德都红花-7味散原药方与优化方对肝纤维化大鼠Ⅰ,Ⅲ型胶原mRNA表达的影响

Effect of Dedu Honghua-7 Powder and Its Optimized Prescriptions on mRNA Expressions of Type Ⅰ,Ⅲ Collagen in Rats with Liver Fibrosis

目的:探讨德都红花-7味散原药方及优化方对肝纤维化大鼠Ⅰ,Ⅲ型胶原mRNA表达的影响。方法:40只Wistar大鼠分为正常组、模型组、阳性药组、原药组、优化组。大鼠ip 30%CCl4橄榄油溶液建立肝纤维化模型。同时1次/日ig给药,阳性药组给予秋水仙碱片0.4 mg·kg-1;原药组给予德都红花-7味散0.6 g·kg-1;优化组给予德都红花-7味散优化方0.6 g·kg-1。连续40 d后处死大鼠。取肝脏天狼星红染色,观察肝组织纤维化程度和Ⅰ,Ⅲ型胶原分型。酶联免疫吸附法(ELISA)检测肝组织匀浆Ⅲ型前胶原和Ⅳ型胶原,层粘连蛋白(LN),透明质酸(HA)含量。实时荧光定量PCR法检测Ⅰ型胶原,Ⅲ型胶原mRNA表达变化。结果:天狼星红染色模型组Ⅰ,Ⅲ型胶原纤维较正常组增多,阳性药组、原药组、优化组Ⅰ,Ⅲ型胶原纤维较模型组减少。与正常组比较,酶联免疫法检测模型组Ⅳ型胶原,HA含量升高(P<0.05)。与模型组比较,阳性药组、原药组、优化组Ⅲ型前胶原、Ⅳ型胶原、HA含量明显降低(P<0.05)。实时荧光定量PCR法检测模型组Ⅰ型胶原mRNA和Ⅲ型胶原mRNA表达较正常组上调(P<0.01)。阳性药组、原药组、优化组Ⅰ型胶原mRNA和Ⅲ型胶原mRNA表达较模型组下调(P<0.01)。原药组、优化组Ⅰ型胶原mRNA表达较阳性药组下调(P<0.05)。优化组Ⅰ型胶原mRNA表达较原药组下调(P<0.05)。结论:德都红花-7味散原药方与优化方是通过抑制Ⅰ型胶原mRNA和Ⅲ型胶原mRNA的转录,达到阻断或延缓肝纤维化的发生发展,德都红花-7味散优化方优于原药方。

Objective： To observe the effects of Dedu Honghua-7 powder and its optimized prescriptions on mRNA expression of type Ⅰ and Ⅲ collagen in rats with liver fibrosis. Method： Forty Wistar rats were randomly divided into normal group,model group,positive group,original drug group and optimized prescription group. The liver fibrosis models were induced by abdominal cavity injection of olive oil solution containing 30% of CCl4. At the same time,drugs were given 1 time / day by intragastrical administration. Rats in positive group received 0. 4 mg·kg- 1of colchicine pill; rats in the original drug group received 0. 6 g·kg- 1of Dedu Honghua-7 powder; rats in optimized prescription group received 0. 6 g·kg- 1of Dedu Honghua-7 powder optimized product.The rats were sacrificed after 40 days of treatment. The livers were picked out for picrosirius red staining to observe fibrosis degree of liver tissues and type Ⅰ,Ⅲ collagen classifications. ELISA assay was applied to detect the levels of type Ⅲ procollagen,type Ⅳ collagen,hyaluronic acid（ HA） and laminin（ LN） in liver tissue homogenate. The real-time q PCR was used to detect changes in mRNA expression of type Ⅰ collagen and type Ⅲcollagen. Result： In picrosirius red staining,the levels of type Ⅰ and Ⅲ collagenous fibersin model group were higher than those in normal group; the levels of type Ⅰ,Ⅲ collagenous fibersin positive group,original drug group and optimized prescription group were lower than those in model group. In ELISA assay,the levels of type Ⅳcollagen and HA in model group werehigher than those in normal group（ P〈 0. 05）. The levels of type Ⅲprocollagen,type Ⅳ collagen,and HA contents in positive group,original drug group,and optimized prescription group were lower than those in model group（ P〈 0. 05）. In real-time q PCR method,mRNA expressions of type Ⅰand Ⅲ collagen in model group were higher than those in normal group（ P〈 0. 01）. The mRNA expressions of type Ⅰ and Ⅲ collagen in positive group,original drug group and optimized prescription group were lower rhan those in model group（ P〈 0. 01）. The mRNA expressions of type Ⅰ collagen in original drug group and optimized prescription group were lower than those in positive group（ P〈 0. 05）. The mRNA expression of type Ⅰ collagen in optimized prescription group was lower than that in original drug group（ P〈 0. 05）. Conclusion： Dedu Honghua-7 powder and its optimized prescription can block or delay the occurrence and development of liver fibrosis by inhibiting the mRNA transcription of type Ⅰ,Ⅲ collagen,and the efficacy of optimized prescription is better than that of original powder.