Single-cell transcriptome atlas identified ERO1A+ angiogenesis-associated macrophage with a central potential in promoting hepatocellular carcinoma

Abstract Hepatocellular carcinoma (HCC) is the most prevalent form of liver cancer and remains a global health challenge due to its complexity and limited therapeutic options. Within the HCC tumor microenvironment, tumor-associated macrophages (TAMs) are the most abundant immune cells and critical mediators of immunosuppression and angiogenesis, but their heterogeneity and clinical relevance remain incompletely characterized. In this study, we investigated TAMs in clinical HCC by integrative analysis of single-cell RNA-sequencing (scRNA-seq) data from 46 HCC and 13 adjacent liver samples across five independent cohorts (256,236 cells). The resulting unified macrophage atlas identified eight subsets based on functional gene modules, with monocyte-derived angiogenesis-associated macrophages (Angio-Mac) emerging as the most clinically significant subset. Crucially, the identification of this cluster was robustly reproduced by a cross-platform validation using an independent human HCC single-nucleus RNA-sequencing (snRNA-seq) dataset and a sensitivity re-analysis with a strict 20% mitochondrial filtration cutoff. Angio-Mac expanded in advanced HCC, correlated with poor prognosis, and exhibited pro-tumorigenic transcriptomic features, including enhanced mTORC1 signaling and suppressed antigen presentation. Angio-Mac overexpressed SPP1 and was suggested to promote interactions with T/NK cells via the SPP1-CD44 axis. Unbiased risk analysis further revealed ERO1A as an Angio-Mac-specific marker linked to advanced HCC and poor prognosis. In contrast, Kupffer cell-derived TAMs displayed a “stalled” phenotype with limited pro-tumor activity. These findings highlight Angio-Mac as a macrophage subset in HCC characterized by prominent expression of genes associated with angiogenesis and immunosuppression. Consequently, this macrophage subset and its signature genes represent valuable candidates for further mechanistic investigation and clinical validation.