反义肽核酸阻断CXCR3表达延长大鼠肝脏移植物存活期

Prolongation of liver allograft survival with antisense peptide nucleic acid by means of blockade of CXCR3 expression in rats

目的研究反义肽核酸(antisense peptide nucleic acid,asPNA)阻断CXC趋化因子受体3(CXCR3)表达延长大鼠肝脏移植物存活期的作用和机制,探讨asPNA技术治疗肝移植急性排斥反应(acute rejection,AR)的潜在临床价值。方法采用改良"二袖套"法建立大鼠原位肝移植模型。分为4组:肝移植急性排斥组,肝移植免疫抑制剂组,asPNA组,asPNA对照组。RT-PCR法检测T细胞内CXCR3-mRNA表达水平,Western-blotting免疫印迹法检测CXCR3蛋白表达水平,流式细胞仪测定外周血CD3+T细胞表面CXCR3表达水平。AR诊断参照Banff标准。结果asPNA在体外能有效抑制T细胞内CXCR3-mRNA和CXCR3蛋白表达水平;显著延长大鼠肝脏移植物存活期;asPNA组大鼠肝移植术后第7天外周血和肝脏细胞内CXCR3的表达较AR组及asPNA对照组明显减少(P<0.05),与FK506组呈现出类似的趋势。结论asPNA能有效抑制T细胞内CXCR3基因和蛋白表达水平,降低大鼠AR发生率并延长肝脏移植物存活期,具有潜在临床应用价值。

Objective To explore the effect and mechanisms of the blockade of CXC pathway with antisense peptide nucleic acid （asPNA） on prolongation of liver allograft survival in rats. Methods The models of orthotopic liver transplantation （OLT） were established with two-cuff technique. Rats were devided into 4 groups： group of OLT with AR, group of OLT treated with FK506, group treated with asPNA and group treated with control asPNA. The post-OLT survival time was recorded and pathological changes of grafts were evaluated by Banff＇s standard. The changes of expression of CXCR3 were measured 1 day prior to and 1, 3, 5, 7 days after OLT respectively. Results asPNA could availably restrain the mRNA expression of CXCR3 in T cells, and could obviously debase the expression level of CXCR3 protein in T cells in vitro. The survival time of recipient rats in asPNA group was significantly longer than that in AR group. The expression of CXCR3-mRNA of liver grafts in asPNA group was reduced significantly than that of AR group and asPNA- control group in 7 days after OLT. Conclusion asPNA can availably restrain the gene and protein expression of CXCR3 in T cell, which reduces the incidence of AR and orolongs the graft survival time.