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

GM-CSF-Producing Th Cells in Rats Sensitive and Resistant to Experimental Autoimmune Encephalomyelitis

Given that granulocyte macrophage colony-stimulating factor (GM-CSF) is identified as the key factor to endow auto-reactive Th cells with the potential to induce neuroinflammation in experimental autoimmune encephalomyelitis (EAE) models, the frequency and phenotype of GM-CSF-producing (GM-CSF+) Th cells in draining lymph nodes (dLNs) and spinal cord (SC) of Albino Oxford (AO) and Dark Agouti (DA) rats immunized for EAE were examined. The generation of neuroantigen-specific GM-CSF+ Th lymphocytes was impaired in dLNs of AO rats (relatively resistant to EAE induction) compared with their DA counterparts (susceptible to EAE) reflecting impaired CD4+ lymphocyte proliferation and less supportive of GM-CSF+ Th cell differentiation dLN cytokine microenvironment. Immunophenotyping of GM-CSF+ Th cells showed their phenotypic heterogeneity in both strains and revealed lower frequency of IL-17+ IFN-gamma+, IL-17+ IFN-gamma-, and IL-17-IFN-gamma+ cells accompanied by higher frequency of IL-17-IFN-gamma- cells among them in AO than in DA rats. Compared with DA, in AO rats was also found (i) slightly lower surface density of CCR2 (drives accumulation of highly pathogenic GM-CSF+ IFN-gamma+ Th17 cells in SC) on GM-CSF+ IFN-gamma+ Th17 lymphocytes from dLNs, and (ii) diminished CCL2 mRNA expression in SC tissue, suggesting their impaired migration into the SC. Moreover, dLN and SC cytokine environments in AO rats were shown to be less supportive of GM-CSF+ IFN-gamma+ Th17 cell differentiation (judging by lower expression of mRNAs for IL-1 beta, IL-6 and IL-23/p19). In accordance with the (i) lower frequency of GM-CSF+ Th cells in dLNs and SC of AO rats and their lower GM-CSF production, and (ii) impaired CCL2 expression in the SC tissue, the proportion of proinflammatory monocytes among peripheral blood cells and their progeny (CD45(hi) cells) among the SC CD11b+ cells were reduced in AO compared with DA rats. Collectively, the results indicate that the strain specificities in efficacy of several mechanisms controlling (auto) reactive CD4+ lymphocyte expansion/differentiation into the cells with pathogenic phenotype and migration of the latter to the SC contribute to AO rat resistance to EAE

Aging diminishes the resistance of AO rats to EAE: putative role of enhanced generation of GM-CSF Expressing CD4+T cells in aged rats

Background: Aging influences immune response and susceptibility to EAE in a strain specific manner. The study was designed to examine influence of aging on EAE induction in Albino Oxford (AO) rats. Results: Differently from 3-month-old (young) rats, which were resistant to EAE induction, the majority of aged (24-26-month-old) rats developed mild chronic form of EAE. On 16th day post-immunization, when in aged rats the neurological deficit reached plateau, more mononuclear cells, including CD4+ T lymphocytes was retrieved from spinal cord of aged than young rats. The frequencies of IL-17+ and GM-CSF+ cells within spinal cord infiltrating CD4+ lymphocytes were greater in aged rats. To their increased frequency contributed the expansion of GM-CSF + IL-17 + IFN-gamma+ cells, which are highly pathogenic in mice. The expression of the cytokines (IL-1 beta and IL-23/p19) driving GM-CSF + IL-17 + IFN-gamma + cell differentiation in mice was also augmented in aged rat spinal cord mononuclear cells. Additionally, in aged rat spinal cord the expansion of GM-CSF + IL-17-IFN-gamma- CD4+ T lymphocytes was found. Consistently, the expression of mRNAs for IL-3, the cytokine exhibiting the same expression pattern as GM-CSF, and IL-7, the cytokine driving differentiation of GM-CSF + IL-17-IFN-gamma- CD4 + lymphocytes in mice, was upregulated in aged rat spinal cord mononuclear cells, and the tissue, respectively. This was in accordance with the enhanced generation of the brain antigen-specific GM-CSF+ CD4+ lymphocytes in aged rat draining lymph nodes, as suggested by (i) the higher frequency of GM-CSF+ cells (reflecting the expansion of IL-17-IFN-gamma- cells) within their CD4+ lymphocytes and (ii) the upregulated GM-CSF and IL-3 mRNA expression in fresh CD4+ lymphocytes and MBP-stimulated draining lymph node cells and IL-7 mRNA in lymph node tissue from aged rats. In agreement with the upregulated GM-CSF expression in aged rats, strikingly more CD11b + CD45(int) (activated microglia) and CD45(hi) (mainly proinflammatory dendritic cells and macrophages) cells was retrieved from aged than young rat spinal cord. Besides, expression of mRNA for SOCS1, a negative regulator of proinflammatory cytokine expression in innate immunity cells, was downregulated in aged rat spinal cord mononuclear cells. Conclusions: The study revealed that aging may overcome genetic resistance to EAE, and indicated the cellular and molecular mechanisms contributing to this phenomenon in AO rats

