Feng Jing, Chen Bao-Yuan, Guo Mei-Nan, Cao Jie, Zhao Hai-Yan, Liang Dong-Chun, Zuo Ai-Jun
Respiratory Department, Tianjin Medical University General Hospital, Tianjin 300052, China.
Zhonghua Yi Xue Za Zhi. 2007 Mar 20;87(11):774-7.
To investigate the damage of different patterns of intermittent hypoxia (IH) and continuous hypoxia (CH) on endothelial cells.
Human umbilical vein endothelial cells of the line ECV304 were cultured in a program-controlled gas delivery system newly developed and divided into 8 groups to undergo different IH/reoxygenation (ROX) cycles so as to simulate the patterns of hypoxic episode seen in recurrent apnea and chronic obstructive pulmonary disease: intermittent normoxia (IN) group (exposed to 21% O2 15 s/21% O2 225 s for 60 cycles), IH group (exposed to 1.5% O2 15 s/21% O2 225 s for 30 or 60 cycles), IH hypercapnia group (exposed to 1.5% O2 and 20% CO2 15 s/21% O2 and 5% CO2 225 s, for 60 cycles), continuous hypoxia (CH group, exposed to 1.5% or 10% O2 for 15, 30 or 60 min), CH hypercapnia group (exposed to 10% O2 and 10% CO2 for 15, 30 or 60 min), CH added to IH group (exposed to 1.5% O2 15 s/10% O2 225 s for 60 cycles), different intermittent hypoxia extent group (exposed to 1.5% or 10% O2 15 s/21% O2 225 s for 60 cycles), different intermittent hypoxia frequency group (exposed to 1.5% O2 15 s/21% O2 225 s 315 s, 495 s or 105 s for 60 cycles), and different intermittent hypoxia duration group (exposed to 1.5% O2 15 s or 30 s/21% O2 225 s for 60 cycles). Then ELISA was conducted to examine the levels of interleukin-6 (IL-6) and tumor necrosis factor-alpha (TNFalpha) Bicinchoninic acid method was used to standardize the cell total protein level.
The levels of IL-6 and TNFalpha levels in the IH group were (770.40 +/- 21.60) and (126.93 +/- 2.58) pg.ml(-1).100 mg protein(-1) respectively, both significantly higher than those in the IN group [(374.06 +/- 38.10) and (31.96 +/- 13.64) pg.ml(-1).100 mg protein(-1) respectively, U = 0.000, P = 0.002], but significantly lower than those in the IH hypercapnia group [(829.27 +/- 7.16) and (78.77 +/- 4.00) pg.ml(-1).100 mg protein(-1) respectively, U = 0.000, P = 0.002]. The IL-6 levels of the CH hypercapnia 15, 30, and 60 min subgroups were all significantly higher than those of the corresponding CH subgroup (U = 0.000, P = 0.002). The IL-6 and TNFalpha levels of the CH added to IH group were (536.74 +/- 14.97) and (51.10 +/- 6.80) pg.ml(-1).100 mg protein(-1) respectively, both were significantly higher than those of the IN group, but significantly lower than those of the IH group (chi(2) = 23.4, P < 0.05). The levels of IL-6 and TNFalpha increased along with the increase of the IH degree (chi(2) = 23.4, P < 0.05). The level changes of IL-6 and TNFalpha of the groups with different intermittent hypoxia frequency and with different intermittent hypoxia duration were complicated.
IH and CH significantly damage the endothelial cells dose-dependently, especially combined with hypercapnia. In IH/ROX, the inflammatory damage comes from ROX phase but not IH phase. Hypoxia duration and hypoxia frequency are also important parameters in the activation of inflammation.
研究不同模式的间歇性缺氧（IH）和持续性缺氧（CH）对内皮细胞的损伤。
将ECV304人脐静脉内皮细胞培养于新开发的程序控制气体输送系统中，分为8组，进行不同的IH/复氧（ROX）循环，以模拟复发性呼吸暂停和慢性阻塞性肺疾病中所见的缺氧发作模式：间歇性常氧（IN）组（暴露于21%氧气15秒/21%氧气225秒，共60个循环），IH组（暴露于1.5%氧气15秒/21%氧气225秒，共30或60个循环），IH高碳酸血症组（暴露于1.5%氧气和20%二氧化碳15秒/21%氧气和5%二氧化碳225秒，共60个循环），持续性缺氧（CH组，暴露于1.5%或10%氧气15、30或60分钟），CH高碳酸血症组（暴露于10%氧气和10%二氧化碳15、30或60分钟），CH加IH组（暴露于1.5%氧气15秒/10%氧气225秒，共60个循环），不同间歇性缺氧程度组（暴露于1.5%或10%氧气15秒/21%氧气225秒，共60个循环），不同间歇性缺氧频率组（暴露于1.5%氧气15秒/21%氧气225秒315秒、495秒或105秒，共60个循环），以及不同间歇性缺氧持续时间组（暴露于1.5%氧气15秒或30秒/21%氧气225秒，共60个循环）。然后进行酶联免疫吸附测定（ELISA）以检测白细胞介素-6（IL-6）和肿瘤坏死因子-α（TNFα）的水平。采用二辛可宁酸法标准化细胞总蛋白水平。
IH组中IL-6和TNFα水平分别为（770.40±21.60）和（126.93±2.58）pg·ml⁻¹·100mg蛋白⁻¹，均显著高于IN组[分别为（374.06±38.10）和（31.96±13.64）pg·ml⁻¹·100mg蛋白⁻¹，U = 0.000，P = 0.002]，但显著低于IH高碳酸血症组[分别为（829.27±7.16）和（78.77±4.00）pg·ml⁻¹·100mg蛋白⁻¹，U = 0.000，P = 0.002]。CH高碳酸血症15、30和60分钟亚组的IL-6水平均显著高于相应的CH亚组（U = 0.000，P = 0.002）。CH加IH组的IL-6和TNFα水平分别为（536.74±14.97）和（51.10±6.80）pg·ml⁻¹·100mg蛋白⁻¹，均显著高于IN组，但显著低于IH组（χ² = 23.4，P < 0.05）。IL-6和TNFα水平随IH程度的增加而升高（χ² = 23.4，P < 0.05）。不同间歇性缺氧频率组和不同间歇性缺氧持续时间组的IL-6和TNFα水平变化较为复杂。
IH和CH均能显著地、剂量依赖性地损伤内皮细胞，尤其是合并高碳酸血症时。在IH/ROX中，炎症损伤来自复氧阶段而非IH阶段。缺氧持续时间和缺氧频率也是炎症激活的重要参数。
