荣肝汤对二甲基亚硝胺大鼠肝纤维化的干预作用研究

Preventive Effects of Ronggan Decoction on Dimethylnitrosamin-induced Hepatic Fibrosis in Rats

目的:观察荣肝汤对二甲基亚硝胺(DMN)诱导的大鼠肝纤维化的干预作用。方法:35只Wista雄性大鼠随机分为对照组和造模组,以0.5%DMN按2 mL/kg体质量腹腔注射4周复制大鼠肝纤维化模型,后将造模组大鼠随机分为模型组,荣肝汤组,灌胃相应药物3周。7周末处死动物,留取标本,检测血清丙氨酸氨基转移酶(ALT)和天门冬氨酸氨基转移酶(AST)活性和白蛋白(Alb)、总胆红素(TBIL)含量;测定肝组织中羟脯氨酸(Hyp)含量。HE染色和天狼猩红染色观察肝组织病理学改变。蛋白印迹法检测大鼠肝组织I型胶原和alpha平滑肌肌动蛋白(α-SMA)表达水平。结果:与正常组比较,模型组大鼠血清TBIL、ALT、AST水平明显升高(P<0.05),Alb含量显著下降(P<0.05),经荣肝汤干预后,血清学肝功能指标显著改善。HE染色显示模型组肝脏内有大量炎性细胞浸润,肿胀、变性坏死,有明显出血现象;天狼猩红染色显示模型组大鼠肝脏胶原增生明显,形成假小叶,胶原面积明显增加。模型组肝组织Hyp含量较正常组明显升高[(1412.42±367.68)μg/g VS(530.20±62.37)μg/g,P<0.01],肝组织Ⅰ型胶原抗体、α-SMA蛋白表达水平显著升高;药物干预后肝组织炎症和纤维化程度较模型组均有所改善,肝组织Hyp含量明显降低[(1092.83±146.19)μg/g]。统计分析显示荣肝汤组与模型组比较肝组织胶原面积明显减轻[(358 810±235 058)×10~4μm^2VS(358 810±235 058)×10~4μm^2,P<0.05],肝组织Ⅰ型胶原抗体、α-SMA表达水平明显降低。结论:荣肝汤对DMN诱导的大鼠肝纤维化有较好的逆转作用。

Objective To observe the intervention effects of Ronggan decoction in dimethylnitrosamin（DMN）-induced chronic hepatic fibrosis in rat.Methods A total of 35 Wister male rats were randomly divided into the normal control group and the model group. The model group was used to establish the hepatic fibrosis model with 0.5% DMN by intraperitoneal injection of 2 mL/kg of body mass for 4 weeks, and then by randomly divided the rats into model group and Ronggan group. The two groups were treated with corresponding drug intragastrically every day for 3 weeks. Rats were then sacrificed for sampling at 7 the weekend, and the serum levels of alanine aminotransferase（ALT）, aspartate aminotransferase（AST）,albumin（Alb）, total bilirubin（TBIL）,as well as hepatic levels of hydroxyproline（Hyp） were measured. Pathological changes of liver tissue was evaluated by Sirius Red staining and HE staining. Collagen type Ⅰ and smooth muscle actin（α-SMA） protein level inliver was determined by western blotting.Results Compared with the normal control group, serum levels of TBIL, ALT, AST were significant increased and Alb were significant descend in the model group（P〈0.05）. After the treatment of the Ronggan decoction, serological index of liver function improved significantly. HE staining showed that the model group had a large number of inflammatory cell infiltration, swelling, degeneration andnecrosis, and bleeding phenomenon was significant in the liver tissue. Sirius red staining showed that the livercollagen of the model group was significantly hyperplasia, forming a pseudo lobule, collagen area was significantly increased. The Hyp content of the liver tissue of the model group were significantly increased than the normalcontrol group（[1412.42±367.68）μg/g VS（530.20±62.37）μg/g,P〈0.01], and the expression levels of collagen type Ⅰand α-SMA has a significant increased. The degree of inflammation and fibrosis in liver tissue wereimproved in the model group after the drug intervention, and the Hyp content of the liver tissue were decreasedobviously（[1092.83 ± 146.19） μg/g]. Statistical analysis shows that the liver tissue collagen area of Ronggan group were significantly descend than the model group [（358 810±235 058） ×10^4μm^2 VS（358 810±235 058）×10^4μm^2, P〈0.05], and the expression levels of collagen type Ⅰ antibodies and α-SMA were decreased obviously.Conclusion Ronggan decoction has better reverse action to liver fibrosis induced by DMN.