瘦素对大鼠肝纤维化星状细胞的作用及相关ERK信号转导机制  

Effect of leptin on hepatic stellate cells in liver fibrosis rats and its relation to ERK signal transduction pathway

作　　者：周光耀[1] 金玲湘[1] 林巍[1] 潘陈为[1] 诸葛璐[1] 方佩佩[1] 

机构地区：[1]温州医科大学附属第二医院感染内科,325027

出　　处：《浙江医学》2014年第8期659-662,共4页Zhejiang Medical Journal

基　　金：温州市科技局基金项目(Y20090296)

摘　　要：目的探讨瘦素(leptin)对大鼠肝纤维化星状细胞(HSC)的作用及相关信号转导机制。方法采用改良原位灌注、Optiprep密度梯度离心法分离纯化大鼠HSC。通过台盼蓝拒染试验评估细胞存活率,α-平滑肌肌动蛋白(α-SMA)免疫组化鉴定,光镜观察形态学变化。将40只SD大鼠分为4个处理组:对照组、血管紧张素Ⅱ(AngⅡ)组(加入10^(-7)mol/L AngⅡ)、Leptin组(加入100ng/ml Leptin)、Leptin+AngⅡ组(加入100ng/ml Leptin+10^(-7)mol/L AngⅡ)。分别采用~3H-TdR和~3H-Pro掺入法进行HSC增殖和胶原合成的测定。Western blot检测各处理组p-ERK1/ERK1、p-ERK2/ERK2、AngⅡ蛋白表达情况。结果大鼠HSC存活率>90%,传代1次后HSC纯度>95%。与对照组相比,AngⅡ组、Leptin组、Leptin+AngⅡ组的HSC增殖和胶原合成均明显增加(均P<0.05);与AngⅡ组、Leptin组相比,Leptin+AngⅡ组HSC增殖和胶原合成明显增加(均P<0 05)。Western blot显示,AngⅡ组、Leptin组、Leptin+AngⅡ组的p-ERK1/ERK1、p-ERK2/ERK2、AngⅡ蛋白水平均较对照组明显升高(均P<0.05);与AngⅡ组、Leptin组相比,Leptin+AngⅡ组p-ERK1/ERK1、p-ERK2/ERK2、AngⅡ蛋白水平明显升高(均P<0 05。结论瘦素可通过上调AngⅡ水平,激活ERK信号转导通路。刺激HSC增殖和胶原合成,导致肝纤维化。Objective To investigate the effect of leptin on hepatic stel ate cells（HSC） in liver fibrosis rats and its relation to ERK signal transduction pathway. Methods The purified HSCs were obtained by the modified in situ perfusion and Optiprep density gradient centrifugation. The survival rate of HSCs was evaluated by trypan blue exclusion test, the identification of HSCs was tested by α- SMA immunocytochemical staining, and the morphological changes were observed by light microscope. The cultured HSCs were divided into four groups： the control group, the angiotensin II group （Ang II, 10-7mol/L）, the Leptin group （Leptin, 100ng/ml）, and the Leptin＋Ang II group [Leptin （100ng/ml）＋AngⅡ（10-7mol/L）]. The HSCs proliferation and col agen syn-thesis were measured by 3H- TdR and 3H- Pro incorporation methods. The protein levels of p- ERK1/ERK1, p- ERK2/ERK2 and Ang II were detected by Western blot. Results The survival rate of rat HSCs was＆gt;90%and the purity of HSCs was＆gt;95%after passage 1. The findings of HSCs proliferation and col agen synthesis showed that compared with control group, the proliferation and col agen synthesis of HSCs were significantly increased in the Ang II, Leptin and Leptin＋Ang II groups （P＆lt;0.05）;compared with the Ang II, Leptin groups, the HSCs proliferation and col agen synthesis were significantly increased in leptin＋Ang II group （P＆lt;0.05）. Western blot analysis showed that compared with the control group, the protein levels of p- ERK1/ERK1, p- ERK2/ERK2 and Ang II were significantly increased in Ang II, Leptin and Leptin＋Ang II groups（P＆lt;0.05）;compared with Ang II, Leptin groups, the protein levels of p- ERK1/ERK1, p- ERK2/ERK2 and Ang II were significantly increased in Leptin＋Ang II group （P＆lt;0.05）. Conclusion Leptin may up- regulate the Ang II levels, activate the ERK signal transduction pathway, stimulate the HSCs prolifera-tion and col agen synthesis, and then result in the liver fibrosis.

关 键 词：瘦素 肝纤维化 信号转导机制 

分 类 号：R575.2[医药卫生—消化系统]