Jean-Pierre Scheerlinck

Hypoxia Potentiates Endotoxin-Induced Allopregnanolone Concentrations in the Newborn Brain

Publisher: S. Karger AG

Date: 2006

DOI: 10.1159/000094146

Abstract: i Background: /i Allopregnanolone is a neurosteroid produced in the brain that can alter the excitability of the CNS. Neurosteroids have neuroprotective properties, and their elevation in response to stress may protect the newborn brain following infection or hypoxia. Infection, particularly of the respiratory tract, may lead to episodes of hypoxia. Infection and hypoxia have been identified as factors contributing to neonatal morbidity and mortality. i Objectives: /i To determine the effect of acute episodes of hypoxia alone or in combination with lipopolysaccharide (LPS) exposure on plasma and brain allopregnanolone concentrations in lambs 10–21 days old. Also, to examine plasma levels of cortisol and the cytokines, tumour necrosis factor-α and interleutkin-6 after these challenges. i Results: /i Allopregnanolone concentrations in the brain were markedly increased after hypoxia. Hypoxia following prior LPS treatment resulted in greater increases in brain allopregnanolone concentrations compared to either the LPS or hypoxia treatment alone. Importantly, brain regions unaffected by LPS or hypoxia alone (thalamus/hypothalamus, cerebellum) showed significant increases of allopregnanolone content following the combined LPS and hypoxia treatments. Plasma tumour necrosis factor-α and interleukin-6 concentrations were increased after LPS treatment with and without hypoxia, but not by hypoxia alone. In contrast, plasma cortisol concentrations were increased after both stressors. i Conclusions: /i These results show that the brain of young lambs readily responds to physiological stress by increased production of allopregnanolone. This response may protect the developing brain from the cytotoxicity following hypoxic and infectious episodes.

Activation of NF-κB transcription factor in the preterm ovine brain and placenta after acute LPS exposure

Publisher: Wiley

Date: 2006

DOI: 10.1002/JNR.20757

Abstract: Intrauterine infection may be causally related to inflammation and injury of the fetal brain, however the mechanisms by which this occurs are unclear. We have investigated whether nuclear factor (NF)-kappaB, a transcription factor for proinflammatory cytokines, is activated in the fetal brain after acute LPS-exposure. At 95 days of gestation (term = approximately 147 days), 5 fetuses received a single intravenous bolus dose of LPS (1 microg/kg) 6 fetuses served as controls. Fetal blood s les were taken hourly for 6 hr post LPS-exposure to assess physiological status. Ewes and fetuses were then euthanased, placental and brain tissue examined histologically, and NF-kappaB activation assessed in several regions of the fetal brain using an electromobility shift assay (EMSA). Oxidative stress was measured using lipid peroxidation and 8-isoprostane biochemical assays and brain cytokine concentrations analysed by enzyme linked immunosorbent assay (ELISA). LPS-exposed fetuses (relative to controls) were hypoxemic and the haematocrit and lactate levels had increased. In the brains of LPS-exposed fetuses compared to controls, NF-kappaB binding activity was elevated in the hippoc us and the thalamus/basal ganglia 8-isoprostane levels were elevated overall (P < 0.05) in the parietal/occipital/temporal lobes and thalamus/basal ganglia. TNF-alpha and IL-6 concentrations were not elevated, however, there was a tendency for an elevation of IFN-gamma concentrations in the thalamus/basal ganglia. IFN-gamma concentration was elevated (P < 0.05) in the plasma 4 hr after LPS-exposure. In the placenta, NF-kappaB binding activity was increased (P < 0.05). We conclude that acute systemic administration of LPS leads to increased binding activity of NF-kappaB subunits in specific regions of the fetal brain and in the placenta, but that there is no clear-cut relationship between this elevation and vulnerability to endotoxic damage.

Knocking down schistosomes – promise for lentiviral transduction in parasites

Publisher: Elsevier BV

Date: 07-2015

DOI: 10.1016/J.PT.2015.03.009

Abstract: Underpinned by major advances in our understanding of the genomes of schistosomes, progress in the development of functional genomic tools is providing unique prospects to gain insights into the intricacies of the biology of these blood flukes, their host relationships, and the diseases that they cause. This article reviews some key applications of double-stranded RNA interference (RNAi) in Schistosoma mansoni, appraises delivery systems for transgenesis and stable gene silencing, considers ways of increasing efficiency and specificity of gene silencing, and discusses the prospects of using a lentivirus delivery system for future functional genomic-phenomic explorations of schistosomes and other parasites. The ability to achieve effective and stable gene perturbation in parasites has major biological implications and could facilitate the development of new interventions.

