AIM To explore the role and mechanism of total flavone of Abelmoschus manihot(TFA) on epithelial-mesenchymal transition(EMT) progress of Crohn's disease(CD) intestinal fibrosis.METHODS First,CCK-8 assay was perfor...AIM To explore the role and mechanism of total flavone of Abelmoschus manihot(TFA) on epithelial-mesenchymal transition(EMT) progress of Crohn's disease(CD) intestinal fibrosis.METHODS First,CCK-8 assay was performed to assess TFA on the viability of intestinal epithelial(IEC-6) cells and select the optimal concentrations of TFA for our further studies.Then cell morphology,wound healing and transwell assays were performed to examine the effect of TFA on morphology,migration and invasion of IEC-6 cells treated with TGF-β1.In addition,immunofluorescence,real-time PCR analysis(q RT-PCR) and western blotting assays were carried out to detect the impact of TFA on EMT progress.Moreover,western blotting assay was performed to evaluate the function of TFA on the Smad and MAPK signaling pathways.Further,the role of co-treatment of TFA and si-Smad or MAPK inhibitors has been examined by q RTPCR,western blotting,morphology,wound healing andtranswell assays.RESULTS In this study,TFA promoted transforming growth factor-β1(TGF-β1)-induced(IEC-6) morphological change,migration and invasion,and increased the expression of epithelial markers and reduced the levels of mesenchymal markers,along with the inactivation of Smad and MAPK signaling pathways.Moreover,we revealed that si-Smad and MAPK inhibitors effectively attenuated TGF-β1-induced EMT in IEC-6 cells.Importantly,co-treatment of TFA and si-Smad or MAPK inhibitors had better inhibitory effects on TGF-β1-induced EMT in IEC-6 cells than either one of them.CONCLUSION These findings could provide new insight into the molecular mechanisms of TFA on TGF-β1-induced EMT in IEC-6 cells and TFA is expected to advance as a new therapy to treat CD intestinal fibrosis.展开更多
AIM: To investigate the effect of various concentrations of tetrandrine on activation of quiescent rat hepatic stellate cells (HSCs) and transforming growth factor-β (TGF-β) signaling in vitro.METHODS: HSCs were iso...AIM: To investigate the effect of various concentrations of tetrandrine on activation of quiescent rat hepatic stellate cells (HSCs) and transforming growth factor-β (TGF-β) signaling in vitro.METHODS: HSCs were isolated from rats by in situperfusion of liver and 18% Nycodenz gradient centrifugation, and primarily cultured on uncoated plastic plates for 24 hwith DMEM containing 20% fetal bovine serum (FBS/DMEM) before the culture medium was substituted with 2% FBS/DMEM for another 24 h. Then, the HSCs were cultured in 2% FBS/DMEM with tetrandrine (0.25, 0.5, 1,2 mg/L, respectively). Cell morphological features were observed under an inverted microscope, smooth muscleα-actin (α-SMA) was detected by immunocytochemistry and image analysis system, laminin (LN) and type Ⅲprocollagen (PCⅢ) in supernatants were determined byradioimmunoassay. TGF-β1 mRNA, Smad 7 mRNA and Smad 7 protein were analyzed with RT-PCR and Western blotting, respectively.RESULTS: Tetrandrine at the concentrations of 0.25-2 mg/L prevented morphological transformation of HSC from the quiescent state to the activated one, while α-SMA, LN and PCⅢ expressions were inhibited. As estimated by gray values, the expression of α-SMA in tetrandrine groups (0.25, 0.5, 1, 2 mg/L) was reduced from 21.3% to 42.2%(control: 0.67, tetrandrine groups: 0.82, 0.85, 0.96, or 0.96, respectively, which were statistically different from the control, P<0.01), and the difference was more significant in tetrandrine at 1 and 2 mg/L. The content of LN in supernatants was significantly decreased in tetrandrine groups to 58.5%, 69.1%, 65.8% or 