RESUMO
BACKGROUND AND AIMS: Bacterially derived factors from the gut play a major role in the activation of inflammatory pathways in the liver and in the pathogenesis of alcoholic liver disease. The intestinal brush-border enzyme intestinal alkaline phosphatase (IAP) detoxifies a variety of bacterial pro-inflammatory factors and also functions to preserve gut barrier function. The aim of this study was to investigate whether oral IAP supplementation could protect against alcohol-induced liver disease. METHODS: Mice underwent acute binge or chronic ethanol exposure to induce alcoholic liver injury and steatosis ± IAP supplementation. Liver tissue was assessed for biochemical, inflammatory, and histopathological changes. An ex vivo co-culture system was used to examine the effects of alcohol and IAP treatment in regard to the activation of hepatic stellate cells and their role in the development of alcoholic liver disease. RESULTS: Pretreatment with IAP resulted in significantly lower serum alanine aminotransferase compared to the ethanol alone group in the acute binge model. IAP treatment attenuated the development of alcohol-induced fatty liver, lowered hepatic pro-inflammatory cytokine and serum LPS levels, and prevented alcohol-induced gut barrier dysfunction. Finally, IAP ameliorated the activation of hepatic stellate cells and prevented their lipogenic effect on hepatocytes. CONCLUSIONS: IAP treatment protected mice from alcohol-induced hepatotoxicity and steatosis. Oral IAP supplementation could represent a novel therapy to prevent alcoholic-related liver disease in humans.
Assuntos
Fosfatase Alcalina/administração & dosagem , Suplementos Nutricionais , Fígado Gorduroso Alcoólico/prevenção & controle , Alanina Transaminase/sangue , Animais , Técnicas de Cocultura , Citocinas/análise , Citocinas/sangue , Etanol , Fígado Gorduroso Alcoólico/sangue , Fígado Gorduroso Alcoólico/enzimologia , Feminino , Células Estreladas do Fígado/enzimologia , Hepatócitos/enzimologia , Intestinos/enzimologia , Lipogênese , Lipopolissacarídeos/sangue , Fígado/química , Camundongos , Camundongos Endogâmicos C57BL , Permeabilidade , Ativador de Plasminogênio Tecidual , Triglicerídeos/análiseRESUMO
The study evaluated the potential protective effect and underlying mechanism of Cucurbitacin E (CuE) in both thioacetamide-induced hepatic fibrosis and activated HSCs. CuE inhibited the proliferation of activated HSC/T-6 cells in a concentration- and time-dependent manner; triggered the activation of caspase-3, cleaved PARP, altered ratio of bcl-2-to-bax, and affected cytochrome C protein in a time- and concentration-dependent manner. CuE arrested activated HSCs at the G2/M phase. Furthermore, CuE reduced levels of p-Erk/MAPK and also inhibited the protein and mRNA expressions of α-SMA, TIMP-1 and collagen I in activated HSC-T6 cells. CuE inhibited PI3K and Akt phosphorylation, and reduced the levels of p-mTOR and p-P70S6K and increased the expression of p-AMPK, which is similar with AICAR and metformin. C57BL/6 mice were intraperitoneally injected with thioacetamide (TAA) for five continuous weeks (100 or 200mg/kg, three times per week) along with daily administration of CuE (5 or 10mg/kg/d) and curcumin (Cur, 20mg/kg). CuE treatments significantly reduced serum ALT/AST levels, α-SMA, TIMP-1, and collagen I protein expressions. HE, Masson trichrome, Sirius red and immunohistochemical staining also suggested that CuE could ameliorate hepatic fibrosis. Our findings suggest that CuE induces apoptosis of activated HSC and ameliorates TAA-induced hepatic fibrosis through activation of AMPK and blocking mTOR-dependent signaling pathway.
