recent event. The SINE in the dystrophin 3' UTR did not have an apparent influence on dystrophin mRNA concentrations. ( Am J Vet Res 2001 :1964–1968)
Publisher: Springer Science and Business Media LLC
Date: 29-01-2006
DOI: 10.1038/NM1345
Abstract: For the majority of Duchenne muscular dystrophy (DMD) mutations, antisense oligonucleotide (AON)-mediated exon skipping has the potential to restore a functional protein. Here we show that weekly intravenous injections of morpholino phosphorodiamidate (morpholino) AONs induce expression of functional levels of dystrophin in body-wide skeletal muscles of the dystrophic mdx mouse, with resulting improvement in muscle function. Although the level of dystrophin expression achieved varies considerably between muscles, antisense therapy may provide a realistic hope for the treatment of a majority of in iduals with DMD.
Publisher: American Society of Civil Engineers (ASCE)
Date: 02-2017
Publisher: Elsevier BV
Date: 10-2009
Publisher: Frontiers Media SA
Date: 28-02-2020
Publisher: Springer Science and Business Media LLC
Date: 21-04-2020
DOI: 10.1038/S41598-020-63461-2
Abstract: Pompe disease is caused by mutations in the GAA gene, resulting in deficient lysosomal acid-α-glucosidase activity in patients, and a progressive decline in mobility and respiratory function. Enzyme replacement therapy is one therapeutic option, but since not all patients respond to this treatment, alternative interventions should be considered. One GAA mutation, c.-32-13T G, impacts upon normal exon 2 splicing and is found in two-thirds of late-onset cases. We and others have explored a therapeutic strategy using splice modulating phosphorodiamidate morpholino oligomers to enhance GAA exon 2 inclusion in the mature mRNA of patients with one c.-32-13T G allele. We designed 20 oligomers and treated fibroblasts derived from five patients to identify an oligomer sequence that maximally increased enzyme activity in all fibroblasts. The most effective splice correcting oligomer was chosen to treat forced-myogenic cells, derived from fibroblasts from nine patients carrying the c.-32-13T G mutation. After transfection, we show increased levels of the full-length GAA transcript, acid-α-glucosidase protein, and enzyme activity in all patients’ myogenic cells, regardless of the nature of the mutation in the other allele. This data encourages the initiation of clinical trials to assess the therapeutic efficacy of this oligomer for those patients carrying the c.-32-13T G mutation.
Publisher: Springer International Publishing
Date: 2016
Publisher: Springer International Publishing
Date: 2016
Publisher: Elsevier BV
Date: 12-2020
Publisher: Informa UK Limited
Date: 23-11-2016
Publisher: Elsevier BV
Date: 07-2001
Publisher: Cambridge University Press (CUP)
Date: 2017
DOI: 10.1017/S175275620000017X
Abstract: Genetic parameters of mastitis are required in genetic selection for mastitis resistance. Excluding cows that are culled at an early stage of lactation due to mastitis from genetic parameter estimation may introduce culling bias. The use of linear models, suitable for continuous traits, is inappropriate for analysis of mastitis records because it is recorded as an all or none (binary) trait. Here, a Bayesian-threshold model with Markov chain Monte Carlo (MCMC) techniques was used to analyze mastitis data. The objective was to estimate genetic parameters of mastitis in first and all-lactation cows using complete or incomplete records on disease.
Publisher: Informa UK Limited
Date: 10-06-2013
Publisher: Elsevier BV
Date: 08-2011
Publisher: Wiley
Date: 02-2020
DOI: 10.14814/PHY2.14359
Publisher: Mary Ann Liebert Inc
Date: 15-11-2015
Abstract: Facioscapulohumeral muscular dystrophy (FSHD) is associated with an activation of the double homeobox 4 (DUX4) gene, which we previously identified within the D4Z4 repeated elements in the 4q35 subtelomeric region. The pathological DUX4 mRNA is derived from the most distal D4Z4 unit and extends unexpectedly within the flanking pLAM region, which provides an intron and polyadenylation signal. The conditions that are required to develop FSHD are a permissive allele providing the polyadenylation signal and hypomethylation of the D4Z4 repeat array compared with the healthy muscle. The DUX4 protein is a 52-kDa transcription factor that initiates a large gene deregulation cascade leading to muscle atrophy, inflammation, differentiation defects, and oxidative stress, which are the key features of FSHD. DUX4 is a retrogene that is normally expressed in germline cells and is submitted to repeat-induced silencing in adult tissues. Since DUX4 mRNAs have been detected in human embryonic and induced pluripotent stem cells, we investigated whether they could also be expressed in human mesenchymal stromal cells (hMSCs). We found that DUX4 mRNAs were induced during the differentiation of hMSCs into osteoblasts and that this process involved DUX4 and new longer protein forms (58 and 70 kDa). A DUX4 mRNA with a more distant 5' start site was characterized that presented a 60-codon reading frame extension and encoded the 58-kDa protein. Transfections of hMSCs with an antisense oligonucleotide targeting DUX4 mRNAs decreased both the 52- and 58-kDa protein levels and confirmed their identity. Gain- and loss-of-function experiments in hMSCs suggested these DUX4 proteins had opposite roles in osteogenic differentiation as evidenced by the alkaline phosphatase activity and calcium deposition. Differentiation was delayed by the 58-kDa DUX4 expression and it was increased by 52-kDa DUX4. These data indicate a role for DUX4 protein forms in the osteogenic differentiation of hMSCs.
Publisher: Springer Science and Business Media LLC
Date: 05-2019
Publisher: Elsevier BV
Date: 12-2010
Publisher: Colegio Brasileiro de Reproducao Animal - CBRA
Date: 2017
Publisher: Elsevier BV
Date: 11-2022
Publisher: Wiley
Date: 15-01-2019
Abstract: The Hippo pathway has emerged as a major eukaryotic signalling pathway and is increasingly the subject of intense interest, as are the key effectors of canonical Hippo signalling, YES-associated protein (YAP) and TAZ. The Hippo pathway has key roles in erse biological processes, including network signalling regulation, development, organ growth, tissue repair and regeneration, cancer, stem cell regulation and mechanotransduction. YAP and TAZ are multidomain proteins and function as transcriptional coactivators of key genes to evoke their biological effects. YAP and TAZ interact with numerous partners and their activities are controlled by a complex set of processes. This review provides an overview of Hippo signalling and its role in growth. In particular, the functional domains of YAP and TAZ and the complex mechanisms that regulate their protein stability and activity are discussed. Notably, the similarities and key differences are highlighted between the two paralogues including which partner proteins interact with which functional domains to regulate their activity.
Publisher: American Physiological Society
Date: 07-2020
DOI: 10.1152/PHYSIOLGENOMICS.00027.2020
Abstract: Modifications of the endometrial transcriptome at day 7 of the estrus cycle are crucial to maintain gestation after transfer of in vitro-produced (IVP) embryos, although these changes are still largely unknown. The aim of this study was to identify genes, and their related biological mechanisms, important for pregnancy establishment based on the endometrial transcriptome of recipient lactating dairy cows that become pregnant in the subsequent estrus cycle, upon transfer of IVP embryos. Endometrial biopsies were taken from Holstein Friesian cows on day 6–8 of the estrus cycle followed by embryo transfer in the following cycle. Animals were classified retrospectively as pregnant (PR, n = 8) or nonpregnant (non-PR, n = 11) cows, according to pregnancy status at 26–47 days. Extracted mRNAs from endometrial s les were sequenced with an Illumina platform to determine differentially expressed genes (DEG) between the endometrial transcriptome from PR and non-PR cows. There were 111 DEG (false discovery rate 0.05), which were mainly related to extracellular matrix interaction, histotroph metabolic composition, prostaglandin synthesis, transforming growth factor-β signaling as well as inflammation and leukocyte activation. Comparison of these DEG with DEG identified in two public external data sets confirmed the more fertile endometrial molecular profile of PR cows. In conclusion, this study provides insights into the key early endometrial mechanisms for pregnancy establishment, after IVP embryo transfer in dairy cows.
Publisher: Springer Basel
Date: 2013
Publisher: Frontiers Media SA
Date: 28-07-2015
Publisher: Wiley
Date: 2006
Publisher: S. Karger AG
Date: 1996
DOI: 10.1159/000134206
Abstract: Sequence-tagged sites (STSs) were developed for the human α-tropomyosin gene TPM4. One STS was used to lify DNA from somatic cell hybrids to localize TPM4 to chromosome 19. The other, a product from a long-range PCR, was used directly as a probe to refine the localization of TPM4 to 19p13.1 by fluorescence in situ hybridization to metaphase chromosome spreads.
Publisher: Elsevier BV
Date: 06-2014
Publisher: Springer US
Date: 19-11-2022
DOI: 10.1007/978-1-0716-2772-3_14
Abstract: The mutation c.-32-13T>G in the GAA gene impacts normal exon 2 splicing and is found in two-thirds of late-onset Pompe disease cases. We have explored a therapeutic strategy using splice modulating phosphorodiamidate morpholino oligomers to enhance GAA exon 2 inclusion in the mature mRNA of patients carrying this common mutation. We performed in silico analysis of the GAA gene transcript for potential splicing silencers and designed oligomers targeting motifs predicted to enhance exon 2 retention in the mature mRNA. Two patient-derived fibroblasts were obtained from Coriell Institute for Medical Research, and seven fibroblast strains from unrelated patients were supplied by Westmead Hospital in Sydney, Australia. Both fibroblasts and forced-myogenic cells were treated with optimized phosphorodiamidate morpholino oligomers supplied by Sarepta Therapeutics. Total RNA and protein were extracted from the cells after incubation with phosphorodiamidate morpholino oligomers, and RT-PCR and RT-qPCR were performed to confirm exon 2 inclusion is enhanced. Acid α-glucosidase activity and expression levels were also assessed to confirm therapeutic potential.
Publisher: Elsevier BV
Date: 04-1991
DOI: 10.1016/0378-1135(91)90010-D
Abstract: DNA lification using the polymerase chain reaction technique was evaluated for rapid identification of Mycobacterium bovis. Two oligonucleotide primers of 20 bases in length were constructed to target a region of the gene encoding the M. bovis secretory protein, MPB70. The lification reaction produced a single product 372 bp in size which was readily detected by agarose gel electrophoresis. All 84 strains of M. bovis tested produced a positive signal in the lification reaction. In addition all isolates fro the M. tuberculosis complex tested, with the exception of M. microti, gave a single band at 372 bp. No lified product was detected when 24 other species of mycobacteria and species from four other genera were tested. The sensitivity of the test was such that a single viable cell could be detected in the reaction. This technique provides a simple and extremely sensitive method of identifying isolates of M. bovis and other pathogenic M. tuberculosis complex organisms.
Publisher: Springer Science and Business Media LLC
Date: 16-11-2019
Publisher: Oxford University Press (OUP)
Date: 24-11-2015
DOI: 10.1093/HMG/DDV472
Publisher: Springer Science and Business Media LLC
Date: 10-07-2013
Publisher: Wiley
Date: 03-2017
DOI: 10.1002/MRD.22771
Publisher: Cold Spring Harbor Laboratory
Date: 05-1992
DOI: 10.1101/GR.1.4.269
Abstract: A comparison of the DNA sequence of the 16S rRNA revealed a region in which there was a minor variation between the species of mycobacteria. This information was used to develop a multiplex lification system that could identify the genus Mycobacterium and then distinguish between M. avium and M. intracellulare, two commonly encountered mycobacteria other than tuberculosis. The combination of these rRNA gene primers together with primers aimed at the MPB70 gene of M. tuberculosis complex organisms permits the detection and identification of clinically significant mycobacteria in a single tube. An lification product of 1030 bp is indicative of the genus Mycobacterium and smaller fragments of 850, 372, and 180 bp are the positive signals for M. intracellulare, M. tuberculosis complex, and M. avium, respectively.
Publisher: Springer Science and Business Media LLC
Date: 18-10-2018
DOI: 10.1038/TPJ.2016.68
Abstract: Selective serotonin reuptake inhibitors (SSRIs) are the most widely used antidepressants, but the efficacy of the treatment varies significantly among in iduals. It is believed that complex genetic mechanisms play a part in this variation. We have used a network based approach to unravel the involved genetic components. Moreover, we investigated the potential difference in the genetic interaction networks underlying SSRI treatment response over time. We found four hub genes (ASCC3, PPARGC1B, SCHIP1 and TMTC2) with different connectivity in the initial SSRI treatment period (baseline to week 4) compared with the subsequent period (4-8 weeks after initiation), suggesting that different genetic networks are important at different times during SSRI treatment. The strongest interactions in the initial SSRI treatment period involved genes encoding transcriptional factors, and in the subsequent period genes involved in calcium homeostasis. In conclusion, we suggest a difference in genetic interaction networks between initial and subsequent SSRI response.
Publisher: Springer Science and Business Media LLC
Date: 03-2017
DOI: 10.1038/NBT.3819
Publisher: S. Karger AG
Date: 1995
DOI: 10.1159/000133905
Abstract: The human tropomyosin 3 (TPM3) gene was previously localized to chromosome 1. The non-muscle isoform of the TPM3 gene product becomes fused to a gene product from the tyrosine kinase receptor gene (NTRKl), previously localized to 1q23→q24, to generate an active oncogene. Two sequence tagged sites spanning three exons and two introns in the carboxy coding region of the gene were used to localize TPM3 to 1q22→q23 by fluorescence in situ hybridization. This localization now places the NTRKl and TPM3 genes in close proximity, so that a gene fusion rearrangement would not be cytologically detected. The 1q22→q23 localization of TPM3 is within the NEM1 locus associated with autosomal dominant nemaline myopathy, making TPM3 a candidate for this disorder.
Publisher: Informa UK Limited
Date: 02-11-2017
DOI: 10.1080/14712598.2017.1250880
Abstract: Antisense nucleic acid analogues can interact with pre-mRNA motifs and influence exon or splice site selection and thereby alter gene expression. Design of antisense molecules to target specific motifs can result in either exon exclusion or exon inclusion during splicing. Novel drugs exploiting the antisense concept are targeting rare, life-limiting diseases however, the potential exists to treat a wide range of conditions by antisense-mediated splice intervention. Areas covered: In this review, the authors discuss the clinical translation of novel molecular therapeutics to address the fatal neuromuscular disorders Duchenne muscular dystrophy and spinal muscular atrophy. The review also highlights difficulties posed by issues pertaining to restricted participant numbers, variable phenotype and disease progression, and the identification and validation of study endpoints. Expert opinion: Translation of novel therapeutics for Duchenne muscular dystrophy and spinal muscular atrophy has been greatly advanced by multidisciplinary research, academic-industry partnerships and in particular, the engagement and support of the patient community. Sponsors, supporters and regulators are cooperating to deliver new drugs and identify and define meaningful outcome measures. Non-conventional and adaptive trial design could be particularly suited to clinical evaluation of novel therapeutics and strategies to treat serious, rare diseases that may be problematic to study using more conventional clinical trial structures.
Publisher: American Society of Civil Engineers
Date: 14-11-2012
Publisher: American Society of Civil Engineers
Date: 14-11-2012
Publisher: BMJ
Date: 23-01-2014
DOI: 10.1136/JMEDGENET-2013-102119
Abstract: The LMNA gene gives rise to at least three isoforms (lamin A, C, lamin AΔ10) as a result of normal alternative splicing, regulated by cis- and trans-acting regulatory factors, as well as the 5' and 3' untranslated regions of the gene. The two main isoforms, lamin A and C, are constitutive components of the fibrous nuclear lamina and have erse physiological roles, ranging from mechanical nuclear membrane maintenance to gene regulation. The clinical spectrum of diseases (called 'laminopathies') caused by LMNA mutations is broad, including at least eight well-characterised phenotypes, some of which are confined to the skeletal muscles or skin, while others are multisystemic. This review discusses the different alternatively spliced isoforms of LMNA and the regulation of LMNA splicing, as well as the subgroup of mutations that affect splicing of LMNA pre-mRNA, and also seeks to bridge the mis-splicing of LMNA at transcript level and the resulting clinical phenotypes. Finally, we discuss the manipulation of LMNA splicing by splice-switching antisense oligonucleotides and its therapeutic potential for the treatment of some laminopathies.