Co-delivery of plasmid-encoded cytokines modulates the immune response to a DNA vaccine delivered by in vivo electroporation

Publisher: Elsevier BV

Date: 03-2007

DOI: 10.1016/J.VACCINE.2006.12.025

Abstract: In this study, in vivo electroporation of a DNA vaccine adjuvanted with plasmids encoding different cytokines was investigated in large animals. Sheep were injected intramuscularly with a DNA vaccine encoding an antigen of Haemonchus contortus (pNPA) and plasmids encoding different cytokines followed by in vivo electroporation. Plasmids (pCI) carrying the genes of different cytokines including ovine IL-4(pCI-IL4), IL-10(pCI-IL10), GM-CSF(pCI-GMCSF), and MCP-1alpha(pCI-MCP1alpha), and pCI-IL4+pCI-GMCSF were co-delivered with pNPA. The results showed that co-delivery of pCI-GMCSF or pCI-IL4+pCI-GMCSF significantly enhanced both antibody responses and T cell proliferation responses to the antigen after two DNA immunisations compared to co-delivery of pCI. In contrast, antibody responses of the sheep that received pCI-IL10 were decreased significantly. Other cytokine expressing plasmids did not significantly alter the measured immune responses. Furthermore, co-delivery of pCI-GMCSF increased IgG2 response more than IgG1 responses, suggesting a Th1 bias. However, the increase in IgG2 over IgG1 was less apparent when co-delivery of pCI-IL4 with pCI-GMCSF. Interestingly, the co-delivery of pCI-IL4 alone did not increase the IgG1 titre, suggesting that both pCI-GMCSF and pCI-IL4 are required for optimal IgG1 production. Thus, co-delivery of plasmid-encoded cytokine genes with in vivo electroporation has the ability to effectively modulate immune responses to a DNA vaccine in a large animal.

Local immune responses following nasal delivery of an adjuvanted influenza vaccine

Publisher: Elsevier BV

Date: 05-2006

DOI: 10.1016/J.VACCINE.2006.02.032

Abstract: A key barrier to producing effective nasal immunisations is the low efficiency of uptake of vaccines across the nasal mucosa. Using a recently developed cannulation system, we examined the antibody response induced by nasal immunisation with an ISCOMATRIX influenza vaccine. This showed for the first time, that following nasal vaccination, specific antibodies enter the circulation of primed animals via the draining lymphatics as a wave that peaks approximately 5-6 days after vaccination. These antibodies included some of the IgA isotype and possessed functional haemagglutination inhibition activity. These responses, though small, were induced using a very simple delivery system, emphasising the applicability of this cannulation model for evaluation of excipients and adjuvants aimed at improving intranasal vaccine efficacy.

Microdiversity of Echinococcus granulosus sensu stricto in Australia

Publisher: Cambridge University Press (CUP)

Date: 04-04-2016

DOI: 10.1017/S0031182016000445

Abstract: Echinococcus granulosus ( sensu lato ) is now recognized as an assemblage of cryptic species, which differ considerably in morphology, development, host specificity (including infectivity athogenicity for humans) and other aspects. One of these species, E. granulosus sensu stricto (s.s.), is now clearly identified as the principal agent causing cystic echinococcosis in humans. Previous studies of a small section of the cox1 and nadh1 genes identified two variants of E. granulosus s.s. to be present in Australia however, no further work has been carried out to characterize the micro ersity of the parasite in its territory. We have analysed the sequence of the full length of the cox1 gene (1609 bp) from 37 isolates of E. granulosus from different hosts and geographic regions of Australia. The analysis shows that seven haplotypes of E. granulosus s.s. not previously described were found, together with five haplotypes known to be present in other parts of the world, including the haplotype EG01 which is widespread and present in all endemic regions. These data extend knowledge related to the geographical spread and host range of E. granulosus s.s. in a country such as Australia in which the parasite established around 200 years ago.

Rel-deficient T cells exhibit defects in production of interleukin 3 and granulocyte-macrophage colony-stimulating factor.

Publisher: Proceedings of the National Academy of Sciences

Date: 16-04-1996

DOI: 10.1073/PNAS.93.8.3405

Abstract: The c-rel protooncogene encodes a subunit of the NF-kappa B-like family of transcription factors. Mice lacking Rel are defective in mitogenic activation of B and T lymphocytes and display impaired humoral immunity. In an attempt to identify changes in gene expression that accompany the T-cell stimulation defects associated with the loss of Rel, we have examined the expression of cell surface activation markers and cytokine production in mitogen-stimulated Rel-/- T cells. The expression of cell surface markers including the interleukin 2 receptor alpha (IL-2R alpha) chain (CD25), CD69 and L-selectin (CD62) is normal in mitogen-activated Rel-/- T cells, but cytokine production is impaired. In Rel-/- splenic T cell cultures stimulated with phorbol 12-myristate 13-acetate and ionomycin, the levels of IL-3, IL-5, granulocyte- macrophage colony-stimulating factor (GM-CSF), tumor necrosis factor alpha (TNF-alpha), and gamma interferon (IFN-gamma) were only 2- to 3-fold lower compared with normal T cells. In contrast, anti-CD3 and anti-CD28 stimulated Rel-/- T cells, which fail to proliferate, make little or no detectable cytokines. Exogenous IL-2, which restitutes the proliferative response of the anti-CD3- and anti-CD28-treated Rel-/- T cells, restores production of IL-5, TNF-alpha, and IFN-gamma, but not IL-3 and GM-CSF expression to approximately normal levels. In contrast to mitogen-activated Rel-/- T cells, lipopolysaccharide-stimulated Rel-/- macrophages produce higher than normal levels of GM-CSF. These findings establish that Rel can function as an activator or repressor of gene expression and is required by T lymphocytes for production of IL-3 and GM-CSF.

