间歇性高浓度胰岛素促进HUECV304细胞eNOS蛋白的表达(1)
3.国家康复辅具研究中心附属康复医院，北京 100049)
摘要：目的 以人脐静脉内皮细胞(HUECV304细胞)为研究对象，研究持续性高浓度胰岛素及间歇性高浓度胰岛素对体外培育的内皮细胞内皮型一氧化氮合酶eNOS(Endothelial Nitric Oxide Synsase)蛋白表达的作用，从而进一步研究间歇性高浓度胰岛素血症导致动脉粥样硬化形成中的原因及机制。方法 采用体外培育的人脐静脉内皮细胞(HUECV304细胞)为研究对象，研究间歇性高浓度胰岛素对HUECV304细胞eNOS(Endothelial Nitric Oxide Synsase)蛋白表达水平的影响。实验分为4组，即间歇性高浓度胰岛素组(含100 nmol/L胰岛素的条件培养液及含1 nmol/L胰岛素的条件培养液轮换培养)与持续性高浓度胰岛素组(持续含100 nmol/L的胰岛素)，含正常浓度胰岛素组(持续含1nmol/L的胰岛素)、阴性对照组(不含胰岛素)。实验期间每隔8h四组同时更换新鲜条件培养液一次，四组共作用72h。采用Western immunoblotting方法检测人脐静脉内皮细胞eNOS(Endothelial Nitric Oxide Synsase)蛋白表达。结果 人脐静脉内皮细胞经过不同浓度的胰岛素作用72h后，结果显示：含正常浓度胰岛素组内皮细胞的eNOS蛋白的表达水平、间歇性高浓度胰岛素组内皮细胞的eNOS蛋白的表达水平与持续性高浓度胰岛素组内皮细胞的eNOS蛋白的表达水平均不同程度的增加，分别为阴性对照组的1.60倍(P<0.001)、3.37倍(P<0.001)和2.74倍(P<0.001)。其中以间歇性高浓度胰岛素组内皮细胞eNOS蛋白的表达水平上升更加明显，显著高于持续性高浓度胰岛素组内皮细胞的eNOS蛋白的表达水平(P<0.001)。结论 间歇性高浓度胰岛素相对于持续性高浓度胰岛素能增强人脐静脉内皮细胞(HUECV304细胞)eNOS蛋白的表达。提示间歇性高浓度胰岛素可能通过影响人脐静脉内皮细胞eNOS(Endothelial Nitric Oxide Synsase)蛋白的表达而影响血管内皮细胞的功能，从而进一步促进糖尿病慢性血管并发症的产生并发展。
, http://www.100md.com
关键词：间歇性高浓度胰岛素；人脐静脉内皮细胞；内皮型一氧化氮合酶；蛋白表达
Abstract：Objective To compare the effects of persistent and fluctuate high insulin media on protein expressions of eNOS(Endothelial Nitric Oxide Synsase)in cultured human umbilical endothelial cells line(HUECV304 cell)so as to explore the mechanism of fluctuate hyperinsulinemia on diabetic vascular complication.Methods Human umbilical vein endothelial cells(EC)line were exposed for 72 hours to 0 nmol/L insulin(control)，1 nmol/L insulin(nomal insulin)，100nmol/L alternating with 1nmol/L insulin per 8 hours(fluctuate high insulin)and 100 nmol/L insulin(persistent high insulin) respectively.The eNOS protein expression of eNOS(Endothelial Nitric Oxide Synsase)was detected by western immunoblotting.Results After HUECVs were exposed to nomal insulin，fluctuate high insulin and persistent high insulin for 72 hours，it caused a significant increase in the protein expressions of eNOS(Endothelial Nitric Oxide Synsase)by the method of western immunoblotting，the expression of eNOS protein increased to 1.60-folds(P<0.001)，3.37-folds(P<0.001)and 2.74-folds(P<0.001)of control group respectively compared with control，and the expression level of eNOS(Endothelial Nitric Oxide Synsase)protein in fluctuate high insulin was higher than that in persistent high insulin(P<0.05). Conclusion Fluctuate high insulin further enhanced protein expressions of eNOS(Endothelial Nitric Oxide Synsase)in HUECVs in vitro compared with persistent high insulin.Our data suggest that by affecting the protein expressions of eNOS(Endothelial Nitric Oxide Synsase)in HUECVs insulin fluctuation may damage endothelial cells and then facilitate the development of chronic vascular complication of diabetes., 百拇医药(黄凌云 廖斌 杨敏等)
