ORCID Profile
0000-0002-3462-7501
Current Organisation
University of Western Australia
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Research Topics
In Research Link Australia (RLA), "Research Topics" refer to ANZSRC FOR and SEO codes. These topics are either sourced from ANZSRC FOR and SEO codes listed in researchers' related grants or generated by a large language model (LLM) based on their publications.
Top 5 Research Topics
ANZSRC Field of Research (FoR)
Marine and Estuarine Ecology (incl. Marine Ichthyology) | Animal Neurobiology | Physiology | Crop and Pasture Improvement (Selection and Breeding) | Plant Physiology | Animal Physiology—Cell | Zoology | Parasitology
ANZSRC Socio-Economic Objective (SEO)
Winter Grains and Oilseeds not elsewhere classified | Animal Production and Animal Primary Products not elsewhere classified | Marine Flora, Fauna and Biodiversity | Biological sciences | Scientific instrumentation |
Related Links
Publications
Enteral branched‐chain amino acids increase the specific activity of jejunal glutaminase and reduce jejunal atrophy
Publisher: Wiley
Date: 06-1997
DOI: 10.1111/J.1440-1746.1997.TB00461.X
Abstract: Branched-chain amino acid (BCAA)-enriched nutrient solutions reduce gut atrophy associated with parenteral nutrition. We hypothesized that this effect was mediated by phosphate-dependent glutaminase. Thirty male Wistar rats (300-350 g) underwent a standardized surgical procedure and were then randomized into three groups to receive 6 days of ad libitum enteral nutrition. The animals were fed a solution of conventional nutrients, a solution of conventional nutrients enriched with 2.0% BCAA or a solution of conventional parenteral nutrients enriched with 2.5% glutamine. When compared with rats fed conventional nutrients, rats fed BCAA and glutamine had less jejunal atrophy (P < 0.05) and a greater specific activity of phosphate-dependent glutaminase in the jejunum (131% P < 0.05). It is concluded that enteral BCAA reduce atrophy of the jejunum via the generation of glutamine.
The role of glutaminase in the small intestine
Publisher: Elsevier BV
Date: 03-1999
DOI: 10.1016/S1357-2725(98)00121-6
Abstract: Glutaminase is the enzyme which hydrolyses glutamine, the main respiratory fuel of the intestine, to yield glutamate and ammonia. Glutaminase has a central role in intestinal metabolism: the products of the reaction catalyzed by glutaminase can be transaminated, catabolized to yield energy or used for the biosynthesis of pyrimidine nucleotides. Experimental treatments which deprive the intestine of glutamine induce intestinal atrophy. In this review, attention is paid to the role of glutaminase in intestinal metabolism. Background information on the structure, kinetics and distribution of glutaminase precede a discussion of the metabolism of glutamine within the intestine. In closing, we review the factors known to regulate glutaminase activity and emphasise that the regulation of glutaminase within the intestine is poorly understood.
Analysis of human chromosomes by imaging flow cytometry
Publisher: Wiley
Date: 25-05-2021
DOI: 10.1002/CYTO.B.22023
Abstract: Chromosomal analysis is traditionally performed by karyotyping on metaphase spreads, or by fluorescent in situ hybridization (FISH) on interphase cells or metaphase spreads. Flow cytometry was introduced as a new method to analyze chromosomes number (ploidy) and structure (telomere length) in the 1970s with data interpretation largely based on fluorescence intensity. This technology has had little uptake for human cytogenetic applications primarily due to analytical challenges. The introduction of imaging flow cytometry, with the addition of digital images to standard multi‐parametric flow cytometry quantitative tools, has added a new dimension. The ability to visualize the chromosomes and FISH signals overcomes the inherent difficulties when the data is restricted to fluorescence intensity. This field is now moving forward with methods being developed to assess chromosome number and structure in whole cells (normal and malignant) in suspension. A recent advance has been the inclusion of immunophenotyping such that antigen expression can be used to identify specific cells of interest for specific chromosomes and their abnormalities. This capability has been illustrated in blood cancers, such as chronic lymphocytic leukemia and plasma cell myeloma. The high sensitivity and specificity achievable highlights the potential imaging flow cytometry has for cytogenomic applications (i.e., diagnosis and disease monitoring). This review introduces and describes the development, current status, and applications of imaging flow cytometry for chromosomal analysis of human chromosomes.
A standardised protocol for the evaluation of small extracellular vesicles in plasma by imaging flow cytometry
Publisher: Elsevier BV
Date: 05-2019
Dysregulation of the intrinsic apoptotic pathway mediates megakaryocytic hyperplasia in myeloproliferative neoplasms
Publisher: BMJ
Date: 08-04-2016
Optimising multiplex immunofluorescence staining for characterising the tumour immune micro-environment
Publisher: Elsevier BV
Date: 11-2023
Mutational Analysis of BRAF Inhibitor–Associated Squamoproliferative Lesions
Publisher: Elsevier BV
Date: 11-2015
DOI: 10.1016/J.JMOLDX.2015.05.009
Abstract: In recent years, there has been increasing use of BRAF-inhibiting drugs for the treatment of various malignancies, including melanoma. However, these agents are associated with the development of other nonmelanoma skin lesions, in particular squamoproliferative lesions such as keratoacanthomas (KAs), squamous cell carcinomas, and BRAF inhibitor-associated verrucous keratoses. The molecular pathogenesis of these lesions is of interest, not only for therapeutic reasons, but also for the insight it might provide into the development of similar lesions in a sporadic setting. We used next-generation sequencing to compare the mutational profiles of lesions after treatment with a BRAF inhibitor, with similar lesions arising sporadically. HRAS mutations were common among the BRAF inhibitor-induced lesions, being identified in 56%, compared with 14% of lesions in the sporadic group (P = 0.002). Thus, despite similar histomorphological appearances, the underlying molecular mechanisms may be different. In addition, within the BRAF inhibitor-associated group, the lesions designated as KAs and BRAF inhibitor-associated verrucous keratoses had a similar mutational profile (mutations in PIK3CA, APC, and HRAS), which was distinct to that seen in squamous cell carcinomas (FGFR3, CDKN2A, and STK11). We have previously noted histological overlap between KAs and BRAF inhibitor-associated verrucous keratoses, and this finding supports the notion that they may represent morphological or temporal variants of a single lesion type.
The pathobiology of peritonitis
Publisher: Elsevier BV
Date: 1998
DOI: 10.1016/S0016-5085(98)70646-8
Abstract: The peritoneum is more than a mechanical covering that allows for the easy gliding of opposed peritoneal surfaces. The peritoneal mesothelial cells facilitate the action of powerful innate immune mechanisms. In addition, the peritoneal-associated lymphoid tissues contain unique cells that may play a crucial role in the localization of intraperitoneal infection. A clearer understanding of the molecular and cellular events underlying peritoneal functions in both the unstimulated and stimulated state will aid future treatment of peritonitis.
The Influence of Ischemia/Reperfusion Injury on the Jejunum
Publisher: Ovid Technologies (Wolters Kluwer Health)
Date: 06-1998
DOI: 10.1097/00000637-199806000-00008
Abstract: Ischemia/reperfusion injury (IRI) after free tissue transfer of the small intestine results in transmural tissue damage. This study examined the effects of IRI on the jejunum. Wistar rats served either as controls (N=10) or underwent cl ing of the infrarenal aorta for 1 hour followed by 1 hour of reperfusion (N=10). Both ischemia and reperfusion reduced the protein and deoxyribonucleic acid content of the jejunal mucosa (p < 0.05). Myeloperoxidase activity in the jejunal mucosa remained relatively low. The expression of leukocyte function-associated antigen 1 and intercellular adhesion molecule 1 (ICAM-1) on the surface of mucosal cells was not altered significantly by the ischemic insult, but was reduced after the period of reperfusion (p < 0.05). This coincided with an increase in messenger ribonucleic acid (mRNA) for ICAM-1 within isolated mucosal cells (p < 0.05). The specific activity of glutaminase in isolated jejunal mucosal cells was diminished after ischemia and reperfusion (p < 0.05), and this was not associated with an appreciable change in glutaminase mRNA expression. These results have identified some molecular mechanisms underlying IRI of the small intestine that are possible candidates for therapeutic intervention.
