Abstract
The liver is known to possess remarkable regenerative potential, but persistent inflammation or severe acute injury can lead to liver fibrosis and incomplete regeneration, ultimately resulting in liver failure. Recent studies have shown that the axis of two types of CXCL12 receptors, CXCR4 and CXCR7, plays a crucial role in liver fibrosis and regeneration. The present study aimed to investigate the regulatory factors involved in CXCR4 expression in injured liver. Immunohistochemical screening of liver tissue samples collected during liver transplantation revealed a reciprocal expression pattern between CXCR4 and MeCP2. An in vitro system involving cultured cell lines and H2O2 treatment was established to study the impact of oxidative stress on signaling pathways and epigenetic alterations that affect CXCR4 mRNA expression. Operating through distinct signaling pathways, H2O2 treatment induced a dose-dependent increase in CXCR4 expression in both hepatocyte- and intrahepatic cholangiocyte-derived cells. Treatment of the cells with trichostatin and azacytidine modulated CXCR4 expression in hepatocytes by modifying the methylation status of CpG dinucleotides located in a pair of TA repeats adjacent to the TATA box of the CXCR4 gene promoter. Only MeCP2 bound to oligonucleotides representing the TATA box region when the cytosine residues within the sequence were methylated, as revealed by electrophoretic mobility shift assay (EMSA). Methylation-specific PCR analysis of microdissected samples revealed a correlation between the loss of CpG methylation and the upregulation of CXCR4 in injured hepatocytes, replicating the findings from the in vitro study. Besides the conventional MEK/ERK and NF-κB signaling pathways that activate CXCR4 in intrahepatic cholangiocytes, the unique epigenetic modifications observed in hepatocytes might also contribute to a shift in the CXCR4–CXCR7 balance towards CXCR4, leading to irreversible liver injury and fibrosis. This study highlights the importance of epigenetic modifications in regulating CXCR4 expression in liver injury and fibrosis.
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Acknowledgements
We express our appreciation to Mss. Yuki Takaoka and Mariko Hashimoto for their valuable assistance.
Funding
This work was supported in part by a Grant-in-Aid for Scientific Research from the Ministry of Education, Culture, Sports, Science and Technology, Japan (20H03458 and 21K19354 to S. K.).
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SK participated in the experimental design and managed whole project. CI participated in all experiments. RH participated in MSP assays. RK participated in cell culture studies. All authors reviewed and approved the manuscript.
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This study was approved by the ethics committee of Ehime University of Medical School (approval no. 2307003).
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Ito, C., Haraguchi, R., Ogawa, K. et al. Demethylation in promoter region of severely damaged hepatocytes enhances chemokine receptor CXCR4 gene expression. Histochem Cell Biol 160, 407–418 (2023). https://doi.org/10.1007/s00418-023-02229-x
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DOI: https://doi.org/10.1007/s00418-023-02229-x