Defining immune memory resilience: implications for vaccine development

Publisher: Informa UK Limited

Date: 04-2010

DOI: 10.1586/ERV.10.14

Lymphatic System

Publisher: Wiley

Date: 14-05-2015

DOI: 10.1002/9780470015902.A0000523.PUB3

Techniques for the Diagnosis of Fasciola Infections in Animals

Publisher: Elsevier

Date: 2014

DOI: 10.1016/B978-0-12-800182-0.00002-7

Efficacy of DNA vaccination by different routes of immunisation in sheep

Publisher: Elsevier BV

Date: 11-2002

DOI: 10.1016/S0165-2427(02)00221-0

Abstract: DNA vaccination, delivered through various routes, has been used extensively in laboratory animals. Few studies have focused on veterinary species and while results obtained in laboratory animals can often be extrapolated to veterinary species this is not always the case. In this study we have compared the effect of the route of immunisation with DNA on the induction of immune responses and protection of sheep to challenge with live Corynebacterium pseudotuberculosis. Intramuscular injection of plasmid DNA encoding an inactivated form of the phospholipase D (PLD) antigen linked to CTLA4-Ig resulted in the induction of a strong memory response and sterile immunity following challenge in 45% of the animals. In contrast, gene gun delivery or subcutaneous (SC) injection of the DNA vaccine induced comparatively poor responses and insignificant levels of protection. Thus, DNA vaccine efficacy in sheep is strongly influenced by the route of vaccination. Amongst intramuscular vaccinates, protected sheep had significantly elevated IgG2 responses compared to unprotected animals, while both subgroups had equivalent IgG1 levels. This suggests that the presence of IgG2 antibodies and hence a Th1-like response, induced by the DNA vaccine gave rise to protective immunity against C. pseudotuberculosis.

Modelling the spread of resistance to Varroa in Australian feral honeybee populations

Publisher: Cold Spring Harbor Laboratory

Date: 10-02-2022

DOI: 10.1101/2022.02.10.479847

Abstract: Australia is the only country where Western honeybees, Apis mellifera , are not infested with the mite Varroa destructor . Hence, a collapse in the feral honeybee population, brought about by an incursion and spread of V. destructor , would have serious consequences for Australian horticulture given its dependence on managed and feral honeybees for pollination. Managing Varroa in commercial colonies is well understood and can be achieved, although at significant bee-health costs, by using miticides. Protecting the feral population in the event of a Varroa incursion is much more difficult, but nevertheless imperative. One way to mitigate against collapse of the feral population is to seed it with Varroa -resistant queens, so as to accelerate the spread of resistance. We developed a simulation model of the spread of Varroa -resistance in feral honeybee populations following the introduction of Varroa -resistant queens into the managed population. We show that, compared with a do-nothing scenario, seeding the managed honeybee population with Varroa -resistant queens was only effective in decreasing the length of time for the feral honeybee population to recover when the size of the managed resistant population was large compared to the size of the feral population. This situation may be achievable in some urban areas where numbers of managed colonies are high and habitat for feral colonies is limited.

Prospects for vector-based gene silencing to explore immunobiological features of schistosoma mansoni

Publisher: Elsevier

Date: 2015

DOI: 10.1016/BS.APAR.2015.02.002

Abstract: Schistosomiasis is a prevalent, socioeconomically important disease of humans caused by parasites of the genus Schistosoma (schistosomes or blood flukes). Currently, more than 200 million people worldwide are infected with schistosomes. Despite major research efforts, there is only one drug routinely used for effective treatment, and no vaccine is available to combat schistosomiasis. The purpose of the present article is to (1) provide a background on the parasites and different forms of disease (2) describe key immunomolecular aspects of disease induced in the host and (3) critically appraise functional genomic methods employed to explore parasite biology, parasite-host interactions and disease at the molecular level. Importantly, the article also describes the features and advantages of lentiviral delivery of artificial microRNAs to silence genes. It also discusses the first successful application of such an approach in schistosomes, in order to explore the immunobiological role of selected target proteins known to be involved in egg-induced disease. The lentiviral transduction system provides exciting prospects for future, fundamental investigations of schistosomes, and is likely to have broad applicability to other eukaryotic pathogens and infectious diseases. The ability to achieve effective and stable gene perturbation in parasites has major biotechnological implications, and might facilitate the development of radically new methods for the treatment and control of parasitic diseases.

Genome and transcriptome of the porcine whipworm Trichuris suis

Publisher: Springer Science and Business Media LLC

Date: 15-06-2014

DOI: 10.1038/NG.3012

Transcriptional analysis identifies key genes involved in metabolism, fibrosis/tissue repair and the immune response against Fasciola hepatica in sheep liver

Publisher: Springer Science and Business Media LLC

Date: 25-02-2015

DOI: 10.1186/S13071-015-0715-7

Activin A: From sometime reproductive factor to genuine cytokine

Publisher: Elsevier BV

Date: 10-2005

DOI: 10.1016/J.VETIMM.2005.08.011

Abstract: The growth factor, activin A, was initially characterized as a putative reproductive hormone but is now known to have many other ergent roles. One of these is during inflammation. Following intravenous injection of bacterial lipopolysaccharide (LPS) into sheep, activin A is released extremely rapidly into the circulation. The release of activin A appears to be independent of fever, prostaglandins or other key proinflammatory cytokines such as TNF-alpha or IL-1beta. While the precise roles and function of this factor in inflammation are yet to be elucidated, the activin response occurs in other mammalian species besides the sheep and elevated activin has been documented for a number of clinical inflammatory conditions. Activin A therefore seems to be part of the regulatory component of the innate immune response.

