Li Lingmei, Cao Ce, Guo Hao, Lin Li, Li Lei, Zhang Yehao, Xin Gaojie, Liu Zixin, Xu Shujuan, Han Xiao, Zhang Qiong, Fu Jianhua
Kunshan Hospital of Chinese Medicine, Kunshan, China.
Institute of Basic Medical Sciences of Xiyuan Hospital, Beijing Key Laboratory of Chinese Materia Pharmacology, China Academy of Chinese Medical Sciences, National Clinical Research Center of Traditional Chinese Medicine for Cardiovascular Diseases, Beijing, China.
BMC Complement Med Ther. 2025 Jan 9;25(1):9. doi: 10.1186/s12906-025-04747-8.
This study intended to explore whether the protective effect safflower yellow injection (SYI) on myocardial ischemia-reperfusion (I/R) injury in rats mediated of the NLRP3 inflammasome signaling.
The I/R model was prepared by ligating the left anterior descending coronary artery for 45 min and then releasing the blood flow for 150 min. 96 male Wistar rats were randomly divided into sham group, I/R group, Hebeishuang group (HBS), SYI high-dose group (I/R + SYI-H), SYI medium-dose group (I/R + SYI-M) and SYI low-dose group (I/R + SYI-L). Cell experiments were divided into normal control group (NC), Oxygen glucose deprivation/reoxygenation group (OGD/R), OGD/R + SYI group, OGD/R + SYI + Chloroquine group (OGD/R + SYI + CQ). The area of myocardial ischemia infarction and pathological changes were observed by the Tetrazolium method (TTC) and HE staining. Myocardial enzymes such as aspartate aminotransferase (AST), lactate dehydrogenase (LDH) and creatine kinase (CK) were measured by chemiluminescence (CL) method. The inflammatory factors levels of TNF-α, IL-1β, MCP-1, and IL-6 were detected by ELISA. The expressions of inflammatory-related proteins (Caspase-1, NLRP3, TLR4, NF-κB), autophagosome-related proteins (LC3-I, LC3-II,LC3-II/LC3-I), apoptosis-related proteins (Bax, Bcl-2, Caspase-3, Bcl-2/Bax) and autophagy-related proteins (p62/SQSTM1, PI3K, p-Akt, mTOR) were detected by Western-Blot. Cell morphology and cell viability were detected by transmission electron microscopy and CCK-8.
In vivo, compared with sham group, the percentage of myocardial infarction area was increased and myocardial tissue arrangement was disordered in I/R group. In addition, the activities of myocardial enzymes, the contents of inflammatory factors, the expressions of inflammatory-related proteins, autophagy-related proteins, autophagosome-related proteins, Bax and Caspase-3 were increased, while Bcl-2 and Bcl-2/Bax were decreased. SYI treatment reversed these trends, except for the expression of autophagosome-related proteins. In vitro, SYI decreased the contents of inflammatory factors and the expressions of inflammatory-related proteins, autophagy-related proteins and autophagosome-related proteins caused by OGD/R. However, the contents of inflammatory factors and the expression of inflammatory-related proteins, p62/SQSTM1 and mTOR were increased, while PI3K, p-AKT, LC3-II/LC3-I were significantly decreased in OGD/R + SYI + CQ group.
SYI can promote myocardial tissue autophagy by regulating NLRP3, thereby attenuating the myocardial inflammatory response and protecting damaged myocardium in I/R rats.
本研究旨在探讨红花黄色素注射液（SYI）对大鼠心肌缺血再灌注（I/R）损伤的保护作用是否通过NLRP3炎性小体信号介导。
通过结扎左冠状动脉前降支45分钟，然后再灌注150分钟制备I/R模型。将96只雄性Wistar大鼠随机分为假手术组、I/R组、河北双组（HBS）、SYI高剂量组（I/R + SYI-H）、SYI中剂量组（I/R + SYI-M）和SYI低剂量组（I/R + SYI-L）。细胞实验分为正常对照组（NC）、氧糖剥夺/复氧组（OGD/R）、OGD/R + SYI组、OGD/R + SYI + 氯喹组（OGD/R + SYI + CQ）。采用四氮唑法（TTC）和苏木精-伊红染色（HE染色）观察心肌缺血梗死面积和病理变化。采用化学发光法（CL）检测天冬氨酸转氨酶（AST）、乳酸脱氢酶（LDH）和肌酸激酶（CK）等心肌酶。采用酶联免疫吸附测定法（ELISA）检测肿瘤坏死因子-α（TNF-α）、白细胞介素-1β（IL-1β）、单核细胞趋化蛋白-1（MCP-1）和白细胞介素-6（IL-6）等炎症因子水平。采用蛋白质免疫印迹法（Western-Blot）检测炎症相关蛋白（半胱天冬酶-1（Caspase-1）、NLRP3、Toll样受体4（TLR4）、核因子-κB（NF-κB））、自噬体相关蛋白（微管相关蛋白轻链3-I（LC3-I）、微管相关蛋白轻链3-II（LC3-II）、LC3-II/LC3-I）、凋亡相关蛋白（Bax、Bcl-2、半胱天冬酶-3（Caspase-3）、Bcl-2/Bax）和自噬相关蛋白（p62/ sequestosome 1（SQSTM1）、磷脂酰肌醇-3激酶（PI3K）、磷酸化蛋白激酶B（p-Akt）、哺乳动物雷帕霉素靶蛋白（mTOR））的表达。采用透射电子显微镜和细胞计数试剂盒-8（CCK-8）检测细胞形态和细胞活力。
在体内，与假手术组相比，I/R组心肌梗死面积百分比增加，心肌组织排列紊乱。此外，心肌酶活性、炎症因子含量、炎症相关蛋白、自噬相关蛋白、自噬体相关蛋白、Bax和Caspase-3的表达增加，而Bcl-2和Bcl-2/Bax降低。SYI治疗逆转了这些趋势，但自噬体相关蛋白的表达除外。在体外，SYI降低了OGD/R引起的炎症因子含量以及炎症相关蛋白、自噬相关蛋白和自噬体相关蛋白的表达。然而，在OGD/R + SYI + CQ组中，炎症因子含量以及炎症相关蛋白、p62/SQSTM1和mTOR的表达增加，而PI3K、p-AKT、LC3-II/LC3-I显著降低。
SYI可通过调节NLRP3促进心肌组织自噬，从而减轻I/R大鼠的心肌炎症反应，保护受损心肌。
