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1.
Life Sci ; 251: 117595, 2020 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-32240681

RESUMO

AIMS: The activation of hepatic stellate cells (HSCs) plays a central role in liver fibrosis progression. Phospholipase D (PLD) enzymes participate in multiple cellular activities. However, whether and how PLD regulates HSCs activation remain elusive. MAIN METHODS: The expression of intrahepatic PLD1 and PLD2 was determined in CCl4-induced mouse liver fibrosis models by western blot and immunohistochemistry. Cell model of liver fibrogenesis was constructed using rat HSCs line (HSC-T6) treated with recombinant transforming growth factor ß1 (TGFß1). Fibrogenesis was evaluated on the aspects of proliferation, expression of pro-fibrogenic markers and migration. The effects mediated by PLD1-mTOR axis on TGFß1-induced fibrogenesis were evaluated using HSC-T6 treated with small-molecular PLD1 inhibitors, PLD1-SiRNA, rapamycin (mTOR inhibitor) and MHY1485 (mTOR activator). KEY FINDINGS: Significant increase of PLD1, not PLD2 was documented in CCl4-induced cirrhotic compared to normal liver tissues. Suppression of PLD1 activities by PLD inhibitors or down-regulation of PLD1 expression in HSC-T6 could significantly restrain TGFß1-induced fibrogenesis, as reflected by decreased cell proliferation and reduced expression of pro-fibrogenic markers. Besides, either PLD1 inhibitor or PLD1-SiRNA significantly inhibited mTOR activity of HSC-T6. Moreover, PLD1 inhibitors not only exhibited similar effects with rapamycin in TGFß1-induced fibrogenesis, but also blunted MHY1485 enhanced cell proliferation of HSC-T6. SIGNIFICANCE: The PLD1-mTOR axis of HSCs could be therapeutically targeted in advanced liver fibrosis.


Assuntos
Células Estreladas do Fígado/metabolismo , Cirrose Hepática/fisiopatologia , Fosfolipase D/metabolismo , Serina-Treonina Quinases TOR/metabolismo , Animais , Linhagem Celular , Proliferação de Células/fisiologia , Modelos Animais de Doenças , Progressão da Doença , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Ratos
2.
PLoS One ; 15(2): e0228729, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32053633

RESUMO

BACKGROUND: There is a correlation between the endocannabinoid system and hepatic fibrosis based on the activation of CB1 and CB2 receptors; where CB1 has profibrogenic effects. Gene therapy with a plasmid carrying a shRNA for CB1 delivered by hydrodynamic injection has the advantage of hepatic tropism, avoiding possible undesirable effects of CB1 pharmacological inhibition. OBJECTIVE: To evaluate hydrodynamics-based liver transfection in an experimental model of liver cirrhosis of a plasmid with the sequence of a shRNA for CB1 and its antifibrogenic effects. METHODS: Three shRNA (21pb) were designed for blocking CB1 mRNA at positions 877, 1232 and 1501 (pshCB1-A, B, C). Sequences were cloned in the pENTR™/U6. Safety was evaluated monitoring CB1 expression in brain tissue. The silencing effect was determined in rat HSC primary culture and CCl4 cirrhosis model. Hydrodynamic injection in cirrhotic liver was through iliac vein and with a dose of 3mg/kg plasmid. Serum levels of liver enzymes, mRNA levels of TGF-ß1, Col IA1 and α-SMA and the percentage of fibrotic tissue were analyzed. RESULTS: Hydrodynamic injection allows efficient CB1 silencing in cirrhotic livers and pshCB1-B (position 1232) demonstrated the main CB1-silencing. Using this plasmid, mRNA level of fibrogenic molecules and fibrotic tissue considerably decrease in cirrhotic animals. Brain expression of CB1 remained unaltered. CONCLUSION: Hydrodynamics allows a hepatotropic and secure transfection in cirrhotic animals. The sequence of the shCB1-B carried in a plasmid or any other vector has the potential to be used as therapeutic strategy for liver fibrosis.


Assuntos
Inativação Gênica , Hidrodinâmica , Cirrose Hepática/patologia , RNA Interferente Pequeno/metabolismo , Receptor CB1 de Canabinoide/metabolismo , Actinas/genética , Actinas/metabolismo , Alanina Transaminase/sangue , Alanina Transaminase/metabolismo , Animais , Aspartato Aminotransferases/sangue , Aspartato Aminotransferases/metabolismo , Encéfalo/metabolismo , Células Cultivadas , Modelos Animais de Doenças , Células Estreladas do Fígado/citologia , Células Estreladas do Fígado/metabolismo , Fígado/metabolismo , Masculino , Plasmídeos/metabolismo , RNA Interferente Pequeno/administração & dosagem , Ratos , Ratos Wistar , Receptor CB1 de Canabinoide/antagonistas & inibidores , Receptor CB1 de Canabinoide/genética , Transfecção , Fator de Crescimento Transformador beta1/genética , Fator de Crescimento Transformador beta1/metabolismo
3.
Anticancer Res ; 40(2): 743-749, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-32014916

RESUMO

BACKGROUND/AIM: The hepatic stellate cells (HSCs) have relationship to cancer progression. The aim of this study is to investigate the effect of HSCs and the role of IL-6/Stat3 pathway on hepatocellular carcinoma (HCC) progression. MATERIALS AND METHODS: HCCs were co-cultured with HSCs. The viability and migration ability of cancer cells were detected. Epithelial-mesenchymal transition (EMT) marker (E-cadherin), stem cell marker (CD44) and p-signal transducer and activator of transcription 3 (p-STAT3) of cancer cells were evaluated. Finally, interleukin-6 (IL-6) neutralization was performed. RESULTS: Co-culture of HCCs with HSCs increased cancer cell viability and migration ability. EMT and stemness of cancer cells increased with HSCs. Following IL-6 neutralization, phospho-STAT3 activation, cancer cell viability and migration, as well as EMT, and stemness of cancer cells decreased. CONCLUSION: HSCs promoted HCC progression through the IL-6/STAT3 pathway.


