摘要
目的:探究蛋白酶激活受体2(protease-activated receptor 2,PAR2)对人内皮祖细胞(endothelial progenitor cell,EPC)功能的影响。方法:EPC予以PAR2天然激动剂类胰蛋白酶、合成激动剂SLIGKV-NH2、合成抑制剂FSLLRY-NH2处理,EdU、Transwell实验观察EPC增殖、迁移,实时定量PCR、ELISA分析检测相关细胞因子及受体表达,免疫印迹法评估Ras同源家族成员A(Ras homolog family member A,RhoA)水平;同时给予RhoA特异性抑制剂Y-27632,观察PAR2活化效应可否被取消。结果:PAR2激动剂可剂量依赖性抑制EPC增殖及迁移(P<0.05),下调血管内皮生长因子A、血管内皮生长因子受体2、基质细胞衍生因子1及趋化性细胞因子受体4等表达(P<0.05),该效应可被PAR2抑制剂取消;活化PAR2可显著上调EPC RhoA表达(P<0.05),抑制PAR2活性可取消该效应;Y-27632可逆转PAR2激动剂导致的EPC细胞增殖、迁移抑制(P<0.05)。结论:PAR2经RhoA信号抑制EPC增殖、迁移功能,是极具潜力的内皮再生与血管生成调控靶点。
Objective:The present study aims to investigate the role of protease⁃activated receptor 2(PAR2)in the regulation of human endothelial progenitor cell(EPC)function.Methods:EPCs were stimulated with tryptase(a natural agonist of PAR2),SLIGKV⁃NH2(a synthetic agonist of PAR2)and FSLLRY⁃NH2(an antagonist of PAR2).Cell proliferation and migration were evaluated by EdU incorporation and Transwell model.Expression of the cytokines and receptors were estimated by real⁃time quantitative PCR and ELISA.Level of intercellular Ras homolog family member A(RhoA)was assessed by Western blot analysis.And RhoA antagonist Y⁃27632 was also applied to determine whether the effects of PAR2 activation can be abolished by RhoA inhibition.Results:The agonists of PAR2 dramatically inhibited EPCs proliferation and migration in a dose⁃dependent manner(P<0.05).PAR2 activation markedly suppressed the expression of vascular endothelial growth factor⁃A,vascular endothelial growth factor receptor⁃2,stromal cell⁃derived factor⁃1and C⁃X⁃C chemokine receptor type 4(P<0.05).All these effects can be abolished by the PAR2 antagonist(P<0.05).PAR2 activation increased the level of RhoA in EPCs,which was also repressed by FSLLRY⁃NH2(P<0.05).Y⁃27632 notably reversed the influence of PAR2 activation on EPCs proliferation and migration(P<0.05).Conclusion:The activation of PAR2 blunted EPCs proliferation and migration via RhoA signal,hinting a potential role of PAR2 as a novel target for the modulation of endothelial regeneration and vasculogenesis.
作者
顾圣玮
顾浩
马晴晴
姚欣
曾晓宁
GU Shengwei;GU Hao;MA Qingqing;YAO Xin;ZENG Xiaoning(Department of Pulmonary&Critical Care Medicine,the First Affiliated Hospital of Nanjing Medical University,Nanjing 210029,China)
出处
《南京医科大学学报(自然科学版)》
CAS
北大核心
2023年第5期655-662,共8页
Journal of Nanjing Medical University(Natural Sciences)
基金
国家自然科学基金面上项目(81970016,81001428)。
关键词
蛋白酶激活受体2
内皮祖细胞
增殖
迁移
RHOA
protease⁃activated receptor 2
endothelial progenitor cells
proliferation
migration
RhoA