低温低氧复合应激对大鼠肺微血管内皮细胞的损伤作用研究  

The damage effects of hypothermia combined with hypoxia on rat pulmonary microvascular endothelial cells

作　　者：张竞丹[1] 杨丹凤[2] 曹燕卿[2] 李曦[2] 安玉林[2] 刘嘉瀛[2] 汪海[2] 

机构地区：[1]中南大学公共卫生学院,湖南长沙410078 [2]军事医学科学院卫生学环境医学研究所,天津300050

出　　处：《中国应用生理学杂志》2013年第3期219-223,I0001,共6页Chinese Journal of Applied Physiology

基　　金：基金项目:AWS11J003;2011ZXJ09105-03B

摘　　要：目的:探讨低温与低氧对大鼠肺微血管内皮细胞(PMVECs)的影响及二者的协同作用。方法:采用复合酶消化法培养大鼠PMVECs;免疫荧光检测CD31抗原的表达,植物凝集素结合实验并结合形态学鉴定细胞;按低温和低氧两因素的有无,采用2×2析因设计进行实验,检测LDH活性,硝酸还原酶法检测一氧化氮(NO)释放,RT-PCR法检测血管内皮生长因子(VEGF)、内皮素-1(ET-1)mRNA水平。结果:培养的PMVECs呈典型的鹅卵石样排列,CD31抗原表达呈阳性,与植物凝集素结合呈明亮的绿色荧光,鉴定为PMVECs;与对照组相比,低温、低氧及低温低氧组PMVECs的LDH活力增加、NO释放减少、VEGF和ET-1 mRNA表达上调,并且具有显著的统计学差异。析因方差分析显示在以上几方面低温和低氧之间均存在协同作用。结论:低温及低氧在细胞膜通透性和血管活性物质NO、ET-1释放方面对PMVECs有影响,且低温和低氧存在协同作用。Objective： To study the synergistic effects of hypothermia and hypoxia on the damage of pulmonary microvascular endothelial cells（PMVEC） in rat.Methods： Primary PMVECs were obtained by complex phosphoesterasum digesting from isolated lung tissues of Wistar rats,the PMVECs were identified by phase contrast microscope and immunofluorescence studies for CD31 antigen and bandeiraea simplicifolia isolectin（BSI） binding test.Factorial design was adopted in trial according to hypothermia and hypoxia existing or not.Using corresponding kit measured the levels of lactate dehydrogenase（LDH） activity in cell medium.Level of nitric oxide（NO） concentration was measured by Griess Assay.RT-PCR was used to examine the expression of vascular endothelial growth factor（VEGF）, endothelin-1（ET-1） mRNA in PMVECs.Results： The monolayer of cultured PMVECs displayed the shape of pavingstone.CD31 antigen and binding BSI results by fluorescence microscope identified the cultured cells were PMVECs.Compared to the control group,LDH activity and VEGF,ET-1 expression levels were significantly increased in hypothermia group,hypoxia group and hypoxia combined with hypothermia group.And the levels of NO concentration were reduced in these three groups.The results of One-way ANOVA showed that there was a synergistic effect between hypothermia and hypoxia.Conclusion： Hypothermia and hypoxia both have an effect on PMVECs whether in altering the cell permeability or in releasing of vasoactive substances including NO and ET-1.In addition,there is a synergistic effect between hypothermia and hypoxia.

关 键 词：肺微血管内皮细胞 低温 低氧 协同作用 高原肺水肿 

分 类 号：R329.2[医药卫生—人体解剖和组织胚胎学] Q256[医药卫生—基础医学]