Figure 5. TNFSF9 induces M2 polarization of macrophages and promotes the migration of pancreatic cancer cells. (A) U937 cells were successfully induced into macrophages. (B, C) The effect of wild-type pancreatic cancer cells on the polarization of U937-derived macrophages. (D, E) TNFSF9 knockdown pancreatic cancer cells inhibit the expression of M2 markers (IL-10 and TGF-β) mRNA levels in U937 macrophages. At the same time, it promotes the expression of M1 marker (IL-8 and TNF-α) mRNA level and inhibits the expression of IL-1β mRNA level. (F, G) Co-culture of U937-derived macrophages with TNFSF9 knockdown pancreatic cancer cells inhibited the migration of BXPC-3 and PANC-1 cells. (H, I) The migration of BXPC-3 and PANC-1 cells increased after adding recombinant human protein TGF-β. (J–L) Western blots were used to analyze the expression of inflammatory cytokines IL-8, IL-6, TNF-α and IL-1β on U937-derived macrophages after co-culture with pancreatic cancer cells knocked down by TNFSF9. Compared with the sh-NC group, the expression of IL-8, IL-6 and TNF-α in the sh-TNFSF9-1 and sh-TNFSF9-2 groups were significantly increased, and the expression of IL-1β was significantly reduced. sh-1 is sh-TNFSF9-1. sh-2 is sh-TNFSF9-2. *P < 0.05, **P < 0.01, ***P < 0.001 and #P < 0.001.