Assuntos
Carcinoma Hepatocelular/metabolismo , Células Estreladas do Fígado/metabolismo , Interleucina-6/metabolismo , Neoplasias Hepáticas/metabolismo , Carcinoma Hepatocelular/patologia , Comunicação Celular/fisiologia , Linhagem Celular Tumoral , Movimento Celular/fisiologia , Sobrevivência Celular/fisiologia , Técnicas de Cocultura , Células Hep G2 , Células Estreladas do Fígado/patologia , Humanos , Neoplasias Hepáticas/patologia , Células-Tronco Neoplásicas/metabolismo , Células-Tronco Neoplásicas/patologia , Fator de Transcrição STAT3/metabolismo , Transdução de Sinais
4.
Mol Cell Biochem ; 465(1-2): 115-123, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31893334

RESUMO

Increasing studies have indicated that hypoxia serves as a pivotal microenvironmental factor that facilitates activation of hepatic stellate cells (HSCs). However, the mechanism by which hypoxia activates HSCs is not clear. Here, we demonstrated that plasmacytoma variant translocation 1 (PVT1) and autophagy were overexpressed in liver fibrotic specimens. In primary mouse HSCs, both PVT1 and autophagy were induced by hypoxia. Further study showed that hypoxia-induced autophagy depended on expression of PVT1 and miR-152 in HSCs. Luciferase reporter assay indicated that autophagy-related gene 14 (ATG14) was a direct target of miR-152. In addition, inhibition of autophagy by 3-methyladenine and Beclin-1 siRNA impeded activation of HSCs cultured in 1% O2. Taken together, autophagy induction via the PVT1-miR-152-ATG14 signaling pathway contributes to activation of HSCs under hypoxia condition.


Assuntos
Morte Celular Autofágica , Proteínas Relacionadas à Autofagia/metabolismo , Células Estreladas do Fígado/metabolismo , MicroRNAs/metabolismo , RNA Longo não Codificante/metabolismo , Transdução de Sinais , Proteínas de Transporte Vesicular/metabolismo , Animais , Hipóxia Celular , Células Estreladas do Fígado/patologia , Cirrose Hepática/metabolismo , Cirrose Hepática/patologia , Camundongos
5.
Am J Pathol ; 190(3): 586-601, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-31953035

RESUMO

Galanin (Gal) is a peptide with a role in neuroendocrine regulation of the liver. In this study, we assessed the role of Gal and its receptors, Gal receptor 1 (GalR1) and Gal receptor 2 (GalR2), in cholangiocyte proliferation and liver fibrosis in multidrug resistance protein 2 knockout (Mdr2KO) mice as a model of chronic hepatic cholestasis. The distribution of Gal, GalR1, and GalR2 in specific liver cell types was assessed by laser-capture microdissection and confocal microscopy. Galanin immunoreactivity was detected in cholangiocytes, hepatic stellate cells (HSCs), and hepatocytes. Cholangiocytes expressed GalR1, whereas HSCs and hepatocytes expressed GalR2. Strategies were used to either stimulate or block GalR1 and GalR2 in FVB/N (wild-type) and Mdr2KO mice and measure biliary hyperplasia and hepatic fibrosis by quantitative PCR and immunostaining of specific markers. Galanin treatment increased cholangiocyte proliferation and fibrogenesis in both FVB/N and Mdr2KO mice. Suppression of GalR1, GalR2, or both receptors in Mdr2KO mice resulted in reduced bile duct mass and hepatic fibrosis. In vitro knockdown of GalR1 in cholangiocytes reduced α-smooth muscle actin expression in LX-2 cells treated with cholangiocyte-conditioned media. A GalR2 antagonist inhibited HSC activation when Gal was administered directly to LX-2 cells, but not via cholangiocyte-conditioned media. These data demonstrate that Gal contributes not only to cholangiocyte proliferation but also to liver fibrogenesis via the coordinate activation of GalR1 in cholangiocytes and GalR2 in HSCs.


Assuntos
Subfamília B de Transportador de Cassetes de Ligação de ATP/genética , Colestase/metabolismo , Galanina/metabolismo , Cirrose Hepática/metabolismo , Receptor Tipo 1 de Galanina/metabolismo , Receptor Tipo 2 de Galanina/metabolismo , Animais , Ductos Biliares/metabolismo , Proliferação de Células , Colestase/patologia , Modelos Animais de Doenças , Células Epiteliais/metabolismo , Feminino , Galanina/genética , Células Estreladas do Fígado/metabolismo , Células Estreladas do Fígado/patologia , Fígado/metabolismo , Fígado/patologia , Cirrose Hepática/patologia , Camundongos , Camundongos Knockout , Receptor Tipo 1 de Galanina/genética , Receptor Tipo 2 de Galanina/genética
6.
Cancer Sci ; 111(2): 406-417, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31785057

