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利用我们建立的表达人膜型LIGHT分子的基因转染细胞(L929/LIGHT)探讨LIGHT/HVEM信号体外对Mo-DC诱导分化的影响,并进一步研究其对T细胞活化和抗凋亡的共刺激作用。从健康人外周血中分离的单核细胞经GM-CSF联合IL-4诱导形成Mo-DC,流式细胞术分析Mo-DC诱导过程中HVEM和LIGHT的表达;基因转染细胞L929/mock、L929/LIGHT、L929/CD40L或L929/LIGHT联合L929/CD40L,分别经丝裂霉素处理后,与GM-CSF联合IL-4诱导的Mo-DC共育,流式细胞术检测Mo-DC细胞表面成熟标志CD83和CD86的表达,利用FITC-Dextran分析Mo-DC对抗原的摄取能力;L929/LIGHT或L929/mock经丝裂霉素处理后,与抗人CD3单抗激发的T细胞共育,流式细胞术分析CD4+和CD8+T细胞表面活化标志CD25的表达,Annexin V和PI双标记分析T细胞的凋亡率。结果表明,高表达HVEM的单核细胞在诱导形成成熟Mo-D(iDC)的过程中下调了HVEM的表达,成熟Mo-DC(mDC)又上调表达HVEM,而LIGHT在Mo-DC分化过程中呈短暂的诱导性表达;基因转染细胞L929/LIGHT及其联合L929/CD40L能上调Mo-DC表面共刺激分子CD83和CD86的表达,并下调Mo-DC对FITC-Dextran的摄取能力;L929/LIGHT细胞能上调CD4+、CD8+T细胞CD25的表达,并增强T细胞抗凋亡能力。因此,基因转染细胞L929/LIGHT表面表达的人膜型LIGHT分子介导的LIGHT/HVEM共刺激信号对Mo-DC的诱导成熟和T细胞活化及抗凋亡能力具有促进作用。
To investigate the effect of LIGHT / HVEM signaling on the differentiation of Mo-DC cells induced by L929 / LIGHT, a genetically engineered LIGHT-expressing human lentivirus (L929 / LIGHT) was established and its co-stimulation of T cell activation and anti-apoptosis was further studied effect. Monocytes isolated from healthy human peripheral blood were induced to form Mo-DC by GM-CSF combined with IL-4. The expression of HVEM and LIGHT during the induction of Mo-DC was analyzed by flow cytometry. The expression of L929 / mock, L929 / LIGHT, L929 / CD40L or L929 / LIGHT combined with L929 / CD40L respectively. After treated with mitomycin, the cells were co-cultured with GM-CSF and IL-4 induced by Mo- Surface maturation markers CD83 and CD86 expression, the use of FITC-Dextran Mo-DC analysis of antigen uptake capacity; L929 / LIGHT or L929 / mock after mitomycin treatment, with anti-human CD3 monoclonal antibody-stimulated T cells The expression of CD25 on CD4 + and CD8 + T cells was analyzed by flow cytometry. The apoptosis rate of T cells was analyzed by Annexin V and PI double labeling. The results showed that HVEM-overexpressing monocytes downregulated the expression of HVEM during the induction of mature Mo-D (iDC), and up-regulated the expression of HVEM in mature Mo-DCs (mDC) L929 / LIGHT and L929 / CD40L could up-regulate the expression of costimulatory molecules CD83 and CD86 on Mo-DC surface and down-regulate the uptake of FITC-Dextran by Mo-DC. L929 / LIGHT cells can upregulate the expression of CD25 on CD4 + and CD8 + T cells and enhance the anti-apoptotic ability of T cells. Therefore, the LIGHT / HVEM costimulation signal mediated by LIGHT / HVEM, which is expressed on human L929 / LIGHT surface, can promote the induction of M-DC and the activation and anti-apoptotic ability of T cells.