有氧运动通过激活APP/PS1小鼠大脑皮质和海马组织PI3K/Akt信号通路抑制神经细胞凋亡

Aerobic Exercise Inhibits Neuronal Cell Apoptosis through Activation of the PI3K/Akt Pathway in the Cortex and Hippocampus of APP/PS1 Mice

目的:探讨有氧运动对APP/PS1小鼠大脑皮质和海马组织神经细胞凋亡的影响,以及是否通过PI3K/Akt信号通路发挥作用,为阐明有氧运动防治阿尔茨海默症提供一定的理论依据。方法:40只雄性APP/PS1小鼠随机分为安静组(T-SE,n=10)、运动组(T-EX,n=10)、GNE-317处理安静组(T-SEG,n=10)和GNE-317处理运动组(T-EXG,n=10)。T-EX和T-EXG组小鼠进行为期8周的跑台训练,T-SEG和TEXG组小鼠给予GNE-317处理8周。最后一次训练结束后48小时,分离所有小鼠大脑皮质和海马组织。通过TUNEL染色观察各组小鼠大脑皮质和海马组织细胞凋亡情况;通过Western blot检测各组小鼠大脑皮质和海马组织磷脂酰肌醇3激酶(PI3K)、蛋白激酶B(Akt)、B淋巴细胞瘤-2蛋白(Bcl-2)、Bcl-2相关促凋亡蛋白(Bad)、Bcl-2相关X蛋白(Bax)、剪切形式含半胱氨酸的天冬氨酸蛋白水解酶3(Cleaved Caspase-3)、Caspase-7、Caspase-9、线粒体细胞色素C(Cytochrome C)蛋白含量及磷酸化水平。结果:(1)8周跑台训练后,T-EX组大脑皮质和海马组织中PI3K p110和p85亚基的蛋白含量及Akt Thr308和Ser473位点的磷酸化水平均高于T-SE组(P<0.01);T-EX组TUNEL染色阳性细胞数量低于T-SE组(P<0.05);T-EX组抗凋亡因子Bcl-2的含量高于T-SE组(P<0.05),而促凋亡因子Bad、Bax、Cytochrome C、Caspase-9、Caspase-7和Cleaved Caspase-3的含量均低于T-SE组(P<0.05)。(2)经GNE-317处理后,TSEG和T-EXG组Akt Thr308和Ser473位点的磷酸化水平分别低于T-SE和T-EX组(P<0.05);T-SEG和T-EXG组TUNEL染色阳性细胞数量分别高于T-SE和T-EX组(P<0.05);T-SEG和T-EXG组抗凋亡因子Bcl-2的含量分别低于T-SE和T-EX组(P<0.05),而促凋亡因子Bad、Bax、Cytochrome C、Caspase-9、Caspase-7和Cleaved Caspase-3的含量分别高于T-SE和T-EX组(P<0.05)。结论:有氧运动通过激活APP/PS1小鼠大脑皮质和海马组织中PI3K/Akt信号通路活性,提高了抗凋亡因子Bcl-2的含量,同时降低促凋亡因子Bad、Bax、Cytochrome C、Caspase-9、Caspase-7和Cleaved Caspase-3的含量,从而有效抑制神经细胞凋亡。

Objective To explore the effect of aerobic exercise on the neuronal cell apoptosis and ex amine whether such effect is mediated by the phosphoinositide 3-kinase(PI3K)/protein kinase B(Akt)pathway in the cerebral cortex and hippocampus of APP/PS1 mice,so as to confirm that aero bic exercises can prevent Alzheimer's disease.Methods Forty male APP/PS1 male mice were randomly divided into four groups:a sedentary group(T-SE,n=10),an exercise group(T-EX,n=10),a seden tary with GNE-317 treatment group(T-SEG,n=10)and an exercise with GNE-317 treatment group(TEXG,n=10),each of 10.The mice in the T-EX and T-EXG group underwent an 8-week treadmill training,meanwhile the mice in the T-SEG and T-EXG group were given a treatment with GNE-317.Forty-eight hours after the last exercise,all mice were isolated the cerebral cortex and hippocampus.The TUNEL staining was used to detect cell apoptosis,and Western blotting was employed to assay the protein and phosphorylation levels of PI3K,Akt,B cell lymphoma-2(Bcl-2),Bcl-2 associated death promoter(Bad),Bcl-2 associated x protein(Bax),cleaved cysteinyl aspartate specific proteinase 3(Cleaved Caspase-3),Caspase-7,Caspase-9 and Cytochrome C.Results After 8 weeks of treadmill exercise,the protein levels of PI3K p110 and p85 and the phosphorylation levels of Akt Thr308 and Ser473 in the T-EX group were significantly higher than those of the T-SE group(P<0.01).And the number of TUNEL staining positive cells in the T-EX group was significantly lower than that of the TSE group(P<0.05).In addition,in the T-EX group,the protein levels of anti-apoptotic factor Bcl-2 were significantly higher than those of the T-SE group(P<0.05),while the protein levels of pro-apop totic factors Bad,Bax,Cytochrome C,Caspase-9,caspase-7 and Cleaved Caspase-3 were significant ly lower than those of the T-SE group(P<0.05).After GNE-317 treatment,the phosphorylation levels of Akt Thr308 and Ser473 in the T-SEG and T-EXG group were significantly lower than those of TSE and T-EX group respectively(P<0.05).And the number of TUNEL staining positive cells in the TSEG and T-EXG group were significantly higher than those of T-SE and T-EX group respectively(P<0.05).Moreover,in the T-SEG and T-EXG group,the protein levels of anti-apoptotic factor Bcl-2 were significantly lower than those of T-SE and T-EX group respectively(P<0.05),while the protein levels of pro-apoptotic factors including Bad,Bax,Cytochrome C,Caspase-9,caspase-7 and Cleaved Caspase-3 were significantly higher than those of T-SE and T-EX group respectively(P<0.05).Conclu sion Aerobic exercises can increase the content of anti-apoptotic factor Bcl-2,and reduce that of proapoptotic factors Bad,Bax,Cytochrome C,Caspase-9,Caspase-7 and Cleaved Caspase-3 through ac tivating the PI3K/Akt pathway in the cortex and hippocampus of APP/PS1 mice,so as to inhibit the neuronal cell apoptosis effectively.