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SerpinB3 Promotes Pro-fibrogenic Responses in Activated Hepatic Stellate Cells.

ABSTRACT: SerpinB3 is a hypoxia- and hypoxia-inducible factor-2?-dependent cystein protease inhibitor that is up-regulated in hepatocellular carcinoma and in parenchymal cells during chronic liver diseases (CLD). SerpinB3 up-regulation in CLD patients has been reported to correlate with the extent of liver fibrosis and the production of transforming growth factor-?1, but the actual role of SerpinB3 in hepatic fibrogenesis is still poorly characterized. In the present study we analyzed the pro-fibrogenic action of SerpinB3 in cell cultures and in two different murine models of liver fibrosis. "In vitro" experiments revealed that SerpinB3 addition to either primary cultures of human activated myofibroblast-like hepatic stellate cells (HSC/MFs) or human stellate cell line (LX2 cells) strongly up-regulated the expression of genes involved in fibrogenesis and promoted oriented migration, but not cell proliferation. Chronic liver injury by CCl₄ administration or by feeding a methionine/choline deficient diet to transgenic mice over-expressing human SerpinB3 in hepatocytes confirmed that SerpinB3 over-expression significantly increased the mRNA levels of pro-fibrogenic genes, collagen deposition and ?SMA-positive HSC/MFs as compared to wild-type mice, without affecting parenchymal damage. The present study provides for the first time evidence that hepatocyte release of SerpinB3 during CLD can contribute to liver fibrogenesis by acting on HSC/MFs.

SUBMITTER: Novo E

PROVIDER: S-EPMC5469760 | biostudies-literature | 2017 Jun

REPOSITORIES: biostudies-literature

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SerpinB3 Promotes Pro-fibrogenic Responses in Activated Hepatic Stellate Cells.

Novo Erica E Villano Gianmarco G Turato Cristian C Cannito Stefania S Paternostro Claudia C Busletta Chiara C Biasiolo Alessandra A Quarta Santina S Morello Elisabetta E Bocca Claudia C Miglietta Antonella A David Ezio E Sutti Salvatore S Plebani Mario M Albano Emanuele E Parola Maurizio M Pontisso Patrizia P

Scientific reports 20170613 1

SerpinB3 is a hypoxia- and hypoxia-inducible factor-2α-dependent cystein protease inhibitor that is up-regulated in hepatocellular carcinoma and in parenchymal cells during chronic liver diseases (CLD). SerpinB3 up-regulation in CLD patients has been reported to correlate with the extent of liver fibrosis and the production of transforming growth factor-β1, but the actual role of SerpinB3 in hepatic fibrogenesis is still poorly characterized. In the present study we analyzed the pro-fibrogenic a ...[more]

PMID: 28611447

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Project description:Background & aimsMethyl-CpG binding protein 2, MECP2, which binds to methylated regions of DNA to regulate transcription, is expressed by hepatic stellate cells (HSCs) and is required for development of liver fibrosis in mice. We investigated the effects of MECP2 deletion from HSCs on their transcriptome and of phosphorylation of MECP2 on HSC phenotype and liver fibrosis.MethodsWe isolated HSCs from Mecp2-/y mice and wild-type (control) mice. HSCs were activated in culture and used in array analyses of messenger RNAs and long noncoding RNAs. Kyoto Encyclopedia of Genes and Genomes pathway analyses identified pathways regulated by MECP2. We studied mice that expressed a mutated form of Mecp2 that encodes the S80A substitution, MECP2S80, causing loss of MECP2 phosphorylation at serine 80. Liver fibrosis was induced in these mice by administration of carbon tetrachloride, and liver tissues and HSCs were collected and analyzed.ResultsMECP2 deletion altered expression of 284 messenger RNAs and 244 long noncoding RNAs, including those that regulate DNA replication; are members of the minichromosome maintenance protein complex family; or encode CDC7, HAS2, DNA2 (a DNA helicase), or RPA2 (a protein that binds single-stranded DNA). We found that MECP2 regulates the DNA repair Fanconi anemia pathway in HSCs. Phosphorylation of MECP2S80 and its putative kinase, HAS2, were induced during transdifferentiation of HSCs. HSCs from MECP2S80 mice had reduced proliferation, and livers from these mice had reduced fibrosis after carbon tetrachloride administration.ConclusionsIn studies of mice with disruption of Mecp2 or that expressed a form of MECP2 that is not phosphorylated at S80, we found phosphorylation of MECP2 to be required for HSC proliferation and induction of fibrosis. In HSCs, MECP2 regulates expression of genes required for DNA replication and repair. Strategies to inhibit MECP2 phosphorylation at S80 might be developed for treatment of liver fibrosis.

Project description:IntroductionHepatic stellate cells (HSC) become activated, differentiate to myofibroblasts and produce extracellular fibrillar matrix during liver fibrosis. The hepatic fibrogenic response is orchestrated by reciprocal interactions between HSCs and macrophages and their secreted products. SOCS1 can regulate several cytokines and growth factors implicated in liver fibrosis. Here we investigated the role of SOCS1 in regulating HSC activation.MethodsMice lacking SOCS1 in HSCs (Socs1ΔHSC) were generated by crossing Socs1fl/fl and LratCre mice. Liver fibrosis was induced by carbon tetrachloride and evaluated by Sirius red staining, hydroxyproline content and immunostaining of myofibroblasts. Gene expression of pro-fibrogenic factors, cytokines, growth factors and chemokines were quantified by RT-qPCR. The phenotype and the numbers of intrahepatic leukocyte subsets were studied by flow cytometry. The impact of fibrosis on the development of diethyl nitrosamine-induced hepatocellular carcinoma was evaluated.ResultsSocs1ΔHSC mice developed more severe liver fibrosis than control Socs1fl/fl mice that was characterized by increased collagen deposition and myofibroblast differentiation. Socs1ΔHSC mice showed a significant increase in the expression of smooth muscle actin, collagens, matrix metalloproteases, cytokines, growth factors and chemokines in the liver following fibrosis induction. The fibrotic livers of Socs1ΔHSC mice displayed heightened inflammatory cell infiltration with increased proportion and numbers of Ly6ChiCCR2+ pro-inflammatory macrophages. This macrophage population contained elevated numbers of CCR2+CX3CR1+ cells, suggesting impaired transition towards restorative macrophages. Fibrosis induction following exposure to diethyl nitrosamine resulted in more numerous and larger liver tumor nodules in Socs1ΔHSC mice than in Socs1fl/fl mice.DiscussionOur findings indicate that (i) SOCS1 expression in HSCs is a critical to control liver fibrosis and development of hepatocaellular carcinoma, and (ii) attenuation of HSC activation by SOCS1 regulates pro-inflammatory macrophage recruitment and differentiation during liver fibrosis.

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SerpinB3 Promotes Pro-fibrogenic Responses in Activated Hepatic Stellate Cells.

Publications

SerpinB3 Promotes Pro-fibrogenic Responses in Activated Hepatic Stellate Cells.

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