CD137和CD137配体信号通过低氧诱导因子1α调控糖酵解影响巨噬细胞吞噬及迁移功能

CD137 and CD137 ligand signals affect phagocytosis and migration of macrophages by regulating glycolysis through via HIF-1α

目的探讨CD137-CD137配体(CD137L)信号通过低氧诱导因子1α(HIF-1α)调控糖酵解影响巨噬细胞吞噬及迁移功能。方法通过巯基乙酸盐腹腔灌洗法获取C57BL/6J小鼠腹腔巨噬细胞(PM),采用混合气体饱和法建立细胞低氧环境。PM分为对照组、低氧组、CD137激动组、HIF-1α过表达组和糖酵解激动组。采用Western blot检测各组HIF-1α、葡萄糖转运蛋白1(GLUT1)、糖酵解关键酶己糖激酶(HK2)、果糖-2,6-二磷酸酶3(PFKFB3)表达,葡萄糖和乳酸试剂盒检测各组葡萄糖消耗量及乳酸产量,墨汁吞噬实验检测各组吞噬功能,划痕实验和Transwell实验检测各组迁移功能。结果CD137激动组HIF-1α、GLUT1、HK2和PFKFB3蛋白、葡萄糖消耗、乳酸产量、吞噬率、划痕愈合率和迁移的巨噬细胞数明显低于低氧组(P<0.05,P<0.01);HIF-1α过表达组HIF-1α、GLUT1、HK2和PFKFB3蛋白、葡萄糖消耗、乳酸产量、吞噬率、划痕愈合率、迁移的巨噬细胞数明显高于CD137激动组(P<0.05,P<0.01);糖酵解激动组吞噬率、划痕愈合率和迁移的巨噬细胞数明显高于CD137激动组[(85.89±7.76)%vs(69.01±6.94)%,P<0.05;(17.43±1.76)%vs(1.41±0.52)%,P<0.01;(176±16)个vs(48±10)个,P<0.01]。结论CD137-CD137L信号可能通过HIF-1α调控糖酵解影响巨噬细胞吞噬及迁移功能。

Objective To study whether CD137 and CD137 ligand signals can affect the phagocytosis and migration of macrophages by regulating glycolysis via hypoxia inducible factor-1α(HIF-1α).Methods The peritoneal macrophages(PM)taken from C57BL/6J mice by peritoneal lavage with thioglycolate with their hypoxic condition established by mixed gas saturating were divided into control group,hypoxic group,CD137 excitation group,HIF-1αoverexpression group and glycolysis excitation group.The expressions of HIF-1α,GLUT1,key glycolysis enzymes including hexokinaseⅡ(HK2)and fructose-2,6-bisphosphatase 3(PFKFB3)in different groups were detected by Western blot.The glucose consumption and lactic acid production in different groups were measured with a glucose and lactic acid content detection kit.The phagocytosis of PM in different groups was assayed by ink phagocytosis test.The migration of PM in different groups was tested by scratch test and transwell test respectively.Results The expression levels of HIF-1α,GLUT1,HK2 and PFKFB3 were significantly lower,the volume of glucose consumption and lactic acid production was significantly smaller,the rate of phagocytosis and scratch healing was significantly lower and the number of migrating macrophages was significantly smaller in CD137 excitation group than in hypoxic group(P<0.05,P<0.01).The expression levels of HIF-1α,GLUT1,HK2 and PFKFB3 were significantly higher,the volume of glucose consumption and lactic acid production was significantly larger,the rate of phagocytosis and scratch healing was significantly higher and the number of migrating macrophages was significantly greater in HIF-1αoverexpression group than in CD137 excitation group(P<0.05,P<0.01).The rate of phagocytosis and scratch healing was significantly higher and the number of migrating macrophages was significantly greater in glycolysis excitation group than in CD137 excitation group(85.89%±7.76%vs 69.01%±6.94%,P<0.05;17.43%±1.76%vs 1.41%±0.52%,P<0.01;176±16 vs 48±10,P<0.01).Conclusion CD137 and CD137 ligand signals can affect the phagocytosis and migration of macrophages by regulating the glycolysis via HIF-1α.