VEGFA and APOE regulate distinct functional states of mast cells in hepatocellular carcinoma: A single-cell...

Immune cells play a crucial role in the development of Hepatocellular Carcinoma (HCC), with mast cells displaying unique bidirectional biological characteristics. This study aims to investigate the heterogeneity of hypertrophic cells in HCC and their impact on prognosis and potential dual roles. We performed single-cell RNA sequencing (scRNA-seq) analysis on tumor tissues from 32 HCC patients and adjacent non-tumor tissues from 5 of these patients, with validation through multiplex immunohistochemistry (mIHC) on 90 tumor and 90 adjacent tissues. Additional validation was done using public datasets (TCGA-LIHC, GSE14520) and cellular functional assays. Our findings reveal a mast cell-specific enrichment in tumor tissues, identifying four functionally distinct subpopulations of tumor-associated mast cells (TAMCs): Mast_1 (antigen-presenting), Mast_2 (cholesterol metabolism-associated), Mast_3 (transcription factor-enriched), and Mast_4 (oncogenic and prognostically adverse). Deconvolution analysis showed significant upregulation of TAMCs signature genes (PSMB8, APOE, SOX4, VEGFA), and transcriptional network analysis highlighted JUND, NF-κB, REL, and CREM as potential co-regulators of Mast_2/Mast_4 differentiation. mIHC and survival analysis indicated that high apolipoprotein E (APOE) expression correlated with favorable prognosis (p < 0.05), while high VEGFA levels predicted poor outcomes (p < 0.05). Correlation and cell communication analyses revealed a positive association between Mast_2 abundance and CD8+ T cell infiltration (r = 0.24, p < 0.01), with APOE-LDLR-mediated anti-tumor immunity. Functional assays showed that APOE-overexpressing mast cells enhanced IFN-γ secretion in CD8+ T cells, boosting their tumoricidal activity. Conversely, Mast_4 correlated with endothelial cells (r = 0.26, p < 0.05), interacting via the VEGFA-NRP2 axis. VEGFA-overexpressing mast cells increased endothelial tube formation and migration (p < 0.001). In conclusion, the heterogeneity of TAMCs identified in this study suggests novel therapeutic strategies for HCC.