B和C基因型乙型肝炎病毒对Huh7细胞全基因组表达模式的影响

Influence of HBV genotype B and C on the global gene expression pattern of Huh7 cells

目的探讨B和C基因型乙型肝炎病毒对Huh7细胞基因组表达模式的影响。方法重组B或C基因型乙型肝炎病毒(pHY106-BHBV和pHY106-CHBV)转染Huh7细胞,以pHY106空载体为对照,48h后进行芯片杂交,筛选出表达差异值Gy5/Cy3>2或<0.5的基因。pHY106-BHBV、pHY106-CHBV分别转染Huh7细胞48h,以pHY106空载体为对照,应用real-timePCR对基因芯片差异表达的部分基因进行验证。结果 pHY106-BHBV转染Huh7细胞48h后,从4097个基因中筛选出差异表达基因共60个;pHY106-CHBV转染Huh7细胞48h后,筛选出差异表达基因共120个;real-timePCR证实pHY106-BHBV转染Huh7细胞48h后,IFNGR2、GPR125、DDEF2和PI3KmRNA表达水平均出现不同程度下调,pHY106-CHBV转染Huh7细胞48h后,EEF2和INF592表达下调,与基因芯片结果一致。结论 B和C基因型乙型肝炎病毒对Huh7细胞表达谱改变的影响明显不同,表明其对机体的影响可能存在不同的方式。

Objective To investigate on the changes of global gene expression pattern in Huh7 cells when transfected with HBV genotype B or C. Methods Huh7 cells were transfected with recombinant HBV genotype B or C, namely pHY106-BHBV and pHY106-CHBV, respectively, with pHY106 as the control. After being transfected for 48 h, mRNA of Huh7 cells were extracted and reverse transcripted to cDNA, then hybrided with chip. The fluorescence signal intensity of Cy3 and Cy5 were analyzed with the GenePix Pro 3.0 software, the ratio of CyS/Cy3 above 2 or below 0.5 was regarded as significant. The partial results of microarray were verified by real-time PCR. Results After pHY106-BHBV being transfected into Huh7 cells for 48 h, 60 differentially expressed genes were screened out among 4097 genes arid 120 differentially expressed genes were screened out after pHY106-CHBV being transfected into Huh7 cells. IFNGR2, GPR125, DDEF2 and PI3K which were down-regulatory genes after pHY106-BHBV transfection were verified by real-time PCR, and the relative expression ratio were consistent with the results of microarray. EEF2 and INF592 which were down-regulatory genes after pHY106-CHBV transfection were verified by real-time PCR, the relative expression ratio were also consistent with the results of microarray. Conclusions HBV genotype B and C could change the expression pattern of Huh7 cells, with significant difference, and only DDEF2 was found down-regulated, which demonstrated that HBV genotype B and C may have different influence on host.