摘要
目的探讨缺氧对培养人脐静脉内皮细胞(HUVECs)的损伤及17-β雌二醇(E2)对其保护作用。方法胰蛋白酶灌流消化法培养原代人脐静脉内皮细胞,建立细胞缺氧模型。随机分为五组:对照组、缺氧12h组、缺氧24h组、E2+缺氧12h组、E2+缺氧24h组。对照组为常氧下培养的HUVECs,缺氧组按缺氧模型予缺氧下培养HU-VECs,E2+缺氧组予预先加入17-βE2后再缺氧下培养HUVECs。收集各组细胞培养液,Greiss反应测定一氧化氮(NO),放射免疫法测内皮素-1(ET-1);收集各组HUVECs,进行细胞计数,细胞骨架染色观察细胞形态。结果缺氧12h,内皮细胞数由对照组(4.33±0.24)×106/ml减少至(3.23±0.21)×106/ml(P<0.01),形态无明显变化;其分泌NO由对照组(19.28±2.08)nmol/106cell下降至(12.77±1.90)nmol/106cell(P<0.01),ET-1由对照组(95.7±11.32)pg/ml上升至(138.3±10.37)pg/ml(P<0.01)。缺氧24h,内皮细胞数由对照组(4.33±0.24)×106/ml减少至(2.73±0.19)×106/ml(P<0.01),形态变长体积变小;其分泌NO由对照组(19.28±2.08)nmol/106cell下降至(10.76±1.57)nmol/106cell(P<0.01);ET-1由对照组(95.7±11.32)pg/ml上升至(150.1±15.41)pg/ml(P<0.01)。经预先17-βE2处理后能有效抑制上述改变(P<0.01)。结论缺氧可使内皮细胞结构和功能均受损,17-βE2对这种缺氧损伤有保护作用。
Objective To explore the effects of hypoxia on cultured human umbilical vein endothelial cells (HUVECs) and investigate the protective role of 17-β-estradiol (E_2 ) in this injury. Methods The primary cultures of HUVECs were harvested by trypsin. The hypoxic cell model was established. Five groups were randomly divided: control, hypoxic 12h, hypoxic 24h, E_2+hypoxic 12h, E_2+hypoxic 24h. The control cells were cultured under normoxia, hypoxic group were cultured under hypoxia, E_2+hypoxic group: 17-β-estradiol were added into before hypoxic culture. The culture fluid were collected for measuring nitric oxide (NO) and endothelin (ET-1) respectively by Griess reaction and radioimmunoassay. The HUVECs were subjected to cell counting and cytoskeleton staining. Results After cultured under hypoxic condition for 12h, the endothelial cells (ECs) counting decreased from (4.33±0.24)×106/ml in the control group to ( 3.23± 0.21)×106/ml(P<0.01), NO also decreased from (19.28±2.08)nmol/106cell to(12.77±1.90) nmol/106cell(P< 0.01), but the ET-1 increased from( 95.7±11.32 )pg/ml to (138.3±10.37 )pg/ml(P<0.01), the cells didn't appear significant morphy change. After cultured under hypoxic condition for 24h, the ECs counting decreased from (4.33± 0.24)×106/ml in the control group to(2.73±0.19)×106/ml (P<0.01), NO also decreased from (19.28±2.08)nmol/106cell to(10.76±1.57)nmol/106cell(P<0.01), but the ET-1 increased from( 95.7±11.32)pg/ml to (150.1± 15.41)pg/ml(P<0.01), the cells appeared longer morphy and smaller volume. After pretreated with 17-β-estradiol, these changes were markedly inhibited(P<0.01). Conclusion The structure and function of HUVECs are damaged during hypoxia and 17-β-estradiol has a protective role for this injury.
出处
《中国心血管杂志》
2005年第3期176-179,185,共5页
Chinese Journal of Cardiovascular Medicine
