乐果对大鼠肝细胞凋亡作用的研究(英文)

Studies on the Effects of Dimethoate on Rat Hepatic cell Apoptosis

通过给大鼠肝细胞培养液中加入乐果（染毒终浓度分别为0、3、10、30、100和300μmol·L^-1）,染毒12、24h后,AnnexinV/PI双染法检测肝细胞凋亡率;分别用Fluo-2/AM、双氢-乙酰乙酸二氯荧光黄（DCFH-DA）和罗丹名123检测细胞内Ca^2＋浓度、活性氧（ROS）和线粒体膜电位（Δψm）变化,并在扫描电镜和荧光显微镜下观察凋亡细胞情况,研究乐果对大鼠肝细胞凋亡的影响。结果表明,肝细胞染毒12和24h后,出现了明显的细胞凋亡的形态学变化,细胞凋亡率明显升高,除3μmol·L-1组外,与对照组相比差异显著（P〈0.05或P〈0.01）,且呈时间-剂量效应。3μmol·L^-1组细胞内Ca2＋浓度极显著高于对照组（P〈0.01）,之后随染毒剂量的增加,细胞内Ca^2＋浓度逐渐下降;细胞内ROS水平在3-100μmol·L^-1范围内随染毒剂量的增大和染毒时间的延长而升高,而在300μmol·L^-1组略有下降,除3μmol·L^-1组外,与对照组相比均差异极显著（P〈0.01）;Δψm除24h300μmol·L^-1组外均出现持续下降。表明低剂量乐果染毒可诱导肝细胞发生凋亡,细胞内Ca^2＋、ROS和Δψm可能参与了这一过程。

In order to study the effects of Dimethoate on liver cell apoptosis in vitro, Dimethoate was added to rat hepatocyte culture fluid to get final concentration of 0, 3, 10, 30, 100 and 300μmol·L^-1 in each group. After 12 and 24 hours＇ exposure, apoptotic cells were observed by electron and fluorescence microscope, apoptosis rate was analyzed by Annexin V/PI method; Intracellular Ca^2＋ concentration, reactive oxygen species （ROS） and mitochondria membrane potential （Δψm） changes were analyzed by Fluo-2/AM, Dichlorofluorescin diacetate （DCFH-DA） and Rhodamine 123 method respectively. The results showed that after 12 and 24 hours＇ exposure, microstructural changes of apoptotic cells were observed; hepatic cell apoptosis rate increased, and the difference between experimental groups （apart from 3 μmol·L^-1 group） and control group was statistically significant （P〈0.05 or P〈0.01）; an obvious dosage-effect and time-effect were also observed. Intracellular Ca^2＋ eoncentration of 3μmol·L^-1 group was significantly （P〈0.01） higher than that of control group, but then decreased with increasing dosage. Intracellular ROS level increased with increasing dosage and time of exposure in the range of 3-100 μmol·L^-1; however, ROS level decreased slightly in 300 μmol·L^-1 group, and the difference between experimental groups （apart from 3μmol·L^-1 group） and control group was significant （P〈0.01）. Mitochondria membrane potential （Δψm） decreased continuously after 12 and 24 hours＇ exposure apart from the 300 μmol·L^-1 group with 24 h exposure. It was concluded that low dosage of Dimethoate can induce liver cell apoptosis, and intracellular Ca^2＋, ROS and Δψm changes may participate in this process.