RESUMO

STMN1 has been regarded as an oncogene and its upregulation is closely associated with malignant behavior and poor prognosis in multiple cancers. However, the detailed functions and underlying mechanisms of STMN1 are still largely unknown in hepatocellular carcinoma (HCC) development. Herein, we analyzed STMN1 expression and the related clinical significance in HCC by using well-established Protein Atlas, The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO) cancer databases. Analysis indicated that STMN1 was highly expressed in HCC and closely associated with vascular invasion, higher histological grade, advanced clinical grade and shorter survival time in HCC patients. Overexpressing and silencing STMN1 in HCC cell lines showed that STMN1 could regulate cell proliferation, migration, drug resistance, cancer stem cell properties in vitro as well as tumor growth in vivo. Further experiments showed that STMN1 mediated intricate crosstalk between HCC and hepatic stellate cells (HSC) by triggering the hepatocyte growth factor (HGF)/MET signal pathway. When HSC were cocultured with HCC cells, HSC secreted more HGF to stimulate the expression of STMN1 in HCC cells. Mutually, STMN1 upregulation in HCC cells facilitated HSC activation to acquire cancer-associated fibroblast (CAF) features. The MET inhibitor crizotinib significantly blocked this crosstalk and slowed tumor growth in vivo. In conclusion, our findings shed new insight on STMN1 function, and suggest that STMN1 may be used as a potential marker to identify patients who may benefit from MET inhibitor treatment.


Assuntos
Carcinoma Hepatocelular/patologia , Células Estreladas do Fígado/citologia , Neoplasias Hepáticas/patologia , Transdução de Sinais , Estatmina/genética , Animais , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/metabolismo , Linhagem Celular Tumoral , Técnicas de Cocultura , Resistencia a Medicamentos Antineoplásicos , Retroalimentação Fisiológica , Regulação Neoplásica da Expressão Gênica , Células Estreladas do Fígado/metabolismo , Fator de Crescimento de Hepatócito/metabolismo , Humanos , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/metabolismo , Masculino , Camundongos , Gradação de Tumores , Transplante de Neoplasias , Células-Tronco Neoplásicas/metabolismo , Proteínas Proto-Oncogênicas c-met/metabolismo , Regulação para Cima
7.
Toxicol Lett ; 320: 1-8, 2020 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-31756458

RESUMO

With the spread of hexavalent chromium [Cr(VI)] contamination, risk of exposure in non-occupational populations is increasing. The liver is the main target organ for Cr(VI) accumulation; however, the effect of long-term Cr(VI) exposure on liver toxicity is largely unknown. In this study, we investigated the effect of chronic Cr(VI) exposure on liver fibrosis and its possible mechanism. Mice were injected with Cr(VI) for two months, and our results showed Cr(VI) treatment caused liver toxicity characterized by liver structure disorganization, liver dysfunction, and antioxidant enzyme system inhibition. The development of liver fibrosis was also found via the emergence of collagen fibril deposition, increased expression of extracellular matrix-related genes, activation of hepatic stellate cells (HSCs) and increase the expression levels of Hedgehog (Hh) signaling pathway-related molecules. To demonstrate the role of Hh signaling in the regulation of Cr(VI)-induced liver fibrosis, genetically modified mice with heterozygous deficiency of Shh (Shh+/-) were used. In the Shh+/- mice, Hh signaling, HSCs activation and liver fibrosis development were all ameliorated. In conclusion, we demonstrated that Cr(VI)-induced liver fibrosis development resulted from Hh pathway-mediated HSCs activation. Our findings strongly suggest that inhibition of Hh pathway may help in the development of new strategies for Cr(VI)-associated liver fibrosis.


Assuntos
Doença Hepática Induzida por Substâncias e Drogas/metabolismo , Cromo , Proteínas Hedgehog/metabolismo , Células Estreladas do Fígado/metabolismo , Cirrose Hepática Experimental/metabolismo , Fígado/metabolismo , Dicromato de Potássio , Transdução de Sinais , Animais , Doença Hepática Induzida por Substâncias e Drogas/genética , Doença Hepática Induzida por Substâncias e Drogas/patologia , Modelos Animais de Doenças , Proteínas Hedgehog/deficiência , Proteínas Hedgehog/genética , Células Estreladas do Fígado/patologia , Fígado/patologia , Cirrose Hepática Experimental/induzido quimicamente , Cirrose Hepática Experimental/genética , Cirrose Hepática Experimental/patologia , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout
8.
Food Chem Toxicol ; 135: 111044, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31830547

RESUMO

Hemistepsin A (HsA), isolated from Hemistepta lyrata (Bunge) Bunge, has the ability to ameliorate hepatitis in mice. However, the effects of H. lyrata and HsA on other types of liver disease have not been explored. In this report, we investigated the effects of H. lyrata and HsA on liver fibrosis and the underlying molecular mechanisms in activated hepatic stellate cells (HSCs). Based on cell viability-guided isolation, we found HsA was the major natural product responsible for H. lyrata-mediated cytotoxicity in LX-2 cells. HsA significantly decreased the viability of LX-2 cells and primary activated HSCs, increased the binding of Annexin V, and altered the expression of apoptosis-related proteins, suggesting that HsA induces apoptosis in activated HSCs. HsA reduced the phosphorylation of IKKε and the transactivation of nuclear factor-κB (NF-κB). Moreover, HsA decreased the phosphorylation of Akt and its downstream signaling molecules. Transfection experiments suggested that inhibition of NF-κB or Akt is essential for HsA-induced apoptosis of HSCs. In a CCl4-induced liver fibrosis model, HsA administration significantly decreased ALT and AST activities. Furthermore, HsA attenuated CCl4-mediated collagen deposits and profibrogenic genes expression in hepatic tissue. Thus, HsA may serve as a natural product for managing liver fibrosis through inhibition of NF-κB/Akt-dependent signaling.


