摘要
目的探讨野生型第10号染色体缺失的磷酸酶张力蛋白同源物基因(PTEN)过表达对体外活化肝星状细胞(HSC)内钙离子浓度的影响。方法体外培养活化大鼠肝星状细胞系(HSC-T6),利用腺病毒载体,将野生型PTEN基因转染活化HSC;Western blot及实时荧光定量聚合酶链反应(q RT-PCR)检测HSC的PTEN蛋白及m RNA表达;采用钙离子荧光探针Rhod-2/AM,于激光扫描共聚焦显微镜下检测HSC内钙离子浓度变化。实验分组:(1)Control组:腺病毒转染时以DMEM代替病毒液;(2)Ad-GFP组:转染表达绿色荧光蛋白(GFP)的对照空病毒Ad-GFP;(3)Ad-PTEN组:转染携带野生型PTEN基因并表达GFP的重组腺病毒Ad-PTEN。结果野生型PTEN在活化大鼠HSC大量表达,3组HSC内钙离子浓度比较,差异有统计学意义(P<0.05),Ad-PTEN组HSC内钙离子浓度(251.60±90.88)低于Control组(1 953.95±132.99)及Ad-GFP组(1 937.57±115.17),而Control组与Ad-GFP组之间HSC内钙离子浓度比较,差异无统计学意义(P>0.05)。结论野生型PTEN过表达可降低体外活化大鼠HSC内钙离子浓度。
Objective To investigate the effect of over-expression of wild-type phosphatase and tensinhomology deleted on chromosome ten(PTEN)on the intracellular calcium ion concentration in activated hepaticstellate cells(HSCs)in vitro.Methods Using adenoviral vector,wild-type PTEN gene was transduced intoactivated rat HSC(HSC-T6)in vitro,and the expressions of PTEN mRNA and protein were detected by qRT-PCRand Western blot.Then with the help of laser scanning confocal microscope(LSCM),the changes of intracellularcalcium ion concentration in HSCs were detected using calcium ion fluorescence probe Rhod-2/AM.The cells weregrouped as follows:control group,in which the viral medium was replaced by DMEM when HSCs were transfectedwith adenovirus;Ad-GFP group,in which HSCs were transfected with adenovirus expressing green fluorescentprotein(GFP)alone;and Ad-PTEN group,in which HSCs were transfected with adenovirus harboring both wildtypePTEN and GFP genes.Results The wild-type PTEN was successfully transduced into activated HSCs byadenoviral vector.The intracellular calcium ion concentration of the HSCs in the Ad-PTEN group significantlydecreased compared with the control group and the Ad-GFP group(P<0.05).However,no significant difference was observed in the intracellular calcium ion concentration of HSCs between the control group and the Ad-GFP group(P>0.05).Conclusions The over-expression of wild-type PTEN can significantly reduce the intracellular calciumion concentration in activated hepatic stellate cells in vitro.
作者
郝礼森
宋小杰
王玉兰
刘玉龄
宋洁
张明婷
靳丽敏
张朋垒
Li-sen Hao;Xiao-jie Song;Yu-lan Wang;Yu-ling Liu;Jie Song;Ming-ting Zhang;Li-min Jin;Peng-lei Zhang(Department of Gastroenterology, the Affiliated Hospital of North China University of Science and Technology, Tangshan, Hebei 063000, China)
出处
《中国现代医学杂志》
CAS
北大核心
2017年第28期12-15,共4页
China Journal of Modern Medicine
基金
河北省自然科学基金(No:H2013209327)
中国肝炎防治基金会天晴肝病研究基金资助项目(No:CFHPC20132078)
