Murine brain endothelial cells differently modulate interferon-γ and interleukin-17 production in vitro

Brain endothelial cells (BEC) are the major constituents of the blood-brain barrier (BBB), the structure that controls entrance of immune cells into CNS parenchyma. Our aim was to investigate the influence of BEC on production of IL-17 and IFN-γ-cytokines that are important for CNS inflammation. To that end, co-cultivations of the bEnd.3 brain endothelial cell line and lymph node cells (LNC) were performed, and gene expression and production of IL-17 and IFN-γ were determined. It was found that bEnd.3 cells inhibited expression and production of IFN-γ, but not of IL-17. Additionally, bEnd.3 cells also reduced production of the major IFN-γ-promoting cytokine - IL-12 - in LNC. The observed variation in modulation of pro-inflammatory cytokines by BEC could be of importance for the understanding of CNS inflammation

Micro RNA-155 participates in re-activation of encephalitogenic T cells

MicroRNAs (miR) are small non-coding RNAs involved in the immune response regulation. miR-155 has been attributed a major pro-inflammatory role in the pathogenesis of multiple sclerosis and its animal model experimental autoimmune encephalomyelitis (EAE). Here, a role of miR-155 in re-activation of encephalitogenic CD4(+) T cells was investigated. Dark Agouti rats were immunized with myelin basic protein (MBP) emulsified in complete Freund's adjuvant. CD4(+) T cells were purified from draining lymph node cells (DLNC) obtained in the inductive phase and from spinal cord immune cells (SCIC) isolated at the peak of EAE. CD4(+) T cells obtained from SCIC (i.e., in vivo re-activated cells) had markedly higher expression of miR-155 in comparison to those purified from DLNC (not re-activated). Likewise, in vitro re-activation of DLNC with MBP led to increase in miR-155 expression. Further, DLNC and DLNC CD4(+) T cells were transfected with an inhibitor of miR-155 during in vitro re-activation. As a result, expression of important CD4(+) T cell effector cytokines IFN-gamma and IL-17, but not of regulatory cytokines IL-10 and TGF-beta, was reduced. These results imply that miR-155 supports re-activation of encephalitogenic CD4+ T cells. Our results contribute to a view that miR-155 might be a valuable target in multiple sclerosis therapy

Pentoxifylline Prevents Autoimmune Mediated Inflammation in Low Dose Streptozotocin Induced Diabetes

Xanthine derivative, pentoxifylline (PTX), has been recently shown to exert a protective effects in certain animal models of autoimmunity, including diabetes in NOD mice. In the present study, the immunomodulatory potential of PTX was investigated in autoimmune diabetes induced by multiple low doses of streptozotocin (MLD-SZ) in genetically susceptible CBA/H mice (tested with 40 mg SZ/kg b.w. for 5 days) and DA rats (tested with 20 mg/kg b.w. for 5 days). In both species, 2 – 3 weeks following the MLD-SZ treatment, sustained hyperglycemia developed, as an outcome of inflammatory reaction with endothelial cell activation and accumulation of mononuclear cells. Although there was no evidence of typical insulitis in early disease development (day 10), in both rats and mice, macrophages, CD4+ and CD8+ cells were present in the islets of Langerhans as diffuse mononuclear infiltrates with the expression of IFN-γ and inducible NO synthase (iNOS). Administration of PTX (200 mg/kg/day for 10 days) in combination with MLD-SZ reduced insulitis and the production of mediators tested, and prevented the development of hyperglycemia. These results suggest that beneficial effects of PTX involve down-regulation of local proinflammatory cytokine-mediated NO synthase pathway. They also demonstrate that in addition to ameliorating spontaneous autoimmunity in NOD mice, PTX may be effective in downregulating an inflammatory autoimmune process triggered in susceptible host by an external agents, such as streptozotocin

Correlation of Gut Microbiota Composition with Resistance to Experimental Autoimmune Encephalomyelitis in Rats

Multiple sclerosis is a chronic inflammatory disease of the central nervous system (CNS). It is widely accepted that autoimmune response against the antigens of the CNS is the essential pathogenic force in the disease. It has recently become increasingly appreciated that activated encephalitogenic cells tend to migrate toward gut associated lymphoid tissues (GALTs) and that interrupted balance between regulatory and inflammatory immunity within the GALT might have decisive role in the initiation and propagation of the CNS autoimmunity. Gut microbiota composition and function has the major impact on the balance in the GALT. Thus, our aim was to perform analyses of gut microbiota in experimental autoimmune encephalomyelitis (EAE), an animal model of multiple sclerosis. Albino Oxford (AO) rats that are highly resistant to EAE induction and Dark Agouti (DA) rats that develop EAE after mild immunization were compared for gut microbiota composition in different phases after EAE induction. Microbial analyses of the genus Lactobacillus and related lactic acid bacteria showed higher diversity of Lactobacillus spp. in EAE-resistant AO rats, while some members of Firmicutes and Proteobactene (Undibacterium oligocarboniphilum) were detected only in feces of DA rats at the peak of the disease (between 13 and 16 days after induction). Interestingly, in contrast to our previous study where Turicibacter sp. was found exclusively in non immunized AO, but not in DA rats, in this study it was detected in DA rats that remained healthy 16 days after induction, as well as in four of 12 DA rats at the peak of the disease. Similar observation was obtained for the members of Lachnospiraceae. Further, production of a typical regulatory cytokine interleukin-10 was compared in GALT cells of AO and DA rats, and higher production was observed in DA rats. Our data contribute to the idea that gut microbiota and GALT considerably influence multiple sclerosis pathogenesis

