Changes of human umbilical venous endothelium due to different degree ischemia-reperfusion injury and the interventional effect of qingkailing
- VernacularTitle:缺血缺氧再灌注不同损伤程度人脐静脉内皮细胞变化与清开灵的干预效应
- Author:
Qing TANG
- Publication Type:Journal Article
- From:
Chinese Journal of Tissue Engineering Research
2005;9(29):199-201
- CountryChina
- Language:Chinese
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Abstract:
BACKGROUND: "Toxic impaired brain retinervus" is the important mechanism of acute cerebrovascular diseases, mainly presented by cerebral microvascular impairments.OBJECTIVE: To establish cell model of various degrees ischemia-reperfusion injury and expos to qingkailing medication, in order to observe the protective role of qingkailing injection on human umbilical venous endothelium ECV304 (huveECV304)DESIGN: Randomized controlled study based on cultured cells.SETTING: Department of Combination of Traditional Chinese Medicine and Western Medicine, Union Hospital Affiliated to Tongji Medical College, Huazhong University of Science and Technology.MATERIALS: This study was carried out at Center Laboratory of Dongzhimeu Traditional Chinese Medicine College, Beijing from February to April 2002. huveECV304 were randomly divided into 7 groups, namely normal group (NG), ischemic-ypoxia group (IR group), ischemia 6 hours reperfusion 12 hours control group (I-6-hour-R-12-hour group), ischemia 24 hours reperfusion 12 hours control group (I-24-hour-R-12-hour group),ishcmeia 24 hours reperfuiosn 12 hours low dosage qingkailing group (I24-hour-R-12-hour-LDQ group), ischemia 24 hours reperfusion 12 hours middle dosage qingkailing group (I-24-hour-R-12-hour-MDQ group), ischemia 24 hours reperfusion 12 hours high dosage qingkailing group (I-24hour-R-12-hour-HDQ group) and ischemia 6 hours reperfusion 12 hours middle dosage qingkailing group (I-6-hour-R-12-hour-MDQ group). The qingkailing injection mainly consists of cholaic acid, hyodeoxycholic acid,baikal skullcap root, cape jasmine fruit and so on.were collected, and primary culture medium were removed in I-6-hour-R-12hour group and I-24-hour-R-12-hour group and added with non-glucose Earle's solution, the cells in I-6-hour-R-12-hour group were cultured in ischemic incubator in 37 ℃ for 6 hours, for 24 hours in I-24-hour-R-12-hour group and then reperfusion-cultured with Earle's medium replaced by nonLDQ group, I-24-hour-R-12-hour-MDQ group and I-24-hour-R-12-hour-HDQ group, IR injured cells were exposed to qingkailing in dosage of 5, 10,by using MTT colorimetric assay, lactate dehydrogenase leaking rate was determined by using lactate dehydrogenase reagent kit, cell apoptosis rate was detected with FCM and statistically analyzed with Cellquest software.apoptosis rate.were found gathering closely, well adhering to the wall and completely stretching, cell surface was smooth and more cleavage cells were observed,cell grew over all the bottom displaying typical "slabstone signs"; but no obvious cell morphological changes were observed in control group and Ⅰ24-hour-R-12-hour LDQ group; while in I-24-hour-R-12-hour control group, cell were found scarcely distributed and mostly drop off, a great deal of tumid round injured cells and broken cell pieces were observed,adhering cells were shrunk obviously with pseudopod stretching out, less cleavage cells were observed; In I-24-hour-R-12-hour MDQ group, cells were found well adhering to the wall, the surface was smooth without cell: Comparing to I-24-hour-R-12-hour control group, the survival rate were markedly increased in I-24-hour-R-12-hour LDQ group, I-24-hour-R12-hour MDQ group [(42.5±2.8), (72.9±8.0), (80.3±12.7)%, P ＜ 0.01], but obviously descended to (10.1 ±0.4)% (P ＜ 0.01) in I-24-hour-R-12-hour ing rate of ECV304 cell: Comparing to I-24-hour-R-12-hour control group,it was found markedly increased in I-24-hour-R-12-hour LDQ group and Ⅰ24-hour-R-12-hour MDQ group [(6.6±1.4), (2.4±1.4), (2.4±0.5)%, P ＜ 0.01], but significantly ascended to (16.1±2.2)% in I-24-hour-R-12-hour rate of ECV304 cell: Comparing to I-6-hour-R-12-hour control group, it was obviously increased in comparing to I-6-hour-R-12-hour MDQ group [(2.2±0.5), (1.5±0.4)%, P ＜ 0.05]; comparing to normal group, it was found significantly increased in I-24-hour-R-12-hour control group and I-24-hourR-12-hour MDQ group [(0.9±0.2), (5.5±2.5), (9.9±4.3)%, P ＜ 0.01]; Moreover the increment in I-24-hour-R-12-hour control group was significantly higher than I-24-hour-R-12-hour MDQ group (P ＜ 0.01).ECV304 with various degree IR injury, but which displaying tow opposite cells due to I-6-hour-R-12-hour exposure was mainly presented by lowered cell apoptosis rate; But severer injury of ECV304 cells due to I-24-hour-R12-hour exposure was presented by increased cell apoptosis rate and lowon and cell membrane would not break in apoptostic cells and no internal content effused, therefore the surrounding cells would not be affected,which increasing the survival rate and displaying protective role for most cells.