Assuntos
Proteínas Quinases Ativadas por AMP/metabolismo , Suplementos Nutricionais , Modelos Animais de Doenças , Cirrose Hepática/prevenção & controle , Transdução de Sinais , Serina-Treonina Quinases TOR/antagonistas & inibidores , Triterpenos/uso terapêutico , Proteínas Quinases Ativadas por AMP/química , Animais , Antioxidantes/administração & dosagem , Antioxidantes/uso terapêutico , Linhagem Celular Tumoral , Proliferação de Células , Ativação Enzimática , Fase G2 , Regulação da Expressão Gênica , Células Estreladas do Fígado/citologia , Células Estreladas do Fígado/enzimologia , Células Estreladas do Fígado/metabolismo , Cirrose Hepática/metabolismo , Masculino , Camundongos Endogâmicos C57BL , Distribuição Aleatória , Organismos Livres de Patógenos Específicos , Serina-Treonina Quinases TOR/metabolismo , Tioacetamida , Triterpenos/administração & dosagemRESUMO
ETHNOPHARMACOLOGICAL RELEVANCE: Huangqi Decoction (HQD), a classical traditional Chinese medicine (TCM) formula, is used to treating liver injury in China. The aim of the study is to investigate mechanisms of HQD against dimethylnitrosamine (DMN)-induced liver fibrosis underlying metabolic profiles of bile acids. MATERIALS AND METHODS: DMN-induced liver fibrosis rats were administrated HQD and its compounds, astragalosides (AS), glycyrrhizic acid (GA) and their combination. The anti-fibrosis effects were evaluated and targeted metabolomics by UPLC-MS was used to examine whether HQD had an influence on bile acid metabolism. The levels of mRNAs associated with bile acid metabolism were expressed by RT-PCR. Chenodeoxycholic acid (CDCA)-induced hepatic stellate cells (HSCs) proliferation and activation were examined using MTS assay and Western blot. RESULTS: Histopathological changes and serum liver function in HQD group had significant improvements (P<0.01). Concentrations of free bile acids and taurine conjugates were significantly increased in DMN group (P<0.05). HQD and its compounds restored the increased bile acids to normal levels, and HQD was more effected on parts of bile acids. Furthermore, the levels of mRNAs related bile acid synthesis and reabsorption such as CYP7A1, CYP8B1, CYP27A1, OATP2, OATP3, OATP4 and NTCP were significantly down-regulated in DMN group (P<0.05), mRNAs related excretion such as MRP3 and BESP were up-regulated (P<0.01), and CYP7A1, CYP8B1, OATP3, OATP4, NTCP and MRP3 restored to normal levels by HQD treatment. Moreover, CDCA-induced HSCs proliferation and activation were weaken by HQD (P<0.05) with down-regulated α-SMA, TGF-ß1, p-Smad2 and p-Smad3 expressions. CONCLUSIONS: HQD alleviated DMN-induced liver fibrosis with a better effect than its compounds, which may be involved in the regulation of bile acid metabolism enzyme. Moreover, HQD may inhibit CDCA-induced HSCs proliferation and activation.
Assuntos
Ácidos e Sais Biliares/sangue , Doença Hepática Induzida por Substâncias e Drogas/prevenção & controle , Dimetilnitrosamina , Medicamentos de Ervas Chinesas/farmacologia , Cirrose Hepática Experimental/prevenção & controle , Fígado/efeitos dos fármacos , Substâncias Protetoras/farmacologia , Animais , Astragalus propinquus , Biomarcadores/sangue , Proliferação de Células/efeitos dos fármacos , Doença Hepática Induzida por Substâncias e Drogas/enzimologia , Doença Hepática Induzida por Substâncias e Drogas/genética , Doença Hepática Induzida por Substâncias e Drogas/patologia , Cromatografia Líquida de Alta Pressão , Regulação Enzimológica da Expressão Gênica , Células Estreladas do Fígado/efeitos dos fármacos , Células Estreladas do Fígado/enzimologia , Células Estreladas do Fígado/patologia , Hidroxiprolina/metabolismo , Fígado/enzimologia , Fígado/patologia , Cirrose Hepática Experimental/induzido quimicamente , Cirrose Hepática Experimental/enzimologia , Cirrose Hepática Experimental/genética , Cirrose Hepática Experimental/patologia , Masculino , Espectrometria de Massas , Metabolômica/métodos , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Ratos WistarRESUMO
Plant-based whole foods provide thousands of bioactive metabolites to the human diet that reduce the risk of developing chronic diseases. ß-Caryophyllene (CAR) is a common constituent of the essential oil of numerous plants, vegetables, fruits and medicinal herbs, and has been used as a flavouring agent since the 1930 s. Here, we report the antioxidant activity of CAR, its protective effect on liver fibrosis and its inhibitory capacity on hepatic stellate cell (HSC) activation. CAR was tested for the inhibition of lipid peroxidation and as a free radical scavenger. CAR had higher inhibitory capacity on lipid peroxidation than probucol, α-humulene and α-tocopherol. Also, CAR showed high scavenging activities against hydroxyl radical and superoxide anion. The activity of 5-lipoxygenase, an enzyme that actively participates in fibrogenesis, was significantly inhibited by CAR. Carbon tetrachloride-treated rats received CAR at 2, 20 and 200 mg/kg. CAR significantly improved liver structure, and reduced fibrosis and the expression of Col1a1, Tgfb1 and Timp1 genes. Oxidative stress was used to establish a model of HSC activation with overproduction of extracellular matrix proteins. CAR (1 and 10 µm) increased cell viability and significantly reduced the expression of fibrotic marker genes. CAR, a sesquiterpene present in numerous plants and foods, is as a natural antioxidant that reduces carbon tetrachloride-mediated liver fibrosis and inhibits hepatic cell activation.
