| Hepatic fibrosis is a pathologic process that every kind of disease cause liver injure and inflammation.It is the hallmark of most chronic liver diseases. Its essense is excessive deposition of extracellular matrix (ECM) components in liver, which occurs due to an imbalance between the production and degradation of matrix. Collagen is the most important part of ECM.In previous studies, Hepatic fibrosis is limited to a passive stationary state. With the development of molecular biological technology, People have made large progress in many ways,such as hepatic stellate cell activation, Establishment of Animal Model , cytokine signaling ,the molecular targeted therapyin Hepatic fibrosis and so on . transforming growth factor-β1 ,tissue inhibit of metallo Proteinase and extracellular matrix components have become a hotspot for research.In current research work, anti-hepatic fibrosis drug main contents include the type of cytoprotection, hepatic stellate cell activation, Cytokines activity, suppressed extracellular matrix production and secretion, Promote degradation of extracellular matrix, traditional medicine monomer and Chinese Herbal Compound. But HF is a complex process there is no established therapy for hepatic fibrosis。The characteristics of Traditional Chinese Medicine (TCM) treatment are multiple positions, multiple links and multiple targets. There are much superiority in the prevention and treatment on Diabetes Mellitus. And now, to search the traditional medicine drugs or the extract of these drugs is important in the prevention and treatment of HF. If we can explore the possible mechanisms of these drugs, it will benefit the TCM modernization, the treatment of HF.To study the effect of Huazhuo Jiedu Yiqi'formula on the effect of immunity hepatic fibrosis rats induced by pig's serum. Investigate on Pathological Mechanism of Hepatic fibrosis,so that provide theoretical and experimental basis for treating liver fibrosis.ObjectivesThe aim was to study the effect of The ZhuoDu theory in the position of Therapy for Hepatic Fibrosis, and explore its possible mechanism.. The research included preliminary Study of The ZhuoDu theory on Therapy for Hepatic Fibrosis, effect of Huazhuo Jiedu Yiqi'formula on the levels of serum ALT,AST,ALB,TP and Pathomorphology in hepatic fibrosis rats;effect of Huazhuo Jiedu Yiqi'formula on the levels of serum HA,LN,PCⅢ,CIV and Pathomorphology in hepatic fibrosis rats;Huazhuo Jiedu Yiqi'formula on the expression of TGF-β1 in liver tissue of hepatic fibrosis rats; Huazhuo Yiqi'formula on the expression of TIMP-1 in liver tissue of hepatic fibrosis rats.MethodsTotal 60 clean male Wistar rats weighed 200～220 grams were randomly divided into six groups:namely,the normal group, the model group, colchicine prevention group,(C Prevention group for short),Huazhuo Jiedu Yiqi'formula Prevention group(HJY Prevention group for short), colchicine treatment group(C treatment group), Huazhuo Jiedu Yiqi'formula treatment group(HJY reatment group)with 10 rats for each group .In addition to the normal group,pig's serum( 0.5ml per mouse, twice a week) was established in rats by intraperitoneal injection for eight weeks. The drugs were given through stomach-perfusion to C Prevention group and HJY Prevention group at the beginning of experiment.The drugs are given at the same time every day.The treatment groups were given drug at the end of the eighth week. The rats in the normal control and model groups received intragastric administration of saline(2ml each time).Five rats were killed in each group at the 8th and 12th weekend. We draw blood from abdominal aorta, ALT,AST level were measured with Reit′s method; The contents of TP andAlb were measured using biurea method and bromocresol green method ;HA,LN,PCⅢ,CⅣlevel were measured with radioimmunoassay(RIA). The specimens were taken and the degree of hepatic fibrosis was judged by routine haematoxylin-eosin staining and Masson staining.TGF-β1 protein and mRNA expression were determined using immunohistochemical technique and RT-PCR.TIMP-1 protein and mRNA expression were determined using immunohistochemical technique and RT-PCR .Data was collected and analyzed through t test or analysis of variance. The statistic software was SPSS. Differences with P values <0.05 were considered significant.Results1 Effect of Huazhuo JieDu Yiqi'formula on the levels of serum ALT,AST,ALB,TP and pathomorphology in hepatic fibrosis rats.1.1 Effect of Huazhuo JieDu Yiqi'formula on the levels of serum ALT,AST,ALB,TP in hepatic fibrosis rats.Compared with the normal group, the content of ALT and AST was significantly increased with liver injure in the model group(P<0.05). TP,ALB of serum can be increased from C Prevention group and HJY Prevention group significantly at the end of 8 weeks(P<0.05). The effect of HJY Prevention group was better than C prevention group(P<0.05). Compared with the model group, All medicine intervention group decreased the level of serum ALT and AST and increased the concentration of serum TP,ALB at the end of 12 weeks ,there were statistic differences among the groups(P<0.05) .The effect of Chinese Herbal Compound can reduce the evels of serum ALT,AST,ALB,TP