右美托咪定预处理减轻氧糖剥夺再灌注条件下HUVECs的损伤

Dexmedetomidine preconditioning ameliorates oxygen glucose deprivation and reperfusion injury in human umbilical vein endothelial cells

目的探讨右美托咪定预处理对氧糖剥夺再灌注条件下人脐静脉内皮细胞(human umbilical vein endothelial cell,HUVECs)的保护作用。方法体外培养HUVECs,分为4组:对照组(NC组),ogd/r组(加入等体积的PBS),Dex+ogd/r组(加入50μmol/L的右美托咪定预处理2 h),Dex+YOH+ogd/r组(加入50μmol/L的右美托咪定和100μmol/L育亨宾预处理2 h)。对照组置于正常条件下培养,其他3组细胞置于Hank’s液中于缺氧恒温细胞培养箱内培养12 h,之后换正常培养基置于常氧恒温培养箱内培养4 h。采用CCK-8试剂盒检测细胞活力,取细胞培养基上清液检测LDH活力。用流式细胞仪检测细胞内活性氧水平和钙离子水平,用Rhodamine-123检测细胞线粒体膜电位的高低。结果与NC组相比,ogd/r组细胞活力明显下降(P<0.05),LDH释放水平升高(P<0.05),胞内活性氧和钙离子水平升高(P<0.05),线粒体膜电位降低(P<0.05);与ogd/r组相比,Dex+ogd/r组细胞活力升高(P<0.05),LDH释放水平降低(P<0.05),胞内活性氧和钙离子水平降低(P<0.05),线粒体膜电位升高(P<0.05);与Dex+ogd/r组相比,Dex+YOH+ogd/r组细胞活力下降(P<0.05),LDH释放水平升高(P<0.05),胞内活性氧和钙离子水平升高(P<0.05),线粒体膜电位降低(P<0.05)。结论右美托咪定预处理可以减轻氧糖剥夺再灌注造成的HUVECs损伤,可能与激活α受体有关。

Objective To investigate the effect of dexmedetomidine （Dex） preconditioning on oxygen glucose deprivation/reperfusion （ogd/r）-induced injury in human umbilical vein endothelial cells （HUVECs）. Methods Cultured HUVECs were randomly divided into 4 groups （n=12）, that is, control group （NC）, ogd/r group （PBS）, Dex＋ogd/r group （pretreatment of 50 μmol/L Dex for 2 h）, and Dex＋Yohimbine （YOH） ＋ogd/r group （pretreatment of 50 μmol/L Dex and 100 μmol/L YOH for 2 h）. The control group were cultured under normal condition, while the culture media of the other 3 groups were replaced with Hank’s solution and the cells were placed in a hypoxia incubator for 12 h followed by normal condition for 4 h. CCK-8 assay was used to detect cell viability, and the activity of lactate dehydrogenase （LDH） in the supernatant was measured with LDH kit. Flow cytometry was adopted to detect the intracellular levels of reactive oxygen species （ROS） and calcium ion. Rhodamine-123 was employed to measure the trans-membrane potential of mitochondrion. Results Compared with the NC group, the cells in the ogd/r group had significantly decreased viability （P〈0.05）, increased LDH level （P〈0.05）, elevated intracellular ROS and calcium ion levels （P〈0.05）, and reduced trans-membrane potential of mitochondrion （P〈0.05）. While, the cells in the Dex＋ogd/r group had significantly stronger cell viability （P〈0.05）, less secretion of LDH （P〈0.05）, decreased intracellular levels of ROS and calcium ion （P〈0.05）, and increased trans-membrane potential of mitochondrion （P〈0.05） when compared with the ogd/r group. In the Dex＋YOH＋ogd/r group, cell viability was decreased, LDH level was increased, the intracellular levels of ROS and calcium ion were elevated, but the trans-membrane potential of mitochondrion was significantly decreased when compared with the Dex＋ogd/r group （all P〈0.05）. Conclusion Dex preconditioning can effectively attenuate ogd/r-induced injury in HUVECs, which may be related to activation of alpha 2 adrenergic receptor.