Biological activity of ovine IL-23 expressed using a foot-and-mouth disease virus 2A self-cleaving peptide

Publisher: Elsevier BV

Date: 03-2013

DOI: 10.1016/J.CYTO.2013.01.014

Abstract: Hetero-dimeric cytokines often require equi-molar expression of both subunits to achieve biological activity. Previously, we expressed ovine IL-12 p40 and p35 linked using a self-cleaving 2A peptide from foot-and-mouth disease virus (FMDV). We now generated a new improved vector for the expression of hetero-dimeric cytokines and demonstrate the more general applicability of this strategy by cloning and expressing ovine IL-23 using the 2A peptide to link IL-12/IL-23 p40 and p19. The resulting protein was shown to be biologically active when expressed in mammalian COS cells. IL-23 plays a significant role in the differentiation of Th17 cells as well as autoimmunity and the regulation of inflammatory processes. As such this reagents will be invaluable in the unravelling of regulation of the ovine immune system for both veterinary and human animal model applications.

Fetal Responses to Maternal and Intra-Amniotic Lipopolysaccharide Administration in Sheep1

Publisher: Oxford University Press (OUP)

Date: 05-2003

DOI: 10.1095/BIOLREPROD.102.009688

High intraspecific variability of Echinococcus granulosus sensu stricto in Chile

Publisher: Elsevier BV

Date: 04-2017

DOI: 10.1016/J.PARINT.2016.12.001

Abstract: Echinococcus granulosus sensu stricto is the major cause of cystic echinococcosis in most human and animal cases in the world and the most widespread species within the E. granulosus sensu lato complex. E. granulosus s.s. remains endemic in South America together with other species of the Echinococcus genus, especially in some areas in Argentina, Brazil, Chile and Peru. Except for a single human case caused by E. canadensis (G6) described in the literature, only E. granulosus s.s. has been found in the Chilean territory. In the current study 1609bp of the cox1 gene from 69 Chilean isolates of E. granulosus s.s. from humans and animals were analysed. In total, 26 cox1 haplotypes were found, including the widespread haplotype EG01 (22 isolates) and also EGp1 (5), EgRUS7 (1), EgAus02 (1) and EgAus03 (2). Twenty-one different haplotype not previously described were identified from 38 Chilean isolates designated EgCL1-EgCL21. Previous work had described low variability of E. granulosus s.s. in South America, based on isolates from Peru. Results obtained in this work challenge the previously described idea of the low ersity of the parasite in South America, and warrant future investigation on the origin and spread of the parasite in the continent after the Spanish arrival.

Vaccination against foot-and-mouth disease virus using peptides conjugated to nano-beads

Publisher: Elsevier BV

Date: 05-2008

DOI: 10.1016/J.VACCINE.2008.03.025

Abstract: Vaccination against foot-and-mouth disease virus (FMDV) is a major problem as current vaccines do not allow easy differentiation between infected and vaccinated animals. Furthermore, large scale production of inactivated virus poses significant risks. To address this we investigated the feasibility of using inert nano-beads that target antigen to dendritic cells (DCs) to induce immune responses against FMDV-specific synthetic peptides in sheep. Our results demonstrate that while single peptides induce responses in most sheep, the combination of multiple peptides either conjugated separately to in idual nano-beads or conjugated as a mixture induce significant cell-mediated (CM) and humoral immune responses.

Mucosal-Associated Invariant T Cells Augment Immunopathology and Gastritis in Chronic Helicobacter pylori Infection

Publisher: The American Association of Immunologists

Date: 03-2018

DOI: 10.4049/JIMMUNOL.1701512

Abstract: Mucosal-associated invariant T (MAIT) cells produce inflammatory cytokines and cytotoxic granzymes in response to by-products of microbial riboflavin synthesis. Although MAIT cells are protective against some pathogens, we reasoned that they might contribute to pathology in chronic bacterial infection. We observed MAIT cells in proximity to Helicobacter pylori bacteria in human gastric tissue, and so, using MR1-tetramers, we examined whether MAIT cells contribute to chronic gastritis in a mouse H. pylori SS1 infection model. Following infection, MAIT cells accumulated to high numbers in the gastric mucosa of wild-type C57BL/6 mice, and this was even more pronounced in MAIT TCR transgenic mice or in C57BL/6 mice where MAIT cells were preprimed by Ag exposure or prior infection. Gastric MAIT cells possessed an effector memory Tc1/Tc17 phenotype, and were associated with accelerated gastritis characterized by augmented recruitment of neutrophils, macrophages, dendritic cells, eosinophils, and non-MAIT T cells and by marked gastric atrophy. Similarly treated MR1−/− mice, which lack MAIT cells, showed significantly less gastric pathology. Thus, we demonstrate the pathogenic potential of MAIT cells in Helicobacter-associated immunopathology, with implications for other chronic bacterial infections.

