摘要
目的探讨氧化低密度脂蛋白/β2糖蛋白I/抗β2糖蛋白I抗体(oxLDL/β2GPI/抗β2GPI抗体)复合物对RAW264.7小鼠巨噬细胞自噬的影响及机制。方法分别用培养基、oxLDL、oxLDL/β2GPI复合物、oxLDL/β2GPI/抗β2GPI抗体复合物、oxLDL/抗β2GPI抗体复合物和β2GPI/抗β2GPI抗体复合物处理RAW264.7细胞。采用Western blot法检测自噬相关蛋白微管相关蛋白1轻链3(LC3)、P62的蛋白水平,采用携带双荧光蛋白和LC3基因的腺病毒(mRFP-GFP-LC3)感染巨噬细胞检测自噬体形成以及自噬流情况,采用蛋白激酶B(AKT)通路抑制剂LY294002(50μmol/L)预处理,观察AKT通路在oxLDL/β2GPI/抗β2GPI抗体复合物介导的RAW264.7细胞自噬中的作用。结果oxLDL/β2GPI/抗β2GPI抗体复合物能够降低LC3Ⅱ蛋白水平,减少自噬体数量,增加P62蛋白水平,阻断自噬流;LY294002预处理可部分恢复细胞自噬水平,改善自噬流阻断情况。结论oxLDL/β2GPI/抗β2GPI抗体复合物可以通过激活AKT通路抑制RAW264.7细胞的自噬。
Objective To investigate the effect of oxidative low-density lipoprotein/β2 glycoprotein I/anti-β2 glycoprotein I antibody(oxLDL/β2GPI/anti-β2GPI antibody) complex on autophagy in mouse macrophages(RAW264.7 cells) and its mechanism. Methods RAW264.7 cells were stimulated with medium, oxLDL, oxLDL/β2GPI complex, oxLDL/β2GPI/anti-β2GPI antibody complex, oxLDL/anti-β2GPI antibody complex and β2GPI/anti-β2GPI antibody complex separately. The protein levels of autophagy-associated proteins including LC3 and P62 were detected by Western blot analysis. The tandem mRFP-GFP-LC3 adernovirus was used to detect the patency of autophagosomes and autophagy flow. The AKT pathway inhibitor LY294002(50 μmol/L) was used for investigating the role of AKT pathway in the autophagy in RAW264.7 cells treated by oxLDL/β2GPI/anti-β2GPI antibody complex. Results oxLDL/β2GPI/anti-β2GPI antibody complex reduced the level of LC3-Ⅱ protein and the number of autophagosomes, increased the level of P62 protein and blocked the autophagy flow. In addition, the autophagy in RAW264.7 cells was partially restored by the pre-treatment of LY294002. Conclusion The oxLDL/β2GPI/anti-β2GPI antibody complex can inhibit the autophagy in RAW264.7 cells through activating the AKT pathway.
作者
吴倩倩
周红
何超
张贵婷
姚雨叶
陈芋丹
张鹏
匡铭
WU Qianqian;ZHOU Hong;HE Chao;ZHANG Guiting;YAO Yuye;CHEN Yudan;ZHANG Peng;KUANG Ming(School of Medicine,Jiangsu University,Zhenjiang 212013,China)
出处
《细胞与分子免疫学杂志》
CAS
CSCD
北大核心
2019年第12期1057-1063,共7页
Chinese Journal of Cellular and Molecular Immunology
基金
国家自然科学基金(81370614)。