Publisher: Frontiers Media SA
Date: 24-01-2022
DOI: 10.3389/FGENE.2021.806946
Abstract: Understanding pre-mRNA splicing is crucial to accurately diagnosing and treating genetic diseases. However, mutations that alter splicing can exert highly erse effects. Of all the known types of splicing mutations, perhaps the rarest and most difficult to predict are those that activate pseudoexons, sometimes also called cryptic exons. Unlike other splicing mutations that either destroy or redirect existing splice events, pseudoexon mutations appear to create entirely new exons within introns. Since exon definition in vertebrates requires coordinated arrangements of numerous RNA motifs, one might expect that pseudoexons would only arise when rearrangements of intronic DNA create novel exons by chance. Surprisingly, although such mutations do occur, a far more common cause of pseudoexons is deep-intronic single nucleotide variants, raising the question of why these latent exon-like tracts near the mutation sites have not already been purged from the genome by the evolutionary advantage of more efficient splicing. Possible answers may lie in deep intronic splicing processes such as recursive splicing or poison exon splicing. Because these processes utilize intronic motifs that benignly engage with the spliceosome, the regions involved may be more susceptible to exonization than other intronic regions would be. We speculated that a comprehensive study of reported pseudoexons might detect alignments with known deep intronic splice sites and could also permit the characterisation of novel pseudoexon categories. In this report, we present and analyse a catalogue of over 400 published pseudoexon splice events. In addition to confirming prior observations of the most common pseudoexon mutation types, the size of this catalogue also enabled us to suggest new categories for some of the rarer types of pseudoexon mutation. By comparing our catalogue against published datasets of non-canonical splice events, we also found that 15.7% of pseudoexons exhibit some splicing activity at one or both of their splice sites in non-mutant cells. Importantly, this included seven ex les of experimentally confirmed recursive splice sites, confirming for the first time a long-suspected link between these two splicing phenomena. These findings have the potential to improve the fidelity of genetic diagnostics and reveal new targets for splice-modulating therapies.
Publisher: Springer Science and Business Media LLC
Date: 05-2015
DOI: 10.1038/NM0515-537C
Publisher: Springer Science and Business Media LLC
Date: 26-05-1999
Abstract: This study attempted to corroborate findings on the association between butyrylcholinesterase K variant and Alzheimer's disease. This was performed on an autopsy-confirmed series of patients with Alzheimer's disease and controls. The butyrylcholinesterase K variant was found to be of increased allele frequency in patients with sporadic Alzheimer's disease. When related to APOE epsilon4 typing the association was specific but not sensitive for the diagnosis of Alzheimer's disease.
Publisher: Frontiers Media SA
Date: 31-01-2020
Publisher: S. Karger AG
Date: 1993
DOI: 10.1159/000133467
Abstract: The human gene for slow-twitch skeletal muscle troponin I (TNNI1) has previously been mapped to 1q1 i /i →qter using somatic cell hybrids. The TNNI 1 locus has now been further localised to 1q32 using fluorescence in situ hybridization. This result confirms the previous assignment of this locus and maps the gene to a single chromosome band.
Publisher: Wiley
Date: 21-11-2018
DOI: 10.1002/OBY.22308
Abstract: This study aimed to investigate the effect of a genetic risk score (GRS) comprising 15 single-nucleotide polymorphisms, previously shown to associate with childhood BMI, on the baseline cardiometabolic traits and the response to a lifestyle intervention in Danish children and adolescents. Children and adolescents with overweight or obesity (n = 920) and a population-based control s le (n = 698) were recruited. Anthropometric and biochemical measures were obtained at baseline and in a subgroup of children and adolescents with overweight or obesity again after 6 to 24 months of lifestyle intervention (n = 754). The effects of the GRS were examined by multiple linear regressions using additive genetic models. At baseline, the GRS associated with BMI standard deviation score (SDS) both in children and adolescents with overweight or obesity (β = 0.033 [SE = 0.01] P = 0.001) and in the population-based s le (β = 0.065 [SE = 0.02] P = 0.001). No associations were observed for cardiometabolic traits. The GRS did not influence changes in BMI SDS or cardiometabolic traits following lifestyle intervention. A GRS for childhood BMI was associated with BMI SDS but not with other cardiometabolic traits in Danish children and adolescents. The GRS did not influence treatment response following lifestyle intervention.
Publisher: Public Library of Science (PLoS)
Date: 28-10-2011
Publisher: Elsevier BV
Date: 06-2018
Publisher: American Physiological Society
Date: 10-2019
DOI: 10.1152/PHYSIOLGENOMICS.00125.2018
Abstract: Characterization of genetic variants affecting genome-wide gene expression levels (expression quantitative trait loci or eQTLs) in pig testes may improve our understanding of genetic architecture of boar taint (an animal welfare trait) and helps in genome-assisted or genomic selection programs. The aims of this study were to identify eQTLs associated with androstenone, to find candidate eQTLs for low androstenone, and to validate the top eQTL by reverse transcriptase quantitative PCR (RT-qPCR). Gene expression profiles were obtained by RNA sequencing in testis from Danish cross-bred pigs and genotype data by 80K single nucleotide polymorphism panel. A total of 262 eQTLs [false discovery rate (FDR) 0.05] were identified by using two software packages: Matrix eQTL and Krux eQTL. Of these, 149 cis-acting eQTLs were significantly associated with androstenone concentrations and gene expression (FDR 0.05). The eQTLs were associated with several genes of boar taint relevance including CYP1A2, CYB5D1, and SPHK2. One eQTL gene, AMPH, was differentially expressed (FDR 0.05) and affected by chicory. Five candidate eQTLs associated with low androstenone concentrations were discovered, including the top eQTL associated with CYP1A2. RT-qPCR confirmed target gene expression to be significantly ( P 0.05) different based on eQTL genotypes. Furthermore, eQTLs were enriched as QTLs for 15 boar taint related traits from the PigQTLdb. This is the first study to report eQTLs in testes of commercial crossbred pigs used in pork production and to reveal genetic architecture of boar taint. Potential applications include development of a DNA test and in advanced genomic selection models for boar taint.
Publisher: Portland Press Ltd.
Date: 20-07-2007
DOI: 10.1042/BST0350826
Abstract: The cellular uptake of PMOs (phosphorodiamidate morpholino oligomers) can be enhanced by their conjugation to arginine-rich CPPs (cell-penetrating peptides). Here, we discuss our recent findings regarding (R-Ahx-R)4AhxB (Ahx is 6-aminohexanoic acid and B is β-alanine) CPP–PMO conjugates in DMD (Duchenne muscular dystrophy) and murine coronavirus research. An (R-Ahx-R)4AhxB–PMO conjugate was the most effective compound in inducing the correction of mutant dystrophin transcripts in myoblasts derived from a canine model of DMD. Similarly, normal levels of dystrophin expression were restored in the diaphragms of mdx mice, with treatment starting at the neonatal stage, and protein was still detecTable 22 weeks after the last dose of an (R-Ahx-R)4AhxB–PMO conjugate. Effects of length, linkage and carbohydrate modification of this CPP on the delivery of a PMO were investigated in a coronavirus mouse model. An (R-Ahx-R)4AhxB–PMO conjugate effectively inhibited viral replication, in comparison with other peptides conjugated to the same PMO. Shortening the CPP length, modifying it with a mannosylated serine moiety or replacing it with the R9F2 CPP significantly decreased the efficacy of the resulting PPMO (CPP–PMO conjugate). We attribute the success of this CPP to its stability in serum and its capacity to transport PMO to RNA targets in a manner superior to that of poly-arginine CPPs.
Publisher: Frontiers Media SA
Date: 13-03-2019
Publisher: Wiley
Date: 2018
DOI: 10.1002/JGM.3003
Publisher: MDPI AG
Date: 08-06-2018
DOI: 10.3390/RS10060905
Publisher: Wiley
Date: 15-06-1993
Abstract: A single base change in the 5' splice-site of intron 19 has been identified as the cause of the Becker muscular dystrophy in a family which had previously been deduced to carry both a major deletion and another, at that stage unidentified, mutation in the same dystrophin gene [Laing et al., 1992]. RNA from a muscle biopsy of one of the Becker muscular dystrophy patients in the family was analysed using the reverse transcriptase-polymerase chain reaction (RT-PCR) to study the mature gene transcript. Exon 19 was deleted from the dystrophin mRNA but present at the genomic level. The loss of exon 19 in the mature mRNA was found to be associated with an A to C mutation in the 5' splice site of intron 19. Deletion of exon 19 should alter the reading frame of the mRNA and be associated with a severe form of muscular dystrophy however, low levels of normal-size dystrophin message and dystrophin were present in this patient. The distance between the splice-site mutation and the secondary deletion in the dystrophin gene is such that it would seem unlikely that the initial base change could act as a premutation for the deletion. Specific primers to detect the splice-site mutation have been designed and used to genotype all relatives.
Publisher: IEEE
Date: 07-2012
Publisher: Frontiers Media SA
Date: 21-02-2022
Abstract: ATP Binding Cassette Subfamily A Member 3 (ABCA-3) is a lipid transporter protein highly expressed in type-II alveolar (AT-II) cells. Mutations in ABCA3 can result in severe respiratory disease in infants and children. To study ABCA-3 deficiency in vitro , primary AT-II cells would be the cell culture of choice although s le accessibility is limited. Our aim was to investigate the suitability of primary nasal epithelial cells, as a surrogate culture model for AT-II cells, to study ABCA-3 deficiency. Expression of ABCA3 , and surfactant protein genes, SFTPB and SFTPC , was detected in primary nasal epithelial cells but at a significantly lower level than in AT-II cells. ABCA-3, SP-B, and SP-C were detected by immunofluorescence microscopy in primary nasal epithelial cells. However, SP-B and SP-C were undetectable in primary nasal epithelial cells using western blotting. Structurally imperfect lamellar bodies were observed in primary nasal epithelial cells using transmission electron microscopy. Functional assessment of the ABCA-3 protein demonstrated that higher concentrations of doxorubicin reduced cell viability in ABCA-3 deficient nasal epithelial cells compared to controls in an assay-dependent manner. Our results indicate that there may be a role for primary nasal epithelial cell cultures to model ABCA-3 deficiency in vitro , although additional cell culture models that more effectively recapitulate the AT-II phenotype may be required.
Publisher: Informa UK Limited
Date: 19-01-2016
Publisher: Springer International Publishing
Date: 2021
Publisher: Future Science Ltd
Date: 04-1997
DOI: 10.2144/97224BM14
Abstract: Pure titanium and titanium alloys are widely used in dentistry and orthopedics. However, despite their outstanding mechanical and biological properties, implant failure mainly due to post-operative infection still remains a significant concern. The possibility to develop inherent antibacterial medical devices was here investigated by covalently inserting bioactive ammonium salts onto the surface of titanium metal substrates. Titanium discs have been functionalized with quaternary ammonium salts (QASs) and with oleic acid (OA), affording the Ti-AEMAC Ti-GTMAC, Ti-AUTEAB, and Ti-OA s les, which were characterized by ATR-FTIR and SEM-EDX analyses and investigated for the roughness and hydrophilic behavior. The chemical modifications were shown to deeply affect the surface properties of the metal substrates and, as a consequence, their bio-interaction. The bacterial adhesion tests against the Gram-negative
Publisher: Springer International Publishing
Date: 2021
Publisher: IEEE
Date: 09-2011
Publisher: Institute of Electrical and Electronics Engineers (IEEE)
Date: 10-2014
Publisher: Informa UK Limited
Date: 12-2010
Publisher: American Society of Civil Engineers (ASCE)
Date: 05-2021
Publisher: Springer Science and Business Media LLC
Date: 22-08-2019
DOI: 10.1038/S41439-019-0070-X
Abstract: Duchenne muscular dystrophy is an inherited muscle wasting disease with severe symptoms and onset in early childhood. Duchenne muscular dystrophy is caused by loss-of-function mutations, most commonly deletions, within the DMD gene. Characterizing the junction points of large genomic deletions facilitates a more detailed model of the origins of these mutations and allows for a greater understanding of phenotypic variations associated with particular genotypes, potentially providing insights into the deletion mechanism. Here, we report sequencing of breakpoint junctions for seven patients with intragenic, whole-exon DMD deletions. Of the seven junction sequences identified, we found one instance of a “clean” break, three instances of microhomology (2–5 bp) at the junction site, and three complex rearrangements involving local sequences. Bioinformatics analysis of the upstream and downstream breakpoint regions revealed a possible role of short inverted repeats in the initiation of some of these deletion events.
Publisher: Public Library of Science (PLoS)
Date: 08-09-2015
Publisher: Oxford University Press (OUP)
Date: 11-2017
DOI: 10.2527/JAS2017.1752
Publisher: Wiley
Date: 09-08-2015
DOI: 10.1002/JGM.2834
Publisher: Humana Press
Date: 2012
DOI: 10.1007/978-1-61779-767-5_11
Abstract: We have taken an empirical approach in designing splice-switching oligomers to induce targeted dystrophin exon skipping. The nucleotide sequence of the exon under examination is first analyzed for potential exon recognition motifs and then a set of oligomers complementary to the acceptor and donor splice sites, as well as intra-exonic regions predicted to contain exon splice enhancers, are designed and synthesized as 2'-O-methyl-modified bases on a phosphorothioate backbone (2OMeAOs). The 2OMeAOs can be readily transfected into cultured normal myogenic cells as cationic lipoplexes, and are incubated for 24 h before total RNA extraction and subsequent analysis by semi-quantitative RT-PCR. The lification conditions used for each dystrophin transcript region under investigation minimize preferential production of shorter licons and do not exaggerate the level of induced RT-PCR products, compared to the endogenous dystrophin transcript product. It is imperative that the test oligomers are transfected over a range of concentrations and that the target exon is excised in a reproducible and dose-dependent manner.Once it has been demonstrated that an oligomer can induce some degree of exon skipping, that target region of the pre-mRNA is assumed to be involved in splicing of the exon. A series of overlapping oligomers are prepared and evaluated by transfection into normal myogenic cells at lower concentrations to identify the more effective compounds. Clinical application requires antisense compounds that efficiently modulate splicing at low dosages, delivering the greatest benefits in terms of efficacy, safety, and cost.
Publisher: Frontiers Media SA
Date: 06-04-2022
DOI: 10.3389/FGENE.2022.791416
Abstract: Oligonucleotides and nucleic acid analogues that alter gene expression are now showing therapeutic promise in human disease. Whilst the modification of synthetic nucleic acids to protect against nuclease degradation and to influence drug function is common practice, such modifications may also confer unexpected physicochemical and biological properties. Gapmer mixed-modified and DNA oligonucleotides on a phosphorothioate backbone can bind non-specifically to intracellular proteins to form a variety of toxic inclusions, driven by the phosphorothioate linkages, but also influenced by the oligonucleotide sequence. Recently, the non-antisense or other off-target effects of 2′ O - fully modified phosphorothioate linkage oligonucleotides are becoming better understood. Here, we report chemistry-specific effects of oligonucleotides composed of modified or unmodified bases, with phosphorothioate linkages, on subnuclear organelles and show altered distribution of nuclear proteins, the appearance of highly stable and strikingly structured nuclear inclusions, and disturbed RNA processing in primary human fibroblasts and other cultured cells. Phosphodiester, phosphorodiamidate morpholino oligomers, and annealed complimentary phosphorothioate oligomer duplexes elicited no such consequences. Disruption of subnuclear structures and proteins elicit severe phenotypic disturbances, revealed by transcriptomic analysis of transfected fibroblasts exhibiting such disruption. Our data add to the growing body of evidence of off-target effects of some phosphorothioate nucleic acid drugs in primary cells and suggest alternative approaches to mitigate these effects.
