去甲肾上腺素对大鼠肝星状细胞作用的实验研究

目的 探讨交感神经系统在肝纤维化发生和发展中的作用. 方法采用免疫荧光和RT-PCR检测体外培养的肝星状细胞(HSC)中α1、β2-肾上腺素能受体的表达;用四甲基偶氮唑盐法检测不同浓度的去甲肾上腺素(NE)对HSC增殖活性的影响.同时用RT-PCR检测受NE作用后HSC的活化指标胶原蛋白-1、转化生长因子β(TGF β)及α-平滑肌动蛋白(α-SMA)的表达.用高效液相色谱-电化学法测定活化的HSC中交感神经递质NE的水平. 结果α1和β2-肾上腺素能受体表达于HSC的胞膜和胞质内;NE可呈剂量依赖性地促进HSC增殖,在浓度为100μmol/L时达到最大效应,F=140.464,P<0.05,差异有统计学意义.以NE 100 μmol/L作用细胞24h后,可显著促进反应HSC活化的指标上升,胶原蛋白-1表达为0.3022±0.0610,TGF β表达为2.2080±0.2151,α-SMA mRNA表达为0.5469±0.0108,与对照组胶原蛋白-1(0.1040±0.0556)、TGF β(1.1190±0.0070)、α-SMA mRNA表达(0.0759±0.0449)比较,t值分别为-4.160、-8.763和-17.651,P值均<0.05,差异均有统计学意义.HSC可以合成并释放NE,且受血小板衍生生长因子(10ng/ml)刺激后HSC中NE含量为(14.24±0.21)ng/ml,对照组为(11.34±0.15)ng/ml,两组比较,t=-32.907,P<0.05,差异有统计意义.结论 抑制交感神经系统使HSC活性降低对临床上治疗肝纤维化有一定的指导意义.

Effects of norepinephrine on the proliferation and activation of rat hepatic stellate cells

Objective To elucidate the relationship between rat hepatic stellate cells (HSC) and sympathetic neurotransmitter norepinephrine (NE) during liver fibrosis. Methods Using immunofluores-cence and RT-PCR, the expressions of α 1 and β 2-adrenoceptors in activated HSC were detected. Methyl thiazolyl tetrazolium (MTT) was adopted to investigate the effect of NE on the proliferation of HSC. Meanwhile, the expressions of collagen-1, transforming growth factor beta (TGF β) and smooth muscle α-actin (α-SMA) in NE-stimulated HSC were detected by RT-PCR. The contents of NE in HSC were determined by high performance liquid chromatography-electrochemical detector (HPLC-ECD). Results The α 1 and β 2-adrenoeeptors were expressed in HSC. NE markedly stimulated the proliferation of HSC in a concentration-dependent manner (F = 140.464, P < 0.05). NE induced the mRNA expressions of collagen-1, TGF β and α-SMA in HSC (t = -4.160; t = -8.763; t = -17.651, P < 0.05). HSC were synthesing and releasing NE,especially when stimulated with platelet-dedved growth factor (PDGF) (10 ng/ml) (t = -32.907, P < 0.05).Conclusion Our findings show that HSC are direct targets of NE and HSC are hepatic neuroglial ceils that produce and respond to sympathetic neurotransmitter norepinephrine, suggesting that interrupting sympa-thetic nervous system signaling may be useful in the treatment of liver fibrosis.