Assuntos
Apoptose/efeitos dos fármacos , Células Estreladas do Fígado/efeitos dos fármacos , Lactonas/farmacologia , Cirrose Hepática/prevenção & controle , Sesquiterpenos/farmacologia , Animais , Linhagem Celular Transformada , Clorofórmio/farmacologia , Células Estreladas do Fígado/metabolismo , Humanos , Cirrose Hepática/metabolismo , Cirrose Hepática/patologia , Camundongos , NF-kappa B/metabolismo , Extratos Vegetais/farmacologia , Proteínas Proto-Oncogênicas c-akt/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de Sinais/efeitos dos fármacos
9.
Chemosphere ; 242: 124959, 2020 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-31669990

RESUMO

Long-term exposure to arsenic can cause liver injury and fibrosis. The activation of hepatic stellate cells (HSCs) plays an essential role in the process of liver fibrosis. We found that NaAsO2 caused liver damage and fibrosis in vivo, accompanied by excessive collagen deposition and HSCs activation. In addition, NaAsO2 upregulated autophagy flux, elevated the level of cytoplasmic cathepsin B (CTSB), and activated the NOD-like receptors containing pyrin domain 3 (NLRP3) inflammasome in a subtle way. Consistent with these findings in vivo, we demonstrated that NaAsO2-induced activation of HSCs depended on CTSB-mediated NLRP3 inflammasome activation in HSC-t6 cells and rats primary HSCs. Moreover, inhibition of autophagy decreased the cytoplasmic CTSB and alleviated the activation of the NLRP3 inflammasome, thereby attenuating the NaAsO2-induced HSCs activation. In summary, these results indicated that NaAsO2 induced HSCs activation via autophagic-CTSB-NLRP3 inflammasome pathway. These findings may provide a novel insight into the potential mechanism of NaAsO2-induced liver fibrosis.


Assuntos
Arsênico/toxicidade , Autofagia , Catepsina B/metabolismo , Células Estreladas do Fígado/metabolismo , Inflamassomos/fisiologia , Cirrose Hepática/induzido quimicamente , Animais , Arsênico/metabolismo , Inflamassomos/metabolismo , Fígado/metabolismo , Fígado/patologia , Cirrose Hepática/patologia , Masculino , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Ratos
10.
Acta Biochim Biophys Sin (Shanghai) ; 52(1): 18-25, 2020 Jan 02.
Artigo em Inglês | MEDLINE | ID: mdl-31828297

RESUMO

As a highly malignant tumor, hepatocellular carcinoma (HCC) is the third leading cause of cancer-related deaths worldwide. In most HCC patients, the development of HCC begins with hepatitis, which is followed by fibrosis and cirrhosis before progressing to HCC. Cancer-associated fibroblasts (CAFs), which are generally believed to be derived from activated hepatic stellate cells (HSCs), are highly involved in the development of HCC through the secretion of cytokines and angiogenic factors. The results of our study showed that a considerable number of CAFs highly expressed CD90 and were enriched in HCC tissues. Bioinformatics analysis of the transcriptome of HCC tissues revealed that placental growth factor (PlGF) is significantly correlated with CD90 expression. The isolated primary CAFs and activated HSCs overexpressed PlGF and CD90. In addition, the results of gene expression profiling interactive analysis based on The Cancer Genome Atlas showed that high levels of both PlGF and CD90 are correlated with tumor angiogenesis markers (CD31, CD34, and CD105) and predict poor HCC patient prognosis. In summary, our results suggest that CAFs can generate PlGF and may provide an effective target for CAFs-regulated neoangiogenesis in HCC.


Assuntos
Carcinoma Hepatocelular/irrigação sanguínea , Fibroblastos/metabolismo , Neoplasias Hepáticas/irrigação sanguínea , Neovascularização Patológica/metabolismo , Fator de Crescimento Placentário/metabolismo , Actinas/metabolismo , Antígenos CD34/metabolismo , Biomarcadores Tumorais/metabolismo , Carcinoma Hepatocelular/genética , Linhagem Celular Tumoral , Endoglina/metabolismo , Regulação Neoplásica da Expressão Gênica , Células Estreladas do Fígado/metabolismo , Humanos , Neoplasias Hepáticas/genética , Fator de Crescimento Placentário/genética , Molécula-1 de Adesão Celular Endotelial a Plaquetas/metabolismo , Prognóstico , Antígenos Thy-1/genética , Antígenos Thy-1/metabolismo , Transcriptoma/genética
11.
Chem Biol Interact ; 316: 108917, 2020 Jan 25.
Artigo em Inglês | MEDLINE | ID: mdl-31838050

RESUMO

Stearoyl-CoA desaturase (SCD) generates monounsaturated fatty acids (MUFAs) which contribute to cell growth, survival, differentiation, metabolic regulation and signal transduction. Overexpression of SCD is evident and implicated in metabolic diseases such as diabetes and non-alcoholic fatty liver disease. SCD also stimulates canonical Wnt pathway and YAP activation in support of stemness and tumorigenesis. SCD facilitates metabolic reprogramming in cancer which is mediated, at least in part, by regulation of AKT, AMPK, and NF-kB via MUFAs. Our research has revealed the novel positive loop to amplify Wnt signaling through stabilization of LRP5/6 in both hepatic stellate cells and liver tumor-initiating stem cell-like cells. As such, this loop is pivotal in promoting liver fibrosis and liver tumor development. This review summarizes the mechanisms of SCD-mediated tumor promotion described by recent studies and discusses the future prospect for SCD-mediated signaling crosstalk as a potential therapeutic target for cancer.