Publisher: Cold Spring Harbor Laboratory
Date: 20-03-2020
DOI: 10.1101/2020.03.20.998203
Abstract: Improvement of feed efficiency (FE) is key for sustainability and cost reduction in pig production. Our aim was to characterize the muscle transcriptomic profiles in Danbred Duroc (Duroc) and Danbred Landrace (Landrace), in relation to FE for identifying potential biomarkers. RNA-seq data was analyzed employing differential gene expression methods, gene-gene interaction and network analysis, including pathway and functional analysis. We compared the results with genome regulation in human exercise data. In the differential expression analysis, 13 genes were differentially expressed, including: MRPS11, MTRF1 , TRIM63, MGAT4A, KLH30. Based on a novel gene selection method, the ergent count, we performed pathway enrichment analysis. We found 5 significantly enriched pathways related to feed conversion ratio (FCR). These pathways were mainly mitochondrial, and summarized in the mitochondrial translation elongation (MTR) pathway. In the gene interaction analysis, highlights include the mitochondrial genes: PPIF, MRPL35, NDUFS4and the fat metabolism and obesity genes: AACS, SMPDL3B, CTNNBL1, NDUFS4 and LIMD2 . In the network analysis, we identified two modules significantly correlated with FCR. Pathway enrichment of modules identified MTR, electron transport chain and DNA repair as enriched pathways. In the network analysis, the mitochondrial gene group NDUF was a key hub group, showing potential as biomarkers. Comparing with human transcriptomic exercise studies, genes related to exercise displayed enrichment in our FCR related genes. We conclude that mitochondrial activity is a driver for FCR in muscle tissue, and mitochondrial genes could be potential biomarkers for FCR in pigs. We hypothesize that increased FE mimics processes triggered in exercised muscle.
Publisher: Bentham Science Publishers Ltd.
Date: 03-2010
DOI: 10.2174/138161210790883480
Abstract: In little more than a decade, induced exon skipping as a therapy to treat Duchenne muscular dystrophy (DMD) has progressed from a concept tested in vitro, to pre-clinical evaluation in mouse and dog models, and recent completion of Phase I clinical trials in man. There is no longer any doubt that antisense oligomers can redirect dystrophin gene processing and by-pass protein truncating mutations after direct injection into muscle. Proof-of-concept has been demonstrated in human dystrophic muscle, with trials in Leiden and London showing that two different oligomer chemistries can restore the reading-frame in selected DMD patients by excising dystrophin exon 51. Systemic delivery of both oligomer types into DMD patients has commenced with promising results but it remains to be established if this therapy will have measurable clinical benefits. Targeted removal of exon 51 will only be directly applicable to about one in ten DMD in iduals, and the immediate challenges include development of appropriate and effective delivery regimens, and extending splice-switching therapies to other dystrophin gene lesions. The success of induced exon skipping has spawned a number of "fusion therapies", including vector-mediated dystrophin exon skipping and ex vivo viral delivery of splice-switching antisense molecules into myogenic stem cells, followed by implantation, which may address long term oligomer delivery issues. This review summarizes the pivotal events leading to the completion of the first proof-of-concept trials and speculates on some of the scientific, ethical, regulatory and commercial challenges facing targeted exon skipping for the treatment of DMD.
Publisher: Springer Science and Business Media LLC
Date: 10-09-2020
Publisher: Springer Science and Business Media LLC
Date: 03-06-2010
Publisher: MDPI AG
Date: 13-06-2020
Abstract: DNA methylation of different gene components, including different exons and introns, or different lengths of exons and introns is associated with differences in gene expression. To investigate the methylation of porcine gene components associated with the boar taint (BT) trait, this study used reduced representation bisulfite sequencing (RRBS) data from nine porcine testis s les in three BT groups (low, medium and high BT). The results showed that the methylation levels of the first exons and first introns were lower than those of the other exons and introns. The first exons/introns of CpG island regions had even lower levels of methylation. A total of 123 differentially methylated promoters (DMPs), 194 differentially methylated exons (DMEs) and 402 differentially methylated introns (DMIs) were identified, of which 80 DMPs (DMP-CpGis), 112 DMEs (DME-CpGis) and 166 DMIs (DMI-CpGis) were discovered in CpG islands. Importantly, GPX1 contained one each of DMP, DME, DMI, DMP-CpGi, DME-CpGi and DMI-CpGi. Gene-GO term relationships and pathways analysis showed DMP-CpGi-related genes are mainly involved in methylation-related biological functions. In addition, gene–gene interaction networks consisted of nodes that were hypo-methylated GPX1, hypo-methylated APP, hypo-methylated ATOX1, hyper-methylated ADRB2, hyper-methylated RPS6KA1 and hyper-methylated PNMT. They could be used as candidate biomarkers for reducing boar taint in pigs, after further validation in large cohorts.
Publisher: Elsevier BV
Date: 10-2013
Publisher: Medknow
Date: 2008
Abstract: Duchenne muscular dystrophy (DMD), the most common and serious form of childhood muscle wasting is generally caused by protein-truncating mutations in the large DMD gene. Specific removal of an exon from a defective DMD gene transcript has the potential to allow synthesis of a semi-functional dystrophin, thereby reducing the severity and presumably progression of muscle wasting. The efficacy of this treatment will vary greatly between the different mutations that preclude the synthesis of a functional dystrophin. Restoration of the reading frame from a large multi-exon genomic deletion, typically greater than 36 exons, may lead to synthesis of a protein with only partial function and limited clinical benefit, whereas excising a nonsense mutation in a redundant exon should generate a near normal dystrophin. A clinical trial has recently confirmed proof-of-principle that exclusion of Exon 51 from human dystrophin mRNAs, carrying frame-shifting deletions adjacent to this exon, results in dystrophin expression. No major side-effects after local administration of the antisense oligomer were reported. Additional trials are underway, targeting the same exon but using an oligomer of different backbone chemistry. If functional dystrophin synthesis is demonstrated, and safety issues are addressed, subsequent trials will involve systemic delivery. Great challenges are ahead, some technical establishing an effective delivery regimen, some ethical choosing subsequent targets for therapy, and others of an administrative and regulatory nature.
Publisher: Wiley
Date: 13-06-2013
DOI: 10.1002/MGG3.19
Publisher: Frontiers Media SA
Date: 20-12-2019
Publisher: Elsevier BV
Date: 06-2023
Publisher: Informa UK Limited
Date: 27-09-2021
Publisher: Elsevier BV
Date: 03-1985
DOI: 10.1016/0167-4781(85)90049-1
Abstract: The keratin polypeptides of the epidermis from the leg scale region of 17-day-old embryonic chicks were extracted as S-carboxymethylated derivatives and characterised by electrophoresis on SDS and pH 9.5 urea gels including a combination of both in two dimensions. Proteins were isolated that gave X-ray diffraction patterns typical of alpha- and beta- (avian feather) keratins. An mRNA fraction was isolated from 17-day-old scale tissue by guanidinium chloride extraction and sucrose gradient fractionation. The mRNA was translated in the wheat germ system to give a major product indistinguishable from the molecular weight class (Mr 14 500) of scale beta-keratin polypeptides. A cDNA library was constructed in pBR322 from a 15 S mRNA subfraction and two recombinant clones were selected by their strong hybridisation to cDNA prepared from the 15 S mRNA. The sequencing of these has yielded details of the relatedness of two scale keratin genes including their 3' untranslated regions. Almost half of the protein sequences of the two homologous scale keratins has been deduced and a notable feature of the scale keratin structure appears to be the presence of at least two sequence domains consisting of 13 amino acid repeats.
Publisher: Institute of Electrical and Electronics Engineers (IEEE)
Date: 10-2019
Publisher: Elsevier BV
Date: 09-1995
Publisher: S. Karger AG
Date: 1994
DOI: 10.1159/000133644
Abstract: The human skeletal muscle alpha actin gene (ACTAl) has previously been localized to 1p21→qter using somatic cell hybrids and a specific probe from the 3’ untranslated region of the gene. Using fluorescence in situ hybridization the localization has been confirmed and the ACTAl gene precisely mapped to 1q42.
Publisher: Informa UK Limited
Date: 04-2011
Publisher: Elsevier BV
Date: 12-2014
Publisher: Springer Science and Business Media LLC
Date: 20-10-2015
Publisher: Rockefeller University Press
Date: 06-03-2000
Abstract: Conventionally, nonsense mutations within a gene preclude synthesis of a full-length functional protein. Obviation of such a blockage is seen in the mdx mouse, where despite a nonsense mutation in exon 23 of the dystrophin gene, occasional so-called revertant muscle fibers are seen to contain near-normal levels of its protein product. Here, we show that reversion of dystrophin expression in mdx mice muscle involves unprecedented massive loss of up to 30 exons. We detected several alternatively processed transcripts that could account for some of the revertant dystrophins and could not detect genomic deletion from the region commonly skipped in revertant dystrophin. This, together with exon skipping in two noncontiguous regions, favors aberrant splicing as the mechanism for the restoration of dystrophin, but is hard to reconcile with the clonal idiosyncrasy of revertant dystrophins. Revertant dystrophins retain functional domains and mediate plasmalemmal assembly of the dystrophin-associated glycoprotein complex. Physiological function of revertant fibers is demonstrated by the clonal growth of revertant clusters with age, suggesting that revertant dystrophin could be used as a guide to the construction of dystrophin expression vectors for in idual gene therapy. The dystrophin gene in the mdx mouse provides a favored system for study of exon skipping associated with nonsense mutations.
Publisher: Elsevier BV
Date: 10-2005
DOI: 10.1016/J.NMD.2005.06.009
Abstract: Induction of specific exon skipping during the processing of the dystrophin gene transcript is being pursued as a potential therapy for Duchenne muscular dystrophy. Antisense oligonucleotides directed at motifs involved in pre-mRNA processing can manipulate dystrophin exon incorporation in the mature gene transcript. We have compared the exon skipping ability of oligodeoxyribonucleotides with compounds of the identical sequence incorporating 2'-O-methyl modified bases. Antisense oligonucleotides composed entirely of 2'-O-methyl modified bases on a phosphorothioate backbone were consistently more efficient at inducing exon skipping than comparable oligodeoxyribonucleotides. Chimeric antisense oligonucleotides, mixtures of unmodified and 2'-O-methyl modified bases, induced intermediate levels of exon skipping. In addition, we describe terminal modifications that may be incorporated into the 2'-O-methyl antisense oligonucleotides to further enhance efficiency of exon skipping. Our findings suggest that 2'-O-methyl antisense oligonucleotides should be considered for human clinical trials involving targeted exon skipping in dystrophin gene expression in preference to oligodeoxyribonucleotides.
Publisher: MDPI AG
Date: 15-05-2020
Abstract: Metabolites represent the ultimate response of biological systems, so metabolomics is considered the link between genotypes and phenotypes. Feed efficiency is one of the most important phenotypes in sustainable pig production and is the main breeding goal trait. We utilized metabolic and genomic datasets from a total of 108 pigs from our own previously published studies that involved 59 Duroc and 49 Landrace pigs with data on feed efficiency (residual feed intake (RFI)), genotype (PorcineSNP80 BeadChip) data, and metabolomic data (45 final metabolite datasets derived from LC-MS system). Utilizing these datasets, our main aim was to identify genetic variants (single-nucleotide polymorphisms (SNPs)) that affect 45 different metabolite concentrations in plasma collected at the start and end of the performance testing of pigs categorized as high or low in their feed efficiency (based on RFI values). Genome-wide significant genetic variants could be then used as potential genetic or biomarkers in breeding programs for feed efficiency. The other objective was to reveal the biochemical mechanisms underlying genetic variation for pigs’ feed efficiency. In order to achieve these objectives, we firstly conducted a metabolite genome-wide association study (mGWAS) based on mixed linear models and found 152 genome-wide significant SNPs (p-value 1.06 × 10−6) in association with 17 metabolites that included 90 significant SNPs annotated to 52 genes. On chromosome one alone, 51 significant SNPs associated with isovalerylcarnitine and propionylcarnitine were found to be in strong linkage disequilibrium (LD). SNPs in strong LD annotated to FBXL4, and CCNC consisted of two haplotype blocks where three SNPs (ALGA0004000, ALGA0004041, and ALGA0004042) were in the intron regions of FBXL4 and CCNC. The interaction network revealed that CCNC and FBXL4 were linked by the hub gene N6AMT1 that was associated with isovalerylcarnitine and propionylcarnitine. Moreover, three metabolites (i.e., isovalerylcarnitine, propionylcarnitine, and pyruvic acid) were clustered in one group based on the low-high RFI pigs. This study performed a comprehensive metabolite-based genome-wide association study (GWAS) analysis for pigs with differences in feed efficiency and provided significant metabolites for which there is significant genetic variation as well as biological interaction networks. The identified metabolite genetic variants, genes, and networks in high versus low feed efficient pigs could be considered as potential genetic or biomarkers for feed efficiency.
Publisher: Springer Science and Business Media LLC
Date: 04-04-2018
Publisher: Elsevier BV
Date: 12-2017
Publisher: Springer Science and Business Media LLC
Date: 25-05-2006
Abstract: Manipulation of pre-mRNA splicing by antisense oligonucleotides (AOs) offers considerable potential for a number of genetic disorders. One of these is Duchenne muscular dystrophy (DMD), where mutations in the dystrophin gene typically result in premature termination of translation that causes a loss of functional protein. AOs can induce exon skipping such that the mutation is by-passed and the reading frame restored, producing an internally deleted protein similar to that found in the milder Becker muscular dystrophy. To date, this approach has been applied to the mdx mouse model in vitro and in vivo and in human myoblast cultures. Here, we report the application of AO-directed exon skipping to induce dystrophin expression in vitro in a canine model of DMD, golden retriever muscular dystrophy (GRMD). The efficacy of 2'-O-methyl phosphorothioate (2OMe), phosphorodiamidate morpholino oligomers (PMOs) and peptide-linked PMOs (PMO-Pep) to induce dystrophin expression was assessed. The 2OMe chemistry was only effective for short-term induction of corrected transcript and could not induce detectable dystrophin protein. The PMO chemistry generally induced limited exon skipping at only high concentrations however, a low level of dystrophin protein was produced in treated cells. Use of the PMO-Pep, applied here for the first time to a DMD model, was able to induce high and sustained levels of exon skipping and induced the highest level of dystrophin expression with no apparent adverse effects upon the cells. The induction of dystrophin in the GRMD model offers the potential for further testing of AO delivery regimens in a larger animal model of DMD, in preparation for application in human clinical trials.
Publisher: Elsevier BV
Date: 03-2019
Publisher: MDPI AG
Date: 10-2020
DOI: 10.3390/IJMS21197282
Abstract: Parkin-type autosomal recessive juvenile-onset Parkinson’s disease is caused by mutations in the PRKN gene and accounts for 50% of all autosomal recessive Parkinsonism cases. Parkin is a neuroprotective protein that has dual functions as an E3 ligase in the ubiquitin–proteasome system and as a transcriptional repressor of p53. While genomic deletions of PRKN exon 3 disrupt the mRNA reading frame and result in the loss of functional parkin protein, deletions of both exon 3 and 4 maintain the reading frame and are associated with a later onset, milder disease progression, indicating this particular isoform retains some function. Here, we describe in vitro evaluation of antisense oligomers that restore functional parkin expression in cells derived from a Parkinson’s patient carrying a heterozygous PRKN exon 3 deletion, by inducing exon 4 skipping to correct the reading frame. We show that the induced PRKN transcript is translated into a shorter but semi-functional parkin isoform able to be recruited to depolarised mitochondria, and also transcriptionally represses p53 expression. These results support the potential use of antisense oligomers as a disease-modifying treatment for selected pathogenic PRKN mutations.
