Differences of Gpnmb expression in M1 and M2 bone marrow-derived macrophages in mouse / 中华微生物学和免疫学杂志

ABSTRACT

Objective To investigate the differences of glycoprotein non-metastatic melanoma b (Gpnmb) expression between M1 and M2 bone marrow-derived macrophages (BMMφs) in mouse.Meth-ods Primary BMMφs were cultured and then identified by immunofluorescence staining for F 4/80 and flow cytometry testing of CD11b.Interferon-γand lipopolysaccharide were used to induce differentiation of BMMφs towards M1 macrophages and interleukin-4 was adopted to induce differentiation of M 2 macropha-ges.Realtime PCR was performed to analyze mRNA expressions of tumor necrosis factor (TNF-α), induc-ible NO synthase (iNOS), macrophage mannose receptor (MMR), arginase-1 (Arg-1) and Gpnmb.Pro-teins of Gpnmb and MMR were detected by double immunofluorescence staining , Western blot and flow cy-tometry.Results (1) Immunofluorescence staining showed high expression of F 4/80 in BMMφs and flow cytometry results showed that CD11b was expressed in 92.7%±6.1% of BMMφs, suggesting that primary BMMφs were successfully cultured.(2) Compared with M0 BMMφs, mRNAs of TNF-αand iNOS were highly up-regulated in M1 BMMφs (both P<0.01), and mRNAs of MMR and Arg-1 were highly up-regula-ted in M2 BMMφs (both P<0.01), indicating that differentiation of BMMφs towards M1 and M2 BMMφs were successfully induced .(3) Expressions of Gpnmb mRNA and Gpnmb protein were predominantly up-regulated in M2 BMMφs in comparison with those in M0 and M1 BMMφs (both P<0.01).Gpnmb and MMR were co-expressed in M2 BMMφs and 83.2%±9.7% of MMR positive BMMφs expressed Gpnmb. Conclusion Gpnmb expression is significantly increased in M 2 macrophages than that in M 1 macrophages in vitro, indicating that Gpnmb which takes part in the differentiation of macrophages might be used as a marker for identification of M 1 and M2 macrophages .