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目的观察肝纤维化病理过程中肝脏Smads锚着蛋白(SARA)的表达变化及其与肝纤维化的关系。方法大鼠每kg体重10μl二甲基亚硝胺腹腔注射,1次/d,每周连续3 d,共4周,复制大鼠肝纤维化模型。模型大鼠分别设首次造模后1、3 d、1,2、3、4周末,与造模停止后1、2、4周,共9个时间段为观察组。每组5～8只,另设正常大鼠10只为对照组。天狼猩红染色观察肝组织胶原沉积,盐酸水解法测定肝组织羟脯氨酸(HYP)含量,免疫组织化学染色观察肝组织SARA蛋白表达,Western blot法分析肝组织转化生长因子(TGF)β1、α-平滑肌肌动蛋白(α-SMA)和SARA蛋白的表达,并进行以上指标的相关性分析。结果随二甲基亚硝胺染毒持续,模型大鼠肝脏胶原增生(Hyp含量)与沉积增加,4周末时达到高峰,可见宽大纤维间隔与假小叶；而后随染毒停止,肝脏胶原沉积与纤维间隔有所减轻。模型组4、5、6、8周肝组织Hyp平均含量(μg/g)分别为193．0±39．2、188．5±39．9．174．4±21．2、163．6±31．5,对照组分别为125．6±19．5,t值在3．43～4．9,P值均＜0．01。免疫组织化学染色发现,SARA主要表达于正常与纤维化肝脏的肝窦周围间质细胞,随肝纤维化发展SARA阳性染色细胞数量减少,随染毒停止与肝纤维化恢复,SARA渐恢复至正常组水平。Western blot发现,随肝纤维化形成,模型大鼠肝组织TGFβ1、α-SMA蛋白表达逐渐增加,而SARA蛋白表达逐渐减少；肝纤维化恢复过程中,SARA渐恢复接近正常水平,TGFβ1与α-SMA蛋白表达有所下降。在肝纤维化形成与恢复过程中,SARA蛋白与Hyp含量、TGFβ1与α-SMA表达均呈明显负相关。结论SARA蛋白主要表达于肝脏间质细胞；随大鼠肝纤维化发展,SARA表达减少,SARA蛋白与肝纤维化形成呈负相关关系。
Objective To observe the expression of Smad-anchored protein (SARA) and its relationship with hepatic fibrosis in the pathogenesis of hepatic fibrosis. Methods Rats were injected intraperitoneally with 10μl dimethylnitrosamine per kg body weight once a day for 3 weeks for 4 weeks to establish a rat model of hepatic fibrosis. The rats in the model group were respectively set as 1, 3, and 1, 2, 3 and 4 weeks after the first modeling, and 1, 2, 4 and 4 weeks after the model was stopped. Each group of 5 to 8, another set of 10 normal rats as control group. Sirius red staining was used to observe the deposition of collagen in liver tissue, the content of hydroxyproline (HYP) in liver tissue was determined by hydrochloric acid hydrolysis, the expression of SARA protein in liver tissue was observed by immunohistochemical staining, the expression of transforming growth factor (TGF) α-smooth muscle actin (α-SMA) and SARA protein expression, and the correlation analysis of the above indicators. Results With the continuous exposure to dimethylnitrosamine, the hepatic collagen hyperplasia (Hyp content) and deposition in the model rats increased and peaked at the end of 4 weeks. The wide fiber intervals and the pseudolobules were observed. With the cessation of exposure, the hepatic collagen deposition and Fiber spacing has been reduced. The mean Hyp levels of liver tissue in the model group at 4, 5, 6 and 8 weeks were 193.0 ± 39.2, 188.5 ± 39.9, 174.4 ± 21.2 and 163.6 ± 31.5 in the control group, 125.6 ± 19.5 in the control group and 3.43 ~ 4.9 in the control group, all P <0.01. Immunohistochemical staining revealed that SARA was mainly expressed in the interstitial cells around hepatic sinusoids in normal and fibrotic liver. With the development of hepatic fibrosis, the number of SARA-positive cells decreased, and the SARA gradually recovered to normal with the resumption of liver fibrosis Group level. Western blot analysis showed that with the formation of hepatic fibrosis, the expression of TGFβ1 and α-SMA increased gradually and the expression of SARA gradually decreased in the liver of model rats. During the recovery of hepatic fibrosis, the recovery of SARA approached the normal level, and TGFβ1 and α- SMA protein expression decreased. During the formation and recovery of hepatic fibrosis, there was a negative correlation between SARA protein and Hyp content, TGFβ1 and α-SMA expression. Conclusions SARA protein is mainly expressed in hepatic stromal cells. With the development of hepatic fibrosis in rats, the expression of SARA is decreased, and the expression of SARA protein is negatively correlated with the formation of hepatic fibrosis.