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目的检测高迁移率族蛋白B1(HMGB1)诱导肺泡巨噬细胞(AM)的细胞因子表达谱。方法 RT-PCR扩增HMGB1 cDNA,插入pGEX4T-1原核表达载体并转大肠杆菌,IPTG诱导HMGB1表达,Ni2+-NTA和多粘菌素B亲和层析柱分离纯化。应用LiquiChip液相蛋白芯片检测HMGB1(20ng/ml)诱导AM细胞因子表达谱的变化。结果成功表达纯化的重组HMGB1可诱导AM肿瘤坏死因子-α(TNF-α)、白细胞介素-1β(IL-1β)、白细胞介素-6(IL-6)、巨噬细胞炎性蛋白-1α(MIP-1α)、MIP-2、单核细胞趋化蛋白-1(MCP-1)、粒细胞-巨噬细胞集落刺激因子(GM-CSF)及γ-干扰素(IFN)-γ的表达显著增加(P<0.01)。结论 HMGB1可诱导AM释放多种炎性细胞因子。
Objective To detect the cytokine expression profile of alveolar macrophages (AM) induced by high mobility group box 1 (HMGB1). Methods The HMGB1 cDNA was amplified by RT-PCR, inserted into pGEX4T-1 prokaryotic expression vector and transformed into E.coli. The expression of HMGB1 was induced by IPTG, and purified by Ni2 + -NTA and polymyxin B affinity chromatography. LiquiChip liquid protein chip was used to detect the changes of AM cytokine expression profile induced by HMGB1 (20ng / ml). Results The purified recombinant HMGB1 was successfully expressed and the tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), interleukin-6 (IL-6), macrophage inflammatory protein- MIP-1α, MIP-2, MCP-1, GM-CSF and IFN-γ The expression was significantly increased (P <0.01). Conclusion HMGB1 can induce AM to release a variety of inflammatory cytokines.