Neuroprotection in a Novel Mouse Model of Multiple Sclerosis

The authors acknowledge the support of the Barts and the London Charity, the Multiple Sclerosis Society of Great Britain and Northern Ireland, the National Multiple Sclerosis Society, USA, notably the National Centre for the Replacement, Refinement & Reduction of Animals in Research, and the Wellcome Trust (grant no. 092539 to ZA). The siRNA was provided by Quark Pharmaceuticals. The funders and Quark Pharmaceuticals had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript

CXCL1 can be regulated by IL-6 and promotes granulocyte adhesion to brain capillaries during bacterial toxin exposure and encephalomyelitis

<p>Abstract</p> <p>Background</p> <p>Granulocytes generally exert protective roles in the central nervous system (CNS), but recent studies suggest that they can be detrimental in experimental autoimmune encephalomyelitis (EAE), the most common model of multiple sclerosis. While the cytokines and adhesion molecules involved in granulocyte adhesion to the brain vasculature have started to be elucidated, the required chemokines remain undetermined.</p> <p>Methods</p> <p>CXCR2 ligand expression was examined in the CNS of mice suffering from EAE or exposed to bacterial toxins by quantitative RT-PCR and <it>in situ </it>hybridization. CXCL1 expression was analyzed in IL-6-treated endothelial cell cultures by quantitative RT-PCR and ELISA. Granulocytes were counted in the brain vasculature after treatment with a neutralizing anti-CXCL1 antibody using stereological techniques.</p> <p>Results</p> <p>CXCL1 was the most highly expressed ligand of the granulocyte receptor CXCR2 in the CNS of mice subjected to EAE or infused with lipopolysaccharide (LPS) or pertussis toxin (PTX), the latter being commonly used to induce EAE. IL-6 upregulated CXCL1 expression in brain endothelial cells by acting transcriptionally and mediated the stimulatory effect of PTX on CXCL1 expression. The anti-CXCL1 antibody reduced granulocyte adhesion to brain capillaries in the three conditions under study. Importantly, it attenuated EAE severity when given daily for a week during the effector phase of the disease.</p> <p>Conclusions</p> <p>This study identifies CXCL1 not only as a key regulator of granulocyte recruitment into the CNS, but also as a new potential target for the treatment of neuroinflammatory diseases such as multiple sclerosis.</p

Diazoxide attenuates autoimmune encephalomyelitis and modulates lymphocyte proliferation and dendritic cell functionality

Activation of mitochondrial ATP-sensitive potassium (KATP) channels is postulated as an effective mechanism to confer cardio and neuroprotection, especially in situations associated to oxidative stress. Pharmacological activation of these channels inhibits glia-mediated neuroinflammation. In this way, diazoxide, an old-known mitochondrial KATP channel opener, has been proposed as an effective and safe treatment for different neurodegenerative diseases, demonstrating efficacy in different animal models, including the experimental autoimmune encephalomyelitis (EAE), an animal model for Multiple Sclerosis. Although neuroprotection and modulation of glial reactivity could alone explain the positive effects of diazoxide administration in EAE mice, little is known of its effects on the immune system and the autoimmune reaction that triggers the EAE pathology. The aim of the present work was to study the effects of diazoxide in autoimmune key processes related with EAE, such as antigen presentation and lymphocyte activation and proliferation. Results show that, although diazoxide treatment inhibited in vitro and ex-vivo lymphocyte proliferation from whole splenocytes it had no effect in isolated CD4(+) T cells. In any case, treatment had no impact in lymphocyte activation. Diazoxide can also slightly decrease CD83, CD80, CD86 and major histocompatibility complex class II expression in cultured dendritic cells, demonstrating a possible role in modulating antigen presentation. Taken together, our results indicate that diazoxide treatment attenuates autoimmune encephalomyelitis pathology without immunosuppressive effect

Erythropoietin: A potent inducer of peripheral immuno/inflammatory modulation in autoimmune EAE

