The Histone Demethylase KDM4D Promotes Hepatic Fibrogenesis by Modulating Toll-Like Receptor 4 Signaling Pathway

Abstract

Potential conflict of interest: Nothing to report.Abbreviations: HSC, hepatic stellate cell; H3K9, histone H3 on lysine 9; TLR4, Toll-like receptor 4; α-SMA, α-smooth muscle actin; KMT, lysine methyltransferase; KDM, lysine demethylase; KDM4, lysine (K)-specific demethylase 4; qRT-PCR, quantitative real-time PCR; siRNA, small interfering RNA; FBS, fetal bovine serum; i.p., intraperitoneally; CCl 4 , carbon tetrachloride; TAA, thioacetamide; BDL, bile duct ligation; AAV, adeno-associated virus;shRNA, short-hairpin RNA; ALT, alanine aminotransferase; H&E, hematoxylin-eosin; ChIP, chromatin immunoprecipitation; DMEM, Dulbecco's Modified Eagle's Medium; H3K9me2, histone di-methyl-H3K9; IHC, immunohistochemistry; NF-κB, nuclear factor kappa B; MyD88, myeloid differentiation factor 88; GFP, green fluorescent protein.3 AbstractAccumulating evidence has revealed the pivotal role of epigenetic regulation in the pathogenesis of liver disease. However, the epigenetic mechanism that accounts for hepatic stellate cells (HSCs) activation in liver fibrosis remains largely unknown. In this study, primaryHSCs were used to screen the differentially expressed histone H3 lysine methyltransferases and demethylases during HSC activation. KDM4D was identified as a remarkable upregulated histone H3 demethylase during HSC activation. The overexpression profile of KDM4D was further confirmed in three fibrosis animal models and human fibrotic liver tissues. In vitro genetic silencing of Kdm4d impaired the collagen gel contraction and migration capacity of primary HSCs. In established CCl 4 -induced mice model, Kdm4d knockdown inhibited fibrosis progression, and promoted fibrosis reversal, with enhanced thinning and splitting of fibrotic septa, as well as a dramatic decrease in collagen area. Whole gene transcriptome analysis showed the regulatory role of KDM4D in Toll-Like Receptor (TLR) signaling pathway. Mechanistically, KDM4D catalyzed histone 3 on lysine 9 (H3K9) di-, and tri-demethylation, which promoted TLR4 expression, and subsequently prompted liver fibrogenesis by activating NF-κB signaling pathways. Conclusion: KDM4D facilitates TLR4 transcription through demethylation of H3K9, thus activating TLR4/NF-κB signaling pathways in HSCs, contributing to HSC activation and collagen crosslinking, further, hepatic fibrosis progression.

Scoparone alleviates hepatic fibrosis by inhibiting the TLR‐4/NF‐κB pathway

“…In our in vivo experiments, we found that the liver tissue structure was severely damaged after treatment with CCl 4 (Figure 1d), where collagen was excessively accumulated (Figure 2b), and ALT, AST, ALP, γ‐GT, and TBIL had a significant rise in the activity or there was an elevated content of HA, LN, PC‐III, IV‐C, α‐SMA, and TGF‐β1 (Figures 1–2). The above indicators demonstrate that our in vivo animal model of hepatic fibrosis was successfully established (Dong et al, 2019; Liao et al, 2019; Lin et al, 2018). In addition, HSC‐T6 cells were then treated with TGF‐β1, which represented a mature model of hepatic fibrosis in vitro (He et al, 2016; Qin et al, 2018).…”

IOX1 protects from TGF‑β induced fibrosis in LX‑2 cells via the regulation of extracellular matrix protein expression

“…Proliferation and activation of hepatic stellate cells (HSC), and balancing extracellular matrix (ECM) synthesis and metabolism have become research hotspots of hepatic fibrosis treatment (4,5). There have been several reports that modification of aberrant histone methylation status may inhibit the proliferation and activation of HSCs to prevent the development of hepatic fibrosis (6,7). Yang et al (6) found that the histone H3K27 methyltransferase inhibitor, DZNep, displays anti-hepatic fibrosis activity through alteration of the aberrant H3K27 methyl group in HSCs.…”

“…Yang et al (6) found that the histone H3K27 methyltransferase inhibitor, DZNep, displays anti-hepatic fibrosis activity through alteration of the aberrant H3K27 methyl group in HSCs. Dong et al (7) demonstrated that knockout of lysine demethylase 4D (KDM4D) protein, a demethylase of H3K9 and H3K36, increased methylation levels of H3K9 and H3K36 in HSCs to decrease α-smooth muscle actin (SMA) and Col I expression, thereby inhibiting the proliferation and activation of HSC. The results of these studies indicated that reduction of histone methylation in activated HSCs by corresponding inhibitors or siRNA methods may inhibit HSC proliferation and activation.…”

“…The results of these studies indicated that reduction of histone methylation in activated HSCs by corresponding inhibitors or siRNA methods may inhibit HSC proliferation and activation. It has been demonstrated that the H3K9 methylation level was decreased during HSC activation (7); however, it is unknown whether modification of H3K9 methylation by histone H3K9 demethylase inhibitors affects HSC proliferation, activation and ECM synthesis, and metabolism in the activated HSCs.…”