Assuntos
Antioxidantes/uso terapêutico , Intoxicação por Tetracloreto de Carbono/prevenção & controle , Suplementos Nutricionais , Células Estreladas do Fígado/metabolismo , Fígado/metabolismo , Sesquiterpenos/uso terapêutico , Animais , Antioxidantes/administração & dosagem , Antioxidantes/metabolismo , Antioxidantes/farmacologia , Araquidonato 5-Lipoxigenase/química , Araquidonato 5-Lipoxigenase/metabolismo , Intoxicação por Tetracloreto de Carbono/metabolismo , Intoxicação por Tetracloreto de Carbono/patologia , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Feminino , Aromatizantes/administração & dosagem , Aromatizantes/metabolismo , Aromatizantes/uso terapêutico , Sequestradores de Radicais Livres/administração & dosagem , Sequestradores de Radicais Livres/metabolismo , Sequestradores de Radicais Livres/uso terapêutico , Células Estreladas do Fígado/efeitos dos fármacos , Células Estreladas do Fígado/enzimologia , Peroxidação de Lipídeos/efeitos dos fármacos , Inibidores de Lipoxigenase/administração & dosagem , Inibidores de Lipoxigenase/metabolismo , Inibidores de Lipoxigenase/uso terapêutico , Fígado/efeitos dos fármacos , Fígado/patologia , Sesquiterpenos Monocíclicos , Estresse Oxidativo/efeitos dos fármacos , Sesquiterpenos Policíclicos , Probucol/farmacologia , Probucol/uso terapêutico , Distribuição Aleatória , Ratos , Ratos Wistar , Sesquiterpenos/administração & dosagem , Sesquiterpenos/metabolismo , alfa-Tocoferol/metabolismo , alfa-Tocoferol/uso terapêuticoRESUMO
Diabetes is featured by elevated levels of blood glucose, i.e. hyperglycemia, which might be a risk factor for hepatic fibrogenesis in patients with non-alcoholic steatohepatitis. Hepatic stellate cells (HSCs) are the major effectors during hepatic fibrogenesis. This study was designed to evaluate impacts of high levels of glucose on HSC activation, assess roles of the phytochemical curcumin in attenuating the glucose impacts, and elucidate underlying mechanisms. In this report, levels of intracellular glucose were measured. Contents and gene expression of glucose transporter-2 (GLUT2) in cell fractions were examined. Levels of cellular glutathione and oxidative stress were analyzed. We observed that high levels of glucose induced cell proliferation, type I collagen production and expression of genes relevant to HSC activation, and elevated intracellular glucose levels in cultured HSCs. Curcumin eliminated the stimulatory impacts. Curcumin abrogated the membrane translocation of GLUT2 by interrupting the p38 MAPK signaling pathway. In addition, curcumin suppressed glut2 expression by stimulating the activity of peroxisome proliferator-activated receptor-gamma (PPARγ) and de novo synthesis of glutathione. In conclusion, hyperglycemia stimulated HSC activation in vitro by increasing intracellular glucose, which was eliminated by curcumin by blocking the membrane translocation of GLUT2 and suppressing glut2 expression. The latter was mediated by activating PPARγ and attenuating oxidative stress. Our results presented evidence to impacts of hyperglycemia on stimulating HSC activation and hepatic fibrogenesis, and provided novel insights into the mechanisms by which curcumin eliminated the hyperglycemia-caused HSC activation and potential therapeutic strategies for treatment of diabetes-associated hepatic fibrogenesis.
