姜黄素对小鼠小肠上皮细胞氧化应激损伤的保护作用及机制研究  

Protective Effects and Mechanism of Curcumin on Oxidative Damage ofMice Small Intestinal Epithelial Cells

作　　者：贺莹[1] 张夏夏 黄聪 冯彩平[1] HE Ying;ZHANG Xiaxia;HUANG Cong;FENG Caiping(Department of Biological and Food Engineering,Lvliang University,Lvliang 033000,China)

机构地区：[1]吕梁学院生物与食品工程系,吕梁033000

出　　处：《动物营养学报》2024年第7期4654-4664,共11页CHINESE JOURNAL OF ANIMAL NUTRITION

基　　金：山西省基础研究计划项目(202103021224323);山西省大学生创新项目(S202310812020);山西省大学生创新项目(S202310812029);吕梁学院大学生创新项目(CXCYZD202306)。

摘　　要：本试验旨在研究姜黄素对小鼠小肠上皮细胞(MODE-K细胞)氧化应激损伤的保护作用及机制。使用过氧化氢(H_(2)O_(2))诱导小鼠小肠上皮细胞,建立细胞氧化损伤模型,并确定最佳造模H_(2)O_(2)浓度。对照组使用正常培养液培养细胞,不进行任何处理;模型组使用正常培养液培养细胞后,加入H_(2)O_(2)处理细胞4 h;姜黄素组使用正常培养液培养细胞后,先加入不同浓度(1.25、2.50、5.00、10.00和20.00μmol/L)的姜黄素处理细胞12 h,然后再加入H_(2)O_(2)处理细胞4 h。测定小鼠小肠上皮细胞存活率、凋亡率以及细胞中活性氧(ROS)、丙二醛(MDA)含量和乳酸脱氢酶(LDH)、总超氧化物歧化酶(T-SOD)、谷胱甘肽过氧化物酶(GSH-Px)活性,实时荧光定量PCR检测B细胞淋巴瘤-2(Bcl-2)、B细胞淋巴瘤相关X蛋白(Bax)、半胱天冬蛋白酶-3(Caspase-3)的mRNA相对表达水平,利用分子模拟研究姜黄素与Kelch样环氧氯丙烷关联蛋白1(Keap1)口袋结合的优势构象和相互作用,蛋白质印迹(Western blot)检测核因子E2相关因子2(Nrf2)、Keap1、血红素加氧酶-1(HO-1)的蛋白相对表达水平。结果表明:1)最佳造模H_(2)O_(2)浓度为150μmol/L。2)与对照组相比,模型组的细胞存活率及细胞中SOD、GSH-Px活性显著降低(P<0.05),细胞中ROS、MDA含量和LDH活性显著增加(P<0.05)。与模型组相比,5.00~20.00μmol/L姜黄素组的细胞存活率及细胞中SOD、GSH-Px活性显著增加(P<0.05),细胞中ROS、MDA含量和LDH活性显著降低(P<0.05)。3)与对照组相比,模型组的细胞凋亡率显著增加(P<0.05),细胞中Bcl-2的mRNA相对表达水平显著降低(P<0.05),Bax和Caspase-3的mRNA相对表达水平显著提高(P<0.05)。与模型组相比,10.00μmol/L姜黄素组的细胞凋亡率显著降低(P<0.05),细胞中Bcl-2的mRNA相对表达水平显著提高(P<0.05),Bax和Caspase-3的mRNA相对表达水平显著降低(P<0.05)。4)分子对接结果表明,姜黄素通过与Keap1的Kelch结构域结�This experiment was conducted to study the protective effects and mechanism of curcumin on oxidative damage of mice small intestinal epithelial cells(MODE-K cells).The mice small intestinal epithelial cells were induced by H_(2)O_(2) to establish the oxidative damage model,and the optimal modeling concentration of H_(2)O_(2) was determined.The cells in the control group were cultured in a normal medium without any treatment.The cells in the model group were cultured in a normal medium and treated with H_(2)O_(2) for 4 h.The cells in the curcumin group were cultured in a normal medium and treated with curcumin of different concentrations(1.25,2.50,5.00,10.00 and 20.00μmol/L)for 12 h,and subsequently treated with H_(2)O_(2) for 4 h.The survival rate and apoptosis rate of mice small intestinal epithelial cells and reactive oxygen species(ROS),malondialdehyde(MDA)contents and lactate dehydrogenase(LDH),total superoxide dismutase(T-SOD),glutathione peroxidase(GSH-Px)activities in cells were measured,the mRNA relative expression levels of B-cell lymphoma-2(Bcl-2),B-cell lymphoma-2-associated X protein(Bax)and cysteinyl aspartate specific proteinase-3(Caspase-3)were detected by real-time fluorescence quantitative PCR,the dominant conformation and interaction of curcumin and Kelch-like epichlorohydrin-associated protein 1(Keap1)pocket combination were studied using molecular simulation,and the protein relative expression levels of nuclear factor E2-related factor 2(Nrf2),Keap1 and heme oxygenase-1(HO-1)were detected by Western blot.The results showed as follows:1)the optimal modeling concentration of H_(2)O_(2) was 150μmol/L.2)Compared with the control group,the survival rate and SOD,GSH-Px activities in cells of the model group were significantly decreased(P<0.05),and the ROS,MDA contents and LDH activity in cells were significantly increased(P<0.05).Compared with the model group,the survival rate and SOD,GSH-Px activities in cells of 5.00 to 20.00μmol/L curcumin groups were significantly increased(P<0.05),and the RO

关 键 词：姜黄素 小鼠 小肠上皮细胞 氧化应激 

分 类 号：S816.7[农业科学—饲料科学]