in hepatic fibrosis rats and the effect is better than Colchicine (P<0.05) . But there was no significant difference between the effect of prevention and treatment with using two medicines(P>0.05).1.2 Effect of Huazhuo JieDu Yiqi'formula to pathomorphology in hepatic fibrosis rats.The structure of hepatic lobule were integrated hepatic cord radially ranged as the center of navel vein.There was not degeneration and necrosis in liver tissue. There was not collagen proliferation.Compared with normal group,the model control rats had obvious liver inflammation including degeneration,necrosis,fibrosis and collagen proliferation.Pseudolobule proliferation could be seen in individual animal.At the end of 8th weeks, Hepatic lobule in colchicine prevention group and HJY Prevention group lessed injured than model group. Hepatocellular degeneration and necrosis and inflammatory cell infiltration could be seen in liver tissue.Collagen proliferation in the two groups was less than in model group,it were mainly scattered in hepatic sinusoid and disse clearance. There was not pseudolobule proliferation. Pathological examination in HJY Prevention group was better than C prevention group. At the end of 12th weeks, Pathological examination in the treatment groups less injured than in model group. Pathological examination in C treatment group was similar to that of C prevention group ,the result was same between HJY Prevention group and treatment group.And The effect of Chinese Herbal Compound is better than colchicine.2 Effect of Huazhuo Yiqi'formula on the levels of serum HA,LN,PCⅢ,CIV and pathomorphology in hepatic fibrosis rats2.1 Effect of Huazhuo Yiqi'formula on the levels of serum HA,LN,PCⅢ,CIV and pathomorphology in hepatic fibrosis ratsCompared with the normal group, the levels of serum HA,LN,PCⅢ,CIV could be increased from treament group with the significant difference(P<0.05) . The levels of serum HA,LN,PCⅢ,CIV could be decreased from C Prevention group and HJY Prevention group significantly at the end of 8 weeks(P<0.05). The effect of HJY Prevention group was better than C prevention group(P<0.05).Compared with the model group, All medicine intervention group decreased the level of serum HA,LN,PCⅢ,CIV at the end of 12 weeks,There were statistic differences among the groups(P<0.05).The Effect of Chinese Herbal Compound can reduce the levels of serum HA,LN,PCⅢ,CIV in hepatic fibrosis rats and the effect is better than colchicine (P<0.05) . But there was no significant difference between the effect of prevention and treatment with using two medicines(P>0.05). 2.2 Effect of Huazhuo Yiqi'formula to pathomorphology in hepatic fibrosis rats.The structure of hepatic lobule were integrated hepatic cord radially ranged as the center of navel vein.There was not degeneration and necrosis in liver tissue. There was not collagen proliferation.Compared with normal rats,the model control rats had obvious liver inflammation including degeneration,necrosis,fibrosis and collagen proliferation .Pseudolobule proliferation could be seen in individual animal.At the end of 8th weeks, Hepatic lobule in C prevention group and HJY Prevention group lessed injured than model group. Hepatocellular degeneration and necrosis and inflammatory cell infiltration could be seen in liver tissue. Collagen proliferation in the two groups was less than in model group,it were mainly scattered in hepatic sinusoid and disse clearance. There was not pseudolobule proliferation. Pathological examination in HJY Prevention group was better than C prevention group. At the end of 12th weeks, Pathological examination in the treatment groups less injured than in model group. Pathological examination in C treatment group was similar to that of C prevention group ,the result was same between HJY Prevention group and treatment group.And The effect of Chinese Herbal Compound is better than colchicine.3 Experimental study on Huazhuo Jiedu Yiqi'formula effect on TGF-β1 of the immunity hepatic fibrosis rats3.1 Experimental study on Huazhuo Jiedu Yiqi'Formula effect on TGF-β1 pretine of the immunity hepatic fibrosis ratsThe positive expression cells of normal group was a few. the expression of TGF-β1 was strongly positive in model group.The level was higher than normal group. It could be manifested as heterogeneously vinegar particles.Less positive expression of TGF-β1 cell can be seen in the liver tissue of rats form normal group,the cell proportion of TGF-β1 cell expression in the liver tissue of rats form model group were significantly higer than that of the normal group,which were manifested by the unsymmetric brown yellow particulates(P<0.05). The protein level of TGF-β1 of immunity hepatic fibrosis rats could be decreased from C Prevention group and HJY Prevention group significantly at the end of 8 weeks(P<0.05). The effect of HJY Prevention group was better than C prevention group(P<0.05).Compared with the model group, All medicine intervention group decreased the protein level of TGF-β1 of immunity hepatic fibrosis rats at the end of 12 weeks,There were statistic differences among the groups(P<0.05). The Effect of Chinese Herbal Compound can reduce the expression of TGF-β1 