Quantitation of glutaminase mRNA in enterocytes using competitive RT-PCR
Publisher: Elsevier BV
Date: 10-1998
Abstract: The concentration of mRNA within the intestinal mucosa is usually measured by either Northern blot analysis or semi-quantitative reverse transcription polymerase chain reaction (RT-PCR). However, these methods are limited by a lack of valid internal controls, low sensitivity, and large differences in the concentration of the internal control and target gene. The authors present an alternative method using competitive RT-PCR to measure glutaminase mRNA in isolated enterocytes.
Influence of ischaemia–reperfusion injury on CD44 expression in rat small intestine
Publisher: Oxford University Press (OUP)
Date: 08-1998
DOI: 10.1046/J.1365-2168.1998.00826.X
Abstract: CD44 is an adhesion molecule expressed by neutrophils and lymphocytes which is involved in cell–cell and cell–matrix binding. In this study, the effect of ischaemia–reperfusion injury on CD44 messenger RNA (mRNA) and cell surface immunohistochemical expression of CD44 in the rat small intestine was evaluated. Wistar rats (n = 16) were randomized to either serve as controls (sham surgery) or to be subjected to a standardized ischaemia–reperfusion injury (suprarenal aorta occluded for 1 h followed by 1 h of reperfusion). Standardized segments of jejunum were harvested after ischaemia–reperfusion injury (ischaemic and reperfused s les) to measure the mucosal protein and DNA content, mRNA expression of CD44 and the immunohistochemical expression of CD44. Reperfusion significantly damaged the jejunal mucosa, e.g. mucosal protein content was lower after reperfusion compared with that in the control group (z = −2·31, P = 0·02) and the ischaemic s les (z = −2·52, P = 0·01). The expression of cell surface CD44 protein was also significantly decreased after ischaemic injury (z = −1·99, P = 0·04) this coincided with a decrease in the amount of cytoplasmic CD44 mRNA within isolated enterocytes (z = −2·31, P = 0·02). Ischaemia–reperfusion injury decreases the expression of CD44 within the jejunal mucosa. This may contribute to the failure of the gut barrier after such injury.
Enhanced multi-FISH analysis of immunophenotyped plasma cells by imaging flow cytometry
Publisher: Springer Science and Business Media LLC
Date: 10-05-2023
Peritoneal defences and peritoneum-associated lymphoid tissue
Publisher: Oxford University Press (OUP)
Date: 08-1996
Abstract: The peritoneum is mainly protected by the innate immune system. This consists of mechanical clearance of the peritoneal cavity, activation of complement, and the actions of polymorphonuclear neutrophils and macrophages. The specific immune system, which is mediated by the activity of lymphocytes, provides a secondary lification system that may be of great importance for patients with intraperitoneal sepsis. This review provides an overview of the relevant innate immune mechanisms and explores the possible role of peritoneum-associated lymphoid tissue.
Platelets in myeloproliferative neoplasms have a distinct transcript signature in the presence of marrow fibrosis
Publisher: Wiley
Date: 19-08-2020
DOI: 10.1111/BJH.16152
Chimeric antigen receptor T-cells
Publisher: Wiley
Date: 20-06-2023
DOI: 10.1111/IJLH.14121
Abstract: Chimeric antigen receptor (CAR) T‐cell therapy is a novel adoptive T‐cell immunotherapy for haematological malignancies. First introduced into clinical practice in 2017, CAR T‐cell therapy is now finding its place in the management of lymphoid malignancies, primarily of B‐cell lineage, including lymphoblastic leukaemia, non‐Hodgkin lymphoma and plasma cell myeloma, with remarkable therapeutic outcomes. CAR T‐cells are a customised therapeutic product for each patient. Manufacture commences with collection of autologous T‐cells, which are then genetically engineered ex vivo to express transmembrane CARs. These chimeric proteins consist of an antibody‐like extracellular antigen‐binding domain, to recognise specific antigens on the surface of tumour cells (e.g. CD19), linked to the intracellular co‐stimulatory signalling domains of a T‐cell receptor (e.g. CD137). The latter is required for in vivo CAR T‐cell proliferation, survival, and durable efficacy. Following reinfusion, CAR T‐cells harness the cytotoxic capacity of a patient's immune system. They overcome major mechanisms of tumour immuno‐evasion and have potential to generate robust cytotoxic anti‐tumour responses. This review discusses the background to CAR T‐cell therapies, including their molecular design, mechanisms of action, methods of production, clinical applications and established and emerging technologies for CAR T‐cell evaluation. It highlights the need for standardisation, quality control and monitoring of CAR T‐cell therapies, to ensure their safety and efficacy in clinical management.
Automated enumeration of lymphoid and plasma cells in bone marrow to establish normal reference ranges
Publisher: BMJ
Date: 06-2018
DOI: 10.1136/JCLINPATH-2018-205168
Abstract: The number of precursor and mature lymphoid cells and plasma cells in normal bone marrow trephine (BMT) biopsies throughout the human lifespan is unknown. Reference ranges have been established from aspirated marrow, but due to haemodilution errors, these do not accurately reflect the native marrow milieu. We aimed to define age-specific, normal reference ranges for lymphoid and plasma cells in BMT biopsy specimens using a combined immunophenotyping and digital enumeration approach. Morphologically normal BMT biopsy specimens (n=483) were obtained from patients aged 1 month to 90 years of age. Immunohistochemistry was performed to identify lymphoid progenitors , T-lymphocytes (CD3), B-lymphocytes (CD20) and plasma cells (CD138 and MUM1). Positive cells were counted using digital enumeration software, and the percent positivity for each antigen was determined per case. Mean values were generated for specific age groups, and age-defined reference ranges were determined for each antigen using normalised data. A mean of 16 609 cells (range: 7210–34 097) were counted per biopsy. Infant marrows showed a predominance of immature lymphoid progenitors and B cells. With increasing age, an increase in mean T cell and plasma cell numbers were observed. The results showed the same trends to flow cytometry references for aspirate material although the absolute values differed. Combined immunohistochemistry and automated enumeration gives an accurate, reproducible number of antigen-positive cells and has generated normal reference ranges for these cell types in BMT biopsies. The method and ranges we have established have the potential to be applied in routine clinical practice.
Morphology of myeloproliferative neoplasms
Publisher: Wiley
Date: 21-05-2023
DOI: 10.1111/IJLH.14086
Abstract: Myeloproliferative neoplasms (MPN) are a group of clonal haematological malignancies first described by Dameshek in 1957. The Philadelphia‐negative MPN that will be described are polycythaemia vera (PV), essential thrombocythaemia (ET), pre‐fibrotic myelofibrosis and primary myelofibrosis (PMF). The blood and bone marrow morphology are essential in diagnosis, for WHO classification, establishing a baseline, monitoring response to treatment and identifying changes that may indicate disease progression. The blood film changes may be in any of the cellular elements. The key bone marrow features are architecture and cellularity, relative complement of in idual cell types, reticulin content and bony structure. Megakaryocytes are the most abnormal cell and key to classification, as their number, location, size and cytology are all disease‐defining. Reticulin content and grade are integral to assignment of the diagnosis of myelofibrosis. Even with careful assessment of all these features, not all cases fit neatly into the diagnostic entities there is frequent overlap reflecting the biological disease continuum rather than distinct entities. Notwithstanding this, an accurate morphologic diagnosis in MPN is crucial due to the significant differences in prognosis between different subtypes and the availability of different therapies in the era of novel agents. The distinction between “reactive” and MPN is also not always straightforward and caution needs to be exercised given the prevalence of “triple negative” MPN. Here we describe the morphology of MPN including comments on changes with disease evolution and with treatment.
An efficient high-throughput flow cytometric method for estimating DNA ploidy level in plants
Publisher: Wiley
Date: 20-10-2009
DOI: 10.1002/CYTO.A.20816
Abstract: We present an efficient high-throughput flow cytometric method that builds on previously published methods and permits rapid ploidy discrimination in plants. By using Brassica napus L. microspore-derived plants as an ex le, we describe how 192 leaf tissue s les may be processed and analyzed comfortably by one operator in 6 h from tissue s ling to ploidy determination. The technique involves placing young leaf s les in two 96-well racks, using a bead-beating procedure to release nuclei into a lysis solution, filtering the s les on 96-well filter plates, staining with propidium iodide, and then rapidly estimating DNA ploidy using a plate loader on a BD FACS-Canto II flow cytometer. Throughout the s le preparation process, multichannel pipetting allows faster and less error-prone s le handling. In two 96-well plates of s les, the histogram peaks of DNA content from flow cytometry were wellresolved in 189 of 192 s les tested (98.4%), with CV values ranging from 2.98% to 6.20% with an average CV of 4.35% (SD = 0.68%). This new method is useful in doubled haploid plant breeding programs where early discrimination of haploid and doubled haploid (i.e., diploid) plantlets can confer significantly improved operational efficiencies. We discuss how this method could be further refined including adapting the method to robotic s le processing.
