抑制第10号染色体缺失的磷酸酶和张力蛋白同源等位基因活性对缺氧缺血神经元凋亡的保护作用  

Neuroprotective effect of phosphatase and tensin homolog allele activity deleted in chromosome 10 inhibition on neuronal apoptosis after hypoxia-ischemia

作　　者：赵婧[1] 屈艺[1] 母得志[1] 

机构地区：[1]四川大学华西第二医院儿科,成都610041

出　　处：《中华实用儿科临床杂志》2013年第24期1855-1859,共5页Chinese Journal of Applied Clinical Pediatrics

基　　金：国家自然科学基金（81330016,31171020,81172174,81270724）;国家重大科学研究计划项目（2013CB967404）;国家教育部科研基金（313037,20110181130002）;国家科技部科研基金（2012BA104804）;四川省科技厅基金（2010SZ0280,2011.1TD0005）;国家临床重点专科（儿科新生儿专业）建议项目（1311200003303）

摘　　要：目的探讨抑制第10号染色体缺失的磷酸酶和张力蛋白同源等位基因（PTEN）活性对缺氧缺血神经元凋亡的保护作用及机制。方法培养新生SD大鼠皮质神经元，建立体外神经元氧糖剥夺（OGD）模型模拟细胞缺氧缺血。细胞分为3组：1．正常对照组：正常培养液孵育的神经元；2．OGD组：模型制备后，取再灌注后0．5、3．0、6．0、12．0、24．0、48．0h神经元进行观察；3．PTEN抑制剂过氧钒酸钠（bpv）干预组：应用bpv处理神经元，再制备OGD模型，于再灌注后24h观察。应用末端脱氧核苷酰基转移酶介导性dUTP切口末端标记法检测神经元凋亡，Westernblot检测PTEN、p-PTEN、蛋白激酶B（Akt）、p-Akt、糖原合成酶激酶-3β（GSK-3β）、p-GSK-3β、抗凋亡蛋白髓细胞淋巴瘤-1（Mcl-1）蛋白表达。结果1．与正常对照组比较，OGD神经元凋亡增加（P〈0．05），PTEN表达增加，p-PTEN、p-Akt、p-GSK-3β及Mcl-1的表达降低（P均〈0．05），而Akt及GSK-3β表达未发生变化（P均〉0．05）。2．与OGD组比较，bpv干预后神经元凋亡减少（P〈0．05），p-PTEN、p-Akt、p-GSK-3β和Mcl-1的表达增加（P均〈0．05），而PTEN、Akt及GSK-3β并未发生变化（P均〉0．05）。结论缺氧缺血诱导神经元凋亡发生，PTEN／Akt／GSK-3β／Mcl-1信号通路参与调控其凋亡。抑制PTEN活性后，Akt及GSK-3β磷酸化水平增加，使Mcl-1泛素化降低，从而减少神经元凋亡。Objective To investigate the mechanisms of neuroprotective roles of phosphatase and tensin homo- log allele activity deleted in chromosome 10 （ PTEN ） inhibition on neuronal apoptosis after hypoxia-ischemia （ HI ） damage. Methods The cerebral cortical neurons of newborn Sprague-Dawley rats were cultured in vitro. Oxygen and glucose deprivation（OGD） model was established to imitate HI environment in vivo. Neurons were divided randomly in- to 3 groups ： control group ： neurons were treated with normal medium; OGD group ： neurons were treated with OGD for 3 h followed by reperfusion at 0.5,3.0,6.0,12.0,24.0,48.0 h ; PTEN inhibition group ： before OGD treatment, neu- rons were pretreated with PTEN inhibitor, and then the neurons were collected at 24 h after reperfusion. Terminal deoxy- nucleotidyl transferase-mediated dUTP nick and labeling staining was used to detect the apoptotic cells. Western blot was used to detect the expression of PTEN, p-PTEN, protein kinase B （Akt） , p-Akt, synthesis of glucose kinase-3 betal （GSK-3β）, p-GSK-3β and myeloid cell ceukemia-1 （Mcl-1）. Results 1. As compared with control group, TUNEL positive cells increased after OGD observed by TUNEL staining （P 〈 0.05 ）. The expression of PTEN was increased ,the expressions of p-PTEN, p-Akt, p-GSK -3β, and Mcl-1 were significantly decreased after OGD （ all P 〈 0.05 ）. However, Akt and GSK-3β remained unchanged at different time points after OGD （ all P 〉 O. 05 ）. 2. As compared with OGD group,TUNEL positive cells were obviously reduced at 24 h after OGD in PTEN group （ P 〈 0.05 ）. Although total PTEN, Akt, and GSK-3β were not obviously changed in PTEN group （all P 〉 0.05 ）, the expressions of p-PTEN, p-Akt, p-GSK-3β,and Mcl-1 were significantly increased after OGD （ all P 〈 0.05 ）. Conclusions HI can induce neuronal apoptosis,and the mechanisms of apoptosis may involve PTEN/Akt/GSK-3β 3/Mcl-1 pathway. The phosphorylation of Akt and GSK-3β can be increased via PTEN acti

关 键 词：第10号染色体缺失的磷酸酶和张力蛋白同源等位基因 蛋白激酶B 糖原合成酶激酶-3Β 抗凋亡蛋白髓细胞淋巴瘤-1 凋亡 神经元 缺氧缺血 

分 类 号：R722.1[医药卫生—儿科]