摘要
Background Experimental autoimmune myocarditis （EAM） in rats is a T-cell-mediated disorder. The initiation and maintenance of autoimmune responses in EAM depend on the maturation state of dendritic cells. IL-10 is a pleiotrophic immunomodulatory cytokine that functions at different levels of the immune response, so it has emerged as a promising therapeutic factor for the treatment of autoimmune/inflammatory diseases. This study was designed to test the hypothesis that IL-10 gene modified bone marrow-derived immature dendritic cells （iDCs） ameliorate EAM and to explore the underlying mechanisms. Methods EAM was induced using the methods of cardiac myosin immunization on day 0 and day 7. Immature and mature bone marrow-derived dendritic cells （BMDCs） were generated without or with the stimulation by lipopolysaccharide （LPS） and the phenotype was analyzed by flow cytometry. Some of the iDCs were transfected by pcDNA3-IL-10 plasmid. 2× 10^6/per rat mature DC （mDC）, immature DC （iDC）, pcDNA3 transfected iDC, pcDNA3-IL-10 transfected iDC or phosphate buffered saline （PBS） were injected intravenously for treatment 5 days after the first immunization. On day 21, HE staining was performed to detect the myocardial inflammation and T lymphocyte proliferation assay was used to determine the effects of IL-10 gene transfected iDC on autoreactive T cell proliferation. Expression of IκB, the inhibitor of NF-κB pathway, was determined by Western blot. Results BMDCs generated in a medium supplemented with granulocyte-macrophage-colony-stimulating factor （GM-CSF） were relatively immature, as determined by flow cytometry. However, stimulation with LPS induced these cells to become mature （m）DCs with higher levels of surface major histocompatibility complex （MHC）-Ⅱ and costimulatory molecules. Intravenous administration of iDCs, especially pcDNA3-IL-10 transfected iDC, ameliorated the histopathological severity of the myosin induced-EAM, and the effect was lost after the DCs underwent maturation induced by in vitro exposure to LPS. IL-10 gene modified iDC inhibited the antigen specific T cell responses towards cardiac myosin. IκB protein was up-regulated significantly in the IL-10 gene modified iDC group. Conclusions IL-10 gene modified iDC induced antigen-specific tolerance in EAM. The underlying mechanisms may be related to costimulatory molecules down-regulation and NF-κB pathway inhibition.
Background Experimental autoimmune myocarditis （EAM） in rats is a T-cell-mediated disorder. The initiation and maintenance of autoimmune responses in EAM depend on the maturation state of dendritic cells. IL-10 is a pleiotrophic immunomodulatory cytokine that functions at different levels of the immune response, so it has emerged as a promising therapeutic factor for the treatment of autoimmune/inflammatory diseases. This study was designed to test the hypothesis that IL-10 gene modified bone marrow-derived immature dendritic cells （iDCs） ameliorate EAM and to explore the underlying mechanisms. Methods EAM was induced using the methods of cardiac myosin immunization on day 0 and day 7. Immature and mature bone marrow-derived dendritic cells （BMDCs） were generated without or with the stimulation by lipopolysaccharide （LPS） and the phenotype was analyzed by flow cytometry. Some of the iDCs were transfected by pcDNA3-IL-10 plasmid. 2× 10^6/per rat mature DC （mDC）, immature DC （iDC）, pcDNA3 transfected iDC, pcDNA3-IL-10 transfected iDC or phosphate buffered saline （PBS） were injected intravenously for treatment 5 days after the first immunization. On day 21, HE staining was performed to detect the myocardial inflammation and T lymphocyte proliferation assay was used to determine the effects of IL-10 gene transfected iDC on autoreactive T cell proliferation. Expression of IκB, the inhibitor of NF-κB pathway, was determined by Western blot. Results BMDCs generated in a medium supplemented with granulocyte-macrophage-colony-stimulating factor （GM-CSF） were relatively immature, as determined by flow cytometry. However, stimulation with LPS induced these cells to become mature （m）DCs with higher levels of surface major histocompatibility complex （MHC）-Ⅱ and costimulatory molecules. Intravenous administration of iDCs, especially pcDNA3-IL-10 transfected iDC, ameliorated the histopathological severity of the myosin induced-EAM, and the effect was lost after the DCs underwent maturation induced by in vitro exposure to LPS. IL-10 gene modified iDC inhibited the antigen specific T cell responses towards cardiac myosin. IκB protein was up-regulated significantly in the IL-10 gene modified iDC group. Conclusions IL-10 gene modified iDC induced antigen-specific tolerance in EAM. The underlying mechanisms may be related to costimulatory molecules down-regulation and NF-κB pathway inhibition.
基金
This study was supported by Harbin Medical University Postgraduate Creation Foundation (2005).