REVIEW: Peyer's patches
Publisher: Wiley
Date: 02-1997
DOI: 10.1111/J.1440-1746.1997.TB00395.X
Abstract: The function of Peyer's patches as antigenic s ling sites involves the complex interplay of a variety of mechanisms that aim to recognize luminal antigens, induce an immunological response and decrease the incidence of antigen translocation across the mucosal epithelium. This is achieved by M cells, which facilitate the uptake of luminal antigens, a vascular architecture that promotes the retention of absorbed antigens within the patch interstitium (allowing for maximal antigenic activation of lymphocytes) and the presence of lymphoid follicles that contain antigen-presenting cells and lymphocytes. Lymphocytes encountering antigen in the Peyer's patches proliferate, differentiate into fully mature antigen-specific effector cells and migrate to the mesenteric lymph nodes where they undergo final maturation. The mature lymphocytes then enter the systemic circulation and migrate throughout the other mucosa-associated lymphoid tissues of the body and "home' into the gut via high endothelial venules and gut-associated lymphoid tissue-specific adhesion molecules, providing antigen-specific lymphocytes at sites likely to re-encounter the antigen.
A mutation in PTPN11 may drive leukemic transformation in a case of essential thrombocythemia
Publisher: Informa UK Limited
Date: 07-06-2018
“Immuno-flowFISH” for the Assessment of Cytogenetic Abnormalities in Chronic Lymphocytic Leukemia
Publisher: Wiley
Date: 24-04-2019
DOI: 10.1002/CYTO.A.23769
Abstract: Imaging flow cytometry is emerging as a diagnostic tool for the assessment of leukemia. It has the functionality of standard flow cytometry and generates high-resolution digital images of each cell with quantifiable numerical data. We demonstrate the use of an automated high-throughput method for performing fluorescence in situ hybridization (FISH) on immunophenotyped whole cells in suspension and analyzed by imaging flow cytometry, a technique called "Immuno-flowFISH". The aim of this study was to demonstrate the application of immuno-flowFISH for the detection of chromosomal abnormalities in CLL, specifically trisomy 12 and del(17p). Mononuclear cells were isolated and immunophenotyped with fluorescently conjugated CD3, CD5, and CD19 monoclonal antibodies. Following fixation, cells were permeabilized, dsDNA denatured and hybridized with chromosome 12 or 17 enumeration (CEP 12 and CEP17) and 17p12 locus-specific FISH probes. Cells were analyzed on the Amnis ImageStream®X Mark II to assess the number and percent FISH-positive CLL cells and the ratio of FISH spot counts for CD5/CD19-positive CLL cells to CD3/CD5-positive T cells (FISH "mean spot ratio"). Deletion of 17p was detected in about 8% of cases to date, with del(17p) ranged from 3.5-22.8% and the FISH "mean spot ratio" 0.86-0.96. Immuno-flowFISH also detected a minimal residual disease case with +12 with a limit of detection of 0.13% and a rare case that presented with atypical phenotype and cytogenetics. Immuno-flowFISH could detect del(17p) in phenotypically identified CD5/CD19-positive B-cells. The 100-fold increase in analyzed cells, as well as the addition of cell phenotype increased the sensitivity and specificity over current clinical FISH testing. Furthermore, immuno-flowFISH analysis demonstrated specific utility in unique clinical scenarios such as residual disease and atypical biology cases which may be of significant benefit with regards to prognostication and MRD analysis. The method will assist in therapeutic decision making and disease monitoring for many hematological malignancies. © 2019 International Society for Advancement of Cytometry.
Megakaryocytes in myeloproliferative neoplasms have unique somatic mutations
Publisher: Elsevier BV
Date: 07-2017
Flow cytometric analysis of lymphocyte subset kinetics in Bali cattle experimentally infected with Jembrana disease virus
Publisher: Elsevier BV
Date: 10-2012
DOI: 10.1016/J.VETIMM.2012.06.013
Abstract: Jembrana disease virus (JDV) is an unusual bovine lentivirus that causes an acute and sometimes fatal disease after a short incubation period in Bali cattle (Bos javanicus). The pathological changes occur primarily in lymphoid tissues, which feature proliferating lymphoblastoid-like cells predominantly throughout parafollicular (T-cell) areas, and atrophy of follicles (B-cell) areas. Five Bali cattle were experimentally infected with JDV and all developed typical clinical signs of Jembrana disease characterised by a transient febrile response, enlargement of superficial lymph nodes and a significant leukopenia. Flow cytometric analysis of PBMC during the acute (febrile) disease phase showed that the reduced number of lymphocytes was due to a significant decrease in both the proportion and absolute numbers of CD4(+) T cells, but not CD8(+) T-cells or CD21(+) B-cells. At the end of the febrile phase, total numbers of both CD8(+) T-cells and CD21(+) B-cells increased significantly, while CD4(+) T-cell numbers remained below normal values, resulting in a significantly reduced CD4(+):CD8(+) ratio. We speculate that the persistent depletion of CD4(+) T cells following JDV infection, through lack of CD4(+) T cell help to B cells, may explain the lack of production of JDV-specific antibodies for several weeks after recovery despite an increase in CD21(+) B cell numbers. Further, our previous data showing that IgG(+) plasma cells are targets for JDV infection, correlated with our current data demonstrating an increase in CD8(+) T cell numbers, supports the suggestion that anti-viral cytotoxic T cell or other cell-mediated immune responses may be critical in the recovery process, although this remains to be formally demonstrated for JDV.
Imaging flow cytometry to assess chromosomal abnormalities in chronic lymphocytic leukaemia
Publisher: Elsevier BV
Date: 02-2018
Testing for human papillomavirus in BRAF inhibitor‐associated verrucous keratoses
Publisher: Wiley
Date: 08-12-2014
DOI: 10.1111/CUP.12403
Applications of imaging flow cytometry in the diagnostic assessment of acute leukaemia
Publisher: Elsevier BV
Date: 2017
DOI: 10.1016/J.YMETH.2016.06.023
Abstract: Automated imaging flow cytometry integrates flow cytometry with digital microscopy to produce high-resolution digital imaging with quantitative analysis. This enables cell identification based on morphology (cell size, shape), antigen expression, quantification of fluorescence signal intensity and localisation of detected signals (i.e. surface, cytoplasm, nuclear). We describe applications of imaging flow cytometry for the diagnostic assessment of acute leukaemia. These bone marrow malignancies are traditionally diagnosed and classified by cell morphology, phenotype and cytogenetic abnormalities. Traditionally morphology is assessed by light microscopy, phenotyping by conventional flow cytometry and genetics by karyotype and fluorescence in situ hybridisation (FISH) on interphase nuclei/metaphase spreads of cells on slides. Imaging flow cytometry adds a new dimension to the diagnostic assessment of these neoplasms. We describe three specific applications: From this we conclude that imaging flow cytometry offers benefits over conventional diagnostic methods. Specifically the ability to visualise the cells of interest, the pattern and localisation of expressed antigens and assess cytogenetic abnormalities in one integrated automated high-throughput test. Imaging flow cytometry presents a new paradigm for the diagnostic assessment of leukaemia.
The effect of minimum luminal nutrition on bacterial translocation and atrophy of the jejunum during parenteral nutrition
Publisher: Wiley
Date: 1996
DOI: 10.1111/J.1440-1746.1996.TB00012.X
Abstract: In situations of catabolic stress, the gut becomes atrophic and may have diminished barrier function as evidenced by an increase in bacterial translocation. The aim of this study was to examine the effect of minimum luminal nutrition during parenteral nutrition on the extent of jejunal atrophy and rate of bacterial translocation. Central venous lines were inserted into 30 rats before they underwent randomization to receive nutritional support with: (a) conventional parenteral nutrition (b) conventional parenteral nutrition with 3 g/day of rat food (i.e., minimum luminal nutrition) or (c) rat food ad libitum. The rats were assessed after 10 days for nutritional status, extent of jejunal atrophy, caecal flora, as well as the extent of bacterial translocation to the mesenteric lymph nodes, liver and spleen. Rats in the rat food ad libitum group lost the smallest amount of weight and had the least amount of jejunal atrophy, yet had a similar rate of bacterial translocation as the parenterally nourished groups. When compared with the conventional parenteral nutrition group, the minimum luminal nutrition group had better preservation of the weight of the small bowel and its isolated mucosa (P < 0.01), but had a similar rate of bacterial translocation. Minimum luminal nutrition reduced the extent of atrophy of the gut but did not affect the incidence of bacterial translocation. It is inferred that there is no direct relationship between the extent of mucosal atrophy and incidence of bacterial translocation.