Publisher: Wiley
Date: 25-02-2019
Publisher: Wiley
Date: 06-08-2020
DOI: 10.1002/MED.21718
Publisher: IEEE
Date: 07-2010
Publisher: MDPI AG
Date: 20-11-2021
DOI: 10.3390/RS13224698
Abstract: This paper proposes a new approach based on an unsupervised deep learning (DL) model for landslide detection. Recently, supervised DL models using convolutional neural networks (CNN) have been widely studied for landslide detection. Even though these models provide robust performance and reliable results, they depend highly on a large labeled dataset for their training step. As an alternative, in this paper, we developed an unsupervised learning model by employing a convolutional auto-encoder (CAE) to deal with the problem of limited labeled data for training. The CAE was used to learn and extract the abstract and high-level features without using training data. To assess the performance of the proposed approach, we used Sentinel-2 imagery and a digital elevation model (DEM) to map landslides in three different case studies in India, China, and Taiwan. Using minimum noise fraction (MNF) transformation, we reduced the multispectral dimension to three features containing more than 80% of scene information. Next, these features were stacked with slope data and NDVI as inputs to the CAE model. The Huber reconstruction loss was used to evaluate the inputs. We achieved reconstruction losses ranging from 0.10 to 0.147 for the MNF features, slope, and NDVI stack for all three study areas. The mini-batch K-means clustering method was used to cluster the features into two to five classes. To evaluate the impact of deep features on landslide detection, we first clustered a stack of MNF features, slope, and NDVI, then the same ones plus with the deep features. For all cases, clustering based on deep features provided the highest precision, recall, F1-score, and mean intersection over the union in landslide detection.
Publisher: MDPI AG
Date: 12-08-2019
DOI: 10.3390/MOLECULES24162922
Abstract: One of the crucial aspects of screening antisense oligonucleotides destined for therapeutic application is confidence that the antisense oligomer is delivered efficiently into cultured cells. Efficient delivery is particularly vital for antisense phosphorodiamidate morpholino oligomers, which have a neutral backbone, and are known to show poor gymnotic uptake. Here, we report several methods to deliver these oligomers into cultured cells. Although 4D-Nucleofector™ or Neon™ electroporation systems provide efficient delivery and use lower amounts of phosphorodiamidate morpholino oligomer, both systems are costly. We show that some readily available transfection reagents can be used to deliver phosphorodiamidate morpholino oligomers as efficiently as the electroporation systems. Among the transfection reagents tested, we recommend Lipofectamine 3000™ for delivering phosphorodiamidate morpholino oligomers into fibroblasts and Lipofectamine 3000™ or Lipofectamine 2000™ for myoblasts/myotubes. We also provide optimal programs for nucleofection into various cell lines using the P3 Primary Cell 4D-Nucleofector™ X Kit (Lonza), as well as antisense oligomers that redirect expression of ubiquitously expressed genes that may be used as positive treatments for human and murine cell transfections.
Publisher: Elsevier BV
Date: 04-2013
Abstract: Duchenne muscular dystrophy (DMD) is a devastating muscle-wasting disease caused by mutations in the dystrophin gene. Utrophin is a homologue of dystrophin that can compensate for its absence when overexpressed in DMD animal models. Utrophin upregulation is therefore a promising therapeutic approach for DMD. Utrophin is regulated at both transcriptional and posttranscriptional levels. Transcriptional regulation has been studied extensively, and assays have been described for the identification of utrophin promoter-targeting molecules. However, despite the profound impact that posttranscriptional regulation has on utrophin expression, screening assays have not yet been described that could be used to discover pharmaceuticals targeting this key phase of regulation. We describe the development and validation of a muscle cell line-based assay in which a stably expressed luciferase coding sequence is flanked by the utrophin 5'- and 3'-untranslated regions (UTRs). The assay was validated using the posttranscriptional regulation of utrophin by miR-206. The assay has a Z' of 0.7, indicating robust performance in high-throughput format. This assay can be used to study utrophin regulatory mechanisms or to screen chemical libraries for compounds that upregulate utrophin posttranscriptionally via its UTRs. Compounds identified via this assay, used alone or in a synergistic combination with utrophin promoter-targeting molecules, would be predicted to have therapeutic potential for DMD.
Publisher: Jenny Stanford Publishing
Date: 03-03-2016
DOI: 10.1201/B20047-7
Publisher: Wiley
Date: 23-04-2020
DOI: 10.1002/MGG3.1259
Publisher: MDPI AG
Date: 23-07-2019
Abstract: Residual feed intake (RFI) is designed to estimate net efficiency of feed use, so low RFI animals are considered for selection to reduce feeding costs. However, metabolic profiling of cows and availability of predictive metabolic biomarkers for RFI are scarce. Therefore, this study aims to generate a better understanding of metabolic mechanisms behind low and high RFI in Jerseys and Holsteins and identify potential predictive metabolic biomarkers. Each metabolite was analyzed to reveal their associations with two RFIs in two breeds by a linear regression model. An integrative analysis of metabolomics and transcriptomics was performed to explore interactions between functionally related metabolites and genes in the created metabolite networks. We found that three main clusters were detected in the heat map and all identified fatty acids (palmitoleic, hexadecanoic, octadecanoic, heptadecanoic, and tetradecanoic acid) were grouped in a cluster. The lower cluster were all from fatty acids, including palmitoleic acid, hexadecanoic acid, octadecanoic acid, heptadecanoic acid, and tetradecanoic acid. The first component of the partial least squares-discriminant analysis (PLS-DA) explained a majority (61.5%) of variations of all metabolites. A good ision between two breeds was also observed. Significant differences between low and high RFIs existed in the fatty acid group (P 0.001). Statistical results revealed clearly significant differences between breeds however, the association of in idual metabolites (leucine, ornithine, pentadecanoic acid, and valine) with the RFI status was only marginally significant or not significant due to a lower s le size. The integrated gene-metabolite pathway analysis showed that pathway impact values were higher than those of a single metabolic pathway. Both types of pathway analyses revealed three important pathways, which were aminoacyl-tRNA biosynthesis, alanine, aspartate, and glutamate metabolism, and the citrate cycle (TCA cycle). Finally, one gene (2-hydroxyacyl-CoA lyase 1 (+HACL1)) associated with two metabolites (-α-ketoglutarate and succinic acid) were identified in the gene-metabolite interaction network. This study provided novel metabolic pathways and integrated metabolic-gene expression networks in high and low RFI Holstein and Jersey cattle, thereby providing a better understanding of novel biochemical mechanisms underlying variation in feed efficiency.
Publisher: Springer Science and Business Media LLC
Date: 29-04-2016
Publisher: American Society of Civil Engineers (ASCE)
Date: 05-2016
Publisher: Elsevier BV
Date: 02-2000
DOI: 10.1016/S0960-8966(99)00063-2
Abstract: We report two siblings with a relatively severe limb-girdle muscular dystrophy. The elder sister presented at 8 years of age with inability to climb and abnormal gait. At 12 years she was barely ambulant. Her sister followed a similar course. Serum creatine kinase was 8500-10000 IU (N 25-200) in the elder sister and 17000-19000 IU in the younger sister. Muscle biopsy of the elder sister at 8 years showed chronic myopathic changes with loss of muscle fibres, active necrosis and regeneration. Immunocytochemistry demonstrated normal spectrin and dystrophin, reduced alpha-sarcoglycan and absent gamma-sarcoglycan--indicating a gamma-sarcoglycanopathy. Haplotype analysis for the markers D13S115, D13S232, D13S292, D13S787, D13S1243 and D13S283 internal to and flanking the gamma-sarcoglycan gene showed the affected sisters shared haplotypes, indicating it was possible they were suffering from a gamma-sarcoglycanopathy. Non-inheritance of paternal alleles for D13S232, D13S292 and D13S1243 suggested the inheritance of a deletion, which was confirmed by FISH, using a genomic probe from the gamma-sarcoglycan gene. The gamma-sarcoglycan cDNA was lified by reverse transcriptase PCR from the muscle biopsy of the elder sister and sequenced. A missense mutation changing codon 69 from GGC glycine to CGC arginine was identified. HhaI digestion of exon 3 genomic PCR products showed the two affected sisters were hemizygous for the mutation, while the mother and grandmother were heterozygotes. The mutation, identified by SSCP analysis, was not observed in 116 unrelated, unaffected in iduals. Previously, only two other missense mutations, the Cys283Tyr missense mutation in Gypsies and the Leu193Ser mutation in a Dutch family, have been described in the gamma-sarcoglycan gene. The fact that the affected in iduals in the current and Gypsy families are gamma-sarcoglycan negative may indicate that codons 69 and 283 are important in gamma-sarcoglycan function.
Publisher: Wiley
Date: 08-1993
Abstract: The polymerase chain reaction (PCR) was used on material from a blighted ovum to confirm indirectly the carrier status of a woman with a family history of Becker muscular dystrophy. Conventional testing including creatine kinase levels, muscle biopsy, and EMG had been inconclusive, and on the basis of one elevated creatine kinase level, the woman had been designated a possible carrier. Ultrasound examination at 10 weeks of pregnancy indicated a blighted ovum, from which DNA was subsequently extracted and subjected to PCR testing for determination of sex and genotypic status with respect to the known familial deletion of the dystrophin gene. The blighted ovum was found to have a Y chromosome and also to be deleted for at least exon 6 of the dystrophin gene, indirectly indicating that the mother most likely carried the family mutation for Becker muscular dystrophy.
Publisher: Frontiers Media SA
Date: 22-03-2022
DOI: 10.3389/FPHAR.2022.868863
Abstract: Introduction: Severity and disease progression in people with Cystic Fibrosis (CF) is typically dependent on their genotype. One potential therapeutic strategy for people with specific mutations is exon skipping with antisense oligonucleotides (AO). CFTR exon 9 is an in-frame exon and hence the exclusion of this exon would excise only 31 amino acids but not alter the reading frame of the remaining mRNA. Splice mutations 1209 + 1 G & C and 1209 + 2 T & G were documented to cause CFTR exon 9 skipping and these variants were reported to manifest as a milder CF disease, therefore exon 9 skipping could be beneficial for people with class I mutations that affect exon 9 such as p.Trp401X. While the impact of exon 9 skipping on gene expression and cellular pathways can be studied in cells in vitro , trace amount of full-length normal or mutated material could confound the evaluation. To overcome this limitation, the impact of CFTR exon 9 skipping on disease phenotype and severity is more effectively evaluated in a small animal model. It was hypothesised that antisense oligonucleotide-mediated skipping this particular exon could result in a “mild mouse CF phenotype”. Methods: Cftr exon 9 deleted mice were generated using homologous recombination. Survival of homozygous ( Cftr Δ9/Δ9 ) and heterozygous ( Cftr Δ9/+ ) mice was compared to that of other CF mouse models, and lung and intestinal organ histology examined for any pathologies. Primary airway epithelial cells (pAECs) were harvested from Cftr Δ9/Δ9 mice and cultured at the Air Liquid Interface for CFTR functional assessment using Ussing Chamber analysis. Results: A Cftr Δ9/Δ9 mouse model presented with intestinal obstructions, and at time of weaning (21 days). Cftr Δ9/Δ9 mice had a survival rate of 83% that dropped to 38% by day 50. Histological sections of the small intestine from Cftr Δ9/Δ9 mice showed more goblet cells and mucus accumulation than s les from the Cftr Δ9/+ littermates. Airway epithelial cell cultures established from Cftr Δ9/Δ9 mice were not responsive to forskolin stimulation. Summary: The effect of Cftr exon 9 deletion on Cftr function was assessed and it was determined that the encoded Cftr isoform did not result in a milder “mouse CF disease phenotype,” suggesting that Cftr exon 9 is not dispensable, although further investigation in human CF pAECs would be required to confirm this observation.
Publisher: Elsevier BV
Date: 08-1996
DOI: 10.1016/S0140-6736(05)64588-6
Abstract: Carotenoids have several crucial biological functions and are part of the cold adaptation mechanism of some bacteria. Some pink-pigmented Arthrobacter species produce the rare C
Publisher: Elsevier BV
Date: 12-1994
Publisher: Wiley
Date: 02-02-2016
DOI: 10.1038/ICB.2016.3
Abstract: Inflammasomes are molecular complexes activated by infection and cellular stress, leading to caspase-1 activation and subsequent interleukin-1β (IL-1β) processing and cell death. The autoimmune NZB mouse strain does not express NLRP3, a key inflammasome initiator mediating responses to a wide variety of stimuli including endogenous danger signals, environmental irritants and a range of bacterial, fungal and viral pathogens. We have previously identified an intronic point mutation in the Nlrp3 gene from NZB mice that generates a splice acceptor site. This leads to inclusion of a pseudoexon that introduces an early termination codon and is proposed to be the cause of NLRP3 inflammasome deficiency in NZB cells. Here we have used exon skipping antisense oligonucleotides (AONs) to prevent aberrant splicing of Nlrp3 in NZB macrophages, and this restored both NLRP3 protein expression and NLRP3 inflammasome activity. Thus, the single point mutation leading to aberrant splicing is the sole cause of NLRP3 inflammasome deficiency in NZB macrophages. The NZB mouse provides a model for addressing a splicing defect in macrophages and could be used to further investigate AON design and delivery of AONs to macrophages in vivo.
Publisher: Wiley
Date: 09-2013
DOI: 10.1002/JGM.2740
Publisher: Proceedings of the National Academy of Sciences
Date: 02-03-1999
Abstract: The congenital nemaline myopathies are rare hereditary muscle disorders characterized by the presence in the muscle fibers of nemaline bodies consisting of proteins derived from the Z disc and thin filament. In a single large Australian family with an autosomal dominant form of nemaline myopathy, the disease is caused by a mutation in the α-tropomyosin gene TPM3 . The typical form of nemaline myopathy is inherited as an autosomal recessive trait, the locus of which we previously assigned to chromosome 2q21.2-q22. We show here that mutations in the nebulin gene located within this region are associated with the disease. The nebulin protein is a giant protein found in the thin filaments of striated muscle. A variety of nebulin isoforms are thought to contribute to the molecular ersity of Z discs. We have studied the 3′ end of the 20.8-kb cDNA encoding the Z disc part of the 800-kDa protein and describe six disease-associated mutations in patients from five families of different ethnic origins. In two families with consanguineous parents, the patients were homozygous for point mutations. In one family with nonconsanguineous parents, the affected siblings were compound heterozygotes for two different mutations, and in two further families with one detected mutation each, haplotypes are compatible with compound heterozygosity. Immunofluorescence studies with antibodies specific to the C-terminal region of nebulin indicate that the mutations may cause protein truncation possibly associated with loss of fiber-type ersity, which may be relevant to disease pathogenesis.
Publisher: Public Library of Science (PLoS)
Date: 23-03-2017
Publisher: MDPI AG
Date: 27-03-2021
DOI: 10.3390/IJMS22073479
Abstract: Marfan syndrome is one of the most common dominantly inherited connective tissue disorders, affecting 2–3 in 10,000 in iduals, and is caused by one of over 2800 unique FBN1 mutations. Mutations in FBN1 result in reduced fibrillin-1 expression, or the production of two different fibrillin-1 monomers unable to interact to form functional microfibrils. Here, we describe in vitro evaluation of antisense oligonucleotides designed to mediate exclusion of FBN1 exon 52 during pre-mRNA splicing to restore monomer homology. Antisense oligonucleotide sequences were screened in healthy control fibroblasts. The most effective sequence was synthesised as a phosphorodiamidate morpholino oligomer, a chemistry shown to be safe and effective clinically. We show that exon 52 can be excluded in up to 100% of FBN1 transcripts in healthy control fibroblasts transfected with PMO52. Immunofluorescent staining revealed the loss of fibrillin 1 fibres with ~50% skipping and the subsequent re-appearance of fibres with % skipping. However, the effect of exon skipping on the function of the induced fibrillin-1 isoform remains to be explored. Therefore, these findings demonstrate proof-of-concept that exclusion of an exon from FBN1 pre-mRNA can result in internally truncated but identical monomers capable of forming fibres and lay a foundation for further investigation to determine the effect of exon skipping on fibrillin-1 function.
