Fig. 5. RAD18^High TNBC promotes M2-like TAM polarization via TGF-β secretion and JNK pathway inhibition, inducing more TGF-β secreted from M2-like TAM reciprocally upregulates RAD18 to form a positive regulatory loop in the TNBC-TAM interaction.

A, B qRT-PCR was used to detect the expression of cytokines in both TNBC cells and macrophages after co-cultivation. The THP1- derived Mφ co-cultured with shNC and shRAD18 MDA-MB-231 for 3 d. The primary THP-1-derived Mφ (M0) was used as control (B). C Gene set enrichment analysis plot showing enriched TGF-β signaling pathway in the TCGA cohorts. D Detection of TGF-β in co-cultured supernatant by ELISA. E WB detection of protein expression levels of CD163, PPAR-δ, NF-κBp65, NF-κBp-p65, JNK, p-JNK, and c-Jun in macrophages. F ELISAs were used to determine the TGF-β concentration in supernatant secreted by macrophage alone after co-cultivation. G, H The tumor sphere formation and protein levels of stemness-related signature genes (CD44, OCT4, SOX2, Nanog) were detected to verify the stemness of shNC/shRAD18 MDA-MB-231 co-cultured with or without macrophages in the absence or presence of a neutralizing antibody specific for TGF-β are presented. *p < 0.05, **p < 0.01, ***p < 0.001. All experiments were performed independently at least three times.