Embryogenesis and plant regeneration of the perennial pasture and medicinal legume Bituminaria bituminosa (L.) C.H. Stirton
Publisher: CSIRO Publishing
Date: 2014
DOI: 10.1071/CP14083
Abstract: Bituminaria bituminosa (common name tedera) is a drought-tolerant perennial pasture species of agronomic and pharmaceutical interest for Mediterranean climates. Considering the importance of this legume, in vitro experiments were conducted to develop protocols for plant regeneration from embryogenic calli of leaves, petioles and anthers to efficiently exploit and maintain selected important clones from the tedera breeding program. The type of explant was a key factor in the frequency of embryogenesis and the number of embryos per callus. For plant regeneration from cultured anthers, appropriate anther physiological state (uninucleate stage of microsporogenesis), stress treatments (electroporation, 25 Ω, 25 µF, 1500 V) and culture conditions were determined. A robust flow-cytometry method was developed to analyse the ploidy status of callus, in vitro shoots and in vivo acclimatised plants derived from anther and leaf explants.
Glutamine
Publisher: Oxford University Press (OUP)
Date: 03-1996
Abstract: Glutamine is the most abundant free amino acid in the circulation. It is a primary fuel for rapidly iding cells and plays a key role in the transport of nitrogen between organs. Although glutamine is absent from conventional regimens aimed at nutritional support, glutamine deficiency can occur during periods of metabolic stress this has led to the reclassification of glutamine as a conditionally essential amino acid. Experiments with various animal models have demonstrated that the provision of glutamine can result in better nitrogen homoeostasis, with conservation of skeletal muscle. There is also considerable evidence that glutamine can enhance the barrier function of the gut. This review concludes by discussing the clinical evidence that supports the inclusion of stable forms of glutamine in solutions of nutrients.
The Role of Enterocytes in Gut Dysfunction
Publisher: Elsevier BV
Date: 1998
Attachment and Invasion of Neisseria meningitidis to Host Cells Is Related to Surface Hydrophobicity, Bacterial Cell Size and Capsule
Publisher: Public Library of Science (PLoS)
Date: 06-02-2013
Cells of Epithelial Lineage Are Present in Blood, Engraft the Bronchial Epithelium, and Are Increased in Human Lung Transplantation
Publisher: Elsevier BV
Date: 06-2009
DOI: 10.1016/J.HEALUN.2009.03.013
Abstract: There is a growing expectation that cell-based therapies will prove effective for a wide range of conditions including lung diseases such as cystic fibrosis. The promise of these therapies will depend largely on effective delivery and engraftment. In this study, in the setting of human lung transplantation, we sought to determine whether exogenous epithelial cells are able to engraft the transplanted organ and if cells of a similar phenotype could be detected in peripheral blood. Cells obtained from bronchial brushings and peripheral blood were analyzed via dual fluorescent in situ hybridization/fluorescent immunohistochemistry (FISH/IHC), short tandem repeat polymerase chain reaction (STR-PCR) and flow cytometry. In 2 of 3 gender-mismatched patients we observed limited (5.9% to 6.8% by STR-PCR and 3.5% to 4% by FISH/IHC) engraftment of the bronchial epithelium by exogenous epithelial cells. Engrafting cells were CD34(-) CD15(-) CD68(-) c-Kit(-), but expressed CXCR4 on the cell surface. Cells with a similar phenotype were also identified in peripheral blood. In 8 patients, at 2 to 66 months post-transplant, 0.57 +/- 0.17% of CD14(-) peripheral blood mononuclear cells were of epithelial lineage. Almost all were CD45(+) and most expressed CXCR4 on the cell membrane. Cells of epithelial lineage were also identified in peripheral blood in healthy in iduals but in much lower numbers (0.08 +/- 0.01%, p < 0.05). Cells of epithelial lineage are detectable in peripheral blood and are able to engraft the bronchial epithelium in humans. Cell numbers are increased in lung transplantation.
TGFα expression in myeloid malignancies
Publisher: BMJ
Date: 16-03-2016
DOI: 10.1136/JCLINPATH-2015-203526
Abstract: Transforming growth factor α (TGFα) is a peptide growth factor known to be expressed in normal haemopoiesis. It is also expressed in a range of epithelial neoplasms but has not been assessed in haemopoietic malignancies. We have performed an immunohistochemical evaluation of TGFα in acute and chronic myeloid malignancies. TGFα expression was semiquantitatively assessed in 69 normal bone marrow trephines and 157 cases of myeloid malignancy using an immunohistochemical approach. Blast cells of myeloid origin in acute myeloid leukaemia (AML), myelodysplasia and accelerated and blast phases of chronic myeloid leukaemia (CML) were TGFα positive. In acute promyelocytic leukaemia the neoplastic cells had significantly weaker TGFα expression than seen in other forms of AML. The blast cells in CML-accelerated and blast phases were positive with similar expression to AML. TGFα is expressed in neoplastic myeloblasts and could, therefore, be used as blast cell biomarker in diagnostic haematopathology. In addition, TGFα immunohistochemistry may be of use in identifying a therapeutic target.
Developments in the immunophenotypic analysis of haematological malignancies
Publisher: Elsevier BV
Date: 07-2013
DOI: 10.1016/J.BLRE.2013.06.005
Abstract: Immunophenotyping is the method by which antibodies are used to detect cellular antigens in clinical s les. Although the major role is in the diagnosis and classification of haematological malignancies, applications have expanded over the past decade. Immunophenotyping is now used extensively for disease staging and monitoring, to detect surrogate markers of genetic aberrations, to identify potential immuno-therapeutic targets and to aid prognostic prediction. This expansion in applications has resulted from developments in antibodies, methodology, automation and data handling. In this review we describe recent advances in both the technology and applications for the analysis of haematological malignancies. We highlight the importance of the expanding repertoire of testing capability for diagnostic, prognostic and therapeutic applications. The impact and significance of immunophenotyping in the assessment of haematological neoplasms are evident.
Circulating tumour DNA in colorectal cancer management
Publisher: Oxford University Press (OUP)
Date: 16-05-2023
DOI: 10.1093/BJS/ZNAD126
Abstract: Circulating tumour DNA analysis can be performed using two opposing paradigms: tumour-informed and tumour-agnostic approaches. The first requires sequencing data from the primary tumour s le to identify tumour DNA in circulation, whereas the latter occurs without previous primary tumour genetic profiling. Several preanalytical and laboratory considerations need to be taken into account before proceeding with in-house circulating tumour DNA analysis. Detection of circulating tumour DNA after curative resection is associated with a significant risk of recurrence. For those with stage II disease and detectable postoperative circulating tumour DNA, administration of adjuvant chemotherapy results in a reduction in the number of patients receiving chemotherapy while providing non-inferior recurrence-free survival compared with standard histopathological decision-making algorithms. Monitoring circulating tumour DNA during post-treatment surveillance may provide a significantly earlier diagnosis of recurrence.
A revised model of platelet aggregation
Publisher: American Society for Clinical Investigation
Date: 15-03-2000
DOI: 10.1172/JCI7569
Quantifying spore viability of the honey bee pathogen Nosema apis using flow cytometry
Publisher: Wiley
Date: 12-12-2014
DOI: 10.1002/CYTO.A.22428
Abstract: Honey bees are hosts to more than 80 different parasites, some of them being highly virulent and responsible for substantial losses in managed honey bee populations. The study of honey bee pathogens and their interactions with the bees' immune system has therefore become a research area of major interest. Here we developed a fast, accurate and reliable method to quantify the viability of spores of the honey bee gut parasite Nosema apis. To verify this method, a dilution series with 0, 25, 50, 75, and 100% live N. apis was made and SYTO 16 and Propidium Iodide (n = 35) were used to distinguish dead from live spores. The viability of spores in each s le was determined by flow cytometry and compared with the current method based on fluorescence microscopy. Results show that N. apis viability counts using flow cytometry produced very similar results when compared with fluorescence microscopy. However, we found that fluorescence microscopy underestimates N. apis viability in s les with higher percentages of viable spores, the latter typically being what is found in biological s les. A series of experiments were conducted to confirm that flow cytometry allows the use of additional fluorescent dyes such as SYBR 14 and SYTOX Red (used in combination with SYTO 16 or Propidium Iodide) to distinguish dead from live spores. We also show that spore viability quantification with flow cytometry can be undertaken using substantially lower dye concentrations than fluorescence microscopy. In conclusion, our data show flow cytometry to be a fast, reliable method to quantify N. apis spore viabilities, which has a number of advantages compared with existing methods.
