PPARδ的激活通过抑制糖酵解减轻PDGF-BB诱导的肺动脉平滑肌细胞表型转化

Activation of PPARδsuppresses phenotypic switching of PDGF-BB-in⁃duced pulmonary artery smooth muscle cells via inhibition of glycolysis

目的:探讨过氧化物酶体增殖物激活受体δ(PPARδ)激动剂GW501516对血小板源性生长因子BB(PDGF-BB)诱导的大鼠肺动脉平滑肌细胞(PASMCs)表型转化和糖酵解的影响及其机制。方法:以20μg/L PDGFBB作为诱导剂,建立PASMCs表型转化及糖酵解的细胞模型,通过GW501516或糖酵解抑制剂2-脱氧葡萄糖(2-DG)干预24 h,检测PASMCs表型和糖酵解的变化。设置缺氧诱导因子1α(HIF-1α)抑制剂LW6为阳性对照组,通过GW501516和(或)HIF-1α稳定剂二甲基草酰甘氨酸(DMOG)干预24 h,测定细胞外乳酸含量及葡萄糖摄取量的变化;检测PPARδ、血管平滑肌细胞表型标志蛋白、HIF-1α及糖酵解相关蛋白表达,探讨GW501516抑制PASMCs表型转化及糖酵解的机制。结果:PDGF-BB刺激可抑制PASMCs内PPARδ表达,而GW501516处理可促进PPARδ表达(P<0.05)。GW501516上调PASMCs收缩表型标志蛋白平滑肌22α蛋白(SM22α)和α-平滑肌肌动蛋白(α-SMA)表达,下调合成表型标志蛋白波形蛋白(vimentin)和骨桥蛋白(OPN)表达,减少细胞外乳酸生成,抑制细胞对葡萄糖的摄取(P<0.05),而2-DG在抑制PASMCs表型转化及糖酵解方面的作用与GW501516相似。LW6和GW501516可抑制糖酵解与细胞表型转化,抑制HIF-1α、葡萄糖转运体1(GlUT1)、丙酮酸脱氢酶激酶1(PDK1)、乳酸脱氢酶A(LDHA)和单羧酸转运体蛋白4(MCT4)表达,同时促进丙酮酸脱氢酶(PDH)表达(P<0.05),而DMOG可逆转GW501516的上述作用(P<0.05)。结论:PPARδ激动剂GW501516可通过抑制糖酵解而抑制PDGF-BB诱导的PASMCs表型转化,其机制可能与抑制HIF-1α的表达有关。

AIM:To investigate the effects of peroxisome proliferator-activated receptorδ(PPARδ)agonist GW501516 on the phenotypic switching and glycolysis in rat pulmonary artery smooth muscle cells(PASMCs)induced by platelet-derived growth factor-BB(PDGF-BB),and to explore its mechanisms.METHODS:PDGF-BB(20μg/L)was used to induce phenotypic switching and glycolysis in PASMCs,and the PASMCs were then treated with GW501516 or glycolysis inhibitor 2-deoxy-D-glucose(2-DG)for 24 h to investigate the phenotypic switching and glycolysis.To explore the molecular mechanisms by which GW501516 suppresses the phenotypic switching and glycolysis of PASMCs,we used GW501516,hypoxia-inducible factor 1α(HIF-1α)inhibitor LW6 and HIF-1αstabilizer dimethyloxalylglycine(DMOG)to treat PASMCs for 24 h.The extracellular lactate level and cellular glucose uptake were determined by an assay kit.The levels of PPARδ,phenotype marker proteins of vascular smooth muscle cells,HIF-1αand glycolysis-related proteins were detected by Western blot.RESULTS:PDGF-BB reduced the expression of PPARδ,while GW501516 promoted the expression of PPARδin PASMCs(P<0.05).In addition,GW501516 increased the expression of contractile phenotype markers smooth muscle 22α(SM22α)andα-smooth muscle actin(α-SMA),but decreased the expression of synthetic phenotype markers vimentin and osteopontin(OPN)in PDGF-BB-exposed PASMCs(P<0.05).Moreover,GW501516 re‐duced extracellular lactate level and suppressed cellular glucose uptake in PDGF-BB-stimulated PASMCs(P<0.05).The glycolysis inhibitor 2-DG had similar effects to GW501516 in inhibiting cell phenotypic switching and glycolysis.Further experiments showed that both GW501516 and LW6 blocked glycolysis and phenotypic switching of PASMCs,reduced the expression levels of glucose transporter 1(GlUT1),pyruvate dehydrogenase kinase 1(PDK1),lactate dehydrogenase A(LDHA)and monocarboxylate transporter protein 4(MCT4),but promoted the pyruvate dehydrogenase(PDH)expres‐sion(P<0.05).Notably,HIF-1αstabilizer DMOG reversed the beneficial effects of GW501516 in PASMCs treated with PDGF-BB(P<0.05).CONCLUSION:The PPARδagonist GW501516 inhibits the phenotypic switching of PDGF-BB-induced PASMCs via inhibition of glycolysis,and its mechanism may be related to suppression of HIF-1α.