Mutant MMP-9 and HGF Gene Transfer Enhance Resolution of CCl4-Induced Liver Fibrosis in Rats: Role of ASH1 and EZH2 Methyltransferases Repression

Abstract

Hepatocyte growth factor (HGF) gene transfer inhibits liver fibrosis by regulating aberrant cellular functions, while mutant matrix metalloproteinase-9 (mMMP-9) enhances matrix degradation by neutralizing the elevated tissue inhibitor of metalloproteinase-1 (TIMP-1). It was shown that ASH1 and EZH2 methyltransferases are involved in development of liver fibrosis; however, their role in the resolution phase of liver fibrosis has not been investigated. This study evaluated the role of ASH1 and EZH2 in two mechanistically different therapeutic modalities, HGF and mMMP-9 gene transfer in CCl4 induced rat liver fibrosis. Liver fibrosis was induced in rats with twice a week intraperitoneal injection of CCl4 for 8 weeks. Adenovirus vectors encoding mMMP-9 or HGF genes were injected through tail vein at weeks six and seven and were sacrificed one week after the second injection. A healthy animal group was likewise injected with saline to serve as a negative control. Rats treated with mMMP-9 showed significantly lower fibrosis score, less Sirius red stained collagen area, reduced hydroxyproline and ALT concentration, decreased transforming growth factor beta 1 (TGF-β1) mRNA and lower labeling indices of α smooth muscle actin (α-SMA) and proliferating cell nuclear antigen (PCNA) stained cells compared with HGF- or saline-treated rats. Furthermore, TIMP-1 protein expression in mMMP-9 group was markedly reduced compared with all fibrotic groups. ASH1 and EZH2 protein expression was significantly elevated in fibrotic liver and significantly decreased in mMMP-9- and HGF-treated compared to saline-treated fibrotic livers with further reduction in the mMMP-9 group. Conclusion: Gene transfer of mMMP-9 and HGF reduced liver fibrosis in rats. ASH1 and EZH2 methyltransferases are significantly reduced in mMMP-9 and HGF treated rats which underlines the central role of these enzymes during fibrogenesis. Future studies should evaluate the role of selective pharmacologic inhibitors of ASH1 and EZH2 in resolution of liver fibrosis.

Kupffer-derived matrix metalloproteinase-9 contributes to liver fibrosis resolution

“…KC-derived MMP9 might also play important role in elimination of active myofibroblasts. A recent study found that transgene of MMP9 resulted in apoptosis of myofibroblasts 23 . Other studies also found that TIMPs and collagen I prevented myofibroblasts from apoptosis, while MMP9 or MMP13 derived from KCs has the able to reduce the levels of TIMPs and collagen I 22 .…”

Targeting histone methyltransferase enhancer of zeste homolog‐2 inhibits renal epithelial‐mesenchymal transition and attenuates renal fibrosis

“…Increasing evidence indicates the importance of EZH2 in mediating tissue and organ fibrosis. EZH2 levels are elevated in the fibrotic liver, and genetic or pharmacologic disruption of this molecule can attenuate liver fibrogenesis by inhibiting fibrotic characteristics of myofibroblasts (57). EZH2-mediated histone hypermethylation has also been found to be involved in idiopathic pulmonary fibrosis (58).…”

The fibrolytic potentials of vitamin D and thymoquinone remedial therapies: insights from liver fibrosis established by CCl4 in rats

“…Both TQ and VitD have been reported to upregulate the protein expression and activity of many MMPs; among them is MMP-9, and to suppress the expression of TIMP-1 [ 23 ]. MMP-9 is known for its high affinity to bind with TIMP-1 thus scavenging and reducing the effects of TIMP-1 on the other MMPs [ 18 , 52 ]. Additionally, the observed fibrolytic effect of combined therapy protocol in our study was associated with improved actions for TQ and VitD on the expression of TGF-β1, IL-6, IL-10, IL-22 and MMP-9 in hepatic tissue.…”