Megakaryocytic hyperplasia in myeloproliferative neoplasms is driven by disordered proliferative, apoptotic and epigenetic mechanisms
Publisher: BMJ
Date: 19-08-2016
DOI: 10.1136/JCLINPATH-2015-203177
Abstract: Myeloproliferative neoplasms (MPN) are a heterogeneous group of clonal proliferative bone marrow diseases characterised by extensive megakaryocytic hyperplasia and morphological atypia. Despite knowledge of genomic defects, the pathobiological processes driving these megakaryocytic abnormalities in MPN remain poorly explained. We have explored the proliferative, apoptotic and epigenetic profiles of megakaryocytes in human MPN. Immunohistochemical staining was performed on bone marrow trephine biopsies of 81 MPN (with and without JAK2(V617F) and CALR mutations) and 15 normal controls to assess the megakaryocytic expression of biomarkers associated with proliferation (Ki67), apoptosis (Bcl-XL, BNIP-3) and epigenetic regulation (EZH2, SUZ12). Myeloproliferative megakaryocytes showed significantly greater expression of proliferative Ki67 and anti-apoptotic Bcl-XL, reduced pro-apoptotic BNIP-3 and increased SUZ12 compared with controls. In essential thrombocythaemia, large-giant megakaryocytes with hyperlobated nuclei showed a trend towards a proliferative signature. In contrast, myelofibrotic megakaryocytes with condensed nuclear chromatin, and cases with CALR mutations, had significant reductions in pro-apoptotic BNIP-3. Uncontrolled megakaryocytic expansion in MPN results from a combination of increased proliferation, attenuated apoptosis and defective epigenetic regulation with CALR mutations favouring apoptotic failure. The higher platelet counts reported to be seen in MPN with CALR mutations may be due to greater dysregulation of megakaryocyte apoptosis.
Measurement of monocyte-platelet aggregates by imaging flow cytometry
Publisher: Wiley
Date: 16-12-2015
DOI: 10.1002/CYTO.A.22587
Abstract: Platelets are subcellular blood elements with a well-established role in haemostasis. Upon activation platelets express P-Selectin (CD62P) on the cell membrane and bind to P-Selectin glycoprotein ligand 1 expressing monocytes, influencing them toward a pro-adhesive and inflammatory phenotype. It is well established that elevated circulating monocyte-platelet aggregates (MPAs) are linked to atherothrombosis in high risk patients. However, whole blood flow cytometry (FCM) has recently shown that circulating MPAs may also occur in the absence of platelet activation, particularly in healthy children. A potential limitation of conventional FCM is the potential for coincident events to resemble monocyte platelet aggregates. Here we report a novel imaging cytometry approach to further characterize monocyte-platelet aggregate formation by P-Selectin dependent and P-Selectin independent mechanisms and distinguish circulating MPAs from coincidental events. Monocytes were identified by expression of the lipopolysachharide receptor (CD14 BV421), while platelets were identified by expression of the glycoprotein Ib (CD42b APC). Differentiation of P-Selectin dependent and P-Selectin independent binding was achieved with AF488 labeled CD62P. Overall analysis of circulating and in vitro generated MPAs by conventional and imaging cytometry methods showed very strong correlation (r(2) = >0.99, P < 0.01). The Bland-Altman bias of -1.72 was not significantly different to zero. However, when measuring only P-Selectin negative MPAs, a lack of correlation (r(2) = 0.27, P = n.s.) likely reflects better discrimination of coincidence events using imaging cytometry. Our data demonstrate that IFC is more accurate in enumerating MPAs than conventional FCM, which over-estimates the number of MPAs due to the presence of coincident events.
Simultaneous flow cytometric characterization of multiple cell types and metabolic states in the rat brain after repeated mild traumatic brain injury
Publisher: Elsevier BV
Date: 07-2021
IGH cytogenetic abnormalities can be detected in multiple myeloma by imaging flow cytometry
Publisher: BMJ
Date: 16-09-2023
Abstract: Cytogenetic abnormalities involving the IGH gene are seen in up to 55% of patients with multiple myeloma. Current testing is performed manually by fluorescence in situ hybridisation (FISH) on purified plasma cells. We aimed to assess whether an automated imaging flow cytometric method that uses immunophenotypic cell identification, and does not require cell isolation, can identify IGH abnormalities. Aspirated bone marrow from 10 patients with multiple myeloma were studied. Plasma cells were identified by CD38 and CD138 coexpression and assessed with FISH probes for numerical or structural abnormalities of IGH . Thousands of cells were acquired on an imaging flow cytometer and numerical data and digital images were analysed. Up to 30 000 cells were acquired and IGH chromosomal abnormalities were detected in 5 of the 10 marrow s les. FISH signal patterns seen included fused IGH signals for IGH/FGFR3 and IGH/MYEOV , indicating t(4 ) and t(11 ), respectively. In addition, three IGH signals were identified, indicating trisomy 14 or translocation with an alternate chromosome. The lowest limit of detection of an IGH abnormality was in 0.05% of all cells. This automated high-throughput immuno-flowFISH method was able to identify translocations and trisomy involving the IGH gene in plasma cells in multiple myeloma. Thousands of cells were analysed and without prior cell isolation. The inclusion of positive plasma cell identification based on immunophenotype led to a lowest detection level of 0.05% marrow cells. This imaging flow cytometric FISH method offers the prospect of increased precision of detection of critical genetic lesions involving IGH and other chromosomal defects in multiple myeloma.
Flow Cytometry as a Rapid and Reliable Method to Quantify Sperm Viability in the Honeybee Apis mellifera
Publisher: Wiley
Date: 04-04-2014
DOI: 10.1002/CYTO.A.22462
Abstract: An important measure of male quality is sperm viability i.e., the percentage of live sperm within an ejaculate, as this provides an accurate measure of the number of sperm potentially available for egg fertilization. Sperm viability is often determined by fluorescence microscopy using dyes that differentially stain viable and nonviable sperm, but the technique has a number of limitations. Here, a flow cytometry (FCM) method was developed, which allows the rapid determination of honeybee sperm viability, facilitating high throughput analyses. Using s les with known sperm viabilities, it was found that data obtained from FCM were more accurate and less variable compared with data obtained for the same s les using fluorescence microscopy. It was also found that a previously reported additional population of honeybee sperm found in datasets using FCM is caused by freeze-thawing s les. In conclusion, the method described here allows to quantify sperm viability of honeybees quickly and with high accuracy. This will be of great value for future scientific research and could also be of value to guide future bee breeding programs, given the agricultural importance of honeybees as pollinators.
Herniation World Federation of Neurosurgical Societies Scale Improves Prediction of Outcome in Patients With Poor-Grade Aneurysmal Subarachnoid Hemorrhage.
Publisher: Ovid Technologies (Wolters Kluwer Health)
Date: 07-2022
DOI: 10.1161/STROKEAHA.121.036699
Abstract: Favorable outcomes are seen in up to 50% of patients with World Federation of Neurosurgical Societies (WFNS) grade V aneurysmal subarachnoid hemorrhage. Therefore, the usefulness of the current WFNS grading system for identifying the worst scenarios for clinical studies and for making treatment decisions is limited. We previously modified the WFNS scale by requiring positive signs of brain stem dysfunction to assign grade V. This study aimed to validate the new herniation WFNS grading system in an independent prospective cohort. We conducted an international prospective multicentre study in poor-grade aneurysmal subarachnoid hemorrhage patients comparing the WFNS classification with a modified version—the herniation WFNS scale (hWFNS). Here, only patients who showed positive signs of brain stem dysfunction (posturing, anisocoric, or bilateral dilated pupils) were assigned hWFNS grade V. Outcome was assessed by modified Rankin Scale score 6 months after hemorrhage. The primary end point was the difference in specificity of the WFNS and hWFNS grading with respect to poor outcomes (modified Rankin Scale score 4–6). Of the 250 patients included, 237 reached the primary end point. Comparing the WFNS and hWFNS scale after neurological resuscitation, the specificity to predict poor outcome increased from 0.19 (WFNS) to 0.93 (hWFNS) (McNemar, P .001) whereas the sensitivity decreased from 0.88 to 0.37 ( P .001), and the positive predictive value from 61.9 to 88.3 (weighted generalized score statistic, P .001). For mortality, the specificity increased from 0.19 to 0.93 (McNemar, P .001), and the positive predictive value from 52.5 to 86.7 (weighted generalized score statistic, P .001). The identification of objective positive signs of brain stem dysfunction significantly improves the specificity and positive predictive value with respect to poor outcome in grade V patients. Therefore, a simple modification—presence of brain stem signs is required for grade V—should be added to the WFNS classification. URL: clinicaltrials.gov Unique identifier: NCT02304328.
