Effect Of Serum Of Total Flavonoids Of Litsea Coreana Level On Tgf-β₁-induced Rat Hepatic Stellate Cell And Its Mechanism

Liver fibrosis is a self-healing process directing towards damage factors,at the same time,is also a common pathological basis and an inevitable stage of chronic liver diseases developing into hepatic cirrhosis;therefore,the process is an important link to influence the prognosis of chronic liver diseases.At present,it is thought that liver fibrosis is reversible while hepatic cirrhosis is irreversible.There is a high incidence rate of liver diseases in our country,and chronic hepatitis B and C are commonly encountered and seriously influence the people's life quality and labor level.Because there is no effective antiviral method,it has become a quite important countermeasure of treating chronic liver diseases to stop or reverse the liver fibrosis progress.The pathological foundation of liver fibrosis includes the increased synthesis and decreased degradation of extracellular matrix(ECM),which leads the excessive deposition of collagen.The activation and proliferation of hepatic stellate cells(HSC)(major collagen-synthesizing cells)are key links of liver fibrosis.In a chronically damaged liver,some stimulators can greatly change HSC's shape,function and phenotype,which are remarkably characterized by the increased HSC proliferation,α-SMA expression and ECM synthesis. The activation of HSC is a process of transformation from still HSC to myofibroblasts(MFB)and dependent on the network adjustment of numerous cytokines(CKs),of which transforming growth factor-β1(TGF-β1)play the most crucial roles. TGF-β1 is the strongest collagen growth stimulating factor. Now transforming growth factor-beta (TGF-β) is considered the key factor couses the liver fibrosis.Connective tissue growth factor(CTGF)may represent a downstream effector molecule of the profibrotic activities of TGF-βin the maintenance and repair of connective tissues and within fibrotic disease settings.With the gradual clarification of pathologic mechanism of liver fibrosis, it is possible to prevent,treat and reverse the liver fibrosis.However,up to now,there is no effective therapeutic method against liver fibrosis in modern medicine;while the good preventing and treating efficacies against liver fibrosis and low toxicity of traditional Chinese medicine(TCM)have brought to the extensive attention of the scholars at home and abroad at present.In recent years,a lot of experiments and clinical researches have confirmed that blood-activating and stasis-eliminating TCM has a better therapeutic efficacy in the treatment of liver fibrosis.Litea corneane levevar,which is distributed in southern part of China,has been widely used as a health promoting tea for several centuries.Previous study has showed that Lissea corneana levevar has hypolipidemic effect in rats fed with high fat diet.It has been identified that flavonoids are the main chemical componens in this plant,which possess antioxidant property.The total flavonoids also have effect on alcoholic steatohepatitis nonalcoholic steatohepatitis and Liver fibrosis.Although the studies on treating liver fibrosis by the use of TCM have made an encouraging progress,the study on active mechanism and ingredients has been a hot point and a difficult point in the new drug development and research of TCM.Compound preparations(especially cruder preparations) contain more impurities.When these preparations are added to a reaction system in vitro,their non-specific physicochemical factors such as pH, osmotic pressure,tannin,inorganic salts and so on seriously interfere the experiments and lead to false positive or false negative results.In 1984,the seropharmacological method was put forward in Japan,which is used to accurately reproduce the active ingredients of compound preparations of TCM playing roles in vivo.In this study, we use the"seropharmacological method"to further research the effect and mechanism of total flavonoids of litsea coreana level(TFLC)on liver fibrosis.The main contents are divided into two sections,as follows:1. Effect of drug serum of total flavonoids of litsea coreana level on the the mRNA expression of the collagenⅠ,Ⅲandα-SMA in hepatic stellate cellAdult healthy male rats were randomly divided into four groups(the normal and the TFLC(1,2,4 g/kg) groups),and the serum was prepared in all the groups. The HSC was cultured with the serum and TGF-β1.MTT colorimetric assay was used to evaluate the proliferation of the HSC. The mRNA expression 0fα-SMA,collagenⅠand collagenⅢin HSC were detected by RT-PCR. The TFLC serum of different groups can inhibit the proliferation of HSC and down-regulating the expression ofα-SMA,collagenⅠ,collagenⅢ. Conclution,the TFLC serum can inhibit the proliferation of HSC and decrease exudation of collagen.2. Effect of drug serum of total flavonoids of litsea coreana level on the expression of the TβR1/CTGF and Smad signaling pathway in TGF-β1-induced hepatic stellate cellAdult healthy male rats were randomly divided into four groups(the normal and the TFLC(1,2,4 g/kg) groups),and the serum was prepared in all the groups. The HSC was cultured with the serum and TGF-β1. The TFLC drug serum of different groups can decreases the expression of TβR1,CTGF,Smad2,Smad3and increase the expression of Smad7 apparently.Conclution,the TFLC`s mechanism of anti-hepatic fibrosis might be associated with down-regulating Smad3,CTGF and up-regulating Smad7 in HSC.