Assuntos
Carcinogênese , Estearoil-CoA Dessaturase/metabolismo , Animais , Proteínas de Ciclo Celular/metabolismo , Células Estreladas do Fígado/citologia , Células Estreladas do Fígado/metabolismo , Humanos , Neoplasias/metabolismo , Neoplasias/patologia , Estearoil-CoA Dessaturase/genética , Via de Sinalização Wnt
12.
Int J Mol Sci ; 20(24)2019 Dec 13.
Artigo em Inglês | MEDLINE | ID: mdl-31847129

RESUMO

4-methylumbelliferone (4MU) is an inhibitor of hyaluronan deposition and an active substance of hymecromone, a choleretic and antispasmodic drug. 4MU reported to be anti-fibrotic in mouse models; however, precise mechanism of action still requires further investigation. Here we describe the cellular and molecular mechanisms of 4MU action on CCl4-induced liver fibrosis in mice using NGS transcriptome, Q-PCR and immunohistochemical analysis. Collagen and hyaluronan deposition were prevented by 4MU. The CCl4 stimulated expression of Col1a and αSMA were reduced, while the expression of the ECM catabolic gene Hyal1 was increased in the presence of 4MU. Bioinformatic analysis identified an activation of TGF-beta and Wnt/beta-catenin signaling pathways, and inhibition of the genes associated with lipid metabolism by CCL4 treatment, while 4MU restored key markers of these pathways to the control level. Immunohistochemical analysis reveals the suppression of hepatic stellate cells (HSCs) transdifferentiation to myofibroblasts by 4MU treatment. The drug affected the localization of HSCs and macrophages in the sites of fibrogenesis. CCl4 treatment induced the expression of FSTL1, which was downregulated by 4MU. Our results support the hypothesis that 4MU alleviates CCl4-induced liver fibrosis by reducing hyaluronan deposition and downregulating FSTL1 expression, accompanied by the suppression of HSC trans-differentiation and altered macrophage localization.


Assuntos
Proteínas Relacionadas à Folistatina/biossíntese , Regulação da Expressão Gênica/efeitos dos fármacos , Ácido Hialurônico/biossíntese , Himecromona/farmacologia , Cirrose Hepática , Via de Sinalização Wnt/efeitos dos fármacos , Actinas/biossíntese , Animais , Intoxicação por Tetracloreto de Carbono/metabolismo , Intoxicação por Tetracloreto de Carbono/patologia , Intoxicação por Tetracloreto de Carbono/prevenção & controle , Transdiferenciação Celular/efeitos dos fármacos , Feminino , Células Estreladas do Fígado/metabolismo , Células Estreladas do Fígado/patologia , Hialuronoglucosaminidase/biossíntese , Cirrose Hepática/induzido quimicamente , Cirrose Hepática/metabolismo , Cirrose Hepática/patologia , Cirrose Hepática/prevenção & controle , Camundongos , Camundongos Endogâmicos BALB C , Miofibroblastos/metabolismo , Miofibroblastos/patologia
13.
Int J Mol Sci ; 20(24)2019 Dec 13.
Artigo em Inglês | MEDLINE | ID: mdl-31847284

RESUMO

: Salacia chinensis L. (SC) stems have been used as an ingredient in Thai traditional medicine for treating patients with hepatic fibrosis and liver cirrhosis. However, there is no scientific evidence supporting the antifibrotic effects of SC extract. Therefore, this study aimed to determine the antifibrotic activity of SC stem extract in human hepatic stellate cell-line called LX-2. We found that upon TGF-ß1 stimulation, LX-2 cells transformed to a myofibroblast-like phenotype with a noticeable increase in α-SMA and collagen type I production. Interestingly, cells treated with SC extract significantly suppressed α-SMA and collagen type I production and reversed the myofibroblast-like characteristics back to normal. Additionally, TGF-ß1 also influenced the development of fibrogenesis by upregulation of MMP-2, TIMP-1, and TIMP-2 and related cellular signaling, such as pSmad2/3, pErk1/2, and pJNK. Surprisingly, SC possesses antifibrotic activity through the suppression of TGF-ß1-mediated production of collagen type 1, α-SMA, and the phosphorylation status of Smad2/3, Erk1/2, and JNK. Taken together, the present study provides accumulated information demonstrating the antifibrotic effects of SC stem extract and revealing its potential for development for hepatic fibrosis patients.


Assuntos
Células Estreladas do Fígado/metabolismo , Cirrose Hepática/tratamento farmacológico , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Extratos Vegetais/farmacologia , Caules de Planta/química , Salacia/química , Proteína Smad2/metabolismo , Proteína Smad3/metabolismo , Fator de Crescimento Transformador beta1/metabolismo , Linhagem Celular , Células Estreladas do Fígado/patologia , Humanos , Cirrose Hepática/metabolismo , Cirrose Hepática/patologia , Extratos Vegetais/química
14.
Life Sci ; 239: 117010, 2019 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-31672578