Assuntos
Membrana Celular/metabolismo , Curcumina/farmacologia , Curcumina/uso terapêutico , Regulação da Expressão Gênica/efeitos dos fármacos , Transportador de Glucose Tipo 2/genética , Células Estreladas do Fígado/metabolismo , Hiperglicemia/tratamento farmacológico , Animais , Membrana Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Relação Dose-Resposta a Droga , Glucose/metabolismo , Transportador de Glucose Tipo 2/metabolismo , Glutamato-Cisteína Ligase/metabolismo , Glutationa/metabolismo , Células Estreladas do Fígado/efeitos dos fármacos , Células Estreladas do Fígado/enzimologia , Células Estreladas do Fígado/patologia , Hiperglicemia/genética , Hiperglicemia/patologia , Espaço Intracelular/efeitos dos fármacos , Espaço Intracelular/metabolismo , Peroxidação de Lipídeos/efeitos dos fármacos , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Masculino , Estresse Oxidativo/efeitos dos fármacos , PPAR gama/metabolismo , Subunidades Proteicas/metabolismo , Transporte Proteico/efeitos dos fármacos , Ratos , Ratos Sprague-Dawley , Espécies Reativas de Oxigênio/metabolismo , Fatores de Tempo , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismoRESUMO
OBJECTIVE: To investigate the effects of Shaoqiduogan (SQDG) on the expression of matrix metalloproteinase 13 (MMP-13) and tissue inhibitor of metalloproteinase 1 (TIMP-1) in carbon tetrachloride (CCl4) induced hepatic fibrosis rats and transforming growth factor beta1 (TGF-beta1) irritated hepatic stellate cells (HSC), and to explore its possible mechanisms. METHOD: The model of chemical hepatic fibrosis induced by CCl4 was prepared. The rats were randomly divided into 5 groups, including normal control group, liver fibrosis model group and SQDG (42. 5, 85, 170 mg x kg(-1)) treated groups. The level of collagen type 1 (C-1) in serum was determined by radioimmunoassay. Masson stain was used to examine the histopathological change. MMP-13 and TIMP-1 ex-pression in liver tissues were assayed by immunohistochemistry. In vitro, effects of SQDG on the expression of MMP-13, TIMP-1 and C-1 in HSC-T6 stimulated by TGF-beta were measured by Western-blot. RESULT: The results showed that SQDG significantly decreased the elevated level of C-1 in serum of hepatic fibrosis rats induced by CCl4. Pathological examination showed that SQDG could remarkably alleviate the degree of liver fibrogenesis and formation of pseudolobulus. The results of immunohistochemistry demonstrated that SQDG significantly increased MMP-13 expression and decreased TIMP-1 expression in liver tissues. Furthermore, SQDG (20-160 mg x L(-1)) could facilitate MMP-13 expression, inhibit TIMP-1 expression and significantly inhibit the C-I production of HSC stimulated with TGF-beta1 in vitro. CONCLUSION: The anti-fibrotic effects of SQDG may be associated with its action of promoting collagen degradation via controlling the levels of MMP-13 and TIMP-1 in liver.
Assuntos
Medicamentos de Ervas Chinesas/administração & dosagem , Expressão Gênica/efeitos dos fármacos , Células Estreladas do Fígado/metabolismo , Cirrose Hepática/tratamento farmacológico , Cirrose Hepática/genética , Metaloproteinase 13 da Matriz/genética , Inibidor Tecidual de Metaloproteinase-1/genética , Animais , Modelos Animais de Doenças , Células Estreladas do Fígado/efeitos dos fármacos , Células Estreladas do Fígado/enzimologia , Humanos , Cirrose Hepática/enzimologia , Cirrose Hepática/metabolismo , Masculino , Metaloproteinase 13 da Matriz/metabolismo , Ratos , Ratos Sprague-Dawley , Inibidor Tecidual de Metaloproteinase-1/metabolismoRESUMO
BACKGROUND AND AIMS: In chronic liver injury, hepatic stellate cells (HSCs) acquire an activated phenotype, migrate to the injured region in response to chemotactic factors, and produce extracellular matrix proteins including collagen. In this study, we investigated the effects of rhubarb (Rheum palmatum L.) on transforming growth factor (TGF)-beta1-induced expressions of alpha-smooth-muscle actin (SMA) and collagen, and the migration of HSCs. METHODS: HSC-T6, a cell line of rat HSCs, was used in the in vitro experiments. An enzyme-linked immunosorbent assay and Sircol red assay were used to detect the expressions of alpha-SMA and collagen, respectively. HSC-T6 migration was assayed with a transwell apparatus. Phosphorylations of Smad2/3 and mitogen-activated protein kinases (MAPKs), including extracellular signal-regulated kinase (ERK) 1/2, p38, and c-jun N-terminal kinase (JNK), were analyzed with Western blotting. Matrix metalloproteinase (MMP)-2 activity was examined by gelatin zymography. RESULTS: The results revealed that a rhubarb extract concentration-dependently attenuated TGF-beta1-induced alpha-SMA and collagen expressions and migration of HSCs. The inhibitory effect of rhubarb was associated with (i) down-regulation of the phosphorylation of Smad2/3 and JNK, and (ii) attenuation of MMP-2 activity. Within the working concentrations used, the rhubarb extract did not affect cell viability of HSCs. CONCLUSION: The results suggest that rhubarb attenuated TGF-beta1-mediated migration of HSCs possibly by interfering with Smad2/3 phosphorylation, the MAPK pathway, and MMP-2 activity.