Single-cell profiling of cross-tissue features of fetal pig macrophages and their remodeling in fetal growth restriction

Macrophages are highly heterogeneous immune cells with context-dependent functions, yet their diversity across prenatal tissues of the whole body, sexes, and stages remains elusive. During fetal development, macrophages play essential roles in tissue formation and maintaining maternal-fetal immune balance, but their specific phenotypic and functional changes in the context of fetal growth restriction remain relatively underexplored. Herein, we constructed a cross-tissue macrophage atlas of 85 473 cells from 54 tissues of late-gestation fetal pigs. We identified 15 macrophage subtypes, among which two Hofbauer cell (HBC) subtypes were distinguished by their distinct transcriptional and functional characteristics between the amniotic and chorioallantoic membranes: HBC GPNMB+ cells upregulated glycolytic enzymes and extracellular matrix regulators such as LDHA and GPNMB, while HBC PEG10+ cells expressed immunomodulators and developmental regulators, including A2M and PEG10. Two dichotomous subtypes, LYVE1^highMHCII^low and LYVE1^lowMHCII^high, were delineated across 27 tissues, with a conserved transcriptional signature in pigs and humans. Transcriptional profiling of metabolic pathways revealed oxidative phosphorylation and the tricarboxylic acid cycle as core pathways across tissues, alongside tissue-specific adaptations such as enhanced glycolysis in amniotic macrophages. Cross-stage comparisons showed fetal macrophages mainly maintained developmental programs, whereas adult macrophages globally exhibited heightened immune activity. Cross-sex and cross-species analyses revealed high transcriptomic similarity in fetal macrophages. In growth-restricted fetuses, chorioallantoic HBCs showed profound remodeling, including increased inflammation, hypoxia response, and vascular dysfunction signaling, with altered intercellular communication. Collectively, this work provides a comprehensive resource for understanding fetal macrophage heterogeneity and offers insights into the potential mechanisms underlying fetal growth restriction.