1572-P: Lipotoxicity-Induced TET1 Mediates CD36 DNA Demethylation in Liver Macrophages to Aggravate NAFLD Progression

Aims: DNA methylation is dysregulated in nonalcoholic fatty liver disease (NAFLD), yet the dominant genes and regulatory mechanisms remain obscure. Here, whole-genome bisulfite (WGBS) and RNA-seq revealed significantly downregulated DNA methylation and upregulated mRNA expression of Cd36 in a high fat diet (HFD)-induced murine fatty liver, particularly in macrophages. Meanwhile, ten-eleven translocation-1 (TET1), a DNA demethylase, was upregulated. We investigated whether lipotoxicity-induced TET1 erases the methylation at Cd36 promoter in liver macrophages and aggravates NAFLD progression.

Methods: The livers of 16-week HFD-induced male C57BL/6J mice were subjected to WGBS, RNA-seq and pyrosequencing. Hydroxymethylated DNA immunoprecipitation- and chromatin immunoprecipitation-qPCR were conducted to detect the level of 5-hydroxymethylcytosine on DNA, and the binding efficiency to the Cd36 promoter, respectively. TET1 expression was knocked down by adeno-associated virus in HFD-induced NAFLD mice. In vitro, Palmitate acid was applied to simulate lipotoxicity in Kupffer cell- and THP-1 cell line. CRISPR/Cas9 sg-Tet1 knockout lentivirus and TETs enzyme inhibitor were applied, and CD36 expression was knocked down by siRNA.

Results: In macrophages of fatty liver, the expression of CD36 and TET1 is markedly increased. Lipotoxicity enhances TET1 localization and demethylase activity at the Cd36 promoter. RXRα is recruited to the Cd36 promoter following the DNA demethylation of Cd36 by TET1 and increases Cd36 expression. Moreover, lipotoxicity-induced CD36 promotes M1 macrophage polarization and exacerbates inflammation. And the deficient TET1 activity restores DNA methylation of the Cd36 promoter, reducing its expression and M1 polarization.

Conclusion: TET1-mediated DNA demethylation has a crucial role in lipotoxicity-driven CD36 transcription in liver macrophage that boosts steatosis-to-nonalcoholic steatohepatitis progression.