Ghrelin 预处理对 H9C2心肌细胞缺氧/复氧损伤的影响

目的：探讨 Ghrelin 预处理对 H9C2心肌细胞缺氧/复氧损伤的影响。方法培养 H9C2心肌细胞并建立心肌细胞缺氧(21 h)/复氧(6 h)模型。细胞随机分成4组：对照组,缺氧/复氧组(H/ R 组),缺氧/复氧+ Ghrelin 组(H/ R +G 组),缺氧/复氧+ Ghrelin +生长激素促分泌素受体(GH-SR)-siRNA 组(H/ R + G + G-siRNA 组)；采用 siRNA 干扰技术阻断 Ghrelin 受体 GHSR 的表达以及 Hoechst 33258染色剂和流式细胞术检测各组细胞凋亡率,Western blot 法检测各组凋亡相关蛋白表达情况。结果 GHSR 存在于正常H9C2心肌细胞中,GHSR-siRNA 可以显著抑制 H9C2心肌细胞中 GHSR 的表达。与 H/ R 组相比,在 H/ R + G 组中缺氧/复氧损伤所导致心肌细胞的凋亡率明显下降(P <0．05),B淋巴细胞瘤-2蛋白/ Bcl-2相关 X 蛋白(Bcl-2/ Bax)比率明显上调(P <0．05),含半胱氨酸的天冬氨酸蛋白水解酶-3(caspase-3)表达水平被显著抑制(P <0．05)。而与 H/ R + G组相比,H/ R + G + G-siRNA 组心肌细胞凋亡率增加( P <0．05),Bcl-2/ Bax 比率显著下降(P <0．05),caspase-3表达上调(P <0．05)。结论 Ghrelin 具有减轻缺氧/复氧所引起的心肌细胞损伤、抑制心肌细胞凋亡的作用。

Effect of pretreatment with Ghrelin on hypoxia/reoxygenation injury in H9C2 cells

Objective To determine the effect of pretreatment with Ghrelin on hypoxia / reoxygenation (H/ R) inju-ry in rat cardiomyocytes. Methods Cultured H9C2 cells were randomly divided into 4 groups, including control group , H / R group , H / R + Ghrelin group , and H / R + Ghrelin + growth secretagogue receptor (GHSR)-siRNA group. H/ R injury was established by 21 hours of hypoxia and subsequent 6 hours of reoxygenation. Expression of growth GHSR in H9C2 cells was reduced by siRNA. Apoptosis of H9C2 cardiomyocytes under different treatments following H/ R injury was observed by Hoechst 33258 staining and apoptotic rate was detected by flow-cytometry a-nalysis. Besides, Western blot analysis was performed to detect the expression of the apoptosis related proteins in each group. Results GHSR was expressed in H9C2 cells and the expression of GHSR could be significantly knocked down by GHSR-siRNA. Compared with H/ R group, in H/ R + Ghrelin group the myocardium apoptotic rate was markedly decreased (P < 0. 05), the Bcl-2 / Bax ratio was increased (P < 0. 05) and the expression of caspase-3 protein was decreased (P < 0. 05). However, compared with H/ R + Ghrelin group, the apoptotic rate was increased(P < 0. 05), the Bcl-2 / Bax ratio was decreased (P < 0. 05) and the expression of caspase-3 protein was up-regulated (P < 0. 05) in H/ R + Ghrelin + GHSR-siRNA group. Conclusion Ghrelin provides concrete pro-tection against H/ R injury by inhibiting apoptosis in cardiomyocytes.