60.0% that of the control respectively, and that of PCⅢ to 84.6%, 81.5%,75.7% or 80.7% respectively (P<0.05 vs control), with no significant difference among tetrandrine groups. RTPCR showed that TGF-β1 mRNA expression was reduced by tetrandrine treatments from 56.56% to 87.90% in comparison with the control, while Smad 7 mRNA was increased 1.4-4.8 times. The TGF-β1 mRNA and Smad 7 mRNA expression was in a significant negative correlation (r= -0.755, P<0.01), and both were significantly correlated with α-SMA protein expression (r = -0.938, P<0.01;r = 0.938, P<0.01, respectively). The up-regulation of Smad 7 protein by tetrandrine (1 mg/L)was confirmed by Western blotting as well.CONCLUSION: Tetrandrine has a direct inhibiting effect on the activation of rat HSCs in culture. It up-regulates the expression of Smad 7 which in turn blocks TGF-β1 expression and signaling.展开更多
According to data from 2020,Slovakia has long been among the top five countries with the highest incidence rate of colorectal cancer(CRC)worldwide,and the rate is continuing to rise every year.In approximately 80%of C...According to data from 2020,Slovakia has long been among the top five countries with the highest incidence rate of colorectal cancer(CRC)worldwide,and the rate is continuing to rise every year.In approximately 80%of CRC cases,allelic loss(loss of heterozygosity,LOH)occurs in the long arm of chromosome 18q.The most important genes that can be silenced by 18q LOH or mutations are small mothers against decapentaplegic homolog(SMAD)2 and SMAD4,which are intracellular mediators of transforming growth factor(TGF)-βsuperfamily signals.TGF-βplays an important role in the pro-oncogenic processes,including such properties as invasion,epithelial-mesenchymal transition(commonly known as EMT),promotion of angiogenesis,and immunomodulatory effects.Several recent studies have reported that activation of TGF-βsignaling is related to drug resistance in CRC.Because the mechanisms of drug resistance are different between patients in different stages of CRC,personalized treatment is more effective.Therefore,knowledge of the activation and inhibition of factors that affect the TGF-βsignaling pathway is very important.展开更多
OBJECTIVE:To examine the effects of moxibustion on myocardial injury and myocardial metabolomics in rats with rheumatoid arthritis(RA)based on the transforming growth factor beta1(TGF-β1)/Smads signaling pathway.METH...OBJECTIVE:To examine the effects of moxibustion on myocardial injury and myocardial metabolomics in rats with rheumatoid arthritis(RA)based on the transforming growth factor beta1(TGF-β1)/Smads signaling pathway.METHODS:One hundred rats were treated with saline[normal control(NC)group]or complete Freund’s adjuvant(CFA)by right plantar injection for the RA model group,and the latter were randomly divided into 4 groups.Tripterygium wilfordii polyglycoside tablets(雷公藤多苷片,TPT)have anti-inflammatory and are widely used in the clinical treatment of RA,therefore serving as a positive control group.Three days post injection rats were given TPT tablet(TPT group),acupuncture therapy(APT group),and moxibustion treatment(MOX group)for 15 consecutive days,while NC group and model group were equally grasped and fixed and received normal saline.Rat joint swelling scores and arthritis index(AI)were evaluated in each group before the CFA challenge,therapy and after receiving therapy.Myocardial ultrastructure was observed by electron microscope.Enzyme-linked immunosorbent assay was used to detect cardiac troponin I(cTnI)levels in rat myocardial tissue.Quantitative reverse transcription polymerase chain reaction and Western blotting analysis were used to measure the mRNA and protein levels of TGF-βsignaling molecules