目的:探究王不留行黄酮苷(VAC)对2型糖尿病(T2DM)内皮功能障碍的治疗作用及机制研究.方法:(1)通过腹腔注射链脲佐菌素和喂饲高脂饲料(21.8 kJ/kg,60%能量为脂肪)构建T2DM小鼠模型.将30只C57BL/6小鼠随机分为对照组、T2DM组和T2DM+VAC组,每组10只.T2DM+VAC组小鼠给予1 mg/kg VAC灌胃6周,对照组和T2DM组小鼠均给予等体积PBS.RT-qPCR和Western blot检测胸主动脉中BCL2相互作用蛋白3(BINIP3)、PTEN诱导激酶1(PINK1)和parkin的mRNA和蛋白表达.(2)在体外通过高糖(HG)刺激人脐静脉内皮细胞(HUVECs),用JC-1染色检测线粒体膜电位变化,吖啶橙(AO)染色检测自噬溶酶体变化,MitoSOX染色检测线粒体超氧化物的积累.结果:与对照组相比,T2DM小鼠胸主动脉BNIP3、PINK1和parkin的mRNA和蛋白表达水平显著升高(P<0.05);与T2DM组相比,T2DM+VAC组小鼠胸主动脉中BNIP3、PINK1和parkin的mRNA和蛋白表达水平显著降低(P<0.05).JC-1、AO和MitoSOX染色结果显示,VAC可抑制HG诱导的HUVECs线粒体膜电位降低、自噬溶酶体增多和线粒体超氧化物水平升高.VAC也可减轻BNIP3过表达后HG诱导的HUVECs线粒体损伤.微小RNA-570-3p(miR-570-3p)mimic与VAC有相似的减轻线粒体损伤作用.RT-qPCR和Western blot结果显示,miR-570-3p mimic和VAC都使BNIP3、PINK1和parkin的mRNA和蛋白水平显著降低.使用miR-570-3p抑制剂后,VAC的上述作用均被抑制.结论:VAC通过miR-570-3p/BNIP3减轻HG诱导的线粒体损伤,从而缓解T2DM内皮功能障碍.
AIM:To investigate the effect of vaccarin(VAC)on endothelial dysfunction in type 2 diabetes mellitus(T2DM),and to uncover the underlying mechanisms.METHODS:(1)C57BL/6 mice received intraperitoneal injection of streptozotocin and were fed with a high-fat diet(21.8 kJ/kg,60%of the energy source was fat)to construct a T2DM mouse model.Thirty mice were randomly divided into control,T2DM and T2DM+VAC groups,with 10 mice in each group.The mice in T2DM+VAC group were given 1 mg/kg VAC via oral gavage for 6 weeks,while those in control and T2DM groups were given the same volume of PBS.The mRNA and protein expression levels of BCL2-interacting pro-tein 3(BNIP3),PTEN-induced kinase 1(PINK1)and parkin in the thoracic aorta were detected by RT-qPCR and West-ern blot.(2)Human umbilical vein endothelial cells(HUVECs)were stimulated by high glucose(HG;35 mmol/L glu-cose).Mitochondrial membrane potential,autophagy and mitochondrial superoxide levels were detected using JC-1,acri-dine orange(AO)and MitoSOX staining,respectively.RESULTS:Compared with control group,the mRNA and protein levels of BNIP3,PINK1 and parkin were significantly increased in the thoracic aorta of T2DM mice(P<0.05).Compared with T2DM group,the mRNA and protein levels of BNIP3,PINK1 and parkin in the thoracic aorta were significantly re-duced in T2DM+VAC group(P<0.05).The results of JC-1,AO and MitoSOX staining showed that VAC attenuated the decrease in mitochondrial membrane potential and the increase in autophagy and mitochondrial superoxide levels in HG-in-duced HUVECs.Treatment with VAC also inhibited HG-mediated mitochondrial damage in HUVECs after BNIP3 overex-pression.The effect of miR-570-3p mimic on mitochondrial damage was similar to VAC.RT-qPCR and Western blot showed that both miR-570-3p mimic and VAC significantly reduced the mRNA and protein levels of BNIP3,PINK1 and parkin.In contrast,inhibition of miR-570-3p exhibited the opposite effects.CONCLUSION:Treatment with VAC alle-viated endothelial dysfunction in T2DM by inhibiting HG-induced mitochondrial dysfunction through miR-570-3p/BNIP3.
赵晨阳;朱雪雪;陈欣雨;陈天笑;徐锦朋;李泰悦;曹幸予;张源;邱丽颖
江南大学无锡医学院,江苏 无锡 214122
临床医学
王不留行黄酮苷;2型糖尿病;内皮功能障碍;BNIP3蛋白;微小RNA-570-3p;线粒体损伤
vaccarin;type 2 diabetes mellitus;endothelial dysfunction;BNIP3 protein;microRNA-570-3p;mitochondrial damage
《中国病理生理杂志》 2024 (005)
872-881 / 10
国家自然科学基金资助项目(No.82300414);江苏省研究生科研与实践创新计划(No.KYCX24_2651)
10.3969/j.issn.1000-4718.2024.05.012
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