巴马小型猪内源性反转录病毒的整合对HEK293细胞周期、细胞凋亡和相关基因表达的影响

检测广西巴马小型猪内源性反转录病毒(PERV)整合到HEK293细胞基因组(HEK293-PERV-BM)后,宿主细胞的细胞周期、细胞凋亡及cyclin D1、CDK4、c-Myc、p53、p16和k-Ras等相关基因表达的改变,推测相关作用机制。检测发现P10代巴马小型猪内源性反转录病毒感染HEK293细胞模型(HEK293-PERV-BM)的细胞周期主要被阻滞在S期和G2/M期,细胞凋亡受到抑制,P25代细胞周期主要被阻滞在S期,细胞凋亡明显受到诱导。通过实时荧光定量PCR和Western blot检测发现细胞周期相关基因的表达与细胞感染模型传代次数(P1~P35)有密切关系,其中cyclin D1、c-Myc和k-Ras基因表现为先降低后升高,而CDK4和p16基因则恰好相反,同时,p53基因表达的改变不明显,据此可以推测P10代细胞周期的改变可能由Rb调控通路诱导发生,P25代细胞周期的改变可能由Rb调控通路与p53调控通路协同诱导发生。上述结果提示PERV的感染可能存在潜在的危险。

Effects of the integraton of PERV from inbreeded Bama miniature pigs on cell cycle and apoptosis as well as the expresson of some related genes in HEK293 cells

The cell cycle and apoptosis as well as the expression of some related genes were altered after the integration of PERV-BM,and differed with different passages of HEK293-PERV-BM cells(P1-P35).In HEK293-PERV-BM P10,the cell cycle was arrested in S phase and G2/M phase as well as the cell apoptosis was inhibited.In HEK293-PERV-BM P25,the cell cycle was arrested in S phase and the apoptosis was induced.To find the mechanisms,qPCR was applied to detect the mRNA expression of cyclin D1,CDK4,c-Myc,p53,p16 and k-Ras genes in different passages of HEK293-PERV-BM.Meanwhile,the protein expressions of cyclin D1,CDK4,c-Myc and other genes was detected by Western blot.The expression level of cyclin D1,c-Myc and k-Ras genes exhibited decrease and then increase,while the expression of CDK4 and p16 genes did just the opposite,and p53 did not significantly change.The expressions of the related genes suggested that the cell cycle of lower passages might be regulated by Rb passway,while the cell cycle of higher passages might be regulated by Rb passway and p53 passway.These results provide the clues of potential infection and effects of PERV-BM.