Objective To investigate effects of Kv1.5 protein on LPS-induced peroxidation injury in endothelial cells.Methods Human umbilical vein endothelial cells(HUVECs)were cultured in vitro.Sepsis model was established by stim-ulating HUVECs with LPS(5 μ/mL).The experiment was divided into four groups :Control group ,LPS group ,LPS+ MT1 group(250 nmol/L of MT was used for pretreatment for 30 min to block Kv1.5 channels)and LPS+ MT2(500 nmol/L MT). Then the survival rate of endothelial cells was detected by MTT assay.ELISA was used to detect the changes of E-selectin and ICAM-1 in the supernatant ,and to measure MDA and SOD activity. Results Compared with the control group ,the survival rate of endothelial cells in LPS group was significantly decreased [(32.54 ± 1.87)% vs. (98.64 ± 0.92)%,P＜0.01] ,while the sur-vival rate of LPS+ MT1 group and LPS+ MT2 group was(77.39 ± 1.15)% and(93.22 ± 1.15)%,which was significantly higher than that of LPS group(P＜0.01).At the same time ,the secretion of E-selectin and ICAM-1 in endothelial cells of LPS group increased(P＜0.01) ,and decreased after pretreatment with 250 and 500 nmol/L MT(P＜0.01).By observation with la-ser confocal microscopy ,intracellular ROS level in endothelial cells in LPS group were increased to (3.39 ± 0.42)times as much as that of the blank control group(P＜ 0.05) ,and after pretreatment with 250 and 500 nmol/L MT ,ROS level was(1.88 ± 0.36)times and(1.46 ± 0.47)times as much as that of the blank control group ,respectively ,which was significantly different from that of LPS group(P＜0.05).Compared with the blank control group ,MDA level was significantly increased and SOD ac-tivity decreased in LPS group(P＜0.01) ,while MT pretreatment significantly reduced MDA content and increased SOD activity (P＜0.01).Conclusion Kv1.5 protein is closely related to LPS-induced injury of vascular endothelial cells.MT can reduce lipid peroxidation injury of endothelial cells by blocking Kv1.5 channel.%目的 研究Kv1.5蛋白对内毒素(LPS)所致血管内皮细胞氧化应激损伤的影响.方法 体外培养人脐静脉内皮细胞(HUVECs) ,给予5 μg/mL LPS刺激建立脓毒症细胞模型,并分为:空白对照组、LPS组、LPS+ MT1组(以Kv1.5特异性通道阻滞剂MT 250 nmol/L预处理30 min)及LPS+MT2组(MT浓度为500 nmol/L).采用MTT法检测内皮细胞存活率;ELISA法检测细胞上清液中内皮细胞E-选择素(E-selectin)、细胞间粘附分子(ICAM-1)浓度变化;激光共聚焦显微镜观察内皮细胞内ROS水平;比色法检测脂质过氧化物丙二醛(MDA)含量及超氧化物歧化酶(SOD)活性.结果 与空白对照组比较,LPS组内皮细胞存活率明显下降[(32.54 ± 1.87)% vs. (98.64 ± 0.92)%,P＜0.01] ,而LPS+MT1组及LPS+MT2组内皮细胞存活率分别为(77.39 ± 1.15)%、(93.22 ± 1.15)%,均较LPS组上升(均 P＜0.01);LPS组内皮细胞 E-selectin 、ICAM-1分泌增加(均 P＜0.01) ,予以250 、500 nmol/LMT 预处理后,E-selectin 、ICAM-1分泌减少(均 P＜0.01);激光共聚焦显微镜观察内皮细胞内ROS水平发现,LPS组细胞内ROS水平升高为空白对照组的(3.39 ± 0.42)倍(P＜ 0.05) ,予以250 、500 nmol/LMT 预处理后,ROS水平分别为空白对照组的(1.88 ± 0.36)倍、(1.46 ± 0.47)倍,与LPS组比较,差异有统计学意义(均 P＜0.05);且LPS组较空白对照组MDA 水平上升、SOD活性降低(均 P＜0.01) ,而MT预处理可降低MDA含量、增加SOD活性(均 P＜0.01).结论 Kv1.5蛋白与内毒素诱导的血管内皮细胞损伤密切相关;MT通过阻断Kv1.5通道可减轻内皮细胞脂质过氧化损伤.
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