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

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

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pSilencer4．1-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IL4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IL4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。

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-γ 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-γ 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.