Getting the most out of parasitic helminth transcriptomes using HelmDB: Implications for biology and biotechnology

Publisher: Elsevier BV

Date: 12-2013

DOI: 10.1016/J.BIOTECHADV.2012.12.004

Abstract: Compounded by a massive global food shortage, many parasitic diseases have a devastating, long-term impact on animal and human health and welfare worldwide. Parasitic helminths (worms) affect the health of billions of animals. Unlocking the systems biology of these neglected pathogens will underpin the design of new and improved interventions against them. Currently, the functional annotation of genomic and transcriptomic sequence data for socio-economically important parasitic worms relies almost exclusively on comparative bioinformatic analyses using model organism- and other databases. However, many genes and gene products of parasitic helminths (often >50%) cannot be annotated using this approach, because they are specific to parasites and/or do not have identifiable homologs in other organisms for which sequence data are available. This inability to fully annotate transcriptomes and predicted proteomes is a major challenge and constrains our understanding of the biology of parasites, interactions with their hosts and of parasitism and the pathogenesis of disease on a molecular level. In the present article, we compiled transcriptomic data sets of key, socioeconomically important parasitic helminths, and constructed and validated a curated database, called HelmDB (www.helmdb.org). We demonstrate how this database can be used effectively for the improvement of functional annotation by employing data integration and clustering. Importantly, HelmDB provides a practical and user-friendly toolkit for sequence browsing and comparative analyses among ergent helminth groups (including nematodes and trematodes), and should be readily adaptable and applicable to a wide range of other organisms. This web-based, integrative database should assist 'systems biology' studies of parasitic helminths, and the discovery and prioritization of novel drug and vaccine targets. This focus provides a pathway toward developing new and improved approaches for the treatment and control of parasitic diseases, with the potential for important biotechnological outcomes.

The Expression and Biologic Effects of Ovine Interleukin-4 on T and B Cell Proliferation

Publisher: Mary Ann Liebert Inc

Date: 04-2000

DOI: 10.1089/107999000312360

Abstract: Using the reverse-transcriptase polymerase chain reaction (RT-PCR), cDNA encoding ovine (Ov) interleukin-4 (OvIL-4) was generated from mitogen-stimulated peripheral blood mononuclear cells (PBMC). Two identical clones generated from separate RT-PCR reactions differed from a published OvIL-4 sequence, although they had a high degree of identity with the bovine and human homologs. We show by sequence analysis that the OvIL-4 cDNA retained the four alpha-helix structure and disulfide bonds identified in human IL-4 (HuIL-4). Moreover, the cDNA encoding OvIL-4 was expressed in insect cells using the baculovirus Autographa californica nuclear polyhedrosis virus (AcMNPV) as a vector. Supernatants from insect cells infected with the recombinant virus secreted an additional protein with a relative molecular mass of 17,000. This protein was recognized by an anticervine IL-4 monoclonal antibody (mAb) in a Western blot and did not react with any proteins in supernatants from uninfected insect cells or cells infected with the wild-type AcMNPV. Supernatants from insect cells infected with the recombinant virus induced the proliferation of activated B cells in a dose-dependent manner and typically demonstrated 5 x 105 dilution U/ml of activity. However, OvIL-4 had no effect on the proliferation of resting T cells isolated from efferent lymph and actually inhibited the ability of a mitogen to stimulate these resting lymphocytes. In contrast, OvIL-4 induced the proliferation of mitogen-activated lymphoblast, demonstrating the complex role(s) OvIL-4 plays in the regulation of B and T cells.

Comparing Sugar Shake to Alcohol Wash: Is Alcohol Wash the Gold Standard?

Publisher: Informa UK Limited

Date: 07-03-2022

DOI: 10.1080/0005772X.2022.2046367

Characterisation of local immune responses induced by a novel nano-particle based carrier-adjuvant in sheep

Publisher: Elsevier BV

Date: 09-2013

DOI: 10.1016/J.VETIMM.2013.05.015

Abstract: Most adjuvants require danger signals to promote immune responses against vaccine antigens. Our previous studies have characterised a powerful nano-particulate antigen delivery system, which by itself does not induce inflammation, and which further appears to induce substantial immune responses in mice and sheep without the requirement for added stimulators of toll like receptors or other pathogen recognition receptors. In the present study we dissect the nature of the early induction phase of the immune response stimulated by such a vaccine comprising 40 nm polystyrene nano-particles conjugated to the antigen. We analyse the kinetics of export from an in idual draining lymph node from the sheep, of antibodies and cytokines as well as antigen responsive CD4 and CD8 T cells. Our results indicate that simple inert nano-bead based antigen delivery into the draining area of the lymph node is highly efficient at priming combined humoral and T cell antigen specific immunity without the need for added 'danger signals'. Furthermore this nano-bead adjuvant is a potent agent capable of promoting cross-priming for CD8 T cell induction in sheep. Interestingly, using nano-beads, similarly to what has been observed with natural pathogen based lymph node stimulation, a phase of CD4 T cell priming and export preceded CD8 T cell induction, suggesting the engagement of natural priming processes and kinetics.

Phenotypic analysis of ovine antigen presenting cells loaded with nanoparticles migrating from the site of vaccination

Publisher: Elsevier BV

Date: 05-2013

DOI: 10.1016/J.YMETH.2013.02.008

Abstract: Virus-sized particulate adjuvants such as ISCOMs, polystyrene nanoparticles and virus-like particles have been shown to target dendritic cells, resulting in the activation of T and B cells in vivo. Using an ovine pseudo-afferent lymph cannulation model to capture APC that traffic from the site of injection to the local lymph node, we show that 40-50 nm nanoparticles are taken up at the site of injection by dendritic cells (DCs) migrating to the draining lymph node. These DCs can express CD11c, CD1b, CD5, MHC class II and CD8. Nanoparticles transported by DCs migrating from the site of injection to the local lymph node therefore needs to be considered as a new mechanism underlying the immunogenicity of virus-sized vaccine delivery systems.

