| Objective: To investigate the effects and mechanisms of Rougan Decoction resisting hepatic fibrosis induced by CCL4 in rats, provide theoretical basis for clinical medicine.Methods: One hundred and five male Wistar rats with 200g士20g in weight were randomly divided into blank control group, model control group, colchicines group, Dahuangzhechongwan group, and high,middle and low-dose group of Chinese Medicine, N=15。After one week's adaptive breeding, fasting 12 hours and weighting. Using the CCL4 composite approach to make the rat modle. Excepting normal blank control group, other six groups firstly use 0.5ml/100g mixture of 50% CCL4 and oliver oil to do hypodermic injection in the back, afterthen inject in the back every other 3 days. Weighing the rats every week to adjust the medicine dosage. Rougan Decoction low, medium and high dosage of adults, the amount of clinical equivalence(20g/d)for conversion, respectively, 3.6g/kg, 7.2g/kg, 14.4g/kg, diluted with different concentrations,1ml/100g.Likewise,by(6g/d)conversion, Dahuangzhechongwan is coincidently 0.108g /kg,1ml/100g. Each colchicine 1mg, referring to adult dosage of 1mg/d, calculation for 0.18mg/kg/d, double of human's dose according to the body surface. Animal sausage: normal blank group and model control group infuse in stomach with the same volume distilled water, oberseving the ordinary conditions(behavior,hair,stools and so on). First two weeks feeding the rats with high fat and low protein food(80% corn meal, 0.5% cholesterin and 20% lard), only beverage of 5% alcohol, transform commom feed after two weeks. The normal blank group accept hypodermic injection with equivalent normal saline, normal food and drinks.Take materials and blood to detect AST,ALT,PH,TP,HA,LN,C-IV,PCⅢafter eight weeks. Investigating the variation of liver histology: Hestain and Masson trichrome stain. Meanwhile detecting TGF-β1 and protein expression of Smad3,Smad7 by immunohistochemical method. Results:1. Rougan Decoction effectively reduce the level of AST,ALT,PH,TP of heptic fibrosis in rats, the influnce of high-dose group is the optimal, better than colchicine group, Dahuangzhechong group, low and medium-dose of Rougan Decoction group(P<0.05).Efficacy of low and medium-dose group of Rougan Decoction, colchicine group and Dahuangzhechong group is similar to each other, there is no obvious difference among the four groups(P>0.05).2. Rougan Decction can effectively reduce the plasma level of HA,LN,C-Ⅳ,PCⅢof the heptic fibrosis in rats, the therapeutic efficacy of high-dose group is the best, better than colchicine group, Dahuangzhechong group, low and medium-dose group of Rougan Decoction(P<0.05). Efficacy of low and medium-dose group of Rouganjian, colchicine group and Dahuangzhechong group have equivalent effects, no obvious difference exist among the four groups(P>0.05).3. Observations with light microscope after HE staining: the high-dose group of Rougan Decoction significantly reduce the activity extent of inflammation in hepatic tissue and the degree of hepatic fibrosis, the efficacy of high dose group is better than colchicine group, Dahuangzhechong group, low and medium-dose group of Rougan Decoction(P<0.05). Efficacy of low and medium-dose group of Rougan Decoction, colchicine group and Dahuangzhechong group is almost similar, no obvious difference exist among the four groups(P>0.05).4. After Masson trichrome stain, the result of counting the collagenous area in hepatic tissue and the proportion of the total collagenous area to the visual field shows that: the high-dose group of Rougan Decoction significently reduce the total collagenous area of hepatic tissue and also decrease the proportion of the total field of vision.The effect is markedly better than colchicine group, Dahuangzhechong group, low and medium-dose group of Rougan Decoction(P<0.05). Efficacy of low and medium-dose group of Rougan Decoction, colchicine group and Dahuangzhechong group is almost resembled, no obvious difference exist among the four groups(P>0.05).5.Rougan Decoction can effectively inhibit the expression of TGF-β1,Smad3 and markedly increase the expression of Smad7 in the liver tissue, the efficacy of high-dose group is the optimal.The influnce is better than colchicine group, Dahuangzhechong group, low and medium-dose group of Rougan Decocotion(P<0.05). Efficacy of low and medium-dose group of Rougan Decoction, colchicine group and Dahuangzhechong group has a equivalent effect, there is no markble variance among the four groups(P>0.05).6. Rougan Decoction can effectively reduce the TGF-β1,Smad3mRNA in the liver tissue,and increase the Smad7mRNA in liver tissue levels of high-dose group of Rougan Decoction most obviously was superior to colchicine group, Dahuangzhechong group, low and medium-dose group of Rougan Decoction(P<0.05). Efficacy of low and medium-dose group of Rougan Decoction , colchicine group and Dahuangzhechong group is equivalent ,there is no significant difference among the four groups(P>0.05).7.The therapic efficacy of Rougan Decoction is related to the definite dosage: the high-dose group is superior to the low and medium one, the medical influnce of the medium-dose group is similar to the low one, no significant relationship with treatment course.Conclutions:1. Rougan Decoction effectively reduce the level of AST,ALT,PH and increase TP of heptic fibrosis in rats.It also can effectively reduce the plasma level of HA,LN,C-Ⅳ,PCⅢ.2. Rougan Decoction apparently reduce the total collagenous area in hepatic tissue and the proportion of the total field to vision which points out Rougan Decoction has efficacy of inhibiting collagenous hyperplasis and collagen sendiment in the extracellular matrix of liver.3. Rougan Decoction obviously inhibit the expression of TGF-β1,Smad3 and enhance the expression of Smad7 in the hepatic tissue,accordingly inhibit TGF-β1 by means of Smad3,Smad7 to promote hepatic fibrosis.4. Rougan Decoction can reduce the mRNA expression of TGF-β1 and Smad3 and increase the mRNA expression of Smad7.This may be one mechanism of resist hepatic fibrosis by Rougan Decoction. |
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