胸腺素β4对大鼠皮质神经细胞氧糖剥夺-复氧损伤的保护作用研究  

Protective effect of thymosin β4 on oxygen-glucose deprivation/reoxygenation injury in rat cortical neurons

作　　者：张忠胜[1] 刘双凤[1] 黄四春 Zhang Zhongsheng;Liu Shuangfeng;Huang Sichun(Department of Neurology,The 6th Affiliated Hospital of Guangzhou Medical University(Qingyuan People's Hospital),Qingyuan,Guangdong 511518,China)

机构地区：[1]广州医科大学附属第六医院(清远市人民医院)神经内科

出　　处：《中国脑血管病杂志》2019年第6期296-302,共7页Chinese Journal of Cerebrovascular Diseases

基　　金：广东省医学科学技术研究基金项目(A2016468);广东省清远市科技计划项目(2017A022)

摘　　要：目的研究胸腺素β4对体外培养大鼠皮质神经细胞氧糖剥夺-复氧(OGD/R)损伤的保护作用及其机制。方法分离并鉴定原代培养的皮质神经细胞,通过OGD/R法建立皮质神经细胞模拟缺血-再灌注损伤模型(氧糖剥夺6h,复氧12h)。实验分为对照组、模型组、治疗组(建模前2h加入胸腺素β4)。CCK8法确定胸腺素β4的最佳作用浓度,流式细胞术和末端脱氧核苷酸转移酶脱氧尿苷三磷酸缺口末端标记法(TUNEL)检测细胞凋亡,Western blotting检测78000葡萄糖调节蛋白78(GRP78)、CCAAT增强子结合蛋白同源蛋白(CHOP)、B细胞淋巴瘤-2(Bcl-2)和Bax的表达水平并进行组间比较。应用SPSS19.0软件分析处理数据,正态分布的计量资料3组间比较采用单因素方差分析。结果大鼠皮质神经细胞被正确分离。胸腺素β4最佳作用浓度为10μg/L。与对照组比较,模型组皮质神经细胞存活率明显下降(P=0.002),细胞凋亡率明显增加(P<0.01),GRP78、CHOP、Bax表达水平明显升高(P值分别为0.034、0、0.045),Bcl-2的表达水平明显下降(P=0.006);与模型组比较,治疗组(10μg/L外源性胸腺素β4)细胞活力明显增高(P=0.008),细胞凋亡率明显降低(P=0.002),GRP78、CHOP、Bax表达水平明显下降(P值分别为0.032、0.027、0.019),Bcl-2表达水平明显升高(P=0.028),差异均有统计学意义。结论胸腺素β4能抑制OGD/R诱导的内质网应激依赖性细胞凋亡,为胸腺素β4在脑缺血-再灌注损伤治疗中的应用提供了理论依据。Objective To investigate the protective effect and mechanisms of thymosin beta 4 (Tβ4) on oxygen-glucose deprivation/reoxygenation (OGD/R) injury in rat cortical neurons.Methods Primary cultured cortical neurons were isolated and identified;Ischemia-reperfusion injury model of rat cortical neurons was established (oxygen glucose deprivation 6 h,reoxygenation 12 h) by OGD/R.The rats were divided into the control group,the model group and the treatment group (addition of thymosinβ4 2 h before modeling);The cell counting Kit-8(CCK8) was used to determine the optimal concentration of Tβ4.Flow cytometry and terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) were used to detect apoptosis.Western blotting was used to detect the expression of 78 000 glucose-regulated protein 78 (GRP78),CCAAT/enhancer binding protein-homologous protein (CHOP),B-cell lymphoma-2 (Bcl-2) and Bax.Expression levels were compared among groups.One-way analysis of variance was used to compare the normally distributed measurement data among three groups with the SPSS 19.0 software. Results Cortical neurons were separated correctly.The optimal concentration of Tβ4 was 10 μg/L.Comparing the model group with the control group,the survival rate of cortical neurons decreased significantly ( P =0.002),the apoptotic rate increased significantly ( P <0.01),the expression of GRP78,CHOP and Bax was up-regulated significantly ( P values were 0.034,0 and 0.045,respectively),and the expression of Bcl-2 was reduced significantly ( P =0.006).Comparing the treatment group(10 μg/L exogenous Tβ4) with the model group,cell viability increased significantly ( P =0.008),the apoptotic rate decreased significantly( P =0.002),the expression of GRP78,CHOP and Bax decreased significantly ( P values were 0.032,0.027 and 0.019,respectively),and the expression of Bcl-2 increased significantly ( P = 0.028).The differences were statistically significant.Conclusions Tβ4 inhibits OGD/R- induced endoplasmic reticulum stress-dependent apoptosis.These resul

关 键 词：胸腺素 细胞凋亡 胸腺素Β4 皮质神经元 内质网应激 

分 类 号：R743[医药卫生—神经病学与精神病学]