HCMV基因在人和小鼠胚胎成纤维细胞中表达的差异性研究

人巨细胞病毒（human cytomegalovirus, HCMV）种属特异性机制尚不清楚。研究通过检测HCMVADl69体外感染人胚胎成纤维细胞（Human embryo fibroblast, HEF）和小鼠胚胎成纤维细胞（mouse embryo fibroblast, MEF）后病毒基因的表达情况，探讨HCMV种属特异性的可能分子机制。首先用HCMV AD169（MOI=5）分别感染HEF和MEF，相差显微镜逐日观察细胞的形态学变化；RT—PCR检测HCMV即刻早期（IE1、IE2）、早期（uL84）和晚期基因（UL83）的表达情况；Western—blot和免疫荧光检测病毒基因编码蛋白表达的情况。形态学观察发现HEF感染HCMV后逐渐变大变圆并相互融合，第4天可见典型的HCMV特征性病变效应，而MEF则未出现上述的变化；RT-PCR和Western—blot表明HEF组表达即刻早期基因IE1和IE2、早期基因uL84和晚期基因UL83 mRNA以及各基因所编码的蛋白，且相对表达量显著高于模拟感染组（P〈0．01）；而MEF组仅IEl和IE2mRNA和蛋白相对表达量显著低于HEF组（P〈0．05），而高于模拟感染组（P〈0．01）。免疫荧光检测发现HEF感染72h表达IE和UL83蛋白，而MEF则无明显表达。以上结果表明，HC—MV不能在MEF中复制并产生完整子代病毒颗粒，且病毒基因表达阻止在IE2基因表达之后和UL84基因表达之前，其种属特异性可能与即刻早期蛋白低水平的表达量有关。

HCMV Genes Expression Differences in Mouse Embryo FibroblastsHuman &

So far the specificity mechanism of species and genus of human cytomegalovirus （HCMV） remains unclear. In this study, the possible molecular mechanism of the specificity mechanism of species and genus of HCMV was investigated through detecting the viral genes expression after HCMV AD 169 infection in vitro on human embryo fibroblast （HEF） and mouse embryo fibroblast （MEF）. HCMV AD 169 was first respectively infected to HEF and MEF and the morphological changes were observed day by day with phase contrast microscope; Expressions of HCMV immediate-early gene （IE1, IE2）, early gene （UL84） and late gene （UL83） were detected by RT-PCR; The viral genes encoded proteins was detected by Western blot and immunofluorescence. Morphological observations found that HEF gradually became big and round and fused each other after HCMV infection, and at day 4 typical specific pathology effect of HCMV was visible ; while the above mentioned changes did not appear in MEF. RT-PCR and western blot results showed that HEF group could express IE1 and IE2, UL84, and UL83 mRNA and various genes encoded pro- teins, moreover, the relative expression amount was significantly higher than the analog infection groups （P 〈 0.01 ） ; while in MEF group only IE1 and IE2 mRNA and protein relative expression amount remarkably lower than HEF group （ P 〈 0.01 ）, but higher than analog infection group （P 〈 0.01 ）. Immunofluorescence detection found that 72 h after HEF infection could express IE and UL83 protein, while MEF had no significant expression. The results above indicated that HCMV could not duplicate in MEF and produce complete filial virus particle, moreover, the viral gene expression was obstructed behind the IE2 gene expression and before the UL84 gene expression, indicated that the specificity of species and genus may relate to the low level expression amount of immediate-early protein.