The Role of Antigen Presentation and Innate Immunity During Immune Induction by Particulate Antigens

Publisher: Elsevier

Date: 2017

DOI: 10.1016/B978-0-323-39981-4.00005-1

Biodegradable and Biocompatible Poly(Ethylene Glycol)-based Hydrogel Films for the Regeneration of Corneal Endothelium

Publisher: Wiley

Date: 20-03-2014

DOI: 10.1002/ADHM.201400045

Abstract: Corneal endothelial cells (CECs) are responsible for maintaining the transparency of the human cornea. Loss of CECs results in blindness, requiring corneal transplantation. In this study, fabrication of biocompatible and biodegradable poly(ethylene glycol) (PEG)-based hydrogel films (PHFs) for the regeneration and transplantation of CECs is described. The 50-μm thin hydrogel films have similar or greater tensile strengths to human corneal tissue. Light transmission studies reveal that the films are >98% optically transparent, while in vitro degradation studies demonstrate their biodegradation characteristics. Cell culture studies demonstrate the regeneration of sheep corneal endothelium on the PHFs. Although sheep CECs do not regenerate in vivo, these cells proliferate on the films with natural morphology and become 100% confluent within 7 d. Implantation of the PHFs into live sheep corneas demonstrates the robustness of the films for surgical purposes. Regular slit l examinations and histology of the cornea after 28 d following surgery reveal minimal inflammatory responses and no toxicity, indicating that the films are benign. The results of this study suggest that PHFs are excellent candidates as platforms for the regeneration and transplantation of CECs as a result of their favorable biocompatibility, degradability, mechanical, and optical properties.

Omega-1 knockdown in Schistosoma mansoni eggs by lentivirus transduction reduces granuloma size in vivo

Publisher: Springer Science and Business Media LLC

Date: 17-11-2014

DOI: 10.1038/NCOMMS6375

Abstract: Schistosomiasis, one of the most important neglected tropical diseases worldwide, is caused by flatworms (blood flukes or schistosomes) that live in the bloodstream of humans. The hepatointestinal form of this debilitating disease results from a chronic infection with Schistosoma mansoni or Schistosoma japonicum . No vaccine is available to prevent schistosomiasis, and treatment relies predominantly on the use of a single drug, praziquantel. In spite of considerable research effort over the years, very little is known about the complex in vivo events that lead to granuloma formation and other pathological changes during infection. Here we use, for the first time, a lentivirus-based transduction system to deliver microRNA-adapted short hairpin RNAs (shRNAmirs) into the parasite to silence and explore selected protein-encoding genes of S. mansoni implicated in the disease process. This gene-silencing system has potential to be used for functional genomic–phenomic studies of a range of socioeconomically important pathogens.

Analysis of the transcriptome of adult Dictyocaulus filaria and comparison with Dictyocaulus viviparus, with a focus on molecules involved in host-parasite interactions

Publisher: Elsevier BV

Date: 03-2014

DOI: 10.1016/J.IJPARA.2013.12.003

Ovine Interleukin-12: Analysis of Biologic Function and Species Comparison

Publisher: Mary Ann Liebert Inc

Date: 06-2000

DOI: 10.1089/10799900050044750

Abstract: Interleukin-12 (IL-12) is a heterodimeric cytokine produced mainly by phagocytic and antigen-presenting cells (APC). The cDNA encoding the ovine IL-12 (OvIL-12) subunits, p40 and p35, were generated from concanavalin A (ConA)-stimulated peripheral blood mononuclear cells (PBMC). The ovine genes encoded proteins that had the highest amino acid identity to caprine p40 (99% amino acid identity) and p35 (97% amino acid identity) and also displayed a high degree of identity with human p40 (84%) and p35 (79%) homologs. To ensure the equal expression of both subunits, we used the self-cleaving properties of the 2A oligopeptide from foot-and-mouth disease virus (FMDV) to express IL-12 as a single, long open reading frame (ORF) encoding p402Ap35. Using an in vitro transcription/translation system, we demonstrated that this 2A oligopeptide mediated cleavage of the p402Ap35 into p402A and p35, in a manner similar to the processing of the FMDV polypeptide. Moreover, when expressed in COSm6 cells, this self-processing polypeptide encoded a functional heterodimer, which elicited biologic activities associated with IL-12 in other species.

Exploring local immune responses to vaccines using efferent lymphatic cannulation

Publisher: Informa UK Limited

Date: 16-01-2015

DOI: 10.1586/14760584.2015.1002475

Abstract: The early stages of the induction of a primary immune response to a vaccine can shape the overall quality of the immune memory generated and hence affect the success of the vaccine. This early interaction between a vaccine and the immune system occurs first at the site of vaccination and can be explored using afferent cannulation. Subsequently, the vaccine and adjuvant activates the local draining lymph node. These interactions can be studied in real time in vivo using efferent lymphatic duct cannulation in large animal models and are the subject of this review. Depending on how the vaccine is delivered, the draining lymph nodes of different organs can be accessed, facilitating the testing of tissue-specific vaccinations. The efferent lymphatic cannulation model provides an avenue to study the effect of both adjuvants and antigen on the local immune system, and hence opens a pathway toward developing more effective ways of inducing immunity.