Publisher: Elsevier BV
Date: 08-2014
Publisher: Cold Spring Harbor Laboratory
Date: 13-04-2020
DOI: 10.1101/2020.04.11.037168
Abstract: DNA methylation in gene or promoter or gene body could restrict romote the gene transcription. Moreover, methylation in the gene regions along with CpG island regions could modulate the transcription to undetectable gene expression levels. Therefore, it is necessary to investigate the methylation levels within the gene, gene body, CpG island regions and their overlapped regions and then identify the gene-based differentially methylated regions (GeneDMRs). Here, R package GeneDMRs aims to facilitate computing gene based methylation rate using next generation sequencing (NGS)-based methylome data. The user-friendly R package GeneDMRs is presented to analyze the methylation levels in each gene romoter/exon/intron/CpG island/CpG island shore or each overlapped region (e.g., gene-CpG island romoter-CpG island/exon-CpG island/intron-CpG island/gene-CpG island shore romoter-CpG island shore/exon-CpG island shore/intron-CpG island shore). Here, we used the public reduced representation bisulfite sequencing (RRBS) data of mouse (GSE62392) for evaluating software and found novel biologically significant results to supplement the previous research. The R package GeneDMRs can facilitate computing gene based methylation rate to interpret complex interplay between methylation levels and gene expression differences or similarities across physiological conditions or disease states.
Publisher: Wiley
Date: 31-10-2013
DOI: 10.1111/IEP.12048
Publisher: Springer Science and Business Media LLC
Date: 07-01-2020
DOI: 10.1186/S40104-019-0407-9
Abstract: Genotyping by sequencing (GBS) still has problems with missing genotypes. Imputation is important for using GBS for genomic predictions, especially for low depths, due to the large number of missing genotypes. Minor allele frequency (MAF) is widely used as a marker data editing criteria for genomic predictions. In this study, three imputation methods (Beagle, IMPUTE2 and FImpute software) based on four MAF editing criteria were investigated with regard to imputation accuracy of missing genotypes and accuracy of genomic predictions, based on simulated data of livestock population. Four MAFs (no MAF limit, MAF ≥ 0.001, MAF ≥ 0.01 and MAF ≥ 0.03) were used for editing marker data before imputation. Beagle, IMPUTE2 and FImpute software were applied to impute the original GBS. Additionally, IMPUTE2 also imputed the expected genotype dosage after genotype correction (GcIM). The reliability of genomic predictions was calculated using GBS and imputed GBS data. The results showed that imputation accuracies were the same for the three imputation methods, except for the data of sequencing read depth (depth) = 2, where FImpute had a slightly lower imputation accuracy than Beagle and IMPUTE2. GcIM was observed to be the best for all of the imputations at depth = 4, 5 and 10, but the worst for depth = 2. For genomic prediction, retaining more SNPs with no MAF limit resulted in higher reliability. As the depth increased to 10, the prediction reliabilities approached those using true genotypes in the GBS loci. Beagle and IMPUTE2 had the largest increases in prediction reliability of 5 percentage points, and FImpute gained 3 percentage points at depth = 2. The best prediction was observed at depth = 4, 5 and 10 using GcIM, but the worst prediction was also observed using GcIM at depth = 2. The current study showed that imputation accuracies were relatively low for GBS with low depths and high for GBS with high depths. Imputation resulted in larger gains in the reliability of genomic predictions for GBS with lower depths. These results suggest that the application of IMPUTE2, based on a corrected GBS (GcIM) to improve genomic predictions for higher depths, and FImpute software could be a good alternative for routine imputation.
Publisher: Springer Science and Business Media LLC
Date: 27-04-2012
Publisher: Wiley
Date: 17-02-2009
DOI: 10.1002/MDS.22214
Abstract: Recent whole genome association studies provided little evidence that polymorphisms at the familial Parkinsonism loci influence the risk for Parkinson's disease (PD). However, these studies are not designed to detect the types of subtle effects that common variants may impose. Here, we use an alternative targeted candidate gene approach to examine common variation in 11 genes related to familial Parkinsonism. PD cases (n = 331) and unaffected control subjects (n = 296) were recruited from three specialist movement disorder clinics in Brisbane, Australia and the Australian Electoral Roll. Common genetic variables (76 SNPs and 1 STR) were assessed in all subjects and haplotype, genotype, and allele associations explored. Modest associations (uncorrected P < 0.05) were observed for common variants around SNCA, UCHL1, MAPT, and LRRK2 although none were of sufficient magnitude to survive strict statistical corrections for multiple comparisons. No associations were seen for PRKN, PINK1, GBA, ATP13A2, HTRA2, NR4A2, and DJ1. Our findings suggest that common genetic variables of selected PD-related loci contribute modestly to PD risk in Australians.
Publisher: Elsevier BV
Date: 10-2005
Publisher: Elsevier BV
Date: 02-2010
DOI: 10.1016/J.NMD.2009.10.013
Abstract: Duchenne muscular dystrophy (DMD) is caused by the lack of functional dystrophin protein, most commonly as a result of a range of out-of-frame mutations in the DMD gene. Modulation of pre-mRNA splicing with antisense oligonucleotides (AOs) to restore the reading frame has been demonstrated in vitro and in vivo, such that truncated but functional dystrophin is expressed. AO-induced skipping of exon 51 of the DMD gene, which could treat 13% of DMD patients, has now progressed to clinical trials. We describe here the methodical, cooperative comparison, in vitro (in DMD cells) and in vivo (in a transgenic mouse expressing human dystrophin), of 24 AOs of the phosphorodiamidate morpholino oligomer (PMO) chemistry designed to target exon 53 of the DMD gene, skipping of which could be potentially applicable to 8% of patients. A number of the PMOs tested should be considered worthy of development for clinical trial.
Publisher: Elsevier BV
Date: 2021
Publisher: Colegio Brasileiro de Reproducao Animal - CBRA
Date: 2017
Publisher: Springer Science and Business Media LLC
Date: 17-02-2014
Publisher: Elsevier BV
Date: 09-2021
Publisher: Springer Science and Business Media LLC
Date: 23-07-2021
DOI: 10.1038/S41598-021-94639-X
Abstract: Antisense oligomers (AOs) are increasingly being used to modulate RNA splicing in live cells, both for research and for the development of therapeutics. While the most common intended effect of these AOs is to induce skipping of whole exons, rare ex les are emerging of AOs that induce skipping of only part of an exon, through activation of an internal cryptic splice site. In this report, we examined seven AO-induced cryptic splice sites in six genes. Five of these cryptic splice sites were discovered through our own experiments, and two originated from other published reports. We modelled the predicted effects of AO binding on the secondary structure of each of the RNA targets, and how these alterations would in turn affect the accessibility of the RNA to splice factors. We observed that a common predicted effect of AO binding was disruption of the exon definition signal within the exon’s excluded segment.
Publisher: Public Library of Science (PLoS)
Date: 08-09-2016
Publisher: Colegio Brasileiro de Reproducao Animal - CBRA
Date: 2017
Publisher: Wiley
Date: 06-1997
DOI: 10.1002/(SICI)1097-4598(199706)20:6<728::AID-MUS10>3.0.CO;2-Q
Abstract: The mdx mouse, an animal model used to study Duchenne muscular dystrophy, has a nonsense mutation in exon 23 of the dystrophin gene which should result in a truncated protein that cannot be correctly localized at the sarcolemma of the muscle fibers. Immunohistochemical staining with antidystrophin antibodies has shown that while most of the muscle tissue is dystrophin-negative, a small percentage of muscle fibers is clearly dystrophin-positive and has somehow bypassed the primary nonsense mutation. A sensitive nested polymerase chain reaction-based examination of dystrophin gene transcripts around the mdx mutation has revealed several alternatively processed transcripts. Four mRNA species skipped the mutation in exon 23, were in-frame, and could be translated into a shorter but still functional dystrophin protein. Specific tests for these transcripts demonstrated these were also present in normal mouse muscle tissue.
Publisher: Springer New York
Date: 2018
DOI: 10.1007/978-1-4939-8651-4_13
Abstract: Duplications of one or more dystrophin exons that disrupt the reading frame account for about 15% of all Duchenne cases, and like the more common genomic deletions, most pathogenic duplications of single or multiple dystrophin exons are also amenable to targeted exon skipping. However, additional considerations must be taken into account: (1) skipping of all duplicated exons, and, flanking exons as necessary, will frequently be required to restore the reading frame and generate an in-frame Becker muscular dystrophy-like mRNA, (2) the phosphorodiamidate morpholino oligomer chemistry is more effective than the 2'-O-methyl modified oligonucleotides at inducing multiple exon skipping, and (2) the apparent efficiency of exon skipping can be confounded by the choice of RT-PCR system. Standard RT-PCR systems can preferentially lify the shorter licons, implying more efficient exon skipping than may actually be induced. Unless high fidelity RT-PCR systems are used, strand slippage during annealing/elongation steps will generate normal length transcripts that are artifacts of the lification.
Publisher: S. Karger AG
Date: 1995
DOI: 10.1159/000134070
Abstract: A sequence tagged site (STS) was developed for the human beta tropomyosin gene (TPM2). The STS was used to lify DNA from somatic cell hybrids to localise TPM2 to human chromosome 9. Genomic clones isolated with the STS product were in turn used in fluorescence in situ hybridisatior to metaphase chromosome spreads to further localise TPM2 to 9p13.
Publisher: Wiley
Date: 12-03-2019
DOI: 10.1111/EVA.12783
Publisher: Medknow
Date: 2018
Publisher: Wiley
Date: 1997
DOI: 10.1159/000484768
Publisher: American Physiological Society
Date: 08-2008
DOI: 10.1152/JAPPLPHYSIOL.00068.2008
Abstract: The mdx mouse model of muscular dystrophy has a premature stop codon preventing production of dystrophin. This results in a progressive phenotype causing centronucleation of skeletal muscle fibers, muscle weakness, and fibrosis and kyphosis. Antisense oligonucleotides alter RNA splicing to exclude the nonsense mutation, while still maintaining the open reading frame to produce a shorter, but partially functional dystrophin protein that should ameliorate the extent of pathology. The present study investigated the benefits of chronic treatment of mdx mice by once-monthly deep intramuscular injections of antisense oligonucleotides into paraspinal muscles. After 8 mo of treatment, mdx mice had reduced development of kyphosis relative to untreated mdx mice, a benefit that was retained until completion of the study at 18 mo of age (16 mo of treatment). This was accompanied by reduced centronucleation in the latissimus dorsi and intercostals muscles and reduced fibrosis in the diaphragm and latissimus dorsi. These benefits were accompanied by a significant increase in dystrophin production. In conclusion, chronic antisense oligonucleotide treatment provides clear and ongoing benefits to paralumbar skeletal muscle, with associated marked reduction in kyphosis.
Publisher: CSIRO Publishing
Date: 2016
DOI: 10.1071/RDV28N2AB192
Abstract: Oocytes resume meiosis spontaneously when subjected to in vitro maturation (IVM). Cyclic adenosine monophosphate (cAMP) elevating agents have been used for artificial blocking of meiotic resumption (pre-IVM) to allow the oocyte to prepare for maturation, potentially increasing its developmental competence. However, the ultrastructural effects of this pharmacological approach on oocytes and embryos remain to be addressed. We assessed the effects of pre-IVM with cAMP modulators in oocytes (10 for each group) at the end of IVM and in blastocyst (10 for each group) after 7 days of culture. Cumulus-oocyte complexes (COC) were subjected to pre-IVM for 2 h with forskolin (Sigma, St. Louis, MO, USA 100 μM) and 3-isobutyl-1-methylxanthine) (IBMX, Sigma, 500 μM) followed by 24 h of IVM with FSH-enriched media (IVF Vet Solutions, Adelaide, Australia). Simultaneously, another group of COC was subjected to conventional IVM (con-IVM) for 24 h (EmbryoTransBiotech, Copenhagen, Denmark) with BSA (4 mg mL–1, Sigma), gentamycin (50 mg mL–1), and FSH (0.1 IU mL–1). Matured oocytes were collected for qualitative ultrastructural analysis or followed to IVF. The morphology was carefully evaluated on serially sectioned oocytes and embryos, where each serial section (~60.2-μm section per oocyte/embryo) was analysed under light microscopy. Subsequently, the equatorial section from each oocyte and the section giving the optimal representation of the inner cell mass in each blastocyst was re-embedded and sectioned for electron microcopy as previously described (Hyttel and Madsen 1987 Acta Anat. 129, 12–14). Blastocyst rates did not differ between groups. Ultrastructural analyses revealed subtle ultrastructural differences between pre-IVM and con-IVM conditions. In both groups, oocytes had matured to metaphase II. The perivitelline space of pre-IVM oocytes was significantly narrower than con-IVM. The cytoplasmic vesicles were more abundant and globally distributed in pre-IVM oocytes, whereas at con-IVM a vesicle-free periphery of the ooplasm was frequent, except for cortical granules and clusters of mitochondria associated with smooth endoplasmic reticulum (SER). We observed typical hooded mitochondria and cortical granules either clustered in the periphery or solitarily distributed in the cortical ooplasmic region for both groups. In the blastocysts, differences were noted with respect to especially distribution of ribosomes. In pre-IVM blastocysts, ribosomes were mostly organised in free clusters (polysomes) and peripherally located in cells of the inner cell mass. Con-IVM blastocysts showed ribosomes preferentially associated with the rough ER and often associated with mitochondria. Lipid droplets and rounded mitochondria were observed in both groups as well as apically located tight junctions and desmosomes between adjacent trophectoderm (TE) cells. Pleomorphic and elongated mitochondria were abundant in the TE of pre-IVM blastocysts, whereas the mitochondrial population was more homogenous at con-IVM. These findings suggest that pre-IVM for 2 h affects oocyte and blastocyst ultrastructure. Research was supported by grants 12/50533-2 and 12/23409-9 from FAPESP.
Publisher: Mary Ann Liebert Inc
Date: 03-2013
DOI: 10.1089/HUM.2012.211
Publisher: Ovid Technologies (Wolters Kluwer Health)
Date: 10-2000
DOI: 10.1097/00019052-200010000-00008
Abstract: Gene therapy for inherited muscle disease is an active area of research and development. Initial emphasis has been on gene replacement but alternative approaches are increasingly being considered in order to overcome difficulties, such as the immune rejection of transduced cells, the need for appropriate and tissue-specific control of expression, and the requirement for systemic spread in some conditions. However, the most significant obstacles to the clinical success of gene therapy are still the lack of efficiency and accuracy of gene medicine delivery.
Publisher: Elsevier BV
Date: 10-2011
Publisher: Springer Science and Business Media LLC
Date: 24-03-2017
Publisher: Springer Science and Business Media LLC
Date: 12-2018
Publisher: Elsevier BV
Date: 07-2007
Abstract: Protein-truncating mutations in the dystrophin gene lead to the most common childhood form of muscle wasting, Duchenne muscular dystrophy. Becker muscular dystrophy, a condition that typically arises from dystrophin gene lesions that do not disrupt the reading frame, clearly indicates that substantial domains of the dystrophin protein are not essential. Potential therapeutic intervention exists during pre-mRNA splicing, whereby selected exons are excised to either remove nonsense mutations or restore the reading frame around frame-shifting mutations from the mature mRNA. Appropriately designed antisense oligonucleotides (AOs), directed at amenable splicing motifs across the dystrophin gene transcript, block exon recognition and/or spliceosome assembly so that targeted exons are removed from the mature mRNA. We describe a panel of AOs designed to induce skipping of every exon within the human dystrophin gene transcript, with the exception of the first and last exons. Every exon targeted in vitro could be removed from the dystrophin mRNA, although some exons are more efficiently excluded than others. No single motif has emerged as a universal AO annealing site for redirection of dystrophin pre-mRNA processing, although the general trend is that the most efficient compounds are directed at motifs in the first half of the target exon.