Background: Beneficial effects of short-term erythropoietin (EPO) theraphy have been demonstrated in several animal models of acute neurologic injury, including experimental autoimmune encephalomyelitis(EAE)-the animal model of multiple sclerosis. We have found that EPO treatment substantially reduces the acute clinical paralysis seen EAE mice and this improvements is accompanied by a large reduction in the mononuclear cell infiltration and downregulation of glial MHC class II expression within the inflamed CNS. Other reports have recently indicated that peripherally generated anti-inflammatory CD4 +Foxp3 3 regulatory T cells (Tregs) and the IL17-producing CD4+ T helper cell (Th17) subpopulations play key antagonistic roles in EAE pathogenesis. However, no information regardind the effects of EPO theraphy on the behavior of the general mononuclear-lymphocyte population, Tregs or Th17 cells in EAE has emerged. Methods and Findings: We first determined in vivo that EPO theraphy markedly suppressed MOG specific T cell proliferation and sharply reduced the number of reactive dendritic cells (CD11c positive) in EAE lumph modes during both inductive and later symptomatic phases of MOG 35-55 induced EAE. We then determined the effect in vivo of EPO on numbers of peripheral Treg cells and Th17 cells. We found that EPO treatment modulated immune balance in both the periphery and the inflamed spinal cord by promoting a large expansion in Treg cells, inhibiting Th17 polarization and abrogating proliferation of the antigen presenting dendritic cell population. Finally we utilized tissue culture assays to show that exposure to EPO in vitro similarly downregulated MOG-specific T cell proliferation and also greatly suppressed T cell production of pro-inflammatory cytokines. Conclusions: Taken together, our findings reveal an important new locus whereby EPO induces substantial long-term tissue protection in the host through signalling to several critical subsets of immune cells that reside in the peripheral lymphatic system.published_or_final_versio

Much Ado About the TPP’s Effect on Pharmaceuticals

Ocular antigens are sequestered behind the blood-retina barrier and the ocular environment protects ocular tissues from autoimmune attack. The signals required to activate autoreactive T cells and allow them to cause disease in the eye remain in part unclear. In particular, the consequences of peripheral presentation of ocular antigens are not fully understood. We examined peripheral expression and presentation of ocular neo-self-antigen in transgenic mice expressing hen egg lysozyme (HEL) under a retina-specific promoter. High levels of HEL were expressed in the eye compared to low expression throughout the lymphoid system. Adoptively transferred naïve HEL-specific CD4+ T cells proliferated in the eye draining lymph nodes, but did not induce uveitis. By contrast, systemic infection with a murine cytomegalovirus (MCMV) engineered to express HEL induced extensive proliferation of transferred naïve CD4+ T cells, and significant uveoretinitis. In this model, wild-type MCMV, lacking HEL, did not induce overt uveitis, suggesting that disease is mediated by antigen-specific peripherally activated CD4+ T cells that infiltrate the retina. Our results demonstrate that retinal antigen is presented to T cells in the periphery under physiological conditions. However, when the same antigen is presented during viral infection, antigen-specific T cells access the retina and autoimmune uveitis ensues

Pharmacological prion protein silencing accelerates central nervous system autoimmune disease via T cell receptor signalling

The primary biological function of the endogenous cellular prion protein has remained unclear. We investigated its biological function in the generation of cellular immune responses using cellular prion protein gene-specific small interfering ribonucleic acid in vivo and in vitro. Our results were confirmed by blocking cellular prion protein with monovalent antibodies and by using cellular prion protein-deficient and -transgenic mice. In vivo prion protein gene-small interfering ribonucleic acid treatment effects were of limited duration, restricted to secondary lymphoid organs and resulted in a 70% reduction of cellular prion protein expression in leukocytes. Disruption of cellular prion protein signalling augmented antigen-specific activation and proliferation, and enhanced T cell receptor signalling, resulting in zeta-chain-associated protein-70 phosphorylation and nuclear factor of activated T cells/activator protein 1 transcriptional activity. In vivo prion protein gene-small interfering ribonucleic acid treatment promoted T cell differentiation towards pro-inflammatory phenotypes and increased survival of antigen-specific T cells. Cellular prion protein silencing with small interfering ribonucleic acid also resulted in the worsening of actively induced and adoptively transferred experimental autoimmune encephalomyelitis. Finally, treatment of myelin basic protein1–11 T cell receptor transgenic mice with prion protein gene-small interfering ribonucleic acid resulted in spontaneous experimental autoimmune encephalomyelitis. Thus, central nervous system autoimmune disease was modulated at all stages of disease: the generation of the T cell effector response, the elicitation of T effector function and the perpetuation of cellular immune responses. Our findings indicate that cellular prion protein regulates T cell receptor-mediated T cell activation, differentiation and survival. Defects in autoimmunity are restricted to the immune system and not the central nervous system. Our data identify cellular prion protein as a regulator of cellular immunological homoeostasis and suggest cellular prion protein as a novel potential target for therapeutic immunomodulation

Regulating STING in health and disease.