A sheep cannulation model for evaluation of nasal vaccine delivery

Publisher: Elsevier BV

Date: 02-2006

DOI: 10.1016/J.YMETH.2005.09.011

Abstract: We have developed and validated a novel model to investigate the efficacy of nasal vaccine delivery. Based on lymphatic cannulation of the tracheal lymph trunk of sheep, the model allows collection of lymph draining from the Nasal Associated Lymphoid Tissue. The model is suitable for determining both the amount of material that is absorbed into the lymphatic system, following intra-nasal delivery and the immune response that occurs following vaccination into the nasal area. The cell populations that track in this duct were phenotyped and found to be similar to those previously reported to be present in efferent lymph draining from peripheral lymph nodes. Following intra-nasal spray, we demonstrated that the amount of material recovered in draining lymph is only a very small fraction of the total delivered. Nevertheless, intra-nasal spraying of a vaccine could activate local immune cells. The method described will be invaluable for optimising intra-nasal delivery systems by allowing a separate optimisation of vaccine uptake and immune responses induction.

Bm86 antigen induces a protective immune response against Boophilus microplus following DNA and protein vaccination in sheep

Publisher: Elsevier BV

Date: 11-1999

DOI: 10.1016/S0165-2427(99)00038-0

Abstract: Vaccination of sheep with a plasmid bearing the full length gene for the tick antigen Bm86 either alone or co-administered with plasmid carrying the ovine genes for the cytokines, granulocyte and macrophage colony stimulating factor (GM-CSF) or interleukin (IL)-1beta induced a relatively low level of protection against subsequent tick infestation. This tick damage reached statistical significance only for the groups which were vaccinated with plasmid encoding for Bm86, co-administered with plasmid encoding for ovine GM-CSF. Antibody titres measured against Bm86 were also low in all groups injected with the Bm86 DNA vaccine. Antibody production and anti-tick effect were significantly less than that achieved by two vaccinations with recombinant Bm86 protein. In all cases only a low level of antigen-specific stimulation of peripheral blood lymphocytes was recorded, as measured either by the incorporation of tritiated thymidine or the release of IFN-gamma. Injection of DNA encoding for Bm86, either alone or with co-administered cytokine genes, did however prime for a strong subsequent antibody response following a single injection of recombinant Bm86 protein in adjuvant. Antibody production nevertheless appeared to be slightly less effective than following two vaccinations with recombinant protein. The persistence of antibody following vaccination was the same regardless of the method of primary sensitization. In all cases the half-life of the antibody response was approximately 40-50 days indicating that, in contrast to results reported in mice, DNA vaccination in sheep did not result in sustained antibody production.

Gene gun immunization in a preclinical model is enhanced by B7 targeting.

Publisher: Elsevier BV

Date: 06-2003

DOI: 10.1016/S0264-410X(03)00162-2

Abstract: DNA vaccines have great potential but despite the promise shown in rodent models, responses in large animals, including humans, have been disappointing. Furthermore, gene gun delivery of DNA has been used to improve these responses. However, most cells that are transfected are not the professional antigen presenting cells (APC) which are critical for generating the primary immune response. Here, we show that in the large animal model of the pig, the combination of the use of gene gun delivery and a DNA vector that targets antigen presenting cells by expressing a CTLA4-ovalbumin (OVA) fusion antigen, leads to enhanced ovalbumin specific serum IgG, IgA, IgG1 and IgG2 immune responses.

Assessment of reference genes for reliable analysis of gene transcription by RT-qPCR in ovine leukocytes

Publisher: Elsevier BV

Date: 03-2016

DOI: 10.1016/J.VETIMM.2015.10.010

Abstract: With the availability of genetic sequencing data, quantitative reverse transcription PCR (RT-qPCR) is increasingly being used for the quantification of gene transcription across species. Too often there is little regard to the selection of reference genes and the impact that a poor choice has on data interpretation. Indeed, RT-qPCR provides a snapshot of relative gene transcription at a given time-point, and hence is highly dependent on the stability of the transcription of the reference gene(s). Using ovine efferent lymph cells and peripheral blood mono-nuclear cells (PBMCs), the two most frequently used leukocytes in immunological studies, we have compared the stability of transcription of the most commonly used ovine reference genes: YWHAZ, RPL-13A, PGK1, B2M, GAPDH, HPRT, SDHA and ACTB. Using established algorithms for reference gene normalization "geNorm" and "Norm Finder", PGK1, GAPDH and YWHAZ were deemed the most stably transcribed genes for efferent leukocytes and PGK1, YWHAZ and SDHA were optimal in PBMCs. These genes should therefore be considered for accurate and reproducible RT-qPCR data analysis of gene transcription in sheep.

Time-course study of the transcriptome of peripheral blood mononuclear cells (PBMCs) from sheep infected with Fasciola hepatica

Publisher: Public Library of Science (PLoS)

Date: 20-07-2016

DOI: 10.1371/JOURNAL.PONE.0159194

Systemic immune responses in sheep, induced by a novel nano-bead adjuvant

Publisher: Elsevier BV

Date: 20-02-2006

DOI: 10.1016/J.VACCINE.2005.09.009

Abstract: Although a number of adjuvants are currently approved for use in veterinary species, only alum has been widely used in humans. While it induces strong antibody responses, cell mediated responses are often low and inflammatory reactions at the site of injection are common. We investigated the immunological properties of a novel nano-bead adjuvant in a sheep large-animal model. In contrast to alum, antigen covalently coupled to nano-beads induced substantial cell mediated responses along with moderate humoral responses. No adverse reactions were seen at the site of immunisation in the sheep. Thus, nano-bead adjuvants in veterinary species may be useful for the induction of immunity to viral pathogens, where a cell mediated response is required. These findings also highlight the potential usefulness of nano-bead vaccines for intracellular pathogens in humans.

