【摘要】 观察锌指蛋白265(ZNF265)基因对睾丸Sertoli细胞溶脲脲原体(Ureaplasma urealyticum,UU)感染后IL-1α、IL-6及TGF-β1表达的影响,并探讨其可能的作用机制。构建沉默大鼠ZNF265基因表达的慢病毒载体,体外转染Sertoli细胞,对转染阳性的细胞进行UU感染,在感染后的第1、3、5天分别检测对照组和实验组中IL-1α、IL-6及TGF-β1表达。结果显示,成功构建干扰Sertoli细胞中ZNF265表达的慢病毒载体,在Sertoli细胞中转染效率高达62.8%,在mRNA水平和蛋白质水平干扰ZNF265表达的效率分别为65.73%和94%;干扰Sertoli细胞中ZNF265表达后,在UU感染的第1天和第3天,IL-1α、IL-6及TGF-β1表达升高;在UU感染第5天,ZNF265表达较对照组升高,TGF-β1表达升高,但IL-1α,IL-6表达下降。以上结果说明,构建的慢病毒载体成功地干扰ZNF265基因在大鼠Sertoli细胞中的表达,干扰其表达后,IL-1α和IL-6表达增加;ZNF265作为调控因子,可能通过作用于TGF-β1,在抑制炎症发展、维持睾丸局部免疫微环境的平衡中发挥重要作用。更多还原

【Abstract】 To investigate the influence of ZNF265 on expressions of IL-1α,IL-6 and TGF-β 1 during UU infection of rat Sertoli cells and its mechanism,we first applied lentiviral-mediated RNA interference(RNAi) to inhibit the expression of ZNF265 in Sertoli cells,which were then infected by UU and investigated for expression of IL-1α,IL-6,TGF-β1 at mRNA level and protein level on day 1,3 and 5 post-infection.The infection rate of lentivirus in Sertoli cells was 62.8%,and the interference rate of ZNF265 expression at mRNA level and protein level was 65.73 and 94%,respectively.On day 1 and 3 post-infection,mRNA expression of IL-1α,IL-6,TGF-β1 was higher in the RNAi treated group than the untreated group.On day 5,expression of ZNF265 was higher than that of the control group,and so did TGF-β1,however,the levels of both IL-1α and IL-6 were deceased.These findings indicate that the lentiviral ZNF265 vector we constructed successfully inhibited expression of ZNF265 gene in Sertoli cells resulting in increased expression of IL-1α and IL-6.Therefore,ZNF265 may exerts its regulating role in inhibiting inflammatory and maintaining balance of immune microenvironment in the testis by upregulating expression of ZNF265.As ZNF265 at the fifth day going higher in shRNA-ZNF265 UU infection group than shRNA-NC UU infection group,which is in accordance to TGF-β1,the protein expression of IL-1α,IL-6 was down-regulated,and lower than that without RNAi.In conclusion,we successfully constructed lentiviral vector of RNAi to silence expression of ZNF265,and ZNF265 may combine with TGF-β1 to play a negative regulatory role by down-regulating IL-1α,IL-6 in Sertoli cells during UU infection and control inflammation of Sertoli cell.更多还原