The presence of cytosolic double-stranded DNA molecules can trigger multiple innate immune signalling pathways which converge on the activation of an ER-resident innate immune adaptor named "STimulator of INterferon Genes (STING)". STING has been found to mediate type I interferon response downstream of cyclic dinucleotides and a number of DNA and RNA inducing signalling pathway. In addition to its physiological function, a rapidly increasing body of literature highlights the role for STING in human disease where variants of the STING proteins, as well as dysregulated STING signalling, have been implicated in a number of inflammatory diseases. This review will summarise the recent structural and functional findings of STING, and discuss how STING research has promoted the development of novel therapeutic approaches and experimental tools to improve treatment of tumour and autoimmune diseases

DQB1*0602 rather than DRB1*1501 confers susceptibility to multiple sclerosis-like disease induced by proteolipid protein (PLP)

<p>Abstract</p> <p>Background</p> <p>Multiple sclerosis (MS) is associated with pathogenic autoimmunity primarily focused on major CNS-myelin target antigens including myelin basic protein (MBP), proteolipidprotein (PLP), myelin oligodendrocyte protein (MOG). MS is a complex trait whereby the HLA genes, particularly class-II genes of HLA-DR15 haplotype, dominate the genetic contribution to disease-risk. Due to strong linkage disequilibrium in HLA-II region, it has been hard to establish precisely whether the functionally relevant effect derives from the DRB1*1501, DQA1*0102-DQB1*0602, or DRB5*0101 loci of HLA-DR15 haplotype, their combinations, or their epistatic interactions. Nevertheless, most genetic studies have indicated DRB1*1501 as a primary risk factor in MS. Here, we used 'HLA-humanized' mice to discern the potential relative contribution of DRB1*1501 and DQB1*0602 alleles to susceptibility to "humanized" MS-like disease induced by PLP, one of the most prominent and encephalitogenic target-antigens implicated in human MS.</p> <p>Methods</p> <p>The HLA-DRB1*1501- and HLA-DQB1*0602-Tg mice (MHC-II^-/-), and control non-HLA-DR15-relevant-Tg mice were immunized with a set of overlapping PLP peptides or with recombinant soluble PLP for induction of "humanized" MS-like disease, as well as for ex-vivo analysis of immunogenic/immunodominant HLA-restricted T-cell epitopes and associated cytokine secretion profile.</p> <p>Results</p> <p>PLP autoimmunity in both HLA-DR15-Tg mice was focused on 139-151 and 175-194 epitopes. Strikingly, however, the HLA-DRB1*1501-transgenics were refractory to disease induction by any of the overlapping PLP peptides, while HLA-DQB1*0602 transgenics were susceptible to disease induction by PLP139-151 and PLP175-194 peptides. Although both transgenics responded to both peptides, the PLP139-151- and PLP175-194-reactive T-cells were directed to Th1/Th17 phenotype in DQB1*0602-Tg mice and towards Th2 in DRB1*1501-Tg mice.</p> <p>Conclusions</p> <p>While genome studies map a strong MS susceptibility effect to the region of DRB1*1501, our findings offer a rationale for potential involvement of pathogenic DQ6-associated autoimmunity in MS. Moreover, that DQB1*0602, but not DRB1*1501, determines disease-susceptibility to PLP in HLA-transgenics, suggests a potential differential, functional role for DQB1*0602 as a predisposing allele in MS. This, together with previously demonstrated disease-susceptibility to MBP and MOG in DRB1*1501-transgenics, also suggests a differential role for DRB1*1501 and DQB1*0602 depending on target antigen and imply a potential complex 'genotype/target antigen/phenotype' relationship in MS heterogeneity.</p