Dataset Information

EZH2-mediated inhibition of KLF14 expression promotes HSCs activation and liver fibrosis by downregulating PPARγ.

ABSTRACT:

Objectives

Induction of deactivation and apoptosis of hepatic stellate cells (HSCs) are principal therapeutic strategies for liver fibrosis. Krüppel-like factor 14 (KLF14) regulates various biological processes, however, roles, mechanisms and implications of KLF14 in liver fibrosis are unknown.

Materials and methods

KLF14 expression was detected in human, rat and mouse fibrotic models, and its effects on HSCs were assessed. Chromatin immunoprecipitation assays were utilized to investigate the binding of KLF14 to peroxisome proliferator-activated receptor γ (PPARγ) promoter, and the binding of enhancer of zeste homolog 2 (EZH2) to KLF14 promoter. In vivo, KLF14-overexpressing adenovirus was injected via tail vein to thioacetamide (TAA)-treated rats to investigate the role of KLF14 in liver fibrosis progression. EZH2 inhibitor EPZ-6438 was utilized to treat TAA-induced rat liver fibrosis.

Results

KLF14 expression was remarkably decreased in human, rat and mouse fibrotic liver tissues. Overexpression of KLF14 increased LD accumulation, inhibited HSCs activation, proliferation, migration and induced G2/M arrest and apoptosis. Mechanistically, KLF14 transactivated PPARγ promoter activity. Inhibition of PPARγ blocked the suppressive role of KLF14 overexpression in HSCs. Downregulation of KLF14 in activated HSCs was mediated by EZH2-regulated histone H3 lysine 27 trimethylation. Adenovirus-mediated KLF14 overexpression ameliorated TAA-induced rat liver fibrosis in PPARγ-dependent manner. Furthermore, EPZ-6438 dramatically alleviated TAA-induced rat liver fibrosis. Importantly, KLF14 expression was decreased in human with liver fibrosis, which was significantly correlated with EZH2 upregulation and PPARγ downregulation.

Conclusions

KLF14 exerts a critical anti-fibrotic role in liver fibrosis, and targeting the EZH2/KLF14/PPARγ axis might be a novel therapeutic strategy for liver fibrosis.

SUBMITTER: Du Z

PROVIDER: S-EPMC8249795 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Rationale: As the central hallmark of liver fibrosis, transdifferentiation of hepatic stellate cells (HSCs), the predominant contributor to fibrogenic hepatic myofibroblast responsible for extracellular matrix (ECM) deposition, is characterized with transcriptional and epigenetic remodeling. We aimed to characterize the roles of H3K27 methyltransferase EZH2 and demethylase JMJD3 and identify their effective pathways and novel target genes in HSCs activation and liver fibrosis. Methods: In primary HSCs, we analyzed effects of pharmacological inhibitions and genetic manipulations of EZH2 and JMJD3 on HSCs activation. In HSCs cell lines, we evaluated effects of EZH2 inhibition by DZNep on proliferation, cell cycling, senescence and apoptosis. In CCl4 and BDL murine models of liver fibrosis, we assessed in vivo effects of DZNep administration and Ezh2 silencing. We profiled rat primary HSCs transcriptomes with RNA-seq, screened the pathways and genes associated with DZNep treatment, analyzed EZH2 and JMJD3 regulation towards target genes by ChIP-qPCR. Results: EZH2 inhibition by DZNep resulted in retarded growth, lowered cell viability, cell cycle arrest in S and G2 phases, strengthened senescence, and enhanced apoptosis of HSCs, decreased hepatic collagen deposition and rescued the elevated serum ALT and AST activities of diseased mice, and downregulated cellular and hepatic expressions of H3K27me3, EZH2, α-SMA and COL1A. Ezh2 silencing by RNA interference in vitro and in vivo showed similar effects. JMJD3 inhibition by GSK-J4 and overexpression of wild-type but not mutant Jmjd3 enhanced or repressed HSCs activation respectively. EZH2 inhibition by DZNep transcriptionally inactivated TGF-β1 pathway, cell cycle pathways and vast ECM components in primary HSCs. EZH2 inhibition decreased H3K27me3 recruitment at target genes encoding TGF-β1 pseudoreceptor BAMBI, anti-inflammatory cytokine IL10 and cell cycle regulators CDKN1A, GADD45A and GADD45B, and increased their expressions, while Jmjd3 overexpression manifested alike effects. Conclusions: EZH2 and JMJD3 antagonistically modulate HSCs activation. The therapeutic effects of DZNep as epigenetic drug in liver fibrosis are associated with the regulation of EZH2 towards direct target genes encoding TGF-β1 pseudoreceptor BAMBI, anti-inflammatory cytokine IL10 and cell cycle regulators CDKN1A, GADD45A and GADD45B, which are also regulated by JMJD3. Our present study provides new mechanistic insight into the epigenetic modulation of EZH2 and JMJD3 in HSCs biology and hepatic fibrogenesis.