缺氧诱导大鼠肝星状细胞激活及表达内脂素的实验研究  

Effect of Hypoxia on the Activation and Visfatin Expression in Rat Hepatic Stellate Cells

作　　者：易智慧[1] 门若庭[1] 袁聪[1,2] 朱永军[1] 文茂瑶 杨丽[1] 

机构地区：[1]四川大学华西医院消化内科,成都610041 [2]川北医学院消化内科,南充637000

出　　处：《四川大学学报（医学版）》2014年第4期563-566,共4页Journal of Sichuan University(Medical Sciences)

基　　金：四川省科技厅项目(No.2013FZ0085;No.2014SZ0002);成都市科技局项目(No.13PPYB994SF-014)资助

摘　　要：目的通过观察常氧及缺氧状态下大鼠肝星状细胞(HSCs)表达α-平滑肌肌动蛋白(α-SMA)、缺氧诱导因子-1α(HIF-1α)和内脂素水平的变化,探讨缺氧对HSCs激活及表达内脂素的影响。方法对原代大鼠HSCs进行缺氧(37℃、5%CO2+1%O2+94%N2)处理,检测不同缺氧时间(3、6、12及24h)和常氧条件(5%CO2+21%O2+74%N2)下HSCs中HIF-1α、α-SMA和内脂素的mRNA表达水平及蛋白表达水平。基因表达用RT-PCR检测,蛋白表达用蛋白质印迹法(Western blot)检测。结果缺氧3h即能诱导大鼠HSCs表达HIF-1αmRNA(P<0.05);HSCs缺氧6h后α-SMA mRNA及蛋白表达水平的表达上调(P<0.05);HSCs缺氧12h上调内脂素mRNA的表达(P<0.05),6h上调内脂素蛋白表达(P<0.05);缺氧诱导α-SMA和内脂素表达呈正相关(基因r=0.991,蛋白r=0.968,P<0.05)。结论缺氧可上调HSCs表达α-SMA和内脂素,且二者具有相关性,提示缺氧微环境可能通过内脂素介导HSCs的激活,促进肝纤维化的发生。Objective To investigate the effect of hypoxia on the visfatin and the expression of smooth muscle-actin（α-SMA）and hypoxia-inducible factor-1α（HIF-1α）in rat hepatic stellate cells（HSCs）.Methods Rat primary HSCs were isolated from SD rats by in situ perfusion of collagenase and pronase and single-step Nycodenz density gradient centrifugation,and then cultured and activated.Completely activated primary HSCs were exposed to hypoxic conditions（37℃,5% CO2,1% O2,94%N2）,or normoxic conditions（37℃,5% CO2,21% O2,74%N2）,for 3,6,12or 24 hrespectively.The expression ofα-SMA,the marker of HSC activation,and visfatin were assessed by Real time-PCR and Western blot.The Expression of HIF-1αwas detected by Real time-PCR.Results HIF-1αmRNA in rat HSCs was induced after exposed to hypoxia for 3h,and maintained elevated status up to24h.HSCs exposed to 1% O2 hypoxic conditions for 6h increasedα-SMA mRNA and protein expression.Visfatin mRNA expression was up-regulated after subjected to hypoxia for 12h,and protein level was elevated after 6h hypoxia.A positive linear correlation existed betweenα-SMA and visfatin expression in responsible to hypoxia〔r=0.991（genes）and r=0.968（proteins）,P＆lt;0.05〕.Conclusion Microcirculation impairment could significantly induceα-SMA and visfatin expression in rat HSCs,which might potentate the activation process of HSCs.

关 键 词：肝星状细胞 缺氧诱导因子-1Α 内脂素 Α-平滑肌肌动蛋白 

分 类 号：R575.5[医药卫生—消化系统]