including TGF-β1,Smad2,Smad3,Smad4,and Smad7.Myocardial metabolomics was analyzed using gas chromatography-mass spectrometer.RESULTS:Compared with model group,RA model rats receiving TPT,acupuncture,or moxibustion therapy all showed reduced joint swelling scores and AI(all P<0.01)and improved myocardial damage,whereas rats treated with moxibustion were found to be more marked.Consistently,the expressions of cTnI,TGF-β1,Smad2,Smad3,and Smad4 were found to be elevated in model rat group in contrast to NC rats and were significantly downregulated in TPT,APT and MOX group when compared with model group,while the levels of Smad7 showed the opposite result(all P<0.01).Moreover,the dissection of metabolomics suggested a novel metabolite biomarker panel including D-Xylulose 5-phosphate,dihydroxyacetone phosphate,arachidonic acid,etc was defined and implicated in amino acid,glucose,and fatty acid metabolic processes as revealed by principal component analysis and partial least squares discriminant analysis.CONCLUSION:Moxibustion prevents RA-induced inflammatory response and offers potent therapeutic effects on myocardial dysfunctions.The protective effects might be associated with its role in TGF-β1 inactivation and metabolic reprogramming.展开更多
OBJECTIVE:To evaluate the effect of Wulong Xiaozheng Wan medicated serum on the epithelial-mesenchymal transition (EMT) of BGC823 cell induced by transforming growth factor-β,(TGF-β,) and to explore its mechanism.ME...OBJECTIVE:To evaluate the effect of Wulong Xiaozheng Wan medicated serum on the epithelial-mesenchymal transition (EMT) of BGC823 cell induced by transforming growth factor-β,(TGF-β,) and to explore its mechanism.METHODS:EMT model of BGC823 was stimulated by TGF-β1.Wulong Xiaozheng Wan medicated serum and LY-364947 were used as intervention.The proliferation and adhesion of BGC823 were detected by 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide and flow cytometry was used to detect the apoptosis.The invasion and migration were detected by Transwell.The level of matrix metalloproteins was detected by enzyme-linked immunosorbent assay.The expressions of related proteins and mRNA of EMT marker and TGF-β1/Smad signal pathway were detected by Western blot and reverse transcription-polymerase chain reaction.RESULTS:Compared with the TGF-β1 group,Wulong Xiaozheng Wan medicated serum could inhibit the ability of proliferation,heterogeneous adhesion,invasion,and migration.It also promotes apoptosis and homotypic adhesion in BGC823,with a dose-dependent manner.Meanwhile,Wulong Xiaozheng Wan medicated serum could regulate the expression of related proteins and mRNA of TGF-β1/Smad signaling pathway,and inhibit the expressions of EMT transcription factors and EMT markers.CONCLUSION:Wulong Xiaozheng Wan medicated serum inhibited epithelial-mesenchymal transition by down-regulated the expression of TβRI and the activation of TGF-β1/Smad signaling pathway.展开更多
OBJECTIVE:To investigate the potential mechanism by which Shugan Huoxue Huayu Fang(疏肝活血化瘀方,SGHXHYF)ameliorates liver fibrosis.METHODS:Liver fibrosis was induced in rats by intraperitoneal injection of carbon te...OBJECTIVE:To investigate the potential mechanism by which Shugan Huoxue Huayu Fang(疏肝活血化瘀方,SGHXHYF)ameliorates liver fibrosis.METHODS:Liver fibrosis was induced in rats by intraperitoneal injection of carbon tetrachloride(CCl_(4))in peanut oil solution(40%,3 m L/kg body weight)twice a week for 8 weeks.A normal control group received the same volume of peanut oil alone.During weeks 5-8,the CCl_(4)-injected rat groups were administered saline(vehicle control),colchicine(0.1 mg/mL,1 mg/kg,positive control),or SGHXHYF(0.1 mg/mL;0.3,0.6 and 1.2 mg/kg)once daily by oral gavage.Rats were sacrificed 24 h