中文摘要：

目的 通过建立小鼠慢性睡眠剥夺模型,观察长期睡眠剥夺对小鼠学习记忆以及对神经元细胞自噬和凋亡的影响.方法 C57BL/6小鼠20只,应用随机数字表进行随机分组,睡眠剥夺组和对照组各10只.采用改良的多平台法对睡眠剥夺组小鼠建立慢性睡眠剥夺模型,2个月后通过IntelliCage全自动动物行为学检测系统检测小鼠的学习记忆能力,蛋白免疫印迹法检测微管相关蛋白-1轻链3 （LC3）-Ⅱ和Beclin-1蛋白表达,采用激光共聚焦显微镜观察海马和皮质神经元内的自噬小体,末端脱氧核苷酸转移酶介导的dUTP原位末端标记法（TUNEL）染色观察海马和皮质神经元凋亡.结果 与对照组相比,睡眠剥夺组小鼠表现出明显的空间学习能力下降,其错误的来访率明显增高;此外,睡眠剥夺组小鼠脑组织LC3-Ⅱ（睡眠剥夺组1.681±0.186,对照组1.125±0.048,t=2.892,P=0.027 6）和Beclin-1（睡眠剥夺组1.144±0.048,对照组1.006±0.017,t=2.721,P=0.018 6）蛋白表达显著升高,海马和皮质脑区LC3阳性细胞数增多（海马睡眠剥夺组1.665±0.153,对照组0.819 ±0.071,t=5.024,P=0.002 4;皮质睡眠剥夺组1.925±0.175,对照组1.195±0.111,t=3.521,P=0.012 5）;同时,TUNEL染色显示睡眠剥夺组小鼠海马和皮质凋亡阳性细胞百分比（％）显著升高（海马睡眠剥夺组47.24 ±4.15,对照组19.26±3.72,t=5.025,P=0.007 4;皮质睡眠剥夺组42.25±1.25,对照组27.50±3.23,t=4.262,P=0.005 3）.结论 慢性睡眠剥夺能够造成小鼠学习记忆能力下降,并上调自噬相关蛋白LC3-Ⅱ和Beclin-1的表达,增加自噬小体数目,促进神经元凋亡,提示慢性睡眠剥夺造成的认知损害可能与其诱导自噬和细胞凋亡有关.

英文摘要：

Objective To establish chronic sleep deprivation mouse model,evaluate the learning and memory ability of mice and observe autophagy and apoptosis levels in mouse brain.Methods C57BL/6 mice （n =20） were randomly separated into sleep deprivation group and control group.After 2-month sleep deprivation by using an adapted multiple platform method,the behavioral performance of mice was measured by IntelliCage system.The expression of microtubule associated protein 1 light chain 3-Ⅱ （LC3-Ⅱ） and Beclin-1 was detected by Western blotting.Confocol microscopy was used to observe autophagosome.In addition,terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling （TUNEL） staining was performed to detect neuronal apoptosis level in mouse brain.Results The results of behavioral test showed that the incorrect visit ratio was much higher in sleep deprivation group than that in control group.Moreover,the expression of LC3-Ⅱ （sleep deprivation group 1.681 ± 0.186,control group 1.125 ±0.048,t =2.892,P =0.027 6） and Beclin-1 （sleep deprivation group 1.144 ±0.048,control group 1.006 ± 0.017,t =2.721,P =0.018 6） in mouse hippocampus and cortex was significantly elevated in sleep deprivation group than those in control group.Accordingly,the confocal microscopy observation also revealed an increased nuclear LC3-positive puncta in hippocampus and cortex of sleep deprived mice （hippocampus in sleep deprivation group 1.665 ± 0.153,in control group 0.819 ± 0.072,t =5.024,P =0.002 4;cortex in sleep deprivation group 1.925 ± 0.175,in control group 1.195 ± 0.111,t =3.521,P =0.012 5）.In addition,TUNEL staining showed a much higher percentage of TUNEL-positive nuclei in these brain regions （hippocampus in sleep deprivation group 47.24 ± 4.15,in control group 19.26 ± 3.72,t =5.025,P =0.007 4;cortex in sleep deprivation group 42.25 ± 1.25,in control group 27.50 ± 3.23,t =4.262,P =0.005 3）.Conclusions Chronic sleep deprivation can impair the learning and memory,increase the expression of LC3