重组FoxP3腺相关病毒转染小鼠CD4+CD25 -T细胞的实验研究

目的：研究FoxP3（Forkhead Box P3）基因在小鼠T细胞的表达情况，以及重组FoxP3腺相关病毒（adeno-associatedvirus，AAV）对小鼠CD4^＋CD25^-T细胞功能的影响。方法：采用实时定量PCR检测CD4^＋CD25^-T细胞和CD4^＋CD25^-T细胞FoxP3 mRNA表达情况，利用基因重组技术构建FoxP3腺相关病毒载体，体外转染CD4^＋CD25^-T细胞，通过细胞增殖试验观察转染CD4^＋CD25^-T细胞功能变化。结果：CD4^＋CD25^-T细胞较CD4^＋CD25^-T相比高水平表达FoxP3基因mRNA，重组FoxP3腺相关病毒转染后的CD4^＋CD25^-T细胞对CD3单抗的刺激呈低反应性，并且能抑制新鲜分离CD4^＋CD25^-T细胞的增殖。结论：FoxP3基因转染的CD4^＋CD25^-T细胞在体外具有抑制T细胞活化增殖的功能。

Experimental studies of mouse CD4+CD25 -T cells transduced with Forkhead Box P3 gene recombinant adeno-associated virus vector

Objective:To investigate the expression of Forkhead Box P3(FoxP3) on mouse T cells,and the effects of CD4~+CD25~-T cells transduced with recombinant adeno-associated virus(rAAV) vector carrying FoxP3 gene.Methods:FoxP3 mRNA levels in mouse T cells were analyzed by quantitative real-time PCR.The rAAV vector carrying mouse FoxP3 gene was constructed.CD4~+CD25~-T cells were infected with rAAV-FoxP3 and the functions were analyzed by lymphocyte proliferation assay.Results:CD4~+CD25~+T cells expressed more abundant FoxP3 mRNA than other T cell populations.The rAAV-FoxP3 was successfully constructed.After CD4~+CD25~-T cells were transduced by rAAV-FoxP3,these cells were hyporesponsive to stimulation via their T cell receptor and suppressed the proliferation of freshly isolated CD4~+CD25~-T cells in co-culture experiments.Conclusion:FoxP3-transduced CD4~+CD25~-T cells by rAAV possessed immunosuppression in vitro and will be possibly applied in immune disorders.