摘要：目的 以人脐静脉内皮细胞(HUECV304细胞)为研究对象，研究持续性高浓度胰岛素及间歇性高浓度胰岛素对体外培育的内皮细胞内皮型一氧化氮合酶eNOS(Endothelial Nitric Oxide Synsase)蛋白表达的作用，从而进一步研究间歇性高浓度胰岛素血症导致动脉粥样硬化形成中的原因及机制。方法 采用体外培育的人脐静脉内皮细胞(HUECV304细胞)为研究对象，研究间歇性高浓度胰岛素对HUECV304细胞eNOS(Endothelial Nitric Oxide Synsase)蛋白表达水平的影响。实验分为4组，即间歇性高浓度胰岛素组(含100 nmol/L胰岛素的条件培养液及含1 nmol/L胰岛素的条件培养液轮换培养)与持续性高浓度胰岛素组(持续含100 nmol/L的胰岛素)，含正常浓度胰岛素组(持续含1nmol/L的胰岛素)、阴性对照组(不含胰岛素)。实验期间每隔8h四组同时更换新鲜条件培养液一次，四组共作用72h。采用Western immunoblotting方法检测人脐静脉内皮细胞eNOS(Endothelial Nitric Oxide Synsase)蛋白表达。结果 人脐静脉内皮细胞经过不同浓度的胰岛素作用72h后，结果显示：含正常浓度胰岛素组内皮细胞的eNOS蛋白的表达水平、间歇性高浓度胰岛素组内皮细胞的eNOS蛋白的表达水平与持续性高浓度胰岛素组内皮细胞的eNOS蛋白的表达水平均不同程度的增加，分别为阴性对照组的1.60倍(P<0.001)、3.37倍(P<0.001)和2.74倍(P<0.001)。其中以间歇性高浓度胰岛素组内皮细胞eNOS蛋白的表达水平上升更加明显，显著高于持续性高浓度胰岛素组内皮细胞的eNOS蛋白的表达水平(P<0.001)。结论 间歇性高浓度胰岛素相对于持续性高浓度胰岛素能增强人脐静脉内皮细胞(HUECV304细胞)eNOS蛋白的表达。提示间歇性高浓度胰岛素可能通过影响人脐静脉内皮细胞eNOS(Endothelial Nitric Oxide Synsase)蛋白的表达而影响血管内皮细胞的功能，从而进一步促进糖尿病慢性血管并发症的产生并发展。
, http://www.100md.com
关键词：间歇性高浓度胰岛素；人脐静脉内皮细胞；内皮型一氧化氮合酶；蛋白表达
Abstract：Objective To compare the effects of persistent and fluctuate high insulin media on protein expressions of eNOS(Endothelial Nitric Oxide Synsase)in cultured human umbilical endothelial cells line(HUECV304 cell)so as to explore the mechanism of fluctuate hyperinsulinemia on diabetic vascular complication.Methods Human umbilical vein endothelial cells(EC)line were exposed for 72 hours to 0 nmol/L insulin(control)，1 nmol/L insulin(nomal insulin)，100nmol/L alternating with 1nmol/L insulin per 8 hours(fluctuate high insulin)and 100 nmol/L insulin(persistent high insulin) respectively.The eNOS protein expression of eNOS(Endothelial Nitric Oxide Synsase)was detected by western immunoblotting.Results After HUECVs were exposed to nomal insulin，fluctuate high insulin and persistent high insulin for 72 hours，it caused a significant increase in the protein expressions of eNOS(Endothelial Nitric Oxide Synsase)by the method of western immunoblotting，the expression of eNOS protein increased to 1.60-folds(P<0.001)，3.37-folds(P<0.001)and 2.74-folds(P<0.001)of control group respectively compared with control，and the expression level of eNOS(Endothelial Nitric Oxide Synsase)protein in fluctuate high insulin was higher than that in persistent high insulin(P<0.05). Conclusion Fluctuate high insulin further enhanced protein expressions of eNOS(Endothelial Nitric Oxide Synsase)in HUECVs in vitro compared with persistent high insulin.Our data suggest that by affecting the protein expressions of eNOS(Endothelial Nitric Oxide Synsase)in HUECVs insulin fluctuation may damage endothelial cells and then facilitate the development of chronic vascular complication of diabetes., 百拇医药(黄凌云 廖斌 杨敏等)