Methylprednisolone inhibits interleukin-17 and interferon-gamma expression by both naive and primed T cells

<p>Abstract</p> <p>Background</p> <p>Interleukin-17 (IL-17)-producing cells are increasingly considered to be the major pathogenic population in various autoimmune disorders. The effects of glucocorticoids, widely used as therapeutics for inflammatory and autoimmune disorders, on IL-17 generation have not been thoroughly investigated so far. Therefore, we have explored the influence of methylprednisolone (MP) on IL-17 expression in rat lymphocytes, and compared it to the effect of the drug on interferon (IFN)-γ.</p> <p>Results</p> <p>Production of IL-17 in mitogen-stimulated lymph node cells (LNC) from non-treated rats, as well as in myelin basic protein (MBP)-stimulated draining LNC from rats immunized with spinal cord homogenate and complete Freund's adjuvant was significantly reduced by MP. The reduction was dose-dependent, sustained through the follow-up period of 48 hours, and was not achieved through anti-proliferative effect. Additionally, MP inhibited IL-17 production in purified T cells as well, but to less extent than in LNC. In its influence on IL-17 production MP inhibited Ror-γT transcription factor expression, as well as Jun phosphorylation, but not ERK or p38 activation in mitogen-stimulated LNC. Importantly, MP collaborated with IFN-γ in inhibiting IL-17 generation in LNC.</p> <p>Conclusion</p> <p>The observed difference in the effect of MP on IL-17 and IFN-γ could be important for the understanding of the variability in the efficiency of glucocorticoids in the treatment of autoimmune diseases.</p

Protective effects of carbonyl iron against multiple low-dose streptozotocin-induced diabetes in rodents.

Particulate adjuvants have shown increasing promise as effective, safe, and durable agents for the stimulation of immunity, or alternatively, the suppression of autoimmunity. Here we examined the potential of the adjuvant carbonyl iron (CI) for the modulation of organ-specific autoimmune disease-type 1 diabetes (T1D). T1D was induced by multiple low doses of streptozotocin (MLDS) that initiates beta cell death and triggers immune cell infiltration into the pancreatic islets. The results of this study indicate that the single in vivo application of CI to MLDS-treated DA rats, CBA/H mice, or C57BL/6 mice successfully counteracted the development of insulitis and hyperglycemia. The protective action was obtained either when CI was applied 7 days before, simultaneously with the first dose of streptozotocin, or 1 day after MLDS treatment. Ex vivo cell analysis of C57BL/6 mice showed that CI treatment reduced the proportion of proinflammatory F4/80+ CD40+ M1 macrophages and activated T lymphocytes in the spleen. Moreover, the treatment down-regulated the number of inflammatory CD4+ IFN-γ+ cells in pancreatic lymph nodes, Peyer's patches, and pancreas-infiltrating mononuclear cells, while simultaneously potentiating proportion of CD4+ IL17+ cells. The regulatory arm of the immune system represented by CD3+ NK1.1+ (NKT) and CD4+ CD25+ FoxP3+ regulatory T cells was potentiated after CI treatment. In vitro analysis showed that CI down-regulated CD40 and CD80 expression on dendritic cells thus probably interfering with their antigen-presenting ability. In conclusion, particulate adjuvant CI seems to suppress the activation of the innate immune response, which further affects the adaptive immune response directed toward pancreatic beta cells

Therapeutic effect of nucleoside analogs on experimental autoimmune encephalomyelitis in dark agouti rats

Experimental autoimmune encephalomyelitis (EAE) is a commonly used animal model of the human neurological disorder multiple sclerosis. The purpose of the present study was to investigate the effect of combined treatment with two nucleoside analogs, ribavirin and tiazofurin, on development of EAE actively induced in highly susceptible dark agouti rats. The obtained results showed that ribavirin and tiazofurin applied either separately or in combination from the onset of the firstsymptoms of EAE after its induction (therapeutic treatment) significantly suppressed EAE’s clinical symptoms. However, the most pronounced effect was gained with combined treatment, probably as a result of synergistic/additive action

Comparison of dendritic cells obtained from autoimmunty-prone and resistant rats.

Dendritic cells (DC) are responsible for the initiation and shaping of the adaptive immune response and are in the focus of autoimmunity research. We were interested in comparison of DC obtained from autoimmunity-prone Dark Agouti (DA) rats and autoimmunity-resistant Albino Oxford (AO) rats. DC were generated from bone marrow precursors and matured (mDC) by lipopolysaccharide. Tolerogenic DC (tolDC) obtained by vitamin D3 treatment were studied in parallel. Profile of cytokine production was different in AO and DA mDC and tolDC. Expression of MHC class II molecules and CD86 were higher in DA DC, while vitamin D3 reduced their expression in dendritic cells of both strains. Allogeneic proliferation of CD4+ T cells was reduced by AO tolDC, but not with DA tolDC in comparison to respective mDC. Finally, expression of various genes identified as differentially expressed in human mDC and tolDC was also analyzed in AO and DA DC. Again, AO and DA DC differed in the expression of the analyzed genes. To conclude, AO and DA DC differ in production of cytokines, expression of antigen presentation-related molecules and in regulation of CD4+ T proliferation. The difference is valuable for understanding the divergence of the strains in their susceptibility to autoimmunity