替代活化型巨噬细胞和多发性骨髓瘤早期治疗反应的关系及机制研究

OBJECTIVE: To investigate the relationship between M2-polarized macrophages and early response in multiple myeloma and its molecular mechanism. METHODS: Two hundred and forty bone marrow biopsy tissue were collected and M2-polarized macrophages were stained by anti-CD163 monoclonal antibody. In vitro M2-polarized macrophages were derived from human peripheral blood mononuclear cell or THP-1 cells and identified by flow cytometry. Two myeloma cell lines RPMI 8226 and U266 were co-cultured with M2 macrophages using a transwell system. We measured myeloma cells proliferation through CCK-8 method and the pro-inflammatory cytokines expression (TNF-α and IL-6) by ELISA. Real time PCR was applied to measure chemokines (CCL2 and CCL3), chemokine receptors (CCR2, CCR5), VEGF and their receptors. In addition, flow cytometry was used to analyze the apoptosis of myeloma cells induced by dexamethasone. RESULTS: ①Patients with high percentage of M2 macrophage involvement in bone marrow showed poorer response (23.9% versus 73.0%, χ(2)=60.31, P<0.001). ②In vitro the proliferation of RPMI 8226 cells (P=0.005 at 24 h, P=0.020 at 36 h) or U266 myeloma cells (P= 0.030 at 24h, P=0.020 at 36h) co-cultured with M2-polarized macrophages was higher than control group. ③In vitro the apoptotic rate of RPMI 8226 cells (29.0% versus 71.0%, t=4.97, P=0.008) or U266 myeloma cells (24.9% versus 67.7%, t=6.99, P=0.002) co-cultured with M2-polarized macrophages was lower than control group. ④In vitro M2-polarized macrophages promoted myeloma cells secreting higher level of IL-6, TNF-α and higher expression of CCL2, CCL3, CCR2, CCR5, VEGFA, VEGFR-1,-2 compared with the non-macrophage co-culture system. CONCLUSION: M2-polarized macrophages promote myeloma cells proliferation and inhibit apoptosis through a very complex mechanism involving pro-inflammatory cytokines IL-6 and TNF-α, chemokines and related receptors such as CCL2, CCL3, CCR2, CCR3, and VEGF as well as related VEGFR.