Publisher: Public Library of Science (PLoS)
Date: 03-06-2014
Publisher: Frontiers Media SA
Date: 23-06-2020
Publisher: MDPI AG
Date: 27-03-2020
DOI: 10.3390/ANI10040566
Abstract: Association studies have indicated profound effects of copy number variations (CNVs) on various phenotypes in different species. In this study, we identified the CNV distributions and expression levels of guanylate-binding protein 6 (GBP6) associated with the growth traits of Chinese cattle. The results showed that the phenotypic values of body size and weight of Xianan (XN) cattle were higher than those of Nanyang (NY) cattle. The medium CNV types were mostly identified in the XN and NY breeds, but their CNV distributions were significantly different (adjusted p 0.05). The association analysis revealed that the body weight, cannon circumference and chest circumference of XN cattle had significantly different values in different CNV types (p 0.05), with CNV gain types (Log22−ΔΔCt 0.5) displaying superior phenotypic values. We also found that transcription levels varied in different tissues (p 0.001) and the CNV gain types showed the highest relative gene expression levels in the muscle tissue, consistent with the highest phenotypic values of body weight and cannon circumference among the three CNV types. Consequently, our results suggested that CNV gain types of GBP6 could be used as the candidate markers in the cattle-breeding program for growth traits.
Publisher: Hindawi Limited
Date: 2009
DOI: 10.1002/HUMU.20806
Abstract: Out of three mutations in the dystrophin gene that cause Duchenne muscular dystrophy (DMD), the most common, serious childhood muscle wasting disease, two are genomic deletions of one or more exons that disrupt the reading frame. Specific removal of an exon flanking a genomic deletion using antisense oligonucleotide intervention during pre-RNA processing can restore the reading frame and could potentially reduce disease severity. We describe a rare dystrophin gene rearrangement inversion of approximately 28 kb, flanked by a 10-bp duplication and an 11-kb deletion, which led to the omission of exons 49 and 50 from the mature mRNA and the variable inclusion of several pseudoexons. In vitro transfection of cultured patient cells with antisense oligonucleotides directed at exon 51 induced efficient removal of that exon, as well as one of the more commonly included pseudoexons, suggesting closely coordinated splicing of these exons. Surprisingly, several antisense oligonucleotides (AOs) directed at this pseudoexon had no detectable effect on the splicing pattern, while all AOs directed at the other predominant pseudoexon efficiently excised that target. Antisense oligomers targeting dystrophin exon 51 for removal are currently undergoing clinical trials. Despite the unique nature of the dystrophin gene rearrangement described here, a personalized multiexon skipping treatment is applicable and includes one compound entering clinical trials for DMD.
Publisher: Wiley
Date: 29-08-2022
DOI: 10.1002/MDS.29201
Publisher: Cold Spring Harbor Laboratory
Date: 22-05-2020
DOI: 10.1101/2020.05.21.108050
Abstract: Feed efficiency (FE) is an economically important trait. Thus, reliable predictors would help to reduce the production cost and provide sustainability to the pig industry. We carried out metabolome-transcriptome integration analysis on 40 purebred Duroc and Landrace male uncastrated pigs to identify potential gene-metabolite interactions and explore the molecular mechanism underlying FE. To this end, we applied untargeted metabolomics and RNA-seq approaches to the same animals. After data quality control, we used a linear model approach to integrate the data and find significant differently correlated gene-metabolite pairs separately for the breeds (Duroc and Landrace) and FE groups (low and high FE) followed by a pathway over-representation analysis. We identified 21 and 12 significant gene-metabolite pairs for each group. The valine-leucine-isoleucine biosynthesis/degradation and arginine-proline metabolism pathways were associated with unique metabolites. The unique genes obtained from significant metabolite-gene pairs were associated with sphingolipid catabolism, multicellular organismal process, cGMP, and purine metabolic processes. While some of the genes and metabolites identified were known for their association with FE, others are novel and provide new avenues for further research. Further validation of genes, metabolites, and gene-metabolite interactions in larger cohorts will help us to elucidate the regulatory mechanisms and pathways underlying FE.
Publisher: Informa UK Limited
Date: 03-04-2015
Publisher: Elsevier BV
Date: 08-1992
DOI: 10.1016/0888-7543(92)90054-V
Abstract: A CA dinucleotide repeat polymorphism has been identified for the skeletal muscle alpha-actinin gene ACTN2. The observed heterozygosity is 44% (predicted heterozygosity 50%, PIC 0.47). This polymorphic marker has been localized between D1S74 and D1S103 on the multipoint linkage map of chromosome 1 at a position 44.4 cM from the most distal marker D1S68 at 1 qter.
Publisher: Springer International Publishing
Date: 2016
Publisher: JMIR Publications Inc.
Date: 28-06-2023
DOI: 10.2196/39895
Abstract: On February 25, 2022, Russian forces took control of the Chernobyl power plant after continuous fighting within the Chernobyl exclusion zone. Continual events occurred in the month of March, which raised the risk of potential contamination of previously uncontaminated areas and the potential for impacts on human and environmental health. The disruption of war has caused interruptions to normal preventive activities, and radiation monitoring sensors have been nonfunctional. Open-source intelligence can be informative when formal reporting and data are unavailable. This paper aimed to demonstrate the value of open-source intelligence in Ukraine to identify signals of potential radiological events of health significance during the Ukrainian conflict. Data were collected from search terminology for radiobiological events and acute radiation syndrome detection between February 1 and March 20, 2022, using 2 open-source intelligence (OSINT) systems, EPIWATCH and Epitweetr. Both EPIWATCH and Epitweetr identified signals of potential radiobiological events throughout Ukraine, particularly on March 4 in Kyiv, Bucha, and Chernobyl. Open-source data can provide valuable intelligence and early warning about potential radiation hazards in conditions of war, where formal reporting and mitigation may be lacking, to enable timely emergency and public health responses.
Publisher: Institute of Electrical and Electronics Engineers (IEEE)
Date: 02-2023
Publisher: Springer Science and Business Media LLC
Date: 12-2015
Publisher: MDPI AG
Date: 16-05-2020
DOI: 10.3390/LIFE10050068
Abstract: During the early stages of pregnancy, the uterine endometrium undergoes dramatic morphologic and functional changes accompanied with dynamic variation in gene expression. Pregnancy-stage specific differentially expressed gene (DEG)-transcript-probes were investigated and identified by comparing endometrium transcriptome at 9th day (9D), 12th day (12D) and 16th day (16D) of early pregnancy in Polish large-white (PLW) gilts. Endometrium comparisons between 9D-vs-12D, 9D-vs-16D and 12D-vs-16D of early pregnancy identified 6049, 374 and 6034 highly significant DEG-transcript-probes (p 0.001 FC). GO term enrichment analysis identified commonly shared upregulated endometrial DEG-transcript-probes (p 0.001 FC), that were regulating the gene functions of anatomic structure development and transport (TG), DNA-binding and methyltransferase activity (ZBTB2), ion-binding and kinase activity (CKM), cell proliferation and apoptosis activity (IL1B). Downregulated DEG-transcript-probes (p 0.001 FC) were involved in regulating the gene functions of phosphatase activity (PTPN11), TC616413 gene-transcript and Sus-scrofa LOC100525539. Moreover, blastn comparison of microarray-probes sequences against sus-scrofa11 assembly identified commonly shared upregulated endometrial DEG-transcript-probes (E 0.06 FC), that were regulating the gene functions of reproduction and growth (SELENOP), cytoskeleton organization and kinase activity (CDC42BPA), phosphatase activity (MINPP1), enzyme-binding and cell-population proliferation (VAV3), cancer-susceptibility candidate gene (CASC4), cytoskeletal protein-binding (COBLL1), ion-binding, enzyme regulator activity (ACAP2) Downregulated endometrial DEG-transcript-probes (E 0.06 FC) were involved in regulating the gene functions of signal-transduction (TMEM33), catabolic and metabolic processes (KLHL15). Microarray validation experiment on selected candidate genes showed complementarity to significant endometrial DEG-transcript-probes responsible for the regulation of immune response (IL1B, S100A11), lipid metabolism (FABP3, PPARG), cell-adhesion (ITGAV), angiogenesis (IL1B), intercellular transmission (NMB), cell-adhesion (OPN) and response to stimuli (RBP4) was confirmed by RT-PCR. This study provides a clue that identified pregnancy-stage specific microarray transcript probes could be considered as candidate genes for recognition and establishment of early pregnancy in the pig.
Publisher: Springer Science and Business Media LLC
Date: 21-09-2018
Publisher: Elsevier BV
Date: 2019
Publisher: MDPI AG
Date: 18-10-2020
DOI: 10.3390/IJMS21207705
Abstract: The COL7A1 gene encodes homotrimer fibrils essential for anchoring dermal and epidermal layers, and pathogenic mutations in COL7A1 can cause recessive or dominant dystrophic epidermolysis bullosa. As a monogenic disease gene, COL7A1 constitutes a potential target for antisense oligomer-mediated exon skipping, a therapy applicable to a growing number of other genetic disorders. However, certain characteristics of COL7A1: many exons, low average intron size, and repetitive and guanine-cytosine rich coding sequence, present challenges to the design of specific and effective antisense oligomers. While targeting COL7A1 exons 10 and 73 for excision from the mature mRNA, we discovered that antisense oligomers comprised of 2′-O-methyl modified bases on a phosphorothioate backbone and phosphorodiamidate morpholino oligomers produced similar, but distinctive, splicing patterns including excision of adjacent nontargeted exons and/or retention of nearby introns in some transcripts. We found that the nonsequential splicing of certain introns may alter pre-mRNA processing during antisense oligomer-mediated exon skipping and, therefore, additional studies are required to determine if the order of intron removal influences multiexon skipping and/or intron retention in processing of the COL7A1 pre-mRNA.
Publisher: IEEE
Date: 07-2010
Publisher: Springer Science and Business Media LLC
Date: 1995
DOI: 10.1038/NG0195-75
Abstract: Nemaline myopathies are diseases characterized by the presence in muscle fibres of pathognomonic rod bodies. These are composed largely of alpha-actinin and actin. We have identified a missense mutation in the alpha-tropomyosin gene, TPM3, which segregates completely with the disease in a family whose autosomal dominant nemaline myopathy we had previously localized to chromosome 1p13-q25. The mutation substitutes an arginine residue for a highly conserved methionine in a putative actin-binding site near the N terminus of the alpha-tropomyosin. The mutation may strengthen tropomyosin - actin binding, leading to rod body formation, by adding a further basic residue to the postulated actin-binding motif.
Publisher: Institute of Electrical and Electronics Engineers (IEEE)
Date: 2022
Publisher: Cold Spring Harbor Laboratory
Date: 05-2020
DOI: 10.1101/2020.05.01.071696
Abstract: Many long non-coding RNAs (lncRNA) are highly dysregulated in cancer and are emerging as therapeutic targets. One ex le is NEAT1, which consists of two overlapping lncRNA isoforms, NEAT1_1 (3.7kb) and NEAT1_2 (23kb), that are functionally distinct. The longer NEAT1_2 is responsible for scaffolding gene-regulatory nuclear bodies termed paraspeckles, whereas NEAT1_1 is involved in paraspeckle-independent function. The NEAT1 isoform ratio is dependent on the efficient cleavage and polyadenylation of NEAT1_1 at the expense of NEAT1_2. Here we developed a targeted antisense oligonucleotide (ASO) approach to sterically block NEAT1_1 polyadenylation processing, achieving upregulation of NEAT1_2 and abundant paraspeckles. We have applied these ASOs to cells of the heterogeneous infant cancer, neuroblastoma, as we found higher NEAT1_1:NEAT1_2 ratio and lack of paraspeckles in high-risk neuroblastoma cells. These ASOs decrease NEAT1_1 levels, increase NEAT1_2 araspeckles and concomitantly reduce cell viability in high-risk neuroblastoma specifically. In contrast, overexpression of NEAT1_1 has the opposite effect, increasing cell-proliferation. Transcriptomic analyses of high-risk neuroblastoma cells with altered NEAT1 ratios and increased paraspeckle abundance after ASO treatment showed an upregulation of differentiation pathways, as opposed to the usual aggressive neuroblastic phenotype. Thus, we have developed potential anti-cancer ASO drugs that can transiently increase growth-inhibiting NEAT1_2 RNA at the expense of growth-promoting NEAT1_1 RNA. These ASOs, unlike others that degrade lncRNAs, provide insights into the importance of altering lncRNA polyadenylation events to suppress tumorigenesis as a strategy to combat cancer.
Publisher: Wiley
Date: 14-11-2005
DOI: 10.1002/JGM.838
Abstract: Duchenne and Becker muscular dystrophies are allelic disorders arising from mutations in the dystrophin gene. Duchenne muscular dystrophy is characterised by an absence of functional protein, while Becker muscular dystrophy is usually caused by in-frame deletions allowing synthesis of some functional protein. Treatment options are limited, and we are investigating the potential of transcript manipulation to overcome disease-causing mutations. Antisense oligonucleotides have been used to induce specific exon removal during processing of the dystrophin primary transcript and thereby by-pass protein-truncating mutations. The antisense oligonucleotide chemistry most widely used to alter pre-mRNA processing is 2'-O-methyl-modified bases on a phosphorothioate backbone. The present studies evaluate 2'-O-methylphosphorothioate, peptide nucleic acid and morpholino antisense oligonucleotides in the mdx mouse model of muscular dystrophy, which has a nonsense mutation in exon 23 of the dystrophin gene. We demonstrate dystrophin expression in mdx mouse tissues after localised and systemic delivery of a morpholino antisense oligonucleotide designed to target the dystrophin exon 23 donor splice site. The stability of the morpholino structural type, and the fact that it can be delivered to muscle in the absence of a delivery reagent, render this compound eminently suitable for consideration for therapeutic exon skipping to address dystrophin mutations.
Publisher: Springer Science and Business Media LLC
Date: 25-10-2021
Publisher: Apollo - University of Cambridge Repository
Date: 2019
DOI: 10.17863/CAM.38701
Publisher: Frontiers Media SA
Date: 09-07-2014
Publisher: Elsevier BV
Date: 2016
Publisher: Springer Science and Business Media LLC
Date: 15-02-2017
Publisher: Informa UK Limited
Date: 02-08-2022
Publisher: Wiley
Date: 10-2000
Publisher: IEEE
Date: 11-2012
Publisher: Wiley
Date: 06-03-2015
DOI: 10.1111/JBG.12147
Abstract: Genome-wide association studies are routinely used to identify genomic regions associated with traits of interest. However, this ignores an important class of genomic associations, that of epistatic interactions. A genome-wide interaction analysis between single nucleotide polymorphisms (SNPs) using highly dense markers can detect epistatic interactions, but is a difficult task due to multiple testing and computational demand. However, It is important for revealing complex trait heredity. This study considers analytical methods that detect statistical interactions between pairs of loci. We investigated a three-stage modelling procedure: (i) a model without the SNP to estimate the variance components (ii) a model with the SNP using variance component estimates from (i), thus avoiding iteration and (iii) using the significant SNPs from (ii) for genome-wide epistasis analysis. We fitted these three-stage models to field data for growth and ultrasound measures for subcutaneous fat thickness in Brahman cattle. The study demonstrated the usefulness of modelling epistasis in the analysis of complex traits as it revealed extra sources of genetic variation and identified potential candidate genes affecting the concentration of insulin-like growth factor-1 and ultrasound scan measure of fat depth traits. Information about epistasis can add to our understanding of the complex genetic networks that form the fundamental basis of biological systems.
