摘要
目的 探讨生长转化因子11(GDF11)对巨噬细胞M1极化介导的炎症反应的影响及其作用机制.方法 体外培养RAW264.7巨噬细胞,100 ng/mL LPS + 30 ng/mL IFN-γ诱导巨噬细胞M1极化,并将细胞分成对照组、LPS+IFN组、GDF11组及LPS+IFN+GDF11组.流式细胞术检测巨噬细胞的极化,实时PCR检测IL-6、TNF-α、NF-κB及GDF11mRNA的表达水平,Western blotting检测细胞GDF11、NF-κB及P-NF-κB蛋白表达水平,免疫荧光检测细胞GDF11、NF-κB的蛋白表达和分布.结果 与对照组相比,LPS+IFN组巨噬细胞M1极化比例增加,IL-6、TNF-α及NF-κBmRNA表达升高,GDF11mRNA及蛋白表达降低,NF-κB、P-NF-κB蛋白表达升高,P-NF-κB/NF-κB比值升高,NF-κB入核明显(P<0.05);与对照组相比,GDF11组巨噬细胞M1极化比例、IL-6、TNF-α、NF-κBmRNA表达及P-NF-κB/NF-κB比值无明显变化(P>0.05);与LPS+IFN组相比,LPS+IFN+GDF11组RAW264.7巨噬细胞M1极化比例显著降低,IL-6、TNF-α、NF-κB mRNA表达降低,NF-κB、P-NF-κB蛋白表达降低,P-NF-κB/NF-κB比值降低(P<0.05).结论 GDF11可减轻巨噬细胞M1极化介导的炎症反应,其机制可能与调节NF-κB通路有关.
Abstract
Objective To investigate the effect of GDF11 on the macrophage M1 polarization mediated inflammatory response and its mechanism.Methods RAW264.7 macrophages were cultured in vitro,and M1 polarization was induced by 100 ng/mL LPS + 30 ng/mL IFN-γ.The cells were divided into a control group,LPS+IFN group,GDF11 group,and LPS+IFN+GDF11 group.The polarization of the macrophages was detected by flow cytometry,the mRNA expression levels of IL-6,TNF-α,NF-κB,and GDF11 were detected by RT-qPCR,the protein expressions of GDF11,NF-κB,and P-NF-κB were detected by Western blotting,and the protein expression and distribution of GDF11 and NF-κB were detected by immunofluorescence.Results Compared with the control group,the M1 polariza-tion ratio of macrophages in the LPS+IFN group and the expression of IL-6,TNF-α,and NF-κBwere increased,while the expression of GDF11mRNA and protein were decreased;the expression of NF-κB and P-NF-κB proteins and the ratio of P-NF-κB/NF-κB were higher,and NF-κB nucleation was obvious(P<0.05).Compared with the control group,there were no significant changes in the M1 polarization ratio,IL-6,TNF-α,and NF-κBexpression,and P-NF-κB/NF-κB ratio in the GDF11 group(P>0.05).Compared with the LPS+IFN group,the M1 polarization ratio of RAW264.7 macrophages in the LPS+IFN+GDF11 group was significantly reduced,the expression of IL-6,TNF-α,NF-κB mRNA,and NF-κB,P-NF-κB protein was decreased,and the P-NF-κB/NF-κB ratio was also decreased(P<0.05).Conclusion GDF11 alleviates M1 polarization mediated inflammatory response in macrophages,and its mechanism may be related to regulation of the NF-κB pathway.
基金项目
湖南省自然科学基金科药联合项目(2022JJ80065)
湖南省卫生健康委一般指导项目(202110000505)
湖南省人民医院仁术国自培育项目(BSJJ202103)
国家科技期刊平台
NSTL
万方数据