An active, collaborative approach to learning skills in flow cytometry
Publisher: American Physiological Society
Date: 06-2016
Abstract: Advances in science education research have the potential to improve the way students learn to perform scientific interpretations and understand science concepts. We developed active, collaborative activities to teach skills in manipulating flow cytometry data using FlowJo software. Undergraduate students were given compensated clinical flow cytometry listmode output (FCS) files and asked to design a gating strategy to diagnose patients with different hematological malignancies on the basis of their immunophenotype. A separate cohort of research trainees was given uncompensated data files on which they performed their own compensation, calculated the antibody staining index, designed a sequential gating strategy, and quantified rare immune cell subsets. Student engagement, confidence, and perceptions of flow cytometry were assessed using a survey. Competency against the learning outcomes was assessed by asking students to undertake tasks that required understanding of flow cytometry dot plot data and gating sequences. The active, collaborative approach allowed students to achieve learning outcomes not previously possible with traditional teaching formats, for ex le, having students design their own gating strategy, without forgoing essential outcomes such as the interpretation of dot plots. In undergraduate students, favorable perceptions of flow cytometry as a field and as a potential career choice were correlated with student confidence but not the ability to perform flow cytometry data analysis. We demonstrate that this new pedagogical approach to teaching flow cytometry is beneficial for student understanding and interpretation of complex concepts. It should be considered as a useful new method for incorporating complex data analysis tasks such as flow cytometry into curricula.
A warm-core eddy linking shelf, Leeuwin Current and oceanic waters demonstrated by near-shelf distribution patterns of Synechococcus spp. and Prochlorococcus spp. in the eastern Indian Ocean
Publisher: CSIRO Publishing
Date: 2013
DOI: 10.1071/MF12271
Abstract: In May 2006 (Austral autumn) the distribution and abundance of the cyanobacteria Synechococcus spp. and Prochlorococcus spp. were examined to assess the connectivity of a forming warm-core mesoscale eddy with the Leeuwin Current and shelf waters off south-west Western Australia. Distributions of the cyanobacteria resulted in two broad categories of s les, those dominated by Prochlorococcus spp. from subtropical and Leeuwin Current waters and those with mixed populations from shelf and eddy waters. Water temperature (21.45°C), salinity (35.46) and nitrate (0.33 μM) contributed to these groupings. Synechococcus spp. reached an integrated abundance of 3.3 × 108 cells cm–2 in warm shelf waters, with 60% of cells in G2 phase in the mid-afternoon (~16:00 hours). Cooler, nitrate-poor oceanic waters were almost exclusively inhabited by Prochlorococcus spp., with the highest abundance of 4.2 × 108 cells cm–2 in cool deep waters off the Capes in the south with 40% of cells in G2 phase in the evening (~19:00 hours). The eddy perimeter represented a clear boundary for both species, but showed connectivity between the shelf and eddy centre as both locations had a mixed community, dominated by Synechococcus spp. Eddies of the Leeuwin Current advect shelf waters, and their assemblages and productivity offshore.
Gene expression of CXCL1 (GrO-A) and EGF by platelets in myeloproliferative neoplasms
Publisher: Ovid Technologies (Wolters Kluwer Health)
Date: 26-10-2020
Characterization of columnar cell lesions of the breast: immunophenotypic analysis of columnar alteration of lobules with prominent apical snouts and secretions
Publisher: Elsevier BV
Date: 02-2007
DOI: 10.1016/J.HUMPATH.2006.08.003
Abstract: Columnar cell lesions of the breast are detected with increasing frequency in routine pathology practice, in part as a result of the widespread biopsy of nonpalpable breast abnormalities detected by screening mammography. Immunohistochemical investigation of the lesions in relation to the normal breast or to other breast pathologies is not well characterized, and the malignant potential of this spectrum of lesions has not been examined clinically. In this study, a cohort of 45 breast specimens containing columnar cell lesions, in particular, columnar alteration of lobules with prominent apical snouts and secretions (CAPSS), was investigated for expression of a series of breast tumor biomarkers. Using a semiquantitative immunohistochemical scoring system, up-regulation of estrogen, progesterone, and androgen receptors in CAPSS lesions to levels not significantly different from that in in situ or invasive breast tumors was identified. In four cases where CAPSS within a specimen lacked expression of a steroid hormone receptor, the coexisting in situ or invasive carcinoma also lacked expression of that receptor. In 81% of CAPSS lesions, E-cadherin immunostaining was reduced in isolated foci of cells or was decreased in intensity in all cells within the lesion. Quantitation of Ki-67 immunostaining demonstrated that proliferation of cells within CAPSS lesions was increased, compared with normal breast epithelium, but was lower than that detected in in situ or invasive cancers within the same specimens. Results of these analyses indicate that CAPSS shares immunophenotypic alterations with other premalignant lesions, the clinical implications of which may be investigated using established breast tumor biomarkers.
Immunohistochemical enumeration of plasma cells
Publisher: BMJ
Date: 03-08-2023
Disruption of IKAROS activity in primitive chronic-phase CML cells mimics myeloid disease progression
Publisher: American Society of Hematology
Date: 15-01-2015
DOI: 10.1182/BLOOD-2014-06-581173
Abstract: CML patients with advanced-phase myeloid disease frequently show decreased IKAROS protein in primitive cells. Expression of a dominant-negative IKAROS isoform expands primitive human CML cells and enhances their differentiation into basophils.
Locally performed postoperative circulating tumour DNA testing performed during routine clinical care to predict recurrence of colorectal cancer
Publisher: Wiley
Date: 15-03-2023
DOI: 10.1111/ANS.18385
Abstract: Identifying patients at high risk for colorectal cancer recurrence is essential for improving prognosis. In the postoperative period, circulating tumour DNA (ctDNA) has been demonstrated as a significant prognostic indicator of recurrence. These results have been obtained under the strict rigours of clinical trials, but not validated in a real‐world setting using in‐house testing. We report the outcomes of locally performed postoperative ctDNA testing conducted during routine clinical care and the association with the recurrence of colorectal cancer. We recruited 36 consecutive patients with newly diagnosed colorectal cancer between 2018 and 2020. Postoperative plasma s les were collected at the first outpatient review following resection. Tumour‐informed ctDNA analysis was performed using droplet digital polymerase chain reaction or targeted next‐generation sequencing. At the time of surgery, there were 24 patients (66.7%) with localized cancer, nine (25%) with nodal spread, and three (8.3%) with metastatic disease. The median time from surgery to plasma s le donation was 22 days (IQR 20–28 days). At least one somatic mutation was identified in primary tumour tissue for 28 (77.8%) patients. Postoperative ctDNA was detected in five patients (13.9%). The median duration of follow‐up was 32.0 months (IQR 27.2–38.1 months). Two patients (5.56%) developed metastatic recurrence. However, neither had detectable postoperative ctDNA. There were no instances of loco‐regional recurrence. Analysis of postoperative ctDNA testing can be performed locally, however this study did not reproduce the adverse association between detectable postoperative ctDNA and the development of colorectal cancer recurrence seen in clinical trials.