after the last treatment.Blood samples were collected for measurement of serum alanine aminotransferase(ALT),aspartate aminotransferase(AST),alkaline phosphatase(ALP),albumin(ALB),collagenⅠ,and collagenⅢlevels.Liver samples were analyzed by histopathological staining,Masson's staining of extracellular matrix proteins,and immune-ohistochemical staining ofα-smooth muscle actin(α-SMA).TGF-β1/Smad protein and mRNA levels were analyzed by Western blot and quantitative reverse transcription-polymerase chain reaction analysis,respectively.In vitro experiments were also performed using rat hepatic stellate cells(HSCs).RESULTS:Compared with the control animals,CCl_(4)-exposed rats exhibited elevated serum levels of ALT,AST,ALP,collagenⅠ,and collagenⅢ;reduced serum levels of ALB;and increased collagen deposition andα-SMA expression in liver sections,reflecting liver fibrosis.CCl_(4) also increased expression of TGF-β1 and the activated(phosphorylated)forms of Smad2 and Smad3 but reduced expression of the negative regulator Smad7 in the liver.Notably,concomitant administration of SGHXHYF to CCl_(4)-exposed rats was found to significantly reverse or abolish the pro-fibrotic effects of CCl_(4) in the liver and reduced serum transferase levels.Analysis of HSCs in vitro confirmed that,mechanistically,SGHXHYF inhibited activation of the TGF-β1/Smad signaling pathway by downregulating phosphorylated Smad2 and Smad3 and upregulating Smad7 levels.CONCLUSION:SGHXHYF ameliorated CCl_(4)-induced liver fibrosis by inhibiting the TGF-β1/Smad signaling pathway.These findings suggest that SGHXHYF may have clinical utility for the treatment or prevention of liver fibrosis.展开更多
基金Supported by the Natural Science Foundation of Jiangsu Province,China,No.BK2016157the National Natural Science Foundation of China,No.81673973+1 种基金Phase Ⅱ Project Funded by the Priority Academic Program Development of Jiangsu Higher Education Institutions,No.035062002003Developing Program for Highlevel Academic Talent in Jiangsu Hospital of TCM,No.y2018rc16
文摘AIM To explore the role and mechanism of total flavone of Abelmoschus manihot(TFA) on epithelial-mesenchymal transition(EMT) progress of Crohn's disease(CD) intestinal fibrosis.METHODS First,CCK-8 assay was performed to assess TFA on the viability of intestinal epithelial(IEC-6) cells and select the optimal concentrations of TFA for our further studies.Then cell morphology,wound healing and transwell assays were performed to examine the effect of TFA on morphology,migration and invasion of IEC-6 cells treated with TGF-β1.In addition,immunofluorescence,real-time PCR analysis(q RT-PCR) and western blotting assays were carried out to detect the impact of TFA on EMT progress.Moreover,western blotting assay was performed to evaluate the function of TFA on the Smad and MAPK signaling pathways.Further,the role of co-treatment of TFA and si-Smad or MAPK inhibitors has been examined by q RTPCR,western blotting,morphology,wound healing andtranswell assays.RESULTS In this study,TFA promoted transforming growth factor-β1(TGF-β1)-induced(IEC-6) morphological change,migration and invasion,and increased the expression of epithelial markers and reduced the levels of mesenchymal markers,along with the inactivation of Smad and MAPK signaling pathways.Moreover,we revealed that si-Smad and MAPK inhibitors effectively attenuated TGF-β1-induced EMT in IEC-6 cells.Importantly,co-treatment of TFA and si-Smad or MAPK inhibitors had better inhibitory effects on TGF-β1-induced EMT in IEC-6 cells than either one of them.CONCLUSION These findings could provide new insight into the molecular mechanisms of TFA on TGF-β1-induced EMT in IEC-6 cells and TFA is expected to advance as a new therapy to treat CD intestinal fibrosis.
