氧糖剥夺条件下脑血管内皮细胞对氧化型低密度脂蛋白损伤的易感性及机制研究

发布时间：2019-04-10 07:33

【摘要】：目的:观察缺氧缺糖性损伤条件下,血管内皮细胞对氧化型低密度脂蛋白刺激易感性的考察,以及下游氧化应激信号通路的调控机制。方法:将细胞分为正常对照组、氧糖剥夺(OGD)组6h、100μg/ml氧化型低密度脂蛋白ox-LDL处理24h组(ox-LDL组)、OGD 6h+100μg/ml ox-LDL刺激24h组(OGD+ox-LDL组),分别处理分离的大鼠脑血管内皮细胞(RBECs)。Western blot检测脑血管内皮细胞小凹蛋白1(Cav-1)、凝集素样氧化型低密度脂蛋白受体(LOX-1)变化,及考察下游氧化应激MAPK信号通路和内皮型一氧化氮合酶(eNOS)。免疫荧光检测RBECs中Dil细胞膜荧光标记的ox-LDL(Dil-ox-LDL)荧光强度的变化。比色法和ELISA法检测炎症因子即一氧化氮(NO)、白细胞介素1β(IL-1β)和肿瘤坏死因子-α(TNF-α)的表达。结果 :oxLDL刺激可对增加LOX-1表达和RBECs的Dil-ox-LDL荧光强度,并同时降低Cav-1表达。OGD组LOX-1表达和Dil-ox-LDL荧光强度相较于正常对照组无明显增加,然而Cav-1表达有所降低,然而在OGD+ox-LDL组LOX-1表达和Dil-ox-LDL荧光强度增加,且相较于ox-LDL组,有显著性差异。另外,除正常对照组外,各组RBECs均观察到MAPK信号通路中p38、ERK1/2、JNK氧化应激通路激活和NO、IL-1β、TNF-α炎症因子释放增加,并且OGD+ox-LDL组相较于OGD组、ox-LDL组均有显著性差异。结论:RBECs在OGD和ox-LDL共同刺激条件下,对ox-LDL的刺激具有易感性,可在增加细胞内Dil-ox-LDL荧光强度,并且诱导下游氧化应激反应,机制可能与Cav-1调控氧化应激反应相关,本研究可为以OGD细胞模型为代表的缺血性脑卒中等疾病相关脂质代谢研究提供科学根据。
[Abstract]:Aim: to investigate the susceptibility of vascular endothelial cells to oxidative low density lipoprotein (LDL) stimulation and the regulation mechanism of downstream oxidative stress signaling pathway in hypoxic-glucose-deficient injury. Methods: the cells were divided into normal control group, oxygen glucose deprivation (OGD) group for 6 h, 100 渭 g / ml oxidized low density lipoprotein ox-LDL for 24 h group (ox-LDL group), OGD for 6 h 100 渭 g / ml ox-LDL for 24 h group (OGD ox-LDL group). (RBECs). Western blot of isolated rat cerebral vascular endothelial cells was used to detect the changes of Cav-1 and LOX-1 in rat cerebral vascular endothelial cells, which were divided into two groups, and the control group was divided into control group and control group, and the control group was divided into control group and control group (P < 0. 05). And investigate the downstream oxidative stress MAPK signaling pathway and endothelial nitric oxide synthase (eNOS). The fluorescence intensity of Dil cell membrane labeled ox-LDL (Dil-ox-LDL) in RBECs was detected by immunofluorescence. The expression of inflammatory factors (nitric oxide, (NO), interleukin 1 尾 (IL-1 尾) and tumor necrosis factor-伪 (TNF- 伪) was detected by colorimetric method and ELISA method. Results: oxLDL stimulation could increase the expression of LOX-1 and Dil-ox-LDL fluorescence intensity of RBECs, and decrease the expression of Cav-1 at the same time. Compared with the normal control group, the expression of LOX-1 and the fluorescence intensity of Dil-ox-LDL in OGD group did not increase significantly. However, the expression of Cav-1 decreased, but the expression of LOX-1 and the fluorescence intensity of Dil-ox-LDL increased in OGD ox-LDL group, and there was significant difference compared with ox-LDL group. In addition, except for the normal control group, p38, ERK _ (1) ~ (2), JNK oxidative stress pathway activation and NO,IL-1 尾, TNF- 伪 inflammatory factor release were observed in all groups of RBECs, and compared with OGD group, OGD ox-LDL group was higher than OGD group. There was significant difference in ox-LDL group. Conclusion: under the condition of co-stimulation of OGD and ox-LDL, RBECs is susceptible to the stimulation of ox-LDL, which can increase the fluorescence intensity of Dil-ox-LDL in cells and induce the downstream oxidative stress response. The mechanism may be related to the regulation of oxidative stress response by Cav-1. This study may provide a scientific basis for the study of lipid metabolism related to ischemic stroke such as ischemic stroke represented by OGD cell model.
【作者单位】：南昌大学第一附属医院药学部;南昌大学第一附属医院心血管科;
【基金】：江西省教育厅科学技术研究青年项目(No:GJJ160238) 江西省科技厅青年基金一般项目(No:20171BAB215021)
【分类号】：R743.3

【相似文献】