RESUMO

AIMS: Amlexanox, an inhibitor of nuclear factor κB kinase epsilon (IKKε) and TANK-binding kinase 1(TBK1), was demonstrated to be effective in diabetes and obesity. The aim of this study was to explore the molecular mechanisms of its role in non-alcoholic fatty liver disease (NAFLD). MAIN METHODS: NAFLD mouse models were established by using eight-week-old male C57BL/6 mice fed with high-fat diet (HFD) or (and) lipopolysaccharide (LPS) for 18 weeks. From the beginning of HFD, HFD-induced mice were subjected to amlexanox or vehicle for 18 weeks. HFD + LPS-induced mice were treated with amlexanox or vehicle for the last 6 weeks. Blood biochemistry parameters were determined using automatic biochemistry analyzer. Histological changes of liver tissue were observed by hematoxylin-eosin (H&E) staining and Oil Red O staining. The expressions of IKKε and smooth muscle actin-α (α-SMA) were evaluated through immunohistochemistry. Serum inflammatory mediator was determined by enzyme linked immunosorbent assay (ELISA). Gene expressions involved in glucose and lipid metabolism, insulin signaling pathway were examined using quantitative RT-PCR or Western blotting. KEY FINDINGS: This study demonstrated that amlexanox reversed glucose and lipid metabolic disturbance and hepatic steatosis in NAFLD mice model. IKKε was specific expressed in hepatic stellate cells (HSCs) instead of hepatocytes. This study also found that amlexanox improved insulin signaling (Insulin-IRS-1-Akt) in hepatocytes through inhibiting inflammation (IKKε-NF-κB-TNF-α/IL-1α) in HSCs. SIGNIFICANCE: The present study confirmed that IKKε was specific expressed in HSCs. Inhibition of activated HSCs was responsible for effects of amlexanox on NAFLD, with improving insulin signal pathway in hepatocytes.


Assuntos
Aminopiridinas/uso terapêutico , Células Estreladas do Fígado/efeitos dos fármacos , Quinase I-kappa B/antagonistas & inibidores , Hepatopatia Gordurosa não Alcoólica/tratamento farmacológico , Actinas/antagonistas & inibidores , Aminopiridinas/farmacologia , Animais , Peso Corporal/efeitos dos fármacos , Metabolismo Energético/efeitos dos fármacos , Expressão Gênica/efeitos dos fármacos , Glucose/metabolismo , Células Estreladas do Fígado/metabolismo , Células Estreladas do Fígado/patologia , Hepatócitos/efeitos dos fármacos , Hepatócitos/patologia , Insulina , Metabolismo dos Lipídeos/efeitos dos fármacos , Metabolismo dos Lipídeos/genética , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Hepatopatia Gordurosa não Alcoólica/genética , Hepatopatia Gordurosa não Alcoólica/patologia , Transdução de Sinais/efeitos dos fármacos
15.
Life Sci ; 237: 116902, 2019 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-31610195

RESUMO

AIMS: Insulin-like growth factor binding protein-related protein 1 (IGFBPrP1) promotes hepatic stellate cell (HSC) autophagy and activation. However, the underlying mechanism remains unknown. Noncoding RNAs (ncRNAs) including long noncoding RNAs (lncRNAs) and microRNAs (miRNAs), have received increasing attention. We aimed to investigate the roles of the lncRNA nuclear enriched abundant transcript 1 (NEAT1), miR-29b, and autophagy related protein 9a (Atg9a), and their relationships with each other during IGFBPrP1-induced HSC autophagy and activation. MAIN METHODS: Levels of NEAT1, miR-29b, Atg9a, and autophagy were detected in adenovirus-mediated IGFBPrP1 (AdIGFBPrP1)-treated mouse liver tissue and immortalized mouse hepatic stellate cell line JS1 transfected with either AdIGFBPrP1 or siIGFBPrP1. In AdIGFBPrP1-treated JS1 cells, autophagy and activation were detected after altering NEAT1, miR-29b, or Atg9a levels. In AdIGFBPrP1-treated JS1 cells, relationships among NEAT1, miR-29b, and Atg9a were explored using dual-luciferase reporter assays, Western blot, qRT-PCR, and immunofluorescence. KEY FINDINGS: IGFBPrP1 increased levels of NEAT1, Atg9a, and autophagy while decreasing the level of miR-29b in mouse liver tissues and mouse HSCs. Moreover, NEAT1 increased HSC autophagy and activation while miR-29b decreased both processes. Atg9a also participated in IGFBPrP1-induced HSC autophagy and activation. Importantly, NEAT1, miR-29b, and Atg9a formed a NEAT1/miR-29b/Atg9a regulatory axis for IGFBPrP1-induced HSC autophagy and activation. SIGNIFICANCE: Our study unveiled the new NEAT1/miR-29b/Atg9a regulatory axis involved in IGFBPrP1-induced mouse HSC autophagy and activation. The study thus provides new insights in the pathogenesis and potential therapeutic strategies of liver fibrosis.


Assuntos
Proteínas Relacionadas à Autofagia/metabolismo , Autofagia , Células Estreladas do Fígado/patologia , Proteínas de Ligação a Fator de Crescimento Semelhante a Insulina/metabolismo , Cirrose Hepática/patologia , Proteínas de Membrana/metabolismo , MicroRNAs/genética , RNA Longo não Codificante/genética , Proteínas de Transporte Vesicular/metabolismo , Adenoviridae/genética , Animais , Proteínas Relacionadas à Autofagia/genética , Células Cultivadas , Células Estreladas do Fígado/metabolismo , Proteínas de Ligação a Fator de Crescimento Semelhante a Insulina/genética , Cirrose Hepática/etiologia , Cirrose Hepática/metabolismo , Masculino , Proteínas de Membrana/genética , Camundongos , Camundongos Endogâmicos C57BL , Proteínas de Transporte Vesicular/genética
16.
Life Sci ; 238: 116934, 2019 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-31610205

