Clonorchis sinensis infection remodels chromatin accessibility in hepatocellular carcinoma

Hepatocellular carcinoma (HCC) is a major global health concern, accounting for a significant proportion of liver cancer cases and related deaths. Clonorchis sinensis (C. sinensis) infection, a recognized carcinogen, has been implicated in the progression of liver diseases, including HCC. However, the precise epigenetic mechanisms underlying C. sinensis-associated HCC remain to be elucidated.

To investigate the role of chromatin accessibility in C. sinensis-related HCC progression, we performed an assay for transposase-accessible chromatin with high-throughput sequencing (ATAC-seq) and RNA sequencing (RNA-seq) analyses of C. sinensis-infected (C. sinensis⁺) and non-C. sinensis-infected (C. sinensis^-) HCC tumors. Integrated analyses were conducted to assess chromatin accessibility, transcription factor (TF) motifs, and histone modifications using ATAC-seq, RNA-seq, and classical chromatin immunoprecipitation-sequencing (ChIP-seq) datasets. A scratch wound assay was used to evaluate the effects of C. sinensis excretory/secretory products (CsESPs) on HCC cell migration.

ATAC-seq analysis revealed 9,396 differentially accessible regions (DARs) in C. sinensis⁺ HCC tumors compared with C. sinensis^- HCC tumors. Additionally, several crucial TFs enriched in DARs were identified, including HNF4A, FOXO1, ELF4, and RELA. Combined ATAC-seq and RNA-seq analyses further revealed differentially expressed genes (DEGs) associated with metabolism, immune regulation, and cytoskeletal dynamics. Chromatin accessibility was closely associated with histone modifications such as H3K9ac, H3K4me2, H3K4me3, H3K27ac, H3K4me1, and CTCF binding. Notably, C. sinensis infection significantly increased the migratory capacity of HCC cells, as confirmed by molecular assays and clinical observations.

Our study demonstrates that C. sinensis infection remodels chromatin accessibility and may contribute to HCC progression. Our work offers valuable insights into the pathogenesis of HCC in the context of parasitic infection and lays the groundwork for future biomarker and therapeutic target discovery.