Background: Tumor-associated macrophages (TAMs) and dysregulated
tumor epigenetics contribute to
hepatocellular carcinoma (HCC) progression. However, the mechanistic interactions between TAMs and
tumor epigenetics remain poorly understood. Methods: Immunohistochemistry and multiplexed fluorescence staining were performed to evaluate the correlation between TAMs numbers and UHRF1 expression in human HCC tissues.
PGE2 neutralizing antibody and
COX-2 inhibitor were used to analyze the regulation of TAMs isolated from HCC tissues on UHRF1 expression. Multiple
microRNA prediction programs were employed to identify
microRNAs that target UHRF1
3'UTR.
Luciferase reporter assay was applied to evaluate the regulation of miR-520d on UHRF1 expression.
Chromatin immunoprecipitation (ChIP) assays were performed to assess the abundance of H3K9me2 in the KLF6 promoter and DNMT1 in the CSF1 promoter regulated by UHRF1. The functional roles of TAM-mediated oncogenic network in HCC progression were verified by in vitro colony formation assays, in vivo xenograft experiments and analysis of clinical samples. Results: Here, we find that TAMs induce and maintain high levels of HCC UHRF1, an oncogenic epigenetic regulator. Mechanistically, TAM-derived
PGE2 stimulates UHRF1 expression by repressing miR-520d that targets the 3'-UTR of UHRF1
mRNA. In consequence, upregulated UHRF1 methylates H3K9 to diminish
tumor KLF6 expression, a
tumor inhibitory transcriptional factor that directly transcribes miR-520d.
PGE2 reduces KLF6 occupancy in the promoter of miR-520d, dampens miR-520d expression, and sustains robust UHRF1 expression. Moreover, UHRF1 promotes CSF1 expression by inducing
DNA hypomethylation of the CSF1 promoter and supports TAM accumulation. Conclusions: Capitalizing on studies on HCC cells and tissues, animal models, and clinical information, we reveal a previously unappreciated TAM-mediated oncogenic network via multiple reciprocal enforcing molecular nodes. Targeting this network may be an approach to treat HCC patients.