RESUMO

Proliferation and differentiation of hepatic stellate cells (HSCs) are the most noticeable events in hepatic fibrosis, in which the loss of lipid droplets (LDs) is the most important feature. However, the complex mechanisms of LD disappearance have not been fully elucidated. In the current study, we investigated whether oroxylin A has the pharmacological activity of reversing LDs in activated HSCs, and further examined its potential molecular mechanisms. Using genetic, pharmacological, and molecular biological measure, we found that LD content significantly decreased during HSC activation, whereas oroxylin A markedly reversed LD content in activated HSCs. Interestingly, oroxylin A treatment observably decreased the expression of adipose triglyceride lipase (ATGL) without large differences in classical LD synthesis pathway, LD-related transcription factors, and autophagy pathway. ATGL overexpression could completely impair the effect of oroxylin A on reversing LD content. Importantly, reactive oxygen species (ROS) signaling pathway mediated oroxylin A-induced ATGL downregulation and LD revision in activated HSCs. ROS specific stimulant buthionine sulfoximine (BSO) could dramatically diminish the antioxidant effect of oroxylin A, and in turn, abolish reversal effect of oroxylin A on LD content. Conversely, ROS specific scavenger N-acetyl cystenine (NAC) can significantly enhance the pharmacological effect of oroxylin A on LD revision. Taken together, our study reveals the important molecular mechanism of anti-fibrosis effect of oroxylin A, and also suggests that ROS-ATGL pathway is a potential target for reversing LDs.


Assuntos
Flavonoides/farmacologia , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Células Estreladas do Fígado/efeitos dos fármacos , Lipase/antagonistas & inibidores , Gotículas Lipídicas/metabolismo , Cirrose Hepática/tratamento farmacológico , Animais , Autofagia , Células Cultivadas , Regulação para Baixo , Células Estreladas do Fígado/metabolismo , Células Estreladas do Fígado/patologia , Gotículas Lipídicas/efeitos dos fármacos , Cirrose Hepática/etiologia , Cirrose Hepática/metabolismo , Cirrose Hepática/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Transdução de Sinais
17.
Parasit Vectors ; 12(1): 475, 2019 Oct 14.
Artigo em Inglês | MEDLINE | ID: mdl-31610797

RESUMO

BACKGROUND: Inflammation-induced dysfunction of hepatic stellate cells (HSCs) is involved in schistosomiasis-associated liver fibrosis, and soluble egg antigen (SEA) is a crucial pathogen-associated molecular pattern associated with liver injury in schistosomiasis. In addition, numerous studies have shown that caspase-1-mediated pyroptosis participates in the development of multiple inflammation-related diseases. However, whether pyroptotic cell death of HSCs is involved in SEA-mediated liver damage is not well understood. METHODS: Primary cultured HSCs and Schistosoma japonicum-infected mouse liver tissue were analysed for histological changes and caspase-1 activation, and the role of pyroptosis in the mechanisms underlying SEA-induced HSC death was investigated. Accumulation of reactive oxygen species (ROS) in infected livers and SEA-stimulated HSCs was measured by flow cytometry and immunofluorescence. RESULTS: Caspase-1 activity was elevated in both liver tissues and HSCs of S. japonicum-infected mice. Furthermore, SEA stimulation increased the proportion of pyroptotic HSCs, as shown by lactate dehydrogenase (LDH) release assays and by flow cytometric analysis of propidium iodide (PI) and caspase-1 double staining in cells. In addition, ROS generation was elevated in infected liver tissues and SEA-stimulated HSCs, and ROS inhibition downregulated SEA-induced caspase-1 activation and pyroptosis in HSCs. CONCLUSIONS: Our present study demonstrates that pyroptotic cell death in HSCs induced by SEA via ROS-mediated caspase-1 activation may serve as a significant mechanism to initiate the inflammatory response and thereby exacerbate liver injury during S. japonicum infection.


Assuntos
Antígenos de Helmintos/fisiologia , Células Estreladas do Fígado/fisiologia , Piroptose/fisiologia , Espécies Reativas de Oxigênio/imunologia , Schistosoma japonicum/imunologia , Análise de Variância , Animais , Caspase 1/genética , Caspase 1/metabolismo , Feminino , Imunofluorescência , Células Estreladas do Fígado/citologia , Células Estreladas do Fígado/enzimologia , Células Estreladas do Fígado/metabolismo , Imuno-Histoquímica , Fígado/enzimologia , Fígado/metabolismo , Fígado/parasitologia , Fígado/patologia , Cirrose Hepática/enzimologia , Cirrose Hepática/patologia , Camundongos , Camundongos Endogâmicos C57BL , Espécies Reativas de Oxigênio/metabolismo , Reação em Cadeia da Polimerase em Tempo Real , Schistosoma japonicum/metabolismo , Esquistossomose Japônica/enzimologia , Esquistossomose Japônica/etiologia , Esquistossomose Japônica/patologia , Caramujos/parasitologia
18.
Curr Med Sci ; 39(5): 766-777, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31612395

RESUMO

The activation of hepatic stellate cells (HSCs) is a major event during hepatic fibrogenesis. Restoration of intracellular lipid droplet (LD) formation turns the activated HSC back to a quiescent state. Our previous studies have shown that curcumin suppresses HSC activation through increasing peroxisome proliferator-activated receptor, gamma (PPARγ) and 5' adenosine monophosphate-activated protein kinase (AMPK) activities. This study aims at evaluating the effect of curcumin on lipid accumulation in HSCs and hepatocytes, and further elucidating the underlying mechanisms. Now we showed that curcumin increased LD formation in activated HSCs and stimulated the expression of sterol regulatory element-binding protein and fatty acid synthase, and reduced the expression of adipose triglyceride lipase. Exogenous perilin5 expression in primary HSCs promoted LD formation. Perilipin 5 siRNA eliminated curcumin-induced LD formation in HSCs. These results suggest that curcumin recovers LD formation and lipid accumulation in activated HSCs by increasing perilipin 5 gene expression. Furthermore, inhibition of AMPK or PPARγ activity blocked curcumin's effect on Plin5 gene expression and LD formation. Our results provide a novel evidence in vitro for curcumin as a safe, effective candidate to treat liver fibrosis.


