高渗盐水通过与表皮生长因子受体和白细胞介素-1β水平相关的机制保护脑内皮细胞免受缺氧损伤。
Hypertonic saline protects brain endothelial cells against hypoxia correlated to the levels of epidermal growth factor receptor and interleukin-1β.
作者信息
Chen Sheng-Long, Deng Yi-Yu, Wang Qiao-Sheng, Han Yong-Li, Jiang Wen-Qiang, Fang Ming, Hu Bei, Wu Zhi-Xin, Huang Lin-Qiang, Zeng Hong-Ke
机构信息
Graduate School, Southern Medical University, 1838 North Guangzhou Avenue, Guangzhou 510515, PR China Department of Emergency and Critical Care Medicine, Guangdong General Hospital, Guangdong Academy of Medical Sciences,106 Zhongshan Er Road, Guangzhou 510080, PR China Department of Critical Care Medicine, The First Affiliated Hospital, University of South China, Hengyang, Hunan Province, PR China.
出版信息
Medicine (Baltimore). 2017 Jan;96(1):e5786. doi: 10.1097/MD.0000000000005786.
OBJECTIVE
The aim of this study was to verify the protective effect of hypertonic saline (HS) on brain endothelial cells under hypoxic conditions and the relevant underlying mechanism.
METHODS
bEnd.3 cells were treated with oxygen-glucose deprivation (OGD)-induced injury. To measure HS performance, cell viability was determined using the 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium salt assay, and cell apoptosis was assessed by flow cytometry and Terminal deoxynucleotidyl transferase UTP nick-end labeling staining. RNA-seq was performed to assess the expression profiles and screen the candidate genes that participated in OGD-induced injury and the HS protective effect. Quantitative real-time polymerase chain reaction (qPCR) and western blot analysis were used to confirm the expression of candidate genes, and enzyme-linked immunosorbent assay was used to measure the level of interleukin (IL)-1β. Overexpression analyses were performed to confirm the functions of the differentially expressed genes.
RESULTS
HS with a concentration of 40 mmol/L NaCl had an obvious protective effect on bEnd.3 cells after OGD-induced injury, resulting in increased cell viability and a smaller percentage of apoptotic cells. According to the RNA-seq results, epidermal growth factor receptor (EGFR) was chosen as the differentially expressed gene target in this study. The qPCR and western blot analyses further confirmed that the levels of EGFR/phosphorylated epidermal growth factor receptor and IL-1β were enhanced after OGD-induced injury, but attenuated after treatment with 40 mmol/L of NaCl HS. Overexpressed EGFR reversed the protective effect of HS that caused low viability and high rates of apoptosis in cells.
CONCLUSION
HS can protect endothelial cells against OGD-induced injury, but is affected by the expression of EGFR/p-EGFR and IL-1β.
目的
本研究旨在验证高渗盐水（HS）在缺氧条件下对脑内皮细胞的保护作用及其相关潜在机制。
方法
用氧糖剥夺（OGD）诱导bEnd.3细胞损伤。为检测HS的作用，采用3-(4,5-二甲基噻唑-2-基)-5-(3-羧甲氧基苯基)-2-(4-磺基苯基)-2H-四唑盐法测定细胞活力，通过流式细胞术和末端脱氧核苷酸转移酶介导的dUTP缺口末端标记染色评估细胞凋亡。进行RNA测序以评估基因表达谱并筛选参与OGD诱导损伤和HS保护作用的候选基因。采用定量实时聚合酶链反应（qPCR）和蛋白质免疫印迹分析来确认候选基因的表达，并使用酶联免疫吸附测定法测量白细胞介素（IL）-1β水平。进行过表达分析以确认差异表达基因的功能。
结果
浓度为40 mmol/L NaCl的HS对OGD诱导损伤后的bEnd.3细胞具有明显的保护作用，导致细胞活力增加，凋亡细胞百分比降低。根据RNA测序结果，选择表皮生长因子受体（EGFR）作为本研究中的差异表达基因靶点。qPCR和蛋白质免疫印迹分析进一步证实，OGD诱导损伤后EGFR/磷酸化表皮生长因子受体和IL-1β水平升高，但用40 mmol/L的NaCl HS处理后降低。过表达EGFR可逆转HS对细胞造成的低活力和高凋亡率的保护作用。
结论
HS可保护内皮细胞免受OGD诱导的损伤，但受EGFR/p-EGFR和IL-1β表达的影响。
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