Enrichment of prion protein in exosomes derived from ovine cerebral spinal fluid

Publisher: Elsevier BV

Date: 08-2008

DOI: 10.1016/J.VETIMM.2008.04.002

Abstract: Prion diseases are transmissible neurodegenerative disorders affecting humans and a wide variety of animal species including sheep and cattle. The transmissible agent, the prion, is an abnormally folded form (PrP(Sc)) of the host encoded cellular prion protein (PrP(C)). Distribution of the prion protein in the fluids of species susceptible to these diseases is of importance to human health and the iatrogenic spread of prion disease. Aside from blood which is confirmed to be a source of prion infectivity, it is currently unclear which other body fluids harbor a significant transmission risk. In the current study we examined two ovine fluids pseudo-afferent lymph and cerebral spinal fluid (CSF), for the presence of exosomes and concurrent enrichment of the normal, cellular form of the prion protein (PrP(C)). Here we demonstrate the existence of exosomes in both pseudo-afferent lymph and CSF isolated from sheep. In the CSF derived exosomes we were able to show an enrichment of PrP(C) over unfractionated CSF. This experimental approach suggests that CSF derived exosomes could be used as a novel means of detecting abnormal forms of the prion protein and provide an in vivo link between these vesicles and prion disease pathogenesis.

Identification and characterization of an M cell marker in nasopharynx- and oropharynx-associated lymphoid tissue of sheep

Publisher: Elsevier BV

Date: 02-2019

DOI: 10.1016/J.VETIMM.2018.12.005

Abstract: M cells play a pivotal role in the induction of immune responses within the mucosa-associated lymphoid tissues. M cells exist principally in the follicle-associated epithelium (FAE) of the isolated solitary lymphoid follicles as well as in the lymphoid follicles of nasopharynx-associated lymphoid tissue and gut associated lymphoid tissue (GALT). Through lymphatic cannulation it is possible to investigate local immune responses induced following nasal Ag delivery in sheep. Hence, identifying sheep M cell markers would allow the targeting of M cells to offset the problem of trans-epithelial Ag delivery associated with inducing mucosal immunity. Sheep cDNA from the tonsils of the oropharynx and nasopharynx was PCR lified using Glycoprotein-2 (GP

Molecular Changes in Opisthorchis viverrini (Southeast Asian Liver Fluke) during the Transition from the Juvenile to the Adult Stage

Publisher: Public Library of Science (PLoS)

Date: 29-11-2012

DOI: 10.1371/JOURNAL.PNTD.0001916

Veterinary applications of cytokines

Publisher: Elsevier BV

Date: 10-2005

DOI: 10.1016/J.VETIMM.2005.08.001

Abstract: In the recent past a large variety of cytokines have been cloned for most important veterinary species and more is planned with development of a coordinated approach to cytokine reagents production. Application of these cytokines in veterinary species can be found in the development of effective diagnostics, with the IFN-gamma-based detection of tuberculosis as a prime ex le. In addition, cytokines have been used to determine which immune responses are essential for immune protection with flow-on effects for the development of novel ways to induce these specific immune responses. The realisation that the murine immune system is quite different from the human, together with the increased availability of cytokine reagents for many large animals plus unique experimental approaches only available in these animals, has lead to an explosion in the use of veterinary species as models for human diseases.

The Transcriptome of Trichuris suis - First Molecular Insights into a Parasite with Curative Properties for Key Immune Diseases of Humans

Publisher: Public Library of Science (PLoS)

Date: 24-08-2011

DOI: 10.1371/JOURNAL.PONE.0023590

Linkage Projects - Grant ID: LP0348578

Start Date: 03-2004

End Date: 12-2006

Amount: $300,000.00

Funder: Australian Research Council

Linkage Projects - Grant ID: LP0347058

Start Date: 05-2003

End Date: 12-2006

Amount: $390,000.00

Funder: Australian Research Council

Linkage Projects - Grant ID: LP0561957

Start Date: 02-2006

End Date: 12-2008

Amount: $405,000.00

Funder: Australian Research Council

Discovery Projects - Grant ID: DP0663084

Start Date: 2006

End Date: 12-2008

Amount: $254,000.00

Funder: Australian Research Council

Discovery Projects - Grant ID: DP1094465

Start Date: 2010

End Date: 12-2013

Amount: $360,000.00

Funder: Australian Research Council

Linkage Projects - Grant ID: LP0211604

Start Date: 2002

End Date: 12-2005

Amount: $394,345.00

Funder: Australian Research Council

Discovery Projects - Grant ID: DP110103175

Start Date: 01-2011

End Date: 12-2014

Amount: $420,000.00

Funder: Australian Research Council

Linkage Infrastructure, Equipment And Facilities - Grant ID: LE0775612

Start Date: 2007

End Date: 12-2008

Amount: $700,000.00

Funder: Australian Research Council

Linkage Projects - Grant ID: LP100200728

Start Date: 01-2011

End Date: 01-2014

Amount: $331,521.00

Funder: Australian Research Council