Spatial and functional polarization of cancer-associated fibroblasts with CXCR4-mediated immune modulation in hepatocellular carcinoma

Background: Cancer-associated fibroblasts (CAFs) represent a predominant cell population in the tumor microenvironment (TME) of hepatocellular carcinoma (HCC). How the spatial distribution of CAF subtypes relates to immune modulation within the TME remains incompletely understood. This study aimed to characterize the spatial organization of CAF subtypes in HCC and to explore their potential associations with immune contexture. Methods: Spatial transcriptomic analysis was performed on 6 HCC cases to identify CAF-enriched clusters and their spatial localization. Immunohistochemical analyses were conducted on 132 HCC sections to examine the relationship between CAF-related features, immune cell distribution, and clinical outcomes. Results: Spatial transcriptomic analysis identified distinct CAF sub-clusters, including myofibroblastic CAFs (myCAFs), which were predominantly localized within tumor regions and enriched for extracellular matrix–related gene expression; inflammatory CAFs (iCAFs), which were preferentially localized within tumor septa and associated with inflammatory signaling; and antigen-presenting CAFs (apCAFs), which were enriched in the peritumoral zone and exhibited gene expression patterns suggestive of immune-related functions. Interactome analysis suggested that apCAFs in the peritumoral region showed transcriptional patterns consistent with CXCL12–CXCR4–related interactions with CD8 + T cells accumulating at the tumor periphery. The enrichment of apCAFs on the outer side of tumor septa was associated with reduced intra-tumoral CD8 + T-cell infiltration and poorer prognosis, suggesting a link between apCAF distribution and an immune-excluded TME. Conclusions: Spatial transcriptomic profiling suggests that CAFs exhibit functional and spatial polarization in the TME of HCC. ApCAFs are associated with CXCR4-related immune exclusion at the tumor–stroma interface and may represent a stromal component relevant to immune modulation. These findings provide a framework for CAF–immune interactions and serve as a basis for future functional studies targeting the CXCL12–CXCR4 axis.