基金Supported by the College Science and Technology Developing Foundation of Shanghai, No. 02BK14
文摘AIM: To investigate the effect of various concentrations of tetrandrine on activation of quiescent rat hepatic stellate cells (HSCs) and transforming growth factor-β (TGF-β) signaling in vitro.METHODS: HSCs were isolated from rats by in situperfusion of liver and 18% Nycodenz gradient centrifugation, and primarily cultured on uncoated plastic plates for 24 hwith DMEM containing 20% fetal bovine serum (FBS/DMEM) before the culture medium was substituted with 2% FBS/DMEM for another 24 h. Then, the HSCs were cultured in 2% FBS/DMEM with tetrandrine (0.25, 0.5, 1,2 mg/L, respectively). Cell morphological features were observed under an inverted microscope, smooth muscleα-actin (α-SMA) was detected by immunocytochemistry and image analysis system, laminin (LN) and type Ⅲprocollagen (PCⅢ) in supernatants were determined byradioimmunoassay. TGF-β1 mRNA, Smad 7 mRNA and Smad 7 protein were analyzed with RT-PCR and Western blotting, respectively.RESULTS: Tetrandrine at the concentrations of 0.25-2 mg/L prevented morphological transformation of HSC from the quiescent state to the activated one, while α-SMA, LN and PCⅢ expressions were inhibited. As estimated by gray values, the expression of α-SMA in tetrandrine groups (0.25, 0.5, 1, 2 mg/L) was reduced from 21.3% to 42.2%(control: 0.67, tetrandrine groups: 0.82, 0.85, 0.96, or 0.96, respectively, which were statistically different from the control, P<0.01), and the difference was more significant in tetrandrine at 1 and 2 mg/L. The content of LN in supernatants was significantly decreased in tetrandrine groups to 58.5%, 69.1%, 65.8% or 60.0% that of the control respectively, and that of PCⅢ to 84.6%, 81.5%,75.7% or 80.7% respectively (P<0.05 vs control), with no significant difference among tetrandrine groups. RTPCR showed that TGF-β1 mRNA expression was reduced by tetrandrine treatments from 56.56% to 87.90% in comparison with the control, while Smad 7 mRNA was increased 1.4-4.8 times. The TGF-β1 mRNA and Smad 7 mRNA expression was in a significant negative correlation (r= -0.755, P<0.01), and both were significantly correlated with α-SMA protein expression (r = -0.938, P<0.01;r = 0.938, P<0.01, respectively). The up-regulation of Smad 7 protein by tetrandrine (1 mg/L)was confirmed by Western blotting as well.CONCLUSION: Tetrandrine has a direct inhibiting effect on the activation of rat HSCs in culture. It up-regulates the expression of Smad 7 which in turn blocks TGF-β1 expression and signaling.
文摘According to data from 2020,Slovakia has long been among the top five countries with the highest incidence rate of colorectal cancer(CRC)worldwide,and the rate is continuing to rise every year.In approximately 80%of CRC cases,allelic loss(loss of heterozygosity,LOH)occurs in the long arm of chromosome 18q.The most important genes that can be silenced by 18q LOH or mutations are small mothers against decapentaplegic homolog(SMAD)2 and SMAD4,which are intracellular mediators of transforming growth factor(TGF)-βsuperfamily signals.TGF-βplays an important role in the pro-oncogenic processes,including such properties as invasion,epithelial-mesenchymal transition(commonly known as EMT),promotion of angiogenesis,and immunomodulatory effects.Several recent studies have reported that activation of TGF-βsignaling is related to drug resistance in CRC.Because the mechanisms of drug resistance are different between patients in different stages of CRC,personalized treatment is more effective.Therefore,knowledge of the activation and inhibition of factors that affect the TGF-βsignaling pathway is very important.
