人巨细胞病毒感染对人胚肺成纤维细胞Cdtl与Geminin因子表达的影响

Influence of human cytomegalovirus infection on expressions of cdtl and geminin genes in human embryo lung fibroblasts

目的研究人巨细胞病毒（human cytomegalovirus，HCMV）感染对人胚肺成纤维（human embryonic lung fibroblasts，HEL）细胞Cdtl与Geminin因子表达的影响。方法用血清饥饿法同步化HEL细胞于G0／G1期，流式细胞术测细胞周期；用HCMV感染同步化的HEL细胞作为感染组，同时设模拟感染组为对照组。于感染后12、24、48、72和96h收获细胞，提取总的RNA，采用半定量逆转录-聚合酶链反应检测Cdtl-mRNA和Geminin-mRNA的表达水平。结果（1）血清饥饿后的HEL细胞82．8％处于G0／G1期。（2）12、24、48和72h两组HEL细胞CdtlmRNA表达水平比较均有统计学意义（f值分别为32．231，6．349，-5．846，-1．573；P均为0．000），12和24h时感染组较模拟感染组明显降低，48和72h时感染组较模拟感染组明显升高。（3）12、24、48和72h两组HEL细胞GemininmRNA表达水平比较均有统计学意义（t值分别为-8．891，-6．483，13．010，10．572；P均为0．000），12和24h时感染组较模拟感染组明显增加，48和72h时模拟感染组较感染组明显升高。结论（1）血清饥饿法将绝大部分细胞同步于G0／G1期，血清饥饿法同步化HEL细胞效果较佳。（2）HCMV能诱导Cdtl及Geminin因子表达异常，使Geminin／Cdtl平衡失调，这可能是HCMV感染导致细胞周期阻滞于G1期的机制。

Objective To determine the expressions of Cdtl and Geminin genes in human embryonic lung fibroblasts （HEL） and the influence of human cytomegalovirus （HCMV） infection. Methods The HEL cells were synchronized in G0/G1 by serum starvation. Cell cycle of HEL cells was detected by flow cytometry. Synchronized HEL cells were infected with HCMV, at the same time the mock-infected group was set as control. Infected HEL cells were harvested at 12 h, 24 h, 48 h, 72 h and 96 h post infection. The expressions of Cdtl and Geminin genes were detected with semi- quantitative RT-PCR. Results 82. 80% of the HEL cells were in G0/Ol phase after serum starvation. Analysis showed that the expressions of Cdtl gene in HCMV-infected group were statistically lower （all P = 0. 000） than those in the mock-infected group at 12 h and 24 h post infection, but higher than the latter group at 48 h and 72 h post infection. The expressions of Geminin mRNA in HCMV-infected group were statistically higher than those of the mock-infected group at 12 h and 24 h post infection, but lower than tha latter group at 48 h and 72 h post infection （all P= 0. 000）. Conclusions Serum starvation is a better method to synchronize the HEL cell. HCMV can induce abnormal expressions of Cdtl and Geminin genes and Geminin/Cdtl imbalance, which may play a role in HCMV blocked cell cycle in G1 phase.