高压氧预适应对离体培养大鼠脑星形胶质细胞的缺氧保护作用及机制  

Hyperbaric oxygen preconditioning protects rat brain astrocytes from hypoxia by down-regulating AQP4 and R2X_7 receptor in vitro

作　　者：李雨泓[1] 邹永杰[1] 崔高宇[1] 

机构地区：[1]第三军医大学西南医院神经外科,全军神经外科研究所,全军神经创伤防治重点实验室,重庆400038

出　　处：《第三军医大学学报》2014年第9期893-896,共4页Journal of Third Military Medical University

基　　金：国家自然科学基金面上项目(30973100);国家自然科学基金青年科学基金(81000532)~~

摘　　要：目的研究高压氧预适应(hyperbaric oxygen preconditioning,HBOP)对大鼠乳鼠脑星形胶质细胞的缺氧保护作用及对P2X7受体和AQP4表达的影响。方法离体培养原代星形胶质细胞并纯化传代后,分为对照组、高压氧预适应组(HBOP)、单纯缺氧组(OGD)、高压氧预适应缺氧组(HBOP+OGD)。于预定时相点采用流式细胞仪检测细胞凋亡率,化学发光法检测细胞培养液中的ATP浓度,Western blot检测P2X7受体和AQP4蛋白表达,实时荧光定量PCR检测(RT-qPCR)P2X7受体和AQP4 mRNA的转录水平。结果 HBOP组细胞培养液内ATP浓度在各检测时相点远高于对照组(P<0.01);HBOP+OGD组细胞24 h及48 h凋亡率均低于OGD组(P<0.05);HBOP组细胞AQP4及P2X7受体蛋白表达高于对照组(P<0.05);HBOP+OGD组细胞AQP4及R2X7受体mRNA转录水平在缺氧6 h时低于OGD组(P<0.05)。结论 HBOP可以提高大鼠脑星形胶质细胞线粒体产生ATP的能力,提高细胞对缺氧缺糖的耐受力,减少细胞凋亡,具有明确的细胞保护作用。其保护机制可能与降低了OGD后损伤细胞内P2X7受体及AQP4的mRNA转录水平有关。Objective To determine the protective effect of hyperbaric oxygen preconditioning （HBOP） on rat brain astrocytes against hypoxia and on the expression of R2X7 receptor and AQP4. Methods Cultured primary astrocytes were isolated from neonatal SD rats, primarily cultured and identified by GFAP staining at passage 2. The astrocytes at passage 3 were divided into control group, HBOP group, oxygen glucose deprivation （OGD） group and HBOP＋OGD group. The apoptotic rate was detected by flow cytometry at 24 and 48 h after OGD. The ATP concentration in the culture medium were measured by chemiluminescence detection. The protein expression of R2X7 receptor and AQP4 was detected by Western blotting, and their mRNA levels were determined by RT-qPCR. Results The expression of AQP4 and R2X7 receptor was enhanced in HBOP group than control group （P〈0.05）. The apoptotic rate was significantly lower in the HBOP＋OGD group than the OGD group at 24 and 48 h （P〈0.05）. The protein level of AQP4 and R2X7 receptor was obviously higher in the HBOP group than control group （P〈0.05）. The AQP4 and R2X7 receptor mRNA transcription levels were inhibited in HBOP＋OGD group when compared with those in OGD group at 6 h （P〈0.05）. Conclusion HBOP improves the mitochondrial ATP production by rat brain astrocytes exposure to hypoxia, promotes the cell tolerance to hypoxia and aglycemia, reduces cell apoptosis, and exerts absolutely cytoprotective effect. The underlying mechanism may be due to the down-regulation of AQP4 and R2X7 receptor at mRNA levels after OGD.

关 键 词：高压氧预适应 氧糖剥夺 细胞保护 AQP4 P2X7受体 

分 类 号：R322.81[医药卫生—人体解剖和组织胚胎学] R364.4[医药卫生—基础医学]