Comparative transcriptomic profiling of human hepatocellular carcinoma cell lines Hep3B and HUH7 by RNA sequencing

Hep3B and HUH7 are two widely employed human HCC cell lines that exhibit distinct genetic backgrounds, morphological characteristics, and oncogenic dependencies. To systematically investigate whether the expression levels of orthologous genes differ significantly between these two models, we performed high-throughput RNA sequencing (RNA-seq) on total RNA extracted from exponentially growing Hep3B (experimental group, designated 3B-1/2/3) and HUH7 (control group, designated 7H-1/2/3) cells under identical culture conditions, with three independent biological replicates per cell line.Our data reveal extensive transcriptional heterogeneity between Hep3B and HUH7 cells.

Functional enrichment analyses, including GO and KEGG pathway enrichment (GOSeq v1.34.1) as well as Gene Set Enrichment Analysis (GSEA, gseapy v1.1.9), were further performed to interpret the biological significance of identified DEGs. Additional analyses encompassed alternative splicing events (rMATS v4.1.0), novel transcript prediction (StringTie v1.3.3b and Cuffcompare v2.2.1), single nucleotide variant (SNV) calling (Sentieon v202503), and differential exon usage (DEXSeq v1.20.1).