Chemotherapy Enhances Cross-Presentation of Nuclear Tumor Antigens
Publisher: Public Library of Science (PLoS)
Date: 22-09-2014
Imaging flow cytometry shows monosomy 17 in circulating plasma cells in myeloma
Publisher: Elsevier BV
Date: 12-2022
DNA ploidy variation and distribution in the Lepidosperma costale complex (Cyperaceae): implications for conservation and restoration in a biodiversity hotspot
Publisher: CSIRO Publishing
Date: 2017
DOI: 10.1071/BT16197
Abstract: Intraspecific ploidy variation is an important component of angiosperm bio ersity however, this variation is rarely considered in conservation programs. This is of particular concern when conservation activities include augmentation, reintroduction or ecological restoration because there are potentially negative consequences when ploidy variants are unintentionally mixed within populations. We surveyed regional ploidy variation in the Lepidosperma costale Nees species complex (Schoeneae: Cyperaceae) in the South West Australian Floristic Region, an international bio ersity hotspot. Several L. costale sensu lato populations are threatened by iron-ore extraction, including the rare L. gibsonii R.L.Barrett, and these populations are the subject of ecological restoration programs. The DNA ploidy of 2384 in iduals from 28 populations across the range of the species complex was determined and four DNA ploidy levels were discovered, namely, diploid, triploid, tetraploid and pentaploid. Diploids and tetraploids were the most common cytotypes and were largely geographically segregated, even at an exhaustively studied contact zone. Triploids were found at a low frequency in two populations. The rarity of triploids suggests substantial interploidy sterility, and that mixing of ploidy variants should, therefore, be avoided when restoring L. costale s.l. populations. These data provide a guide for L. costale s.l. germplasm collection and suggest that polyploidy may be an important driver of ersification in these sedges.
Quality of Life After Poor-Grade Aneurysmal Subarachnoid Hemorrhage
Publisher: Ovid Technologies (Wolters Kluwer Health)
Date: 04-01-2023
DOI: 10.1227/NEU.0000000000002332
Abstract: Poor-grade aneurysmal subarachnoid hemorrhage (aSAH) is associated with high mortality and poor disability outcome. Data on quality of life (QoL) among survivors are scarce because patients with poor-grade aSAH are underrepresented in clinical studies reporting on QoL after aSAH. To provide prospective QoL data on survivors of poor-grade aSAH to aid clinical decision making and counseling of relatives. The herniation World Federation of Neurosurgical Societies scale study was a prospective observational multicenter study in patients with poor-grade (World Federation of Neurosurgical Societies grades 4 & 5) aSAH. We collected data during a structured telephone interview 6 and 12 months after ictus. QoL was measured using the EuroQoL - 5 Dimensions - 3 Levels (EQ-5D-3L) questionnaire, with 0 representing a health state equivalent to death and 1 to perfect health. Disability outcome for favorable and unfavorable outcomes was measured with the modified Rankin Scale. Two hundred-fifty patients were enrolled, of whom 237 were included in the analysis after 6 months and 223 after 12 months. After 6 months, 118 (49.8%) patients were alive, and after 12 months, 104 (46.6%) patients were alive. Of those, 95 (80.5%) and 89 (85.6%) reached a favorable outcome with mean EQ-5D-3L index values of 0.85 (±0.18) and 0.86 (±0.18). After 6 and 12 months, 23 (19.5%) and 15 (14.4%) of those alive had an unfavorable outcome with mean EQ-5D-3L index values of 0.27 (±0.25) and 0.19 (±0.14). Despite high initial mortality, the proportion of poor-grade aSAH survivors with good QoL is reasonably large. Only a minority of survivors reports poor QoL and requires permanent care.
The effect on minimum luminal nutrition on mucosal cellularity and immunity of the gut
Publisher: Wiley
Date: 10-1998
DOI: 10.1111/J.1440-1746.1998.TB00563.X
Abstract: Many catabolic patients can only consume small volumes of enteral nutrients. The aim of this study was to evaluate markers of cellularity and immunity in the small intestine of rats randomized to receive 6 days of parenteral nutrition, 25% enteral and 75% parenteral nutrition (i.e. minimum luminal nutrition) or enteral nutrition. The same glutamine-enriched solution was used for both parenteral and enteral nutrition. Enteral nutrition was associated with the least amount of jejunal atrophy (P<0.01), with the results from the minimum luminal nutrition group approximating those of the parenteral nutrition group. Parenteral nutrition was associated with the greatest number of CD2+ cells (P< 0.05) and the lowest CD4/CD8 cell ratio (P< 0.01) in the jejunal mucosa. In essence, we failed to demonstrate that there are any appreciable benefits associated with the enteral consumption of 25% of a nutrient load.
Warm ischaemia time in a model for small bowel transplantation
Publisher: Wiley
Date: 1996
DOI: 10.1002/(SICI)1098-2752(1996)17:8<438::AID-MICR4>3.0.CO;2-A
TGF-α and IL-6 plasma levels selectively identify CML patients who fail to achieve an early molecular response or progress in the first year of therapy
Publisher: Springer Science and Business Media LLC
Date: 22-02-2016
DOI: 10.1038/LEU.2016.34
Abstract: Early molecular response (EMR, BCR-ABL1 (IS)⩽10% at 3 months) is a strong predictor of outcome in imatinib-treated chronic phase chronic myeloid leukemia (CP-CML) patients, but for patients who transform early, 3 months may be too late for effective therapeutic intervention. Here, we employed multiplex cytokine profiling of plasma s les to test newly diagnosed CP-CML patients who subsequently received imatinib treatment. A wide range of pro-inflammatory and angiogenesis-promoting cytokines, chemokines and growth factors were elevated in the plasma of CML patients compared with that of healthy donors. Most of these normalized after tyrosine kinase inhibitor treatment while others remained high in remission s les. Importantly, we identified TGF-α and IL-6 as novel biomarkers with high diagnostic plasma levels strongly predictive of subsequent failure to achieve EMR and deep molecular response, as well as transformation to blast crisis and event-free survival. Interestingly, high TGF-α alone can also delineate a poor response group raising the possibility of a pathogenic role. This suggests that the incorporation of these simple measurements to the diagnostic work-up of CP-CML patients may enable therapy intensity to be in idualized early according to the cytokine-risk profile of the patient.
Use of multiparameter flow cytometry to determine the effects of monoterpenoids and phenylpropanoids on membrane polarity and permeability in staphylococci and enterococci
Publisher: Elsevier BV
Date: 2013
DOI: 10.1016/J.IJANTIMICAG.2012.05.015
Abstract: Monoterpenoids and phenylpropanoids are major components of many plant essential oils and are relatively simple, low-molecular-weight compounds with antimicrobial activity. This study used multiparameter flow cytometry to examine changes in membrane polarity and permeability in Staphylococcus aureus, Staphylococcus epidermidis and Enterococcus faecalis following exposure to the monoterpenoids carvacrol, 1,8-cineole and terpinen-4-ol and the phenylpropanoids eugenol and cinnamaldehyde. Melaleuca alternifolia (tea tree) essential oil was also investigated. The fluorescent dyes DiOC(2)(3) (3,3'-diethyloxacarbocyanine oxide) and TO-PRO(®)-3 were used to assess membrane potential and permeability, respectively, following treatment with the minimum inhibitory concentration (MIC) of each test compound for 5 min and 30 min. Four subpopulations of cells were identified based on polarity and permeability. Eugenol treatment resulted in the greatest depolarisation and permeabilisation at 5 min, followed by carvacrol. Cinnamaldehyde, whilst having the lowest MICs (0.006-0.1%, v/v), did not induce changes in polarity or permeability at the MIC, and substantially higher concentrations were required to induce significant effects. At 30 min, treatment with all six compounds resulted in significant depolarisation (60.9-99.3% of cells), whereas fewer compounds (ranging from two to five per organism) resulted in significant permeabilisation. The extent of permeabilisation was always less than depolarisation, with overall means for all treatments of 46.1% and 89.4% of cells permeabilised and depolarised, respectively, at 30 min. These data demonstrate that several monoterpenoids and phenylpropanoids as well as tea tree oil alter membrane properties by decreasing polarity and increasing permeability in a time- and concentration-dependent manner.