Publisher: Frontiers Media SA
Date: 26-03-2021
DOI: 10.3389/FNAGI.2021.658226
Abstract: There is a critical need to establish genetic markers that explain the complex phenotypes and pathogenicity of ALS. This study identified a polymorphism in the Stathmin-2 gene and investigated its association with sporadic ALS (sALS) disease risk, age-of onset and survival duration. The candidate CA repeat was systematically analyzed using PCR, Sanger sequencing and high throughput capillary separation for genotyping. Stathmin-2 expression was investigated using RT-PCR in patient olfactory neurosphere-derived (ONS) cells and RNA sequencing in laser-captured spinal motor neurons. In a case-control analysis of a combined North American sALS cohort ( n = 321) and population control group ( n = 332), long/long CA genotypes were significantly associated with disease risk ( p = 0.042), and most strongly when one allele was a 24 CA repeat ( p = 0.0023). In addition, longer CA allele length was associated with earlier age-of-onset ( p = 0.039), and shorter survival duration in bulbar-onset cases ( p = 0.006). In an Australian longitudinal sALS cohort ( n = 67), ALS functional rating scale scores were significantly lower in carriers of the long/long genotype ( p = 0.034). Stathmin-2 mRNA expression was reduced in sporadic patient ONS cells. Additionally, sALS patients and controls exhibited variable expression of Stathmin-2 mRNA according to CA genotype in laser-captured spinal motor neurons. We report a novel non-coding CA repeat in Stathmin-2 which is associated with sALS disease risk and has disease modifying effects. The potential value of this variant as a disease marker and tool for cohort enrichment in clinical trials warrants further investigation.
Publisher: Elsevier BV
Date: 12-1999
DOI: 10.1016/S0960-8966(99)00053-X
Abstract: The nemaline myopathies are muscle disorders of variable severity and age of onset, with characteristic nemaline bodies in the sarcoplasm. Genes for dominant (NEM1) and recessive (NEM2A) nemaline myopathy have been localised to chromosomes one and two, respectively. A missense mutation in the alpha-tropomyosin gene (TPM3) has been associated with NEM1 in one family. Probands from 76 other nemaline myopathy families have now been screened for TPM3 mutations. One proband, who was not noted to have any weakness neonatally, but who died at 21 months of age, was shown to be homozygous for a single strand conformation polymorphism (SSCP) in skeletal-muscle-specific exon 1 of TPM3. Sequencing revealed homozygosity for a nonsense mutation at codon 31 (CAG to TAG). The patient should have no functioning alpha-tropomyosin slow protein. The nemaline bodies in this patient were exclusively in type one fibres, consistent with the expression of TPM3 only in type one fibres.
Publisher: Springer Berlin Heidelberg
Date: 2006
DOI: 10.1007/978-3-540-34449-0_8
Abstract: Mutations in the dystrophin gene that prevent synthesis of a functional protein lead to Duchenne muscular dystrophy (DMD), the most common serious childhood muscular dystrophy. The major isoform is produced in skeletal muscle and the size of the dystrophin gene and complexity of expression have posed great challenges to the development of a therapy for DMD. Considerable progress has been made in the areas of gene and cell replacement, yet it appears that any potential therapy for DMD is still some years away. Other approaches are being considered, and one that has generated substantial interest over the last few years is induced exon skipping. Antisense oligonucleotides have been used to block abnormal splice sites and force pre-mRNA processing back to the normal patterns. This approach is re-interpreted to address the more common dystrophin mutations, where normal splice sites are targeted to induce abnormal splicing, resulting in specific exon exclusion. Selected exon removal during processing of the dystrophin pre-mRNA can by-pass nonsense mutations or restore a disrupted reading frame arising from genomic deletions or duplications. Attributes of the dystrophin gene that have h ered gene replacement therapy may be regarded as positive features for induced exon skipping, which may be regarded as a form of by-pass surgery at the molecular level. In humans, antisense oligonucleotides have been more generally applied to down-regulate specific gene expression, for the treatment of acquired conditions such as malignancies and viral infections. From interesting in vitro experiments several years ago, the dystrophin exon-skipping field has progressed to the stage of planning for clinical trials.
Publisher: F1000 Research Ltd
Date: 05-06-2018
DOI: 10.12688/F1000RESEARCH.15091.1
Abstract: Background: Inflammatory bowel disease (IBD) is a group of chronic diseases related to inflammatory processes in the digestive tract generally associated with an immune response to an altered gut microbiome in genetically predisposed subjects. For years, both researchers and clinicians have been reporting increased rates of anxiety and depression disorders in IBD, and these disorders have also been linked to an altered microbiome. However, the underlying pathophysiological mechanisms of comorbidity are poorly understood at the gut microbiome level. Methods: Metagenomic and metatranscriptomic data were retrieved from the Inflammatory Bowel Disease Multi-Omics Database. S les from 70 in iduals that had answered to a self-reported depression and anxiety questionnaire were selected and classified by their IBD diagnosis and their questionnaire results, creating six different groups. The cross-validation random forest algorithm was used in 90% of the in iduals (training set) to retain the most important species involved in discriminating the s les without losing predictive power. The validation set that represented the remaining 10% of the s les equally distributed across the six groups was used to train a random forest using only the species selected in order to evaluate their predictive power. Results: A total of 24 species were identified as the most informative in discriminating the 6 groups. Several of these species were frequently described in dysbiosis cases, such as species from the genus Bacteroides and Faecalibacterium prausnitzii . Despite the different compositions among the groups, no common patterns were found between s les classified as depressed. However, distinct taxonomic profiles within patients of IBD depending on their depression status were detected. Conclusions: The machine learning approach is a promising approach for investigating the role of microbiome in IBD and depression. Abundance and functional changes in these species suggest that depression should be considered as a factor in future research on IBD.
Publisher: Informa UK Limited
Date: 03-03-2016
Publisher: F1000 Research Ltd
Date: 17-04-2019
DOI: 10.12688/F1000RESEARCH.15091.2
Abstract: Background: Inflammatory bowel disease (IBD) is a group of chronic diseases related to inflammatory processes in the digestive tract generally associated with an immune response to an altered gut microbiome in genetically predisposed subjects. For years, both researchers and clinicians have been reporting increased rates of anxiety and depression disorders in IBD, and these disorders have also been linked to an altered microbiome. However, the underlying pathophysiological mechanisms of comorbidity are poorly understood at the gut microbiome level. Methods: Metagenomic and metatranscriptomic data were retrieved from the Inflammatory Bowel Disease Multi-Omics Database. S les from 70 in iduals that had answered to a self-reported depression and anxiety questionnaire were selected and classified by their IBD diagnosis and their questionnaire results, creating six different groups. The cross-validation random forest algorithm was used in 90% of the in iduals (training set) to retain the most important species involved in discriminating the s les without losing predictive power. The validation set that represented the remaining 10% of the s les equally distributed across the six groups was used to train a random forest using only the species selected in order to evaluate their predictive power. Results: A total of 24 species were identified as the most informative in discriminating the 6 groups. Several of these species were frequently described in dysbiosis cases, such as species from the genus Bacteroides and Faecalibacterium prausnitzii . Despite the different compositions among the groups, no common patterns were found between s les classified as depressed. However, distinct taxonomic profiles within patients of IBD depending on their depression status were detected. Conclusions: The machine learning approach is a promising approach for investigating the role of microbiome in IBD and depression. Abundance and functional changes in these species suggest that depression should be considered as a factor in future research on IBD.
Publisher: Cambridge University Press (CUP)
Date: 2017
DOI: 10.1017/S1752756200009182
Abstract: Adulthood of a dairy cow is reached when she is about 60 months old, but cows first calve at a much younger age. Therefore, most heifers are still growing during their first lactation and part of the energy and protein intake will be assigned to this purpose. As the cow grows closer to mature body weight and is able to use more energy and protein for the milk production, milk production will increase from 1 st to 2 nd to 3 rd lactation, but the amount of increase differs among cows. A gradual production increase is believed to be found in trouble-free cows (i.e. cows with good fertility and high disease resistance), and is a measure of the maturity rate (MR), where maturity is defined as reaching mature body weight. Genetic variance for this production increase exists (Krogmeier et al., 2003). The aim of this study was to define MR in a way that enables selection, and to estimate genetic correlations with functional traits.
Publisher: Springer Science and Business Media LLC
Date: 10-08-2014
DOI: 10.1038/NM.3628
Publisher: Informa UK Limited
Date: 29-08-2022
DOI: 10.1080/13816810.2021.1966053
Abstract: Stargardt disease (STGD1) is an autosomal recessive retinal dystrophy due to mutations in ABCA4, characterized by subretinal deposition of lipofuscin-like substances and bilateral centrifugal vision loss. Despite the tremendous progress made in the understanding of STGD1, there are no approved treatments to date. This review examines the challenges in the development of an effective STGD1 therapy. A literature review was performed through to June 2021 summarizing the spectrum of retinal phenotypes in STGD1, the molecular biology of ABCA4 protein, the in vivo and in vitro models used to investigate the mechanisms of ABCA4 mutations and current clinical trials. STGD1 phenotypic variability remains an challenge for clinical trial design and patient selection. Pre-clinical development of therapeutic options has been limited by the lack of animal models reflecting the erse phenotypic spectrum of STDG1. Patient-derived cell lines have facilitated the characterization of splice mutations but the clinical presentation is not always predicted by the effect of specific mutations on retinoid metabolism in cellular models. Current therapies primarily aim to delay vision loss whilst strategies to restore vision are less well developed. STGD1 therapy development can be accelerated by a deeper understanding of genotype-phenotype correlations.
Publisher: BMJ
Date: 08-1999
Abstract: To determine the molecular basis for autosomal dominant intermediate hereditary motor and sensory neuropathy (HMSN) in a four generation family. The gene defects in families with intermediate HMSN are not known, but it has been suggested that most have X linked HMSN. All participating family members were examined clinically. Genomic DNA was obtained from 10 affected and seven unaffected members. Linkage analysis for the known HMSN loci was first performed. Mutations in the peripheral myelin protein zero gene (PMP0) were sought in two affected members, using one unaffected member for comparison, by lification of the six exons of the gene followed by single strand conformation polymorphism (SSCP) analysis, dideoxy fingerprinting (ddF), and sequencing. Subsequently, the mutation was screened for in all affected and unaffected members in the family using Alu I digestion and in 100 unrelated control subjects using "snap back" SSCP analysis. Sequencing of cDNA from a sural nerve biopsy from an affected member was also performed. The clinical phenotype was of variable severity, with motor nerve conduction velocities in the intermediate range. Linkage to PMP0 was demonstrated. Analysis of genomic DNA and cDNA for PMP0 identified a novel codon 35 GAC to TAC mutation. The mutation produces an inferred amino acid change of aspartate to tyrosine at codon six of the processed protein (Asp6Tyr) in the extracellular domain and was present in all affected family members but not in 100 unrelated controls. The present findings further extend the range of phenotypes associated with PMP0 mutations and indicate that families with "intermediate" HMSN need not necessarily be X-linked as previously suggested.
Publisher: Oxford University Press (OUP)
Date: 21-07-2003
DOI: 10.1093/HMG/DDG196
Abstract: The mdx mouse model of muscular dystrophy arose due to a nonsense mutation in exon 23 of the dystrophin gene. We have previously demonstrated that 2'-O-methyl phosphorothioate antisense oligonucleotides (AOs) can induce removal of exon 23 during processing of the primary transcript. This results in an in-frame mRNA transcript and subsequent expression of a slightly shorter dystrophin protein in mdx muscle. Refinement of AO design has allowed efficient exon skipping to be induced in mdx mouse muscle cultures at nanomolar concentrations. In contrast, splicing intervention by morpholino AOs has been applied to the beta-globin gene pre-mRNA in cultured cells to correct aberrant splicing when delivered in the micromolar range. The morpholino chemistry produces a neutral molecule that has exceptional biological stability but poor cellular delivery. We present data showing that exon skipping in mdx cells may be induced by morpholino AOs at nanomolar concentrations when annealed to a sense oligonucleotide or "leash", and delivered as a cationic lipoplex. We have investigated a number of leash designs and chemistries, including mixed backbone oligonucleotides, and their ability to influence delivery and efficacy of the morpholino AO. Significantly, we detected dystrophin protein synthesis and correct sarcolemmal localisation after intramuscular injection of morpholino AO : leash lipoplexes in mdx muscle in vivo. We show enhanced delivery of a morpholino AO, enabling the advantageous properties to be exploited for potentially therapeutic outcomes.
Publisher: Bentham Science Publishers Ltd.
Date: 10-2005
DOI: 10.2174/156652305774329249
Abstract: Antisense oligonucleotides initially offered great hope as specific compounds to modify gene expression, primarily through RNaseH induced degradation of the target transcript. Expansion of the field led to new chemistries capable of invoking different mechanisms, including suppression of protein synthesis by translational blockade, and there is now a major interest in downregulation of gene expression using short interfering RNAs to induce RNA silencing. Naturally occurring microRNAs have been implicated in the regulation of gene expression. This review considers ex les of antisense oligonucleotides redirecting the process of exon recognition and intron removal during gene transcript splicing. While suppression of gene expression is necessary to address some conditions, it appears likely that there may be many more clinical applications for antisense oligonucleotides in re-directing splicing patterns. Pre-mRNA splicing is a tightly co-ordinated, multifactorial process, which can be disrupted by antisense oligonucleotides in a highly specific manner, allowing either suppression of aberrant splicing, by-pass of nonsense or frame-shifting mutations or alteration of spliceoform ratios. Manipulation of splicing patterns has been applied to a erse range of conditions, including beta-thalassemia, Duchenne muscular dystrophy, spinal muscular atrophy and certain cancers. Alternative exon usage has been identified as a major mechanism for generating ersity from a limited repertoire of genes in higher eukaryotes. Considering that up to 75% of all human primary gene transcripts are reported to be alternatively spliced, intervention at the level of pre-mRNA processing is likely to become increasingly significant in the fight against genetic and acquired disorders.
Publisher: Elsevier BV
Date: 10-2017
Publisher: Elsevier BV
Date: 02-2023
Publisher: Public Library of Science (PLoS)
Date: 13-10-2010
Publisher: Springer Science and Business Media LLC
Date: 09-11-2015
Abstract: NUT midline carcinoma (NMC) is a fatal cancer that arises in various tissues along the upper midline of the body. The defining molecular feature of NMC is a chromosomal translocation that joins (in the majority of cases) the nuclear testis gene NUT ( NUTM1 ) to the bromodomain protein family member 4 ( BRD4 ) and thereby creating a fusion oncogene that disrupts cellular differentiation and drives the disease. In this study, we report the case of an adolescent NMC patient presenting with severe facial pain, proptosis and visual impairment due to a mass arising from the ethmoid sinus that invaded the right orbit and frontal lobe. Treatment involved radical resection, including exenteration of the affected eye with the view to consolidate treatment with radiation therapy however, the patient experienced rapid tumor progression and passed away 79 days post resection. Molecular analysis of the tumor tissue identified a novel in-frame BRD4-NUT transcript, with BRD4 exon 15 fused to the last 124 nucleotides of NUT exon 2 ( BRD4-NUT ex15:ex2 Δnt1–585 ). The partial deletion of NUT exon 2 was attributed to a mid-exonic genomic breakpoint and the subsequent activation of a cryptic splice site further downstream within the exon. Inhibition of the canonical 3′ acceptor splice site of NUT intron 1 in cell lines expressing the most common NMC fusion transcripts (PER-403, BRD4-NUT ex11:ex2 PER-624, BRD4-NUT ex15:ex2) induced alternative splicing from the same cryptic splice site as identified in the patient. Detection of low levels of an in-frame BRD4-NUT ex11:ex2 Δnt1–585 transcript in PER-403 confirmed endogenous splicing from this alternative exon 2 splice site. Although further studies are necessary to assess the clinical relevance of the increasing number of variant fusions described in NMC, the findings presented in this case identify alternative splicing as a mechanism that contributes to this pathogenic complexity.
