过表达FGD6对肝干细胞分化的调控作用  

The Effect of Over-Expressing FGD6 on the Differentiation in Hepatic Stem Cells

作　　者：余师师 沙鸥 李宛玲 胡倩 何通川[2] 张秉强[1] YU Shishi;SHA Ou;LI Wanling;HU Qian;HE Tongchuan;ZHANG Bingqiang(Department of Gastroenterology,the First Affiliated Hospital of Chongqing Medical University,Chongqing 400016,China;Molecular Oncology Laboratory,University of Chicago Medical Center,Chicago 600637,USA)

机构地区：[1]重庆医科大学附属第一医院消化内科,重庆400016 [2]美国芝加哥大学医学中心分子肿瘤实验室,芝加哥600637

出　　处：《中国细胞生物学学报》2019年第10期1958-1966,共9页Chinese Journal of Cell Biology

基　　金：重庆市自然科学基金计划项目(批准号:CSTC,2011BB5123)资助的课题~~

摘　　要：该文主要探讨过表达FGD6(faciogenital dysplasia 6)基因对肝干细胞分化的调控作用及其可能的机制。将FGD6基因插入腺病毒载体使其过表达,并包装成腺病毒(Ad-FGD6),感染小鼠胚胎肝干细胞HP14.5,以空载体腺病毒(Ad-null组)和未感染腺病毒组(Blank control组)作为对照;半定量PCR和Western blot分别检测FGD6、肝干细胞标志物(AFP)、肝细胞标志物(ALB)、胆管上皮细胞标志物(CK19、SOX9)及Wnt经典通路中的标志物(β-catenin、wnt3a)的m RNA和蛋白表达水平;并通过糖原染色法(PAS染色)检测细胞内糖原合成的变化情况;CCK8检测各组细胞增殖情况。结果显示,感染Ad-FGD6腺病毒后,与Ad-null组及Blank control组相比,HP14.5细胞中的FGD6的m RNA和蛋白表达水平升高(P<0.05),肝干细胞标志物(AFP)及胆管上皮标志物(CK19、SOX9)的m RNA及蛋白的表达水平降低(P<0.05),而肝细胞标志物ALB及Wnt经典通路中的标志物(β-catenin、wnt3a)的m RNA和蛋白的表达水平均升高(P<0.05);PAS染色发现,Ad-FGD6组存在紫红色颗粒,呈阳性反应,这说明有糖原合成。CCK8检测发现,Ad-FGD6组能促进肝干细胞增殖(P<0.05)。因此,过表达HP14.5细胞的FGD6会使HP14.5细胞向肝细胞分化并促进细胞增殖,而这一过程可能通过Wnt经典信号通路实现。The aim of this study was to investigate the effect of over-expressing FGD6(faciogenital dysplasia 6)on the differentiation of hepatic stem cells and its possible mechanism.The FGD6 gene was inserted into an adenovirus vector to over-express and packaged into an adenovirus(Ad-FGD6)to infect hepatic stem cells HP14.5,where the empty vector adenovirus group(Ad-null group)and the uninfected adenovirus group(Blank control group)were used as comparisons.The mRNA and protein expression levels of FGD6,hepatic stem cell marker(AFP),hepatocyte markers(ALB),bile duct epithelial cell markers(CK19 and SOX9)and the canonical Wnt pathway markers(β-catenin and wnt3 a)were detected by semi-quantitative PCR and Western blot respectively.The change in intracellular glycogen synthesis was detected by glycogen staining(PAS staining),and the cell proliferation was detected by CCK8.The results showed that the mRNA and protein levels of FGD6 when HP14.5 cells were infected by the Ad-FGD6 adenovirus were increased compared with the Ad-null group and the Blank control group(P<0.05).The mRNA and protein levels of hepatic stem cell marker(AFP)and bile duct epithelial markers(CK19,SOX9)were decreased(P<0.05).But the mRNA and protein levels of hepatocyte marker(ALB)and the canonical Wnt pathway markers were elevated(P<0.05).There were the purplish-red granules in the Ad-FGD6 group that represented the positive reaction,which indicated that the HP14.5 had the function of glycogen synthesis.CCK8 detection found that Ad-FGD6 group can promote hepatic stem cell proliferation(P<0.05).Therefore,the up-regulation of FGD6 gene expression can facilitate HP14.5 cells to differentiate into hepatocytes and promote cell proliferation,which may be achieved through the canonical Wnt pathway.

关 键 词：FGD6基因 肝干细胞 分化调控 重组腺病毒 Wnt经典信号通路 

分 类 号：R329.2[医药卫生—人体解剖和组织胚胎学]