Automated digital enumeration of plasma cells in bone marrow trephine biopsies of multiple myeloma
Publisher: BMJ
Date: 24-11-2020
DOI: 10.1136/JCLINPATH-2020-207066
Abstract: Determination of the number of plasma cells in bone marrow biopsies is required for the diagnosis and ongoing evaluation of plasma cell neoplasms. We developed an automated digital enumeration platform to assess plasma cells identified by antigen expression in whole bone marrow sections in multiple myeloma, and compared it with manual assessments. Bone marrow trephine biopsy specimens from 91 patients with multiple myeloma at diagnosis, remission and relapse were stained for CD138 and multiple myeloma oncogene 1 (MUM1). Manual assessment and digital quantification were performed for plasma cells in the entire trephine section. Concordance rates between manual and digital methods were evaluated for each antigen by intraclass correlation analyses (ICC) with associated Spearman’s correlations. The digital platform counted 16 484–1 118 868 cells and the per cent CD138 and MUM1-positive plasma cells ranged from 0.05% to 93.5%. Overall concordance between digital and manual methods was 0.63 for CD138 and 0.89 for MUM1. Concordance was highest with diffuse plasma cell infiltrates (MUM1: ICC=0.90) and lowest when in microaggregates (CD138: ICC=0.13). Manual counts exceeded digital quantifications for both antigens (CD138: mean=26.4% MUM1: mean=9.7%). Diagnostic or relapse threshold counts, as determined by CD138 manual assessments, were not reached with digital counting for 16 cases (18%). Automated digital enumeration of the entire, immunohistochemically stained bone marrow biopsy section can accurately determine plasma cell burden, irrespective of pattern and extent of disease (as low as 0.05%). This increases precision over manual visual assessments which tend to overestimate plasma burden, especially for CD138, and when plasma cells are in clusters.
Imaging flow cytometry in the assessment of leukocyte-platelet aggregates
Publisher: Elsevier BV
Date: 2017
DOI: 10.1016/J.YMETH.2016.10.002
Abstract: Platelets are subcellular blood elements with a well-established role in haemostasis. Upon activation platelets undergo granule exocytosis, resulting in α-granule P-Selectin being expressed on the cell membrane. This allows binding of activated platelets to P-Selectin glycoprotein ligand 1 (PSGL-1) expressing leukocytes, forming leukocyte-platelet aggregates (LPAs). Whole blood flow cytometry (FCM) has demonstrated that elevated circulating LPAs (especially monocyte LPAs) are linked to atherothrombosis in high risk patients, and that activated platelet binding influences monocytes towards a pro-adhesive and pro-atherogenic phenotype. However, a limitation of conventional FCM is the potential for coincident events to resemble LPAs despite no tethering. Imaging cytometry can be used to characterize LPA formation and distinguish circulating MPAs from coincidental events. Platelets and leukocyte subsets are identified by expression of surface markers (e.g. the lipopolysachharide receptor CD14 on monocytes, glycoprotein Ib CD42b on platelets). In conventional FCM, all events with both leukocyte and platelet characteristics are designated as LPAs. However, by using an 'internal' mask based on the brightfield image and the fluorescent platelet identifier, imaging flow cytometry is able to distinguish leukocytes with tethered platelets (genuine LPAs) from leukocyte with coincidental, untethered platelets nearby. Mechanisms (e.g. adhesion molecules) or consequences (e.g. signal transduction) can then be separately analysed in platelet tethered and untethered leukocytes. Imaging flow cytometry therefore provides a more accurate approach for both enumeration and analysis of LPAs than conventional FCM.
Multi-probe FISH Analysis of Immunophenotyped Chronic Lymphocytic Leukemia by Imaging Flow Cytometry
Publisher: Wiley
Date: 10-2021
DOI: 10.1002/CPZ1.260
Abstract: Imaging flow cytometry is an automated method that enables cells and fluorescent signals to be visualized and quantified. Here, we describe a new imaging flow cytometry method whereby fluorescence in situ hybridization (FISH) is integrated with cell phenotyping. The method, called “immuno‐flowFISH,” provides an exciting new dimension for the analysis of genomic changes in cytological s les (e.g., blood, bone marrow). Cells are analyzed in suspension without any requirement for prior cell isolation or separation. Multiple antibodies and FISH probes, each with a unique fluorophore, can be added and many thousands of cells analyzed. Specific cell populations are identified by their antigenic profile and then analyzed for the presence of chromosomal defects. Immuno‐flowFISH was applied to the assessment of chronic lymphocytic leukemia (CLL), a mature B‐cell neoplasm where chromosomal abnormalities predict prognosis and treatment requirements. This integrated immunophenotyping and multi‐probe FISH strategy could detect both structural and numerical chromosomal changes involving chromosomes 12 and 17 in CLL cells. Given that many thousands of cells were analyzed and the leukemic cells were positively identified by their immunophenotype, this multi‐probe method adds precision to the cytogenomic analysis of CLL. © 2021 Wiley Periodicals LLC.
Development of a robust immuno-S-FISH protocol using imaging flow cytometry
Publisher: Wiley
Date: 03-05-2016
DOI: 10.1002/CYTO.A.22852
Abstract: Fluorescence in situ hybridization (FISH) is a microscopy technique which uses a fluorescent probe to detect DNA sequences and is generally performed on metaphase spreads or interphase nuclei of intact cells on a slide. In a diagnostic laboratory, cells are hybridized with fluorescent probes and up to 200 cells counted for the number of cells with probe "spots." Recent modifications to standard FISH include immuno-FISH, where chromosomal abnormalities are detected only in cells by their phenotype, and S-FISH where probe hybridization is performed on whole cells in suspension. Here we describe the development of an immuno-S-FISH method that combines immunophenotyping and FISH analysis of cells in suspension followed by analysis on an imaging flow cytometer. This single platform technique couples microscopy with flow cytometry and "spot" detection of bound FISH probe. Automated immuno-S-FISH enables large numbers of analyzed cells to be identified by phenotype and assessed for specific chromosomal determinants by FISH. This novel robust method enables quantitative cell population analysis and "spot" counting for large numbers of cells. We report method optimization of this imaging immuno-S-FISH flow cytometry protocol which has capability for many clinical applications. © 2016 International Society for Advancement of Cytometry.
The effect of branched-chain amino acid-enriched parenteral nutrition on gut permeability
Publisher: Elsevier BV
Date: 03-1996
DOI: 10.1016/S0899-9007(96)91122-3
Abstract: In situations of catabolic stress, the gut becomes atrophic and has a diminished barrier function as evidenced by an increased permeability to a variety of molecules. It is known that the parenteral administration of branched-chain amino acids (BCAA) reduce gut atrophy. The aim of this study was to examine the effect of BCAA-enriched solutions of parenteral nutrients on gut permeability. A secondary aim was to observe the association between gut permeability and variables that have been used to assess jejunal atrophy. Central venous lines were inserted into 30 rats before randomization to receive nutritional support with: (1) a conventional parenteral solution (CPN), (2) A 2.0% BCAA-enriched solution (BCAA), or (3) rat food ad lib (Rat Food). The rats were assessed after 7 d for nutritional status, gut morphology, and gut permeability ratio (ratio of the permeability to 14C raffinose and 3H mannitol). We found that rats in the Rat Food Group lost the least amount of weight, had the least amount of jejunal atrophy, and had better preservation of barrier function as determined by gut permeability. When compared with the CPN Group, the BCAA Group had better preservation of jejunal morphology and protein content (p < 0.05), but a similar gut permeability. A cross-correlation matrix demonstrated a significant negative correlation between permeability to mannitol and mucosal weight, mucosal protein content and mucosal DNA content. Branched-chain amino acid-enriched parenteral nutrition reduced gut atrophy but not the gut permeability associated with parenteral nutrition. In the parenterally nourished rat model, atrophy of the jejunum is associated with increased permeability to small molecules.
ISCHAEMIA-REPERFUSION INJURY TO THE INTESTINE
Publisher: Wiley
Date: 08-1998
DOI: 10.1111/J.1445-2197.1998.TB02099.X
Abstract: Ischaemia-reperfusion injury (IRI) is of obvious relevance in situations where there is an interruption of blood supply to the gut, as in vascular surgery, or in the construction of free intestinal grafts. It is now appreciated that IRI also underlies the guy dysfunction that occurs in early shock, sepsis, and trauma. The events that occur during IRI are complex. However, recent advances in cellular biology have started to unravel these underlying processes. The aim of this review is to provide an outline of current knowledge on the mechanisms and consequences of IRI. Initially, IRI appears to be mediated by reactive oxygen metabolites and, at a later stage, by the priming and activation of polymorphonuclear neutrophils (PMN). Ischaemia-reperfusion injury can diminish the barrier function of the gut, and can promote an increase in the leakage of molecules (intestinal permeability) or the passage of microbes across the wall of the bowel (bacterial translocation). Ischaemia-reperfusion injury to the gut can result in the generation of molecules that may also harm distant tissues.
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