Kinetics of IFN-gamma and IL-17 expression and production in active experimental autoimmune encephalomyelitis in Dark Agouti rats

Interferon-gamma (IFN-gamma) and interleukin-17 (IL-17) have been involved in the pathogenesis of experimental autoimmune encephalomyelitis (EAE). We have carried out a follow-up study of the expression and production of these cytokines, as well as of cells expressing these cytokines during the course of active EAE in Dark Agouti (DA) rats. As a result, IL-17, but not IFN-gamma expression and production had the peak value in draining lymph nodes (DLN) during the induction phase of the disease, and in spinal cords (SC) at the onset of clinical signs of the disease, and then declined toward the resolution of the disease. Also, a significant proportion of IFN-gamma/IL-17 double-positive cells was observed in SC of DA rats in active EAE. Importantly, the highest proportion of IL-17 single positive and double-positive cells, but not of IFN-gamma single positive cells, was observed at the onset of the disease. The observed difference in the kinetics of IFN-gamma and IL-17 expression during active EAE in DA rats suggests different roles these cytokines might have in the pathogenesis of the disease. (C) 2008 Elsevier Ireland Ltd. All rights reserved.Serbian Ministry of Science [1430296, 1450666]; Alexander von Humboldt Foundation (Bonn, Germany

Kinetics of IFN-gamma and IL-17 expression and production in active experimental autoimmune encephalomyelitis in Dark Agouti rats

Interferon-gamma (IFN-gamma) and interleukin-17 (IL-17) have been involved in the pathogenesis of experimental autoimmune encephalomyelitis (EAE). We have carried out a follow-up study of the expression and production of these cytokines, as well as of cells expressing these cytokines during the course of active EAE in Dark Agouti (DA) rats. As a result, IL-17, but not IFN-gamma expression and production had the peak value in draining lymph nodes (DLN) during the induction phase of the disease, and in spinal cords (SC) at the onset of clinical signs of the disease, and then declined toward the resolution of the disease. Also, a significant proportion of IFN-gamma/IL-17 double-positive cells was observed in SC of DA rats in active EAE. Importantly, the highest proportion of IL-17 single positive and double-positive cells, but not of IFN-gamma single positive cells, was observed at the onset of the disease. The observed difference in the kinetics of IFN-gamma and IL-17 expression during active EAE in DA rats suggests different roles these cytokines might have in the pathogenesis of the disease. (C) 2008 Elsevier Ireland Ltd. All rights reserved.Serbian Ministry of Science [1430296, 1450666]; Alexander von Humboldt Foundation (Bonn, Germany

It is still not for the old iron: adjuvant effects of carbonyl iron in experimental autoimmune encephalomyelitis induction

Experimental autoimmune encephalomyelitis (EAE) is a model of multiple sclerosis. Dark Agouti rats immunized with spinal cord homogenate (SCH) and carbonyl iron (CI), as an adjuvant, develop severe hyperacute form of EAE. They succumb to EAE earlier and have higher clinical scores and lethality rate in comparison to counterparts immunized with SCH + complete Freund's adjuvant. There is no difference in the number of cells or in histological presentation of the CNS infiltrates of rats immunized with the two adjuvants. However, there are more granulocytes, NK and NKT cells, and less CD4(+) T cells in the spinal cord infiltrates of SCH + CI-immunized animals. Nitric oxide (NO)-generating enzyme inducible NO synthase have higher expression in spinal cord of SCH + CI-immunized rats, and this corresponds to more intensive nitrotyrosine formation in the CNS tissue of these rats. Abundant infiltration of granulocytes and NK cells into the CNS and excessive generation of peroxynitrite within the CNS of SCH + CI-immunized rats might account for the severe neurological deficits induced by immunization with CI. These factors should be closely examined in the fulminant forms of multiple sclerosis and acute disseminated encephalomyelitis, as they could represent a promising targets for therapy.Serbian Ministry of Science [173035, 175038

It is still not for the old iron: adjuvant effects of carbonyl iron in experimental autoimmune encephalomyelitis induction

Experimental autoimmune encephalomyelitis (EAE) is a model of multiple sclerosis. Dark Agouti rats immunized with spinal cord homogenate (SCH) and carbonyl iron (CI), as an adjuvant, develop severe hyperacute form of EAE. They succumb to EAE earlier and have higher clinical scores and lethality rate in comparison to counterparts immunized with SCH + complete Freund's adjuvant. There is no difference in the number of cells or in histological presentation of the CNS infiltrates of rats immunized with the two adjuvants. However, there are more granulocytes, NK and NKT cells, and less CD4(+) T cells in the spinal cord infiltrates of SCH + CI-immunized animals. Nitric oxide (NO)-generating enzyme inducible NO synthase have higher expression in spinal cord of SCH + CI-immunized rats, and this corresponds to more intensive nitrotyrosine formation in the CNS tissue of these rats. Abundant infiltration of granulocytes and NK cells into the CNS and excessive generation of peroxynitrite within the CNS of SCH + CI-immunized rats might account for the severe neurological deficits induced by immunization with CI. These factors should be closely examined in the fulminant forms of multiple sclerosis and acute disseminated encephalomyelitis, as they could represent a promising targets for therapy.Serbian Ministry of Science [173035, 175038