Assuntos
Curcumina/farmacologia , Células Estreladas do Fígado/efeitos dos fármacos , Gotículas Lipídicas/efeitos dos fármacos , Perilipina-1/genética , Perilipina-5/genética , Proteínas Quinases Ativadas por AMP/genética , Proteínas Quinases Ativadas por AMP/metabolismo , Animais , Linhagem Celular , Relação Dose-Resposta a Droga , Ácido Graxo Sintase Tipo I/genética , Ácido Graxo Sintase Tipo I/metabolismo , Ácidos Graxos não Esterificados/metabolismo , Regulação da Expressão Gênica , Células Estreladas do Fígado/citologia , Células Estreladas do Fígado/metabolismo , Hepatócitos/citologia , Hepatócitos/efeitos dos fármacos , Hepatócitos/metabolismo , Lipase/genética , Lipase/metabolismo , Gotículas Lipídicas/metabolismo , Camundongos , Especificidade de Órgãos , PPAR gama/genética , PPAR gama/metabolismo , Perilipina-1/agonistas , Perilipina-1/metabolismo , Perilipina-5/agonistas , Perilipina-5/metabolismo , Cultura Primária de Células , Transdução de Sinais , Proteína de Ligação a Elemento Regulador de Esterol 1/genética , Proteína de Ligação a Elemento Regulador de Esterol 1/metabolismo , Triglicerídeos/metabolismo
19.
Int J Mol Sci ; 20(20)2019 Oct 11.
Artigo em Inglês | MEDLINE | ID: mdl-31614491

RESUMO

Non-alcoholic fatty liver disease (NAFLD) affects up to 30% of the adult population and is now a major cause of liver disease-related premature illness and deaths in the world. Treatment is largely based on lifestyle modification, which is difficult to achieve in most patients. Progression of simple fatty liver or steatosis to its severe form non-alcoholic steatohepatitis (NASH) and liver fibrosis has been explained by a 'two-hit hypothesis'. Whilst simple steatosis is considered the first hit, its transformation to NASH may be driven by a second hit. Of several factors that constitute the second hit, advanced glycation end products (AGEs), which are formed when reducing-sugars react with proteins or lipids, have been implicated as major candidates that drive steatosis to NASH via the receptor for AGEs (RAGE). Both endogenous and processed food-derived (exogenous) AGEs can activate RAGE, mainly present on Kupffer cells and hepatic stellate cells, thus propagating NAFLD progression. This review focuses on the pathophysiology of NAFLD with special emphasis on the role of food-derived AGEs in NAFLD progression to NASH and liver fibrosis. Moreover, the effect of dietary manipulation to reduce AGE content in food or the therapies targeting AGE/RAGE pathway on disease progression is also discussed.


Assuntos
Produtos Finais de Glicação Avançada/metabolismo , Hepatopatia Gordurosa não Alcoólica/metabolismo , Receptor para Produtos Finais de Glicação Avançada/metabolismo , Progressão da Doença , Alimentos/efeitos adversos , Células Estreladas do Fígado/metabolismo , Humanos , Macrófagos do Fígado/metabolismo , Hepatopatia Gordurosa não Alcoólica/induzido quimicamente
20.
Int J Mol Sci ; 20(20)2019 Oct 13.
Artigo em Inglês | MEDLINE | ID: mdl-31614930

RESUMO

The tumor microenvironment, which consists of fibroblasts, smooth muscle cells, endothelial cells, immune cells, epithelial cells, and extracellular matrices, plays a crucial role in tumor progression. Hepatic stellate cells (HSCs), a class of unique liver stromal cells, participate in immunomodulatory activities by inducing the apoptosis of effector T-cells, generation of regulatory T-cells, and development of myeloid-derived suppressor cells (MDSCs) to achieve long-term survival of islet allografts. This study provides in vitro and in vivo evidences that HSCs induce the generation of MDSCs to promote hepatocellular carcinoma (HCC) progression through interleukin (IL)-6 secretion. HSC-induced MDSCs highly expressed inducible nitric oxide synthase (iNOS) and arginase 1 mRNA and presented potent inhibitory T-cell immune responses in the tumor environment. Wild-type HSC-induced MDSCs expressed lower levels of CD40, CD86, and MHC II, and a higher level of B7-H1 surface molecules, as well as increased the production of iNOS and arginase I compared with MDSCs induced by IL-6-deficient HSCs in vitro. A murine-transplanted model of the liver tumor showed that HCCs cotransplanted with HSCs could significantly enhance the tumor area and detect more MDSCs compared with HCCs alone or HCCs cotransplanted with HSCs lacking IL-6. In conclusion, the results indicated that MDSCs are induced mainly by HSCs through IL-6 signaling and produce inhibitory enzymes to reduce T-cell immunity and then promote HCC progression within the tumor microenvironment. Therapies targeting the pathway involved in MDSC production or its immune-modulating pathways can serve as an alternative immunotherapy for HCC.


Assuntos
Células Estreladas do Fígado/metabolismo , Interleucina-6/metabolismo , Neoplasias Hepáticas Experimentais/imunologia , Células Supressoras Mieloides/imunologia , Animais , Arginase/metabolismo , Linhagem Celular , Progressão da Doença , Humanos , Neoplasias Hepáticas Experimentais/metabolismo , Neoplasias Hepáticas Experimentais/patologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Monócitos/citologia , Monócitos/metabolismo , Óxido Nítrico Sintase Tipo II/metabolismo , Transdução de Sinais , Linfócitos T/metabolismo , Microambiente Tumoral/imunologia
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