Faciogenital dysplasia 6基因对肝干细胞分化的调控作用

Regulatory effect of faciogenital dysplasia 6 gene on hepatic stem cell differentiation

目的探讨FGD6（faciogenital dysplasia）基因对肝干细胞分化调控的作用。方法选取FGD6基因干扰靶序列，使用AdEasy系统构建腺病毒载体，包装并扩增重组腺病毒载体psEs-FGD6-siRNA，感染HP14．5细胞。细胞免疫荧光检测FGD6蛋白在HP14．5细胞中的表达水平，实时荧光定量PCR检测FGD6、甲胎蛋白（AFP）及白蛋白（Alb）的mRNA水平，Westernblot检测FGD6、AFP及Alb的蛋白表达水平。每组细胞均设置pSES-Ad-RFP腺病毒空载体感染进行对照。所有数据用均数±标准差（x-±s）表示，采用单因素方差分析。结果FGD6蛋白主要表达于HP14.5细胞胞核中。成功构建腺病毒载体pSES-FGD6-siRNA。下调FGD6基因在HP14．5细胞中的表达，可降低AFP的mRNA及蛋白的表达，而升高Alb的mRNA及蛋白的表达（P〈0．01）。结论抑制FGD6基因在HP14．5细胞中的表达，可使HP14．5细胞向肝细胞方向分化，因此，FGD6基因对肝干细胞分化调控可能起着重要作用。

Objective To investigate the regulatory effect of faciogenital dysplasia 6 （FGD6） gene on hepatic stem cell differentiation. Methods FGD6 gene was selected for the co-intervention of target sequence, the AdEasy system was used for the construction of adenovirus vector and the packaging and multiplication of the recombinant adenovirus vector pSES-FGD6-siRNA, and the HP14.5 cells were infected. Immunofluorescence assay was used to measure the expression of FGD6 protein in HP14.5 cells, quantitative real-time PCR was used to measure the mRNA expression of FGD6, alpha-fetoprotein （AFP）, and albumin （Alb）, and Western blot was used to measure the protein expression of FGD6, AFP, and Alb. The empty pSES-Ad-RFP adenovirus vector was constructed as control in each group. All data were expressed as x-±s, and a one-way analysis of variance was performed. Results FGD6 protein was mainly expressed in tahe nucleus of HP14.5 cells. The pSES-FGD6-siRNA adenovirus vector was successfully constructed and it downregulated the expression of FGD6 gene and the mRNA and protein expression of AFP in HPl4.5 cells and upregulated the mRNA and protein expression ofAlb （P〈 0.01）. Conclusion The inhibition of the expression ofFGD6 gene in HP14.5 cells may differentiate HP14.5 cells into hepocytes. Therefore, FGD6 gene plays an important role in the differentiation regulation of hepatic stem cells.