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AQP3 is regulated by PPAR? and JNK in hepatic stellate cells carrying PNPLA3 I148M.

ABSTRACT: Aquaglyceroporins (AQPs) allow the movement of glycerol that is required for triglyceride formation in hepatic stellate cells (HSC), as key cellular source of fibrogenesis in the liver. The genetic polymorphism I148M of the patatin-like phospholipase domain-containing 3 (PNPLA3) is associated with hepatic steatosis and its progression to steatohepatitis (NASH), fibrosis and cancer. We aimed to explore the role of AQP3 for HSC activation and unveil its potential interactions with PNPLA3. HSC were isolated from human liver, experiments were performed in primary HSC and human HSC line LX2. AQP3 was the only aquaglyceroporin present in HSC and its expression decreased during activation. The PPAR? agonist, rosiglitazone, recovered AQP3 expression also in PNPLA3 I148M carrying HSC. When PNPLA3 was silenced, AQP3 expression increased. In liver sections from patients with NASH, the decreased amount of AQP3 was proportional to the severity of fibrosis and presence of the PNPLA3 I148M variant. In PNPLA3 I148M cells, the blockade of JNK pathway upregulated AQP3 in synergism with PPAR?. In conclusion, we demonstrated profound reduction of AQP3 in HSC carrying the PNPLA3 I148M variant in parallel to decreased PPAR? activation, which could be rescued by rosiglitazone and blockade of JNK.

SUBMITTER: Tardelli M

PROVIDER: S-EPMC5676689 | biostudies-other | 2017 Nov

REPOSITORIES: biostudies-other

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AQP3 is regulated by PPARγ and JNK in hepatic stellate cells carrying PNPLA3 I148M.

Tardelli Matteo M Bruschi Francesca V FV Claudel Thierry T Moreno-Viedma Veronica V Halilbasic Emina E Marra Fabio F Herac Merima M Stulnig Thomas M TM Trauner Michael M

Scientific reports 20171107 1

Aquaglyceroporins (AQPs) allow the movement of glycerol that is required for triglyceride formation in hepatic stellate cells (HSC), as key cellular source of fibrogenesis in the liver. The genetic polymorphism I148M of the patatin-like phospholipase domain-containing 3 (PNPLA3) is associated with hepatic steatosis and its progression to steatohepatitis (NASH), fibrosis and cancer. We aimed to explore the role of AQP3 for HSC activation and unveil its potential interactions with PNPLA3. HSC were ...[more]

PMID: 29116096

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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:Secreted phospholipase A2 (sPLA2) is a protein secreted by Clonorchis sinensis and is a component of excretory and secretory products (CsESPs). Phospholipase A2 is well known for its role in liver fibrosis and inhibition of tumour cells. The JNK signalling pathway is involved in hepatic stellate cells (HSCs) activation. Blocking JNK activity with SP600125 inhibits HSCs activation. In a previous study, the protein CssPLA2 was expressed in insoluble inclusion bodies. Therefore, it's necessary to express CssPLA2 in water-soluble form and determine whether the enzymatic activity of CssPLA2 or cell signalling pathways is involved in liver fibrosis caused by clonorchiasis.Balb/C mice were given an abdominal injection of MBP-CssPLA2. Liver sections with HE and Masson staining were observed to detect accumulation of collagen. Western blot of mouse liver was done to detect the activation of JNK signalling pathway. In vitro, HSCs were incubated with MBP-CssPLA2 to detect the activation of HSCs as well as the activation of JNK signalling pathway. The mutant of MBP-CssPLA2 without enzymatic activity was constructed and was also incubated with HSCs to check whether activation of the HSCs was related to the enzymatic activity of MBP-CssPLA2.The recombinant protein MBP-CssPLA2 was expressed soluble and of good enzymatic activity. A mutant of CssPLA2, without enzymatic activity, was also constructed. In vivo liver sections of Balb/C mice that were given an abdominal injection of 50 ?g/ml MBP-CssPLA2 showed an obvious accumulation of collagen and a clear band of P-JNK1 could be seen by western blot of the liver tissue. In vitro, MBP-CssPLA2, as well as the mutant, was incubated with HSCs and it was proved that activation of HSCs was related to activation of the JNK signalling pathway instead of the enzymatic activity of MBP-CssPLA2.Activation of HSCs by CssPLA2 is related to the activation of the JNK signalling pathway instead of the enzymatic activity of CssPLA2. This finding could provide a promising treatment strategy to interrupt the process of liver fibrosis caused by clonorchiasis.

Dataset Information

AQP3 is regulated by PPAR? and JNK in hepatic stellate cells carrying PNPLA3 I148M.

Publications

AQP3 is regulated by PPARγ and JNK in hepatic stellate cells carrying PNPLA3 I148M.

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