Page 54 - 《中国药房》2020年23期

Study on the Anti-apoptotic Effect of Curcumin on H2O2-induced H9c2 Cardiomyocyte Injury and the
Regulation of NF-κB Signaling Pathway
GU Changhai ，LI Zhong ，CHEN Dongling （1. Physical Education Department， Nanyang Institute of
2
1
2
Technology，Henan Nanyang 473004，China；2. Zhang Zhongjing College of Traditional Medical Medicine，
Nanyang Institute of Technology，Henan Nanyang 473004，China）
ABSTRACT OBJECTIVE：To investigate the anti-apoptotic effect of curcumin on H2O2-induced H9c2 cardiomyocyte injury and
the regulation of NF-κB signaling pathway. METHODS：H9c2 cardiomyocyte were randomly divided into normal control group，
injury model group，curcumin low-dose，medium-dose and high-dose groups（25，50，100 μmol/L）. Normal control group didn’t
received any intervention. The cells in injury model group were induced with 50 μmol/L H2O2 for 12 h to establish the injury model.
The cells in curcumin groups were treated with relevant concentration of drugs for 24 h，and then induced with 50 μmol/L H2O2 for
12 h. After cultured for 24 h，survival rate and apoptotic rate of cells were measured by MTT method and TUNEL method；SOD
activity and MDA content were determined by WTS-8 assay and color test；relative fluorescence intensity of LC3 positive
expression was detected by immunofluorescence method；mRNA expression of NF-κB p65 in cells was detected by real-time PCR；
Western blotting assay was used to detect the protein expression of NF-κB p65 and p-NF-κB p65 in cells. RESULTS：Compared
with normal control group，survival rate and SOD activity were decreased significantly in injury model group，while apoptotic rate，
MDA content，relative fluorescence intensity of LC3 positive expression，mRNA expression of NF-κB，protein expression of NF-κ
B p65 and p-NF-κB p65 as well as p-NF-κB/NF-κB were increased significantly（P＜0.05）. Compared with injury model group，
survival rates and SOD activities were increased significantly in curcumin groups，while apoptotic rates，MDA contents，relative
fluorescence intensities of LC3 positive expression，mRNA expression of LC3 positive cells，protein expression of NF-κB p65 and
p-NF-κ B p65 as well as p-NF-κ B p65/NF-κ B p65 were decreased significantly （P＜0.05）. CONCLUSIONS：Curcumin can
increase the survival rate of H2O2-induced H9c2 cardiomyocyte injury，decrease its apoptotic rate，increase SOD activity and
decrease MDA content in cardiomyocytes. Above effects may be related to the regulation of NF-κB signaling pathway.
KEYWORDS Curcumin；H9c2 cells；Cardiomyocyte injury；Anti-apoptotic effect；NF-κB signaling pathway

心血管疾病严重危害人类健康，是目前病死率最高 仪[赛默飞世尔科技（中国）有限公司]；ECO 48型聚合酶
[1]
的疾病 。心血管疾病发生机制非常复杂，心肌细胞损 链式反应（PCR）仪（英国 PCRmax 公司）；MP4 型垂直电
伤是各种心血管疾病的病理生理学基础 。有研究指 泳 仪 、ChemiDocTouch 型 化 学 发 光 成 像 系 统（美 国
[2]
出，氧化应激可引起心肌细胞凋亡，损伤心肌细胞功能 Bio-Rad 公司）；XSP-12CAC 型光学显微镜（上海缔伦光
[2]
并引发心力衰竭等心血管系统疾病 。核因子κB（NF- 学仪器有限公司）；TCS SP5 型激光扫描共聚焦显微镜
κB）信号通路是调节体内多种组织细胞损伤及氧化应激 （德国 Leica 公司）；QL-9001 型振荡器（海门市麒麟医用
状态的重要信号通路。已有研究证实，NF-κB信号通路 仪器厂）。
的异常激活与心肌细胞损伤的发生密切相关 [3-4] 。姜黄 1.2 药品与试剂
素是从姜科、天南星科植物的根茎中提取得到的化合 姜黄素对照品（成都曼思特生物科技有限公司，批
[5]
物，具有抗炎、抗氧化应激等多种药理作用 。近年来研 号：20191022，纯度：≥96％）；MTT 检测试剂盒（北京群
究表明，该化合物对心肌细胞损伤具有显著的改善和修 晓科苑生物技术有限公司，批号：YS171211A）；反转录
[6]
复作用 ，但其对受损心肌细胞的保护作用及 NF-κB 信 试剂盒（日本Takara公司，批号：180223CC）；反转录聚合
号通路的影响目前尚未见明确报道。为此，本研究拟初 酶链反应（RT-PCR）试剂盒（南京威特曼生物技术有限
步探讨姜黄素对 H2O2致心肌细胞损伤的抗凋亡作用以 公司，批号：180115001）；细胞裂解液（南京碧云天生物
及其对NF-κB信号通路的调节作用，旨在为该化合物的 技术研究所，批号：180221A）；胰酶细胞消化液（批号：
临床应用及心肌细胞损伤的药理作用研究提供参考 C0201）、免疫染色洗涤液（批号：P0106）、二喹啉甲酸
依据。 （BCA）蛋白浓度测定试剂盒（批号：P0012）、TUNEL 细
1 材料 胞凋亡检测试剂盒（批号：C1088）、超氧化物歧化酶
1.1 仪器 （SOD）活性检测试剂盒（WST-8法）（批号：S0101）、丙二
Gallios型流式细胞仪（美国Beckman Coulter公司）； 醛（MDA）检测试剂盒（批号：S0131S）、NF-κB p65 兔单
Multiskan FC 型酶标仪（美国 Thermo Fisher Scientific 公 克隆抗体（批号：AF1234）、磷酸化 NF-κB p65（p-NF-κB
司）；HERAcell 型 CO2 培 养 箱 、Multiskan Sky 型 酶 标 p65）兔单克隆抗体（批号：AF5881）和GAPDH兔单克隆

·2864 · China Pharmacy 2020 Vol. 31 No. 23 中国药房 2020年第31卷第23期