基金National Natural Science Foundation of China:a Metabolomic Study of the Effects of Moxibustion on Cardiac Function and Its Intervention in RA Model Rats Based on the TGF-β1/Smads Signaling Pathway (No.81403484)Anhui Province University Natural Science Fund Project of China:Exploring the Mechanism of Action of Moxibustion in AA Rats Based on Intestinal Flora and TLR4/NF-KB Signaling Pathway (No.KJ2019A0448)+2 种基金National Project Cultivation Fund Project Plan:Exploring the Mechanism of Action of Moxibustion in AA Rats Based on Intestinal Flora and TLR4/NF-KB Signaling Pathway (No.2019py01)Anhui Province Clinical Medical Research Center [Anhui Provincial Science and Technology Department Anhui Social Science (2020) No.41]the training Program of Outstanding talents in Colleges and Universities:2021 Domestic Visiting Training Program for Outstanding Young Key Teachers in Colleges and Universities (No.gxgnfx2021122)
文摘OBJECTIVE:To examine the effects of moxibustion on myocardial injury and myocardial metabolomics in rats with rheumatoid arthritis(RA)based on the transforming growth factor beta1(TGF-β1)/Smads signaling pathway.METHODS:One hundred rats were treated with saline[normal control(NC)group]or complete Freund’s adjuvant(CFA)by right plantar injection for the RA model group,and the latter were randomly divided into 4 groups.Tripterygium wilfordii polyglycoside tablets(雷公藤多苷片,TPT)have anti-inflammatory and are widely used in the clinical treatment of RA,therefore serving as a positive control group.Three days post injection rats were given TPT tablet(TPT group),acupuncture therapy(APT group),and moxibustion treatment(MOX group)for 15 consecutive days,while NC group and model group were equally grasped and fixed and received normal saline.Rat joint swelling scores and arthritis index(AI)were evaluated in each group before the CFA challenge,therapy and after receiving therapy.Myocardial ultrastructure was observed by electron microscope.Enzyme-linked immunosorbent assay was used to detect cardiac troponin I(cTnI)levels in rat myocardial tissue.Quantitative reverse transcription polymerase chain reaction and Western blotting analysis were used to measure the mRNA and protein levels of TGF-βsignaling molecules including TGF-β1,Smad2,Smad3,Smad4,and Smad7.Myocardial metabolomics was analyzed using gas chromatography-mass spectrometer.RESULTS:Compared with model group,RA model rats receiving TPT,acupuncture,or moxibustion therapy all showed reduced joint swelling scores and AI(all P<0.01)and improved myocardial damage,whereas rats treated with moxibustion were found to be more marked.Consistently,the expressions of cTnI,TGF-β1,Smad2,Smad3,and Smad4 were found to be elevated in model rat group in contrast to NC rats and were significantly downregulated in TPT,APT and MOX group when compared with model group,while the levels of Smad7 showed the opposite result(all P<0.01).Moreover,the dissection of metabolomics suggested a novel metabolite biomarker panel including D-Xylulose 5-phosphate,dihydroxyacetone phosphate,arachidonic acid,etc was defined and implicated in amino acid,glucose,and fatty acid metabolic processes as revealed by principal component analysis and partial least squares discriminant analysis.CONCLUSION:Moxibustion prevents RA-induced inflammatory response and offers potent therapeutic effects on myocardial dysfunctions.The protective effects might be associated with its role in TGF-β1 inactivation and metabolic reprogramming.