protein of immunity hepatic fibrosis rats and the effect is better than Colchicine (P<0.05) . But there was no significant difference between the effect of prevention and treatment with using two medicines(P>0.05).3.2 Experimental study on Huazhuo Jiedu Yiqi'formula effect on TGF-β1 mRNA of the immunity hepatic fibrosis ratsThe results showed that the expressions of TGF-β1 mRNA in the model groups were increased significantly, respectively compared with normal control group and the difference was significant (P<0.05). The expressions of TGF-β1 mRNA of immunity hepatic fibrosis rats could be decreased from C Prevention group and HJY Prevention group significantly at the end of 8 weeks(P<0.05). The effect of HJY Prevention group was better than C prevention group(P<0.05).Compared with the model group, All medicine intervention group decreased the expressions of TGF-β1 mRNA of immunity hepatic fibrosis rats at the end of 12 weeks,There were statistic differences among the groups(P<0.05).The Effect of Chinese Herbal Compound could reduce the expression of TGF-β1 mRNA of hepatic fibrosis rats and the effect is better than Colchicine (P<0.05) . But there was no significant difference between the effect of prevention and treatment with using two medicines(P>0.05).4 Experimental study on Huazhuo Jiedu Yiqi'formula effect on TIMP-β1 of the immunityhepatic fibrosis rats4.1 Experimental study on Huazhuo Jiedu Yiqi'formula effect on TIMP-1 pretine of the immunity hepatic fibrosis rats The positive expression cells of normal group was a few. the expression of TIMP-1 was strongly positive in model group.The level was higher than normal group. It could be manifested as heterogeneously vinegar particles.Less positive expression of TIMP-1cell can be seen in the liver tissue of rats form normal group,the cell proportion of TIMP-1 cell expression in the liver tissue of rats form model group were significantly higer than that of the normal group,which were manifested by the unsymmetric brown yellow particulates(P<0.05). The protein level of TIMP-1 of immunity hepatic fibrosis rats could be decreased from C Prevention group and HJY Prevention group significantly at the end of 8 weeks(P<0.05). The effect of HJY Prevention group was better than C prevention group(P<0.05).Compared with the model group, All medicine intervention group decreased the protein level of TIMP-1 of immunity hepatic fibrosis rats at the end of 12 weeks,There were statistic differences among the groups(P<0.05). The Effect of Chinese Herbal Compound can reduce the expression of TIMP-1 protein of immunity hepatic fibrosis rats and the effect is better than colchicine (P<0.05) . But there was no significant difference between the effect of prevention and treatment with using two medicines(P>0.05).4.2 Experimental study on Huazhuo Jiedu Yiqi'formula effect on TIMP-1 mRNA of the immunity hepatic fibrosis ratsThe results showed that the expressions of TIMP-1 mRNA in the model groups were increased significantly, respectively compared with normal control group and the difference was significant (P<0.05). The expressions of TIMP-1 mRNA of immunity hepatic fibrosis rats could be decreased from C Prevention group and HJY Prevention group significantly at the end of 8 weeks(P<0.05). The effect of HJY Prevention group was better than C prevention group(P<0.05).Compared with the model group, All medicine intervention group decreased the expressions of TIMP-1 mRNA of immunity hepatic fibrosis rats at the end of 12 weeks,There were statistic differences among the groups(P<0.05).The Effect of Chinese Herbal Compound could reduce the expression of TIMP-1 mRNA of hepatic fibrosis rats and the effect is better than Colchicine (P<0.05) . But there was no significant difference between the effect of prevention and treatment with using two medicines(P>0.05).Conclusions1 ZhuoDu theory has an important effect on etiology and pathogenesis in tcm analysis ,it play an important role in diagnosis and treatment of hepatic fibrosis.2 Huazhuo Jiedu Yiqi'formula can improve the liver pathologic injur and ameliorate hepatocyte injury in hepatic fibrosis rats , exert an anti-hepatic fibrosis effect.3 Huazhuo Jiedu Yiqi'formula decreased the level of serum ALT and AST and increased the concentration of serum TP,ALB, it may play a role in liver-protection,Anti-inflammatory effect of Huazhuo Jiedu Yiqi'formula lead the resistance against oxidant-induced injury anti-hepatic fibrosis,which is better than colchicines.4 Huazhuo Jiedu Yiqi'formula could significantly reduce serum HA, LN,PCⅢ, CⅣand collagen hyperplasia in liver of hepatic fibrosis rats, exert an anti-hepatic fibrosis effect.5 Jiedu Huazhuo Yiqi'formula could reduce the expression of TGF-β1 protein and mRNA of immunity hepatic fibrosis rats, suppresse theaetivation of HSCs, decrease the deposition of ECM.6 Jiedu Huazhuo Yiqi'formula could significantly reduce the expression of TIMP-1 protein and mRNA of immunity pig serum-induced hepatic fibrosis of rat, which might play a role in regulating the disequilibrium between MMPS and TIMPS, promote the degrading of ECM, decrease the deposition of ECM.
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