Publisher: Springer Science and Business Media LLC
Date: 24-01-2019
Publisher: Informa UK Limited
Date: 29-09-2016
Publisher: Cambridge University Press (CUP)
Date: 2017
DOI: 10.1017/S1752756200009625
Abstract: Osteochondrosis (OC) in pigs is an abnormal condition in the bone development which has high economic and welfare implications. Select for OC resistance would be difficult to achieve due to low polygenic heritability (∼0.06 – 0.15 Kadarmideen et al. , 2004), unless a major gene exists. Segregation analysis provides evidence for major gene using only phenotypic data. Bayesian Segregation Analysis (BSA) method via Gibbs s ling is suitable for pedigreed animal population as shown in the application of this methodology to milk flow in dairy cattle (Ilahi and Kadarmideen, 2004). The main objective of this study was to conduct a BSA to determine whether osteochondral disease in pigs has mixed inheritance (major + polygenes).
Publisher: Public Library of Science (PLoS)
Date: 13-02-2018
Publisher: Wiley
Date: 2002
DOI: 10.1002/JGM.295
Abstract: Duchenne muscular dystrophy (DMD) is a fatal genetic disorder caused by dystrophin gene mutations that preclude synthesis of a functional protein. One potential treatment of the disorder has utilised antisense oligoribonucleotides (AOs) to induce removal of disease-associated exons during pre-mRNA processing. Induced in-frame mRNA transcripts encode a shorter but functional dystrophin. We have investigated and improved the design of AOs capable of removing exon 23, and thus the disease-causing nonsense mutation, from mRNA in the mdx mouse model of DMD. H-2K(b)-tsA58 mdx cultures were transfected with complexes of Lipofectin and AOs. Exon skipping was detected by RT-PCR and subsequent protein production was demonstrated by Western blotting. AOs were delivered at a range of doses in order to compare relative efficiencies. We describe effective and reproducible exon 23 skipping with several AOs, including one as small as 17 nucleotides. Furthermore, the location of a sensitive exon 23 target site has been refined, whilst minimum effective doses have been estimated in vitro. These doses are significantly lower than previously reported and were associated with the synthesis of dystrophin protein in vitro. These results demonstrate the increasing feasibility of an AO-based therapy for treatment of DMD. By refining AO design we have been able to reduce the size and the effective dose of the AOs and have dramatically improved the efficiency of the technique.
Publisher: Elsevier BV
Date: 09-2019
Publisher: Springer Science and Business Media LLC
Date: 31-05-2018
Publisher: Bentham Science Publishers Ltd.
Date: 27-03-2018
Publisher: Cambridge University Press (CUP)
Date: 2018
DOI: 10.1017/S1463981500040577
Abstract: A combination of better management and genetic selection for good health and fertility would provide a more effective long term solution for economic loss due to diseases and poor fertility. This would also help to address public concerns about the use of medical treatment in milk production. A balance in the genetic improvement of health and fertility together with milk production could be achieved through their inclusion in national genetic selection indices, for which genetic parameters are needed. One of the main objectives of this study was to estimate genetic parameters for various disease and fertility traits in the UK dairy cattle population, using records from a national recording scheme run by Livestock Services UK Ltd. Genetic analysis of traits recorded as present or absent (binary traits e.g. diseases) requires the use of non-linear threshold models, because linear models require normality assumptions (e.g., Gianola 1982). The other objective of this study was to estimate genetic parameters for binary disease and fertility traits based on threshold animal models and to compare results with those from linear animal models.
Publisher: Springer Science and Business Media LLC
Date: 24-05-2006
Abstract: Duchenne muscular dystrophy is a fatal genetic disorder caused by dystrophin gene mutations that result in premature termination of translation and the absence of functional protein. Despite the primary dystrophin gene lesion, immunostaining studies have shown that at least 50% of DMD patients, mdx mice and a canine model of DMD have rare dystrophin-positive or 'revertant' fibres. Fine epitope mapping has shown that the majority of transcripts responsible for revertant fibres exclude multiple exons, one of which includes the dystrophin mutation. The mdx mouse model of muscular dystrophy has a nonsense mutation in exon 23 of the dystrophin gene. We have shown that antisense oligonucleotides (AOs) can induce the removal of this exon, resulting in an in-frame mRNA transcript encoding a shortened but functional dystrophin protein. To emulate one exonic combination associated with revertant fibres, we target multiple exons for removal by the application of a group of AOs combined as a "cocktail". Exons 19–25 were consistently excluded from the dystrophin gene transcript using a cocktail of AOs. This corresponds to an alternatively processed gene transcript that has been sporadically detected in untreated dystrophic mouse muscle, and is presumed to give rise to a revertant dystrophin isoform. The transcript and the resultant correctly localised smaller protein were confirmed by RT-PCR, immunohistochemistry and western blot analysis. This work demonstrates the feasibility of AO cocktails to by-pass dystrophin mutation hotspots through multi-exon skipping. Multi-exon skipping could be important in expediting an exon skipping therapy to treat DMD, so that the same AO formulations may be applied to several different mutations within particular domains of the dystrophin gene.
Publisher: Springer Berlin Heidelberg
Date: 2012
Publisher: Cold Spring Harbor Laboratory
Date: 13-04-2020
DOI: 10.1101/2020.04.11.036939
Abstract: Metabolites represent the ultimate response of biological systems, so metabolomics is considered to link the genotypes and phenotypes. Feed efficiency is one of the most important phenotypes in sustainable pig production and is the main breeding goal trait. We utilized metabolic and genomic datasets from a total of 108 pigs from our own previously published studies that involved 59 Duroc and 49 Landrace pigs with data on feed efficiency (residual feed intake or RFI), genotype (PorcineSNP80 BeadChip) data and metabolomic data (45 final metabolite datasets derived from LC-MS system). Utilizing these datasets, our main aim was to identify genetic variants (single-nucleotide polymorphisms or SNPs) that affect 45 different metabolite concentrations in plasma collected at the start and end of the performance testing of pigs categorized as high or low in their feed efficiency (based on RFI values). Genome-wide significant genetic variants could be then used as potential genetic or biomarkers in breeding programs for feed efficiency. The other objective was to reveal the biochemical mechanisms underlying genetic variations for pigs’ feed efficiency. In order to achieve these objectives, we firstly conducted a metabolite genome-wide association study (mGWAS) based on mixed linear models and found 152 genome-wide significant SNPs ( P -value 1.06E-06) in association with 17 metabolites that included 90 significant SNPs annotated to 52 genes. On chromosome one alone, 51 significant SNPs associated with isovalerylcarnitine and propionylcarnitine were found to be in strong linkage disequilibrium (LD). SNPs in strong LD annotated to FBXL4 and CCNC consisted of two haplotype blocks where three SNPs (ALGA0004000, ALGA0004041 and ALGA0004042) were in the intron regions of FBXL4 and CCNC . The interaction network revealed that CCNC and FBXL4 were linked by the hub gene N6AMT1 that was associated with isovalerylcarnitine and propionylcarnitine. Moreover, three metabolites (i.e., isovalerylcarnitine, propionylcarnitine and pyruvic acid) were clustered in one group based on the low-high RFI pigs. This study performed a comprehensive metabolite-based GWAS analysis for pigs with differences in feed efficiency and provided significant metabolites for which there is a significant genetic variation as well as biological interaction networks. The identified metabolite genetic variants, genes and networks in high versus low feed efficient pigs could be considered as potential genetic or biomarkers for feed efficiency.
Publisher: Springer Science and Business Media LLC
Date: 15-04-2009
Publisher: Elsevier BV
Date: 02-2013
DOI: 10.1038/MT.2013.4
Publisher: Springer Science and Business Media LLC
Date: 20-10-2011
Abstract: Antisense oligomer induced exon skipping aims to reduce the severity of Duchenne muscular dystrophy by redirecting splicing during pre-RNA processing such that the causative mutation is by-passed and a shorter but partially functional Becker muscular dystrophy-like dystrophin isoform is produced. Normal exons are generally targeted to restore the dystrophin reading frame however, an appreciable subset of dystrophin mutations are intra-exonic and therefore have the potential to compromise oligomer efficiency, necessitating personalised oligomer design for some patients. Although antisense oligomers are easily personalised, it remains unclear whether all patient polymorphisms within antisense oligomer target sequences will require the costly process of producing and validating patient specific compounds. Here we report preclinical testing of a panel of splice switching antisense oligomers, designed to excise exon 25 from the dystrophin transcript, in normal and dystrophic patient cells. These patient cells harbour a single base insertion in exon 25 that lies within the target sequence of an oligomer shown to be effective at removing exon 25. It was anticipated that such a mutation would compromise oligomer binding and efficiency. However, we show that, despite the mismatch an oligomer, designed and optimised to excise exon 25 from the normal dystrophin mRNA, removes the mutated exon 25 more efficiently than the mutation-specific oligomer. This raises the possibility that mismatched AOs could still be therapeutically applicable in some cases, negating the necessity to produce patient-specific compounds.
Publisher: Springer Science and Business Media LLC
Date: 14-11-2006
DOI: 10.1007/S00439-006-0285-Z
Abstract: Quantitative phenotypes correlated with a complex disorder offer increased power to detect linkage in comparison to affected-unaffected classifications. Asthma is a complex disorder characterized by periods of bronchial obstruction and increased bronchial hyper reactivity. In childhood and early adulthood, asthma is frequently associated also with quantitative measures of atopy. Genome wide quantitative multipoint linkage analysis was conducted for serum IgE levels and percentage of positive skin prick test (SPT(per)) using three large groups of families originally ascertained for asthma. In this report, 438 and 429 asthma families were informative for linkage using IgE and SPT(per) which represents 690 independent families. Suggestive linkage (LOD > or = 2) was found on chromosomes 1, 3, and 8q with maximum LODs of 2.34 (IgE), 2.03 (SPT(per)), and 2.25 (IgE) near markers D1S1653, D3S2322-D3S1764, and D8S2324, respectively. The results from chromosomes 1 and 3 replicate previous reports of linkage. We also replicate linkage to 5q with peak LODs of 1.96 (SPT(per)) and 1.77 (IgE) at or near marker D5S1480. Our results provide further evidence implicating chromosomes 1, 3, and 5q. The current report represents one of the biggest genome scans so far reported for asthma related phenotypes. This study also demonstrates the utility of increased s le sizes and quantitative phenotypes in linkage analysis of complex disorders.
Publisher: MDPI AG
Date: 19-07-2023
DOI: 10.3390/IJGI12070288
Abstract: Unlike car navigation, where almost all vehicles can traverse every route, one route might not be optimal or even suitable for all pedestrians. Route geometry information, including tortuosity, twists and turns along roads, junctions, and road slopes, among others, matters a great deal for specific types of pedestrians, particularly those with limited mobility, such as wheelchair users and older adults. Offering practical routing services to these users requires that pedestrian navigation systems provide further information on route geometry. Therefore, this article proposes a novel method for extracting and analyzing the geometry properties of the shortest pedestrian paths, with a focus on open geospatial data across four aspects: (a) similarity, (b) route curviness, (c) road turns and intersections, and (d) road gradients. Deriving from the Hausdorff distance, a metric called the “dissimilarity ratio” was developed, allowing us to determine whether pairs of routes show any tendencies to be similar to each other. Using the “sinuosity index”, a segment-based technique quantified the route curviness based on the number and degree of the road turns along the route. Moreover, relying upon open elevation data, the road gradients were extracted to identify routes offering smoother motion and better accessibility. Lastly, the road turns and intersections were investigated as pedestrian convenience and safety indicators. A local government area of Greater Sydney in Australia was chosen as the case study. The analysis was implemented on OpenStreetMap (OSM) shortest pedestrian paths against Google Maps as a benchmark for real-world commercial applications. The similarity analysis indicated that over 90% of OSM routes were identical or roughly similar to Google Maps. In addition, while Spearman’s rank correlation showed a direct relationship between route curviness and route length, rS(758) = 0.92, p 0.001, OSM, on average, witnessed more tortuous routes and, consequently, shorter straight roads between turns. However, OSM routes could be more suitable for pedestrians when the frequency of intersections and road slopes are at the center of attention. Finally, the devised metrics in this study, including the dissimilarity ratio and sinuosity index, showed their practicability in translating raw values into meaningful indicators.
Publisher: Wiley
Date: 03-04-2014
DOI: 10.1002/HBM.22522
Publisher: Informa UK Limited
Date: 03-2011
DOI: 10.2147/TACG.S8762
Publisher: Springer International Publishing
Date: 2016
Publisher: American Society for Microbiology
Date: 1992
DOI: 10.1128/JCM.30.1.255-258.1992
Abstract: A DNA lification assay using the polymerase chain reaction technique designed for the rapid identification of Mycobacterium bovis organisms was used to test 211 human mycobacterial isolates and 177 clinical specimens previously submitted for routine mycobacterial culture. The procedures described could be used by routine or specialist laboratories for identification of M. tuberculosis complex organisms in 4 h and/or as a rapid screening method for the direct detection of M. tuberculosis complex organisms in specimens.
Publisher: IEEE
Date: 08-2011
Publisher: Wiley
Date: 13-09-2019
DOI: 10.1111/AGE.12842
Abstract: Boar taint (BT) is an offensive flavor observed in non-castrated male pigs that reduces the carcass price. Surgical castration effectively avoids the taint but is associated with animal welfare concerns. The functional annotation of farm animal genomes for understanding the biology of complex traits can be used in the selection of breeding animals to achieve favorable phenotypic outcomes. The characterization of pig epigenomes/methylation changes between animals with high and low BT and genome-wide epigenetic markers that can predict BT are lacking. Reduced representation bisulfite sequencing of DNA methylation patterns based on next-generation sequencing is an efficient technology to identify candidate epigenetic biomarkers associated with BT. Three different BT levels were analyzed using reduced representation bisulfite sequencing data to calculate the methylation levels of cytosine and guanine dinucleotide (CpG) sites. The co-analysis of differentially methylated CpG sites identified by this study and differentially expressed genes identified by a previous study found 32 significant co-located genes. The joint analysis of GO terms and pathways revealed that methylation and gene expression of seven candidate genes were associated with BT in particular, FASN plays a key role in fatty acid biosynthesis, and PEMT might be involved in estrogen regulation and the development of BT. This study is the first to report the genome-wide DNA methylation profiles of BT in pigs using next-generation sequencing and summarize candidate genes associated with epigenetic markers of BT, which could contribute to the understanding of the functional biology of BT traits and selective breeding of pigs against BT based on epigenetic biomarkers.
Publisher: Oxford University Press (OUP)
Date: 20-12-2011
DOI: 10.1093/HMG/DDR600
Publisher: Public Library of Science (PLoS)
Date: 27-12-2011
Publisher: Informa UK Limited
Date: 19-04-2016
Publisher: Springer Science and Business Media LLC
Date: 04-2015
DOI: 10.1038/NM0415-414B
Publisher: Springer International Publishing
Date: 2022
Publisher: MDPI AG
Date: 28-01-2020
Abstract: Pompe disease, or glycogen storage disease II is a rare, progressive disease leading to skeletal muscle weakness due to deficiency of the acid α-1,4-glucosidase enzyme (GAA). The severity of disease and observed time of onset is subject to the various combinations of heterozygous GAA alleles. Here we have characterized two novel mutations: c.2074C T and c.1910_1918del, and a previously reported c.1082C G mutation of uncertain clinical significance. These mutations were found in three unrelated patients with adult-onset Pompe disease carrying the common c.-32-13T G mutation. The c.2074 C T nonsense mutation has obvious consequences on GAA expression but the c.1910_1918del (deletion of 3 amino acids) and c.1082C G missense variants are more subtle DNA changes with catastrophic consequences on GAA activity. Molecular and clinical analyses from the three patients corresponded with the anticipated pathogenicity of each mutation.
Location: Australia
Location: Australia
Start Date: 2015
End Date: 12-2015
Amount: $670,000.00
Funder: Australian Research Council
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