Integrated Single-Cell and Spatial Transcriptomics Analyses Delineate a BAG3-Associated Macrophage Program with Microenvironmental and Prognostic Relevance in Hepatocellular Carcinoma

Background: Tumor-associated macrophages (TAMs) are key components of the hepatocellular carcinoma (HCC) microenvironment, but their spatial heterogeneity remains incompletely characterized. We aimed to assess the biological and prognostic relevance of a BAG3-associated TAM program in HCC. Methods: Public single-cell RNA sequencing (scRNA-seq) datasets were analyzed to characterize TAM heterogeneity, and an integrated validation scRNA-seq dataset was used to assess reproducibility. Spatial transcriptomics was used to provide spatial context in a small treatment-exposed cohort. Pseudotime, regulatory network, and cell–cell communication analyses were performed to characterize state transitions and microenvironmental interactions. Survival modeling evaluated the prognostic relevance of the BAG3-associated program. Results: Five TAM subsets were identified, including MARCO+, MT+ RTM−, MMP9+, UBE2C+, and BAG3+ TAMs. Among them, BAG3+ TAMs, a less well-characterized subset, exhibited coordinated stress-adaptive, proteostasis-related, and matrix-remodeling programs that were reproduced in the validation dataset. Pseudotime analysis suggested a continuum of TAM states, with BAG3+ TAM stress-remodeling features enriched toward late pseudotime. Communication analysis centered on BAG3+ TAMs suggested crosstalk between inflammatory stress cues and angiogenic, stromal-remodeling, and immunomodulatory programs; this pattern was primarily supported by HBV-derived samples and recurrently involved the MIF–CD74 axis. Spatial mapping further supported BAG3+ TAM-enriched niches with elevated AP-1, EGR1, and NFKB1 activity. A BAG3-associated risk score derived from a 10-gene signature remained an independent prognostic factor for overall survival after clinical adjustment. Conclusions: These findings characterize a BAG3-associated TAM program with spatial, immunoregulatory, and prognostic relevance in HCC, and support its further evaluation in biomarker and mechanistic studies.