Cytokine–ontogeny interaction shapes macrophage transcriptional states (combined GM-CSF– and M-CSF–derived macrophages)

We generated eight macrophage states using a cytokine–ontogeny matrix, combining two macrophage ontogenies (M-CSF and GM-CSF) with four cytokines (IL-4, IFN-?, IL-10, TGF-ß). Bulk RNA-seq on five biological replicates per condition revealed that PC1 (54% variance) segregated ontogeny: GM-CSF states loaded negatively, M-CSF states positively. Gene-set enrichment analysis of negative PC1 loadings highlighted NfkB programs, confirming the intrinsic inflammatory bias of GM-CSF macrophages (Hamilton 2008; Hamilton et al. 2016) Overall design: M-CSF–derived and GM-CSF–derived macrophages were stimulated with IL-4, IFN-?, IL-10, TGF-ß, or left untreated (control). Bulk RNA-seq was performed on five biological replicates per condition.