基金Supported by State Administration of Traditional Chinese Medicine Science and Technology Research Projects(Chinese pharmaceutical 2016ZX05)Effect of Wulong Xiaozheng Wan on Intermediate Gastric Carcinoma and its Mechanism
文摘OBJECTIVE:To evaluate the effect of Wulong Xiaozheng Wan medicated serum on the epithelial-mesenchymal transition (EMT) of BGC823 cell induced by transforming growth factor-β,(TGF-β,) and to explore its mechanism.METHODS:EMT model of BGC823 was stimulated by TGF-β1.Wulong Xiaozheng Wan medicated serum and LY-364947 were used as intervention.The proliferation and adhesion of BGC823 were detected by 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide and flow cytometry was used to detect the apoptosis.The invasion and migration were detected by Transwell.The level of matrix metalloproteins was detected by enzyme-linked immunosorbent assay.The expressions of related proteins and mRNA of EMT marker and TGF-β1/Smad signal pathway were detected by Western blot and reverse transcription-polymerase chain reaction.RESULTS:Compared with the TGF-β1 group,Wulong Xiaozheng Wan medicated serum could inhibit the ability of proliferation,heterogeneous adhesion,invasion,and migration.It also promotes apoptosis and homotypic adhesion in BGC823,with a dose-dependent manner.Meanwhile,Wulong Xiaozheng Wan medicated serum could regulate the expression of related proteins and mRNA of TGF-β1/Smad signaling pathway,and inhibit the expressions of EMT transcription factors and EMT markers.CONCLUSION:Wulong Xiaozheng Wan medicated serum inhibited epithelial-mesenchymal transition by down-regulated the expression of TβRI and the activation of TGF-β1/Smad signaling pathway.
基金Supported by the National Natural Science Foundation of China(Study on the Anti-Liver Fibrosis Mechanism of Shugan Huoxue Huayu Fang Based on miRNA-146a Regulating TGF-β/Smads Signaling Pathway No.81373538 and Role of Intracellular Endocytosis of Tight Junction Protein in Intestinal Leakage in Alcoholic Liver Disease No.81570536)。
文摘OBJECTIVE:To investigate the potential mechanism by which Shugan Huoxue Huayu Fang(疏肝活血化瘀方,SGHXHYF)ameliorates liver fibrosis.METHODS:Liver fibrosis was induced in rats by intraperitoneal injection of carbon tetrachloride(CCl_(4))in peanut oil solution(40%,3 m L/kg body weight)twice a week for 8 weeks.A normal control group received the same volume of peanut oil alone.During weeks 5-8,the CCl_(4)-injected rat groups were administered saline(vehicle control),colchicine(0.1 mg/mL,1 mg/kg,positive control),or SGHXHYF(0.1 mg/mL;0.3,0.6 and 1.2 mg/kg)once daily by oral gavage.Rats were sacrificed 24 h after the last treatment.Blood samples were collected for measurement of serum alanine aminotransferase(ALT),aspartate aminotransferase(AST),alkaline phosphatase(ALP),albumin(ALB),collagenⅠ,and collagenⅢlevels.Liver samples were analyzed by histopathological staining,Masson's staining of extracellular matrix proteins,and immune-ohistochemical staining ofα-smooth muscle actin(α-SMA).TGF-β1/Smad protein and mRNA levels were analyzed by Western blot and quantitative reverse transcription-polymerase chain reaction analysis,respectively.In vitro experiments were also performed using rat hepatic stellate cells(HSCs).RESULTS:Compared with the control animals,CCl_(4)-exposed rats exhibited elevated serum levels of ALT,AST,ALP,collagenⅠ,and collagenⅢ;reduced serum levels of ALB;and increased collagen deposition andα-SMA expression in liver sections,reflecting liver fibrosis.CCl_(4) also increased expression of TGF-β1 and the activated(phosphorylated)forms of Smad2 and Smad3 but reduced expression of the negative regulator Smad7 in the liver.Notably,concomitant administration of SGHXHYF to CCl_(4)-exposed rats was found to significantly reverse or abolish the pro-fibrotic effects of CCl_(4) in the liver and reduced serum transferase levels.Analysis of HSCs in vitro confirmed that,mechanistically,SGHXHYF inhibited activation of the TGF-β1/Smad signaling pathway by downregulating phosphorylated Smad2 and Smad3 and upregulating Smad7 levels.CONCLUSION:SGHXHYF ameliorated CCl_(4)-induced liver fibrosis by inhibiting the TGF-β1/Smad signaling pathway.These findings suggest that SGHXHYF may have clinical utility for the treatment or prevention of liver fibrosis.