羟基红花黄色素A减轻大鼠心肌细胞缺氧/复氧损伤

摘要

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 ki-nase 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),HSYA treatment group(A/R+H group),PI3K inhibitor (LY294002)treatment group(A/R+L group)and HSYA+LY294002 treat-ment group (A/R+H+L group),then to collect the supernatant fluid of each group to measure LDH.The flow cy-tometry 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 de-creased LDH release,the apoptosis rate (P<0.001),and the expression of Bax (P<0.001) and increase the ex-pression of Bcl-2,p-Akt(Ser473),p-GSK3β(Ser9)(P<0.001). Compared with the A/R+H group,the expres-sion of Bax was increased (P<0.001),while the expression of Bcl-2, p-Akt(Ser473), p-GSK3β(Ser9)was de-creased (P<0.001) in the A/R+H+L group. Conclusions HSYA protects rats'cardiomyocytes from anoxia/reoxy-genation injury by regulating PI3K/Akt/GSK3β signaling pathway.