Department of Cardiology, Yantaishan Hospital, Yantai 264001, PR China.
Department of Cardiology Group Two, Weihai Municipal Hospital, Weihai 264200, PR China.
Biomed Pharmacother. 2016 Dec;84:106-114. doi: 10.1016/j.biopha.2016.09.014. Epub 2016 Sep 16.
To determine the mechanism by which Tanshinone IIA (Tan IIA) relieves myocardial ischemia reperfusion injury (MIRI) in rats via the PI3K/Akt/mTOR signaling pathway.
Sprague-Dawley (SD) rats received an intravenous injection of Tan IIA and LY294002 and were divided into the sham, control (myocardial ischemia reperfusion), Tan-L (low-dose Tan IIA), Tan-H (high-dose Tan IIA), Tan-L+LY (low-dose Tan IIA+LY294002), Tan-H+LY (high-dose Tan IIA+LY294002) and LY (LY294002) groups. Cardiomyocytes obtained from neonatal rats were treated with hypoxia reoxygenatin, Tan IIA and LY294002 and divided into the blank, control, Tan-L, Tan-H, Tan-L+LY, Tan-H+LY and LY groups. Creatine kinase MB isoenzyme (CK-MB) and lactic dehydrogenase (LDH) levels in serum and cardiomyocytes were measured. Area of necrosis/area at risk (AN/AAR) was determined with double staining of TTC and Evan's blue; viability and apoptosis of cardiomyocytes with MTT and TUNEL assays; SOD, MDA, HO, SDH and COX levels in heart mitochondria together with PI3K/Akt/mTOR and eNOS expressions and phosphorylation with Western blotting.
The Tan-L and Tan-H groups showed a remarkable decrease in AN/AAR, serum CK-MB and LDH, mitochondrial MDA and HO levels but an increase in SOD activity, SDH and COX levels compared with the control group. However, compared with the Tan-L and Tan-H groups, the Tan-L+LY, Tan-H+LY and LY groups indicated an inverse tendency of those indicators. As shown by MTT and TUNEL, the control group had more severe cell damage than the blank group. Furthermore, cell damage and apoptosis were less severe in the Tan-L and Tan-H groups than in the control group, while the Tan-L+LY, Tan-H+LY and LY groups showed an opposite tendency when compared with the Tan-L and Tan-H groups. Meanwhile, the Tan-L and Tan-H groups showed significantly higher expression levels of PI3K, p-Akt/Akt, mTOR and p-eNOS/eNOS than the control group, whereas the Tan-L+LY, Tan-H+LY and LY groups had lower expression levels than the Tan-L and Tan-H groups.
Our study provided evidence that Tan IIA could activate the PI3K/Akt/mTOR signaling pathway to relieve MIRI in rats.
通过 PI3K/Akt/mTOR 信号通路，确定丹参酮 IIA（Tan IIA）缓解大鼠心肌缺血再灌注损伤（MIRI）的机制。
SD 大鼠静脉注射 Tan IIA 和 LY294002，并分为假手术组、对照组（心肌缺血再灌注）、Tan-L 组（低剂量 Tan IIA）、Tan-H 组（高剂量 Tan IIA）、Tan-L+LY 组（低剂量 Tan IIA+LY294002）、Tan-H+LY 组（高剂量 Tan IIA+LY294002）和 LY 组（LY294002）。从新生大鼠中分离的心肌细胞接受缺氧复氧、Tan IIA 和 LY294002 处理，并分为空白组、对照组、Tan-L 组、Tan-H 组、Tan-L+LY 组、Tan-H+LY 组和 LY 组。测定血清和心肌细胞中肌酸激酶同工酶（CK-MB）和乳酸脱氢酶（LDH）水平。用 TTC 和 Evans 蓝双重染色法测定坏死/危险区面积（AN/AAR）；MTT 和 TUNEL 法测定心肌细胞活力和凋亡；Western blot 法测定心脏线粒体中超氧化物歧化酶（SOD）、丙二醛（MDA）、羟自由基（HO）、琥珀酸脱氢酶（SDH）和细胞色素 c 氧化酶（COX）水平以及 PI3K/Akt/mTOR 和 eNOS 的表达和磷酸化。
与对照组相比，Tan-L 组和 Tan-H 组的 AN/AAR、血清 CK-MB 和 LDH、线粒体 MDA 和 HO 水平显著降低，SOD 活性、SDH 和 COX 水平显著升高。然而，与 Tan-L 组和 Tan-H 组相比，Tan-L+LY、Tan-H+LY 和 LY 组的这些指标呈现出相反的趋势。与空白组相比，MTT 和 TUNEL 结果表明对照组的细胞损伤更严重。此外，与对照组相比，Tan-L 组和 Tan-H 组的细胞损伤和凋亡程度较轻，而 Tan-L+LY、Tan-H+LY 和 LY 组与 Tan-L 组和 Tan-H 组相比则呈现出相反的趋势。同时，与对照组相比，Tan-L 组和 Tan-H 组的 PI3K、p-Akt/Akt、mTOR 和 p-eNOS/eNOS 表达水平显著升高，而 Tan-L+LY、Tan-H+LY 和 LY 组的表达水平则低于 Tan-L 组和 Tan-H 组。
本研究提供的证据表明，Tan IIA 可通过激活 PI3K/Akt/mTOR 信号通路来缓解大鼠的 MIRI。
