Protective Effect And Part Of Mechanisms Of Prunella Vulgaris Sulfated Polysaccharide On Liver Fibrosis In Rats

Aim:Prunella vulgaris sulfated polysaccharide?PVSP?,the main bioactive compound of PV that possesses anti-inflammatory,antioxidant,immunoregulatory and anti-tumor activities.However,the antifibrotic effects of PVSP remains poorly understood.The present study was designed to the effect of Prunella vulgaris sulfated polysaccharide?PVSP?on TGF-?1-activated HSC-T6 cell and CCl₄-induced liver fibrosis,and to explore related mechanism.Methods:HSC-T6 cells were stimulated by TGF-?1and treated with different doses of PVSP?50,100,200?g/ml?,the HSC-T6 proliferation were detected by CCK8.The expression levels of?-smooth muscle actin??-SMA?and collagen type I?Col-I?mRNA were detected by qRT-PCR.The production of?-SMA and Col-I protein was measured by Western blot.The liver fibrosis model was established by intraperitoneal injection with 40%CCl₄.The rats were randomly divided into five groups:a nomal control group,a positive control group?colchicines 0.25 mg/kg?,a high-PVSP group?PVSP 300 mg/kg?,a low-PVSP group?100 mg/kg?and a model group.Serum levels of ALT?aspartate aminotransferase?,AST(aspartate aminotransferase,HA?hyaluronic acid?,LN?laminin?,PC-III?procollagen-III-peptide?and C-IV?type IV collagen?were examined by ELISA?enzyme linked immunosorbent assay?in dfferent groups.Hematoxylin and eosin?HE?staining and sinus red staining were used to examine the degree of hepatic fibrosis.RT-PCR was used to detect the expression of?-SMA,Col-I and TGF-?₁ mRNA in liver tissues of rats,and immunohistochemistry Staining was detected the content of?-SMA,Col-I and TGF-?₁ protein in liver tissues.Moreover,the levels of Smad3,Smad2/3 and Smad7in liver tissues were examined by Wertern blot.Results:In vitro,PVSP could inhibit the proliferation of HSC-T6 induced by TGF-?₁.Compared with the TGF-?₁ group,PVSP significantly reduced mRNA and protein expression of?-SMA and Col-I in HSC-T6 cells?P<0.05 or P<0.01?.In vivo,in the model group,serum levels of ALT,AST,HA,LN,PC-III and C-IV increased obviously compare with the normal group?P<0.05 or P<0.01?.Compared with the model group,the levels of ALT,AST,HA,LN,PC-III and C-IV significantly deccreased in the PVSP groups?P<0.05?,and the high-PVSP group had a better effect?P<0.05?.The pathological results showed that PVSP could collagen deposition and the degree of hepatic fibrosis.The expression of?-SMA,Col-I and TGF-?₁ mRNA and protein in liver tissues rats increased obviously compare with the normal group.The expression of Smad3 and Smad2/3 increased in model group compare with the normal group?P<0.05 or P<0.01?,while the content of Smad7decreased?P<0.05?.Compared with the model group,the levels of Smad3 and Smad2/3 were significantly reduced?P<0.05?,and the expression of Smad7decreased?P<0.05?.Conclusion:PVSP treatment could inhibit TGF-?1-activated HSCs.PVSP treatment also could alleviate CCl₄-induced liver fibrosis,and its mechanism may be through regulating the TGF-?₁/Smads signal pathway,thereby inhibiting proliferation and activation of HSCs,downregulating the formation of ECM,finally could reverse the development of hepatic fibrisis.This study provided a basis for using PVSP to treat hepatic fibrosis and may encourage further experimental and clinical trials for developing new drugs to treat this disease.