摘要
目的研究脑源性神经营养因子(BDNF)对内皮细胞Sestrin2表达的影响及机制,并探讨其在血管新生中的作用。方法用100μg/L的BDNF分别处理人脐静脉内皮细胞(HUVEC)1 h、2 h、4 h、6 h、8 h,实时荧光定量PCR检测Sestrin2 mRNA水平,免疫荧光和Western blot检测Sestrin2蛋白表达。将HUVEC分为6组:对照组(Control组)、BDNF组(加BDNF 100μg/L)、BDNF+TrkB-Fc(1 mg/L)组、BDNF+KT-5823(500 nmol/L)组、BDNF+LNAME(10-4 mol/L)组、BDNF+DMSO组,共干预4 h,Western blot检测Sestrin2蛋白表达。将HUVEC分为4组:对照组(Control组)、BDNF组(加BDNF 100μg/L)、BDNF+Sestrin2 siRNA组和BDNF+Control siRNA组,共干预6 h,细胞迁移实验和小管成形实验分别检测HUVEC迁移能力和血管生成能力。结果与0 h和1 h组比较,100μg/L BDNF分别干预HUVEC 2、4及6 h时段,Sestrin2 mRNA水平显著增高(P<0.001),在2、4及8 h时段Sestrin2蛋白表达显著增高(P<0.05);阻断NO/PKG通路可抑制BDNF诱导的Sestrin2表达上调(P<0.001);抑制Sestrin2表达后,HUVEC迁移及小管形成能力较BDNF干预组显著降低(P<0.01)。结论BDNF通过NO/PKG通路促进内皮细胞表达Sestrin2,从而提高内皮细胞血管生成能力。
Aim To explore the effect of brain-derived neurotrophic factor(BDNF) on Sestrin2 expression andangiogenesis-related mechanisms in endothelial cells. Methods Human umbilical vein endothelial cells(HUVEC)were treated with BDNF(100 μg/L) for 1 h, 2 h, 4 h, 6 h, 8 h, and the protein and m RNA expression of Sestrin2 weredetected with immunofluorescent staining, Western blot and quantitative real-time polymerase chain reaction(q RT-PCR),respectively. HUVEC were divided into six groups: control group, BDNF(100 μg/L) group, BDNF+Trk B-Fc(1 mg/L)group, BDNF+KT-5823(500 nmol/L) group, BDNF+L-NAME(NG-nitro-L-arginine methyl ester)(10-4 mol/L) group,BDNF+DMSO(dimethyl sulfoxide)group;after intervention for 4 h, the expression of Sestrin2 was detected with Westernblot. HUVEC were divided into four groups: control group, BDNF(100 μg/L) group, BDNF+Sestrin2 siRNA group,BDNF+control si RNA group;after intervention for 6 h, the capacities of cell migration and tube formation were analysed.Results Sestrin2 m RNA increased in 2 h, 4 h, 6 h group compared with that of the 0 h, 1 h group(P<0. 001), whilethe protein expression of Sestrin2 increased in 2 h, 4 h, 8 h group compared with that of the 0 h, 1 h group(P<0. 05).BDNF-induced increase in Sestrin2 expression was abolished by L-NAME and PKG inhibitor(P< 0. 001). BDNF-induced cell migration and tube formation were completely blocked because of the suppressed expression of Sestrin2 by Ses-trin2 siRNA(P<0. 01). Conclusion BDNF confers certain aspects of its proangiogenic capacity through NO/PKG/Sestrin2 pathway.
作者
王碧蕾
王锦玉
金虹
夏宝妹
WANG Bilei;WANG Jinyu;JIN Hong;XIA Baomei(Faculty of Rehabilitation Science,Nanjing Normal University of Special Education,Nanjing,Jiangsu 210038,China;Department of Rehabilitation,Zhongda Hospital,Medical School of Southeast University,Nanjing,Jiangsu 210009,China;Department of Cardiology,Zhongda Hospital,Medical School of Southeast University,Nanjing,Jiangsu 210009,China)
出处
《中国动脉硬化杂志》
CAS
2021年第10期857-863,共7页
Chinese Journal of Arteriosclerosis
基金
国家自然科学基金项目(81804068)
江苏省自然科学基金项目(BK20190351)。
