可诱导共刺激分子基因表达抑制降低大鼠T淋巴细胞活化增殖能力的研究

目的 RNAi抑制大鼠T淋巴细胞可诱导共刺激分子(ICOS)基因表达,观察其对T淋巴细胞功能的影响.方法 选择4个干扰位点,插入pSilencer 4.1-CMV neo载体,转染大鼠淋巴细胞.RT-PCR检测ICOS基因的抑制程度,流式检测ICOS表达变化.MLR检测细胞的增殖能力,ELISA法检测细胞因子IFN-γ和IL-4分泌水平.结果 转染后,T淋巴细胞ICOS的mRNA和ICOS分子表达水平下降(P＜0.05),淋巴细胞增殖能力明显降低,同时IFN-γ和IL-4的水平也减低.结论 大鼠淋巴细胞ICOS基因的表达抑制,降低了T淋巴细胞活化增殖能力.

Study of reduction rat T-lymphocytes activation and proliferation by inhibition inducible co-stimulator gene expression

OBJECTIVE: To observe the change of activation and proliferation ability of rat T-lymphocytes after suppress ICOS gene expression by RNA interference. METHODS: Four interference sites targeting at rat ICOS gene were designed and four pairs of oligonucleotide fragments were cloned into the pSilencer 4.1-CMV neo plasmid vectors then transfected into rat lymphocytes with cationic liposome. The expression of mRNA and protein of ICOS was detected by RT-PCR and flow cytometry. The alteration of lymphocyte proliferation ability was evaluated by mix lymphocyte reaction, and the secretion levels of IFN-gamma and IL-4 were measured by ELISA procedure. RESULTS: After transfection, the expression of mRNA and protein of ICOS in test groups were lower than that in control groups (P < 0.05). The ability of T-lymphocytes in proliferation was poor and the levels of IFN-gamma and IL-4 were reduced with ICOS gene shut down. CONCLUSIONS: RNA interference plasmid vector can suppress ICOS expression in rat T-lymphocytes significantly, and it may be useful for further study on transplantation immunity tolerance.