羟基红花黄色素A减轻大鼠心肌细胞缺氧/复氧损伤

Hydroxysafflor yellow A reduces anoxia/reoxygenation-induced injury in rat cardiomyocytes

目的观察羟基红花黄色素A(HSYA)对乳鼠原代心肌细胞缺氧复氧(A/R)损伤的保护作用及其与磷脂酰肌醇-3-激酶/蛋白激酶B/糖原合成酶激酶3β(PI3K/Akt/GSK3β)信号通路的关系。方法分离大鼠的乳鼠心肌细胞,孵育48 h,待细胞贴壁后,分为:对照组(Con组)、缺氧复氧组(A/R组)、HSYA处理组(A/R+H组)、PI3K抑制剂(LY294002)处理组(A/R+L组)和HSYA+LY294002处理组(A/R+H+L)。测培养基上清液LDH;流式细胞仪检测凋亡率;Western blot检测Bcl-2、Bax、Akt、p-Akt(Ser473)、GSK3β和p-GSK3β(Ser9)的蛋白表达水平。结果与对照组比较,A/R后LDH释放量和凋亡率增加(P<0.001),促凋亡蛋白Bax表达增加(P<0.001),而抗凋亡蛋白Bcl-2、p-Akt(Ser473)、p-GSK3β(Ser9)蛋白表达减少(P<0.001);HSYA处理后LDH释放量和凋亡率降低(P<0.001),Bax表达减少(P<0.001),而Bcl-2、p-Akt(Ser473)和p-GSK3β(Ser9)蛋白表达增加(P<0.001);与A/R+H组比较,A/R+H+L组Bax表达增加(P<0.001),而Bcl-2、p-Akt(Ser473)和p-GSK3β(Ser9)蛋白表达减少(P<0.001)。结论 HSYA可以通过调控PI3K/Akt/GSK3β信号通路保护大鼠心肌细胞缺氧/复氧损伤。

Objective To observe the protective effect of hydroxysafflor yellow A(HSYA)on anoxia/reoxygenation(A/R)injury of neonatal primary cardiomyocytes,and its relationship with phosphoinositide 3-kinase/protein kinase B/glycogen synthase kinase 3β(PI3K/Akt/GSK3β)signaling pathway.Methods Primary cardiomyocytes of neonatal rats were isolated from the rats and incubated for 48 hours.The cells were adhered to each other and then divided into five groups:control group(Con group),anoxia/reoxygenation group(A/R group),0x09HSYA treatment group(A/R+H group),PI3K inhibitor(LY294002)treatment group(A/R+L group)and HSYA+LY294002 treatment group(A/R+H+L group),then to collect the supernatant fluid of each group to measure LDH.The flow cytometry was used to measure the apoptotic cells.The protein levels of Bcl-2,Bax,Akt,p-Akt(Ser473),GSK3β,p-GSK3β(Ser9)were evalated by Western blot.Results A/R increased LDH release,the apoptosis rate(P<0.001),and the expression of pro-apoptotic protein Bax(P<0.001)with the decrease of anti-apoptotic protein Bcl-2,p-Akt(Ser473),p-GSK3β(Ser9)(P<0.001)as compared with the control group.HSYA treatment decreased LDH release,the apoptosis rate(P<0.001),and the expression of Bax(P<0.001)and increase the expression of Bcl-2,p-Akt(Ser473),p-GSK3β(Ser9)(P<0.001).Compared with the A/R+H group,the expression of Bax was increased(P<0.001),while the expression of Bcl-2,p-Akt(Ser473),p-GSK3β(Ser9)was decreased(P<0.001)in the A/R+H+L group.Conclusions HSYA protects rats cardiomyocytes from anoxia/reoxygenation injury by regulating PI3K/Akt/GSK3βsignaling pathway.