【摘要】 目的 探讨天王补心丹加减方（BXDJJ）改善大鼠睡眠剥夺损伤的作用及机制。方法 （1）将24只SD雄性大鼠随机分为地西泮组(3 mg·kg^-1)和BXDJJ低、中、高剂量组(4.8、9.6、19.2 g·kg^-1)，以24 h基线睡眠作为对照，进行单次灌胃给药，从脑电水平考察各组药物的镇静催眠作用。（2）另取32只SD大鼠随机分为正常对照组、模型组、地西泮组(3 mg·kg^-1)和BXDJJ组(9.6 g·kg^-1)，灌胃给药，每日1次，连续7 d。复制多平台水环境睡眠剥夺大鼠模型，每天睡眠剥夺20 h，连续7 d。实验过程中观察大鼠一般状态，于睡眠剥夺第1、4、7天检测血清超氧化物歧化酶（SOD）、过氧化氢（H₂O₂）、丙二醛（MDA）、游离脂肪酸（NEFA）、总胆固醇（TC）及血糖水平。睡眠剥夺第7天，进行大鼠前肢抓力测试和24 h自发性活动监测；采用ELISA法测定血清中白细胞介素1β（IL-1β）、白细胞介素6（IL-6）、白细胞介素10（IL-10）水平；采用生化反应试剂盒检测前额叶皮层及下丘脑组织中SOD、MDA、H₂O₂水平；Western Blot法检测下丘脑组织中昼夜节律基因BMAL1、PER2的蛋白表达水平。结果 （1）与基线比较，BXDJJ中、高剂量组大鼠的总睡眠时间百分比明显增加（P<0.05）,BXDJJ具有镇静催眠作用，选择BXDJJ中剂量(9.6 g·kg^-1)进行后续实验。（2）与正常对照组比较，模型组大鼠前肢抓力明显降低（P<0.05），在2∶00时的自发性活动总路程显著减少（P<0.01），暗周期活动度明显降低（P<0.05）；第1天的血清SOD活力显著下降（P<0.01）,H₂O₂与MDA含量明显增加（P<0.05,P<0.01），第4天的血清H₂O₂和MDA含量均明显增加（P<0.05,P<0.01），第7天的血清H₂O₂含量显著增加（P<0.01）；皮层H₂O₂和MDA水平均明显上升（P<0.05,P<0.01），下丘脑中SOD活力明显下降（P<0.05）,MDA水平显著上升（P<0.01）；第1、4天的血清NEFA、TC水平均显著升高（P<0.01），第4天的血糖水平显著升高（P<0.01）；血清中IL-1β、IL-6水平显著升高（P<0.01）,IL-10水平明显降低（P<0.05）；下丘脑组织中BMAL1蛋白表达显著下调（P<0.01）,PER2蛋白表达显著上调（P<0.01）。与模型组比较，BXDJJ组大鼠前肢抓力明显升高（P<0.05），在2∶00时的自发性活动总路程明显增加（P<0.05）；第1天的血清SOD活力明显升高（P<0.05）,H₂O₂和MDA含量均明显减少（P<0.05,P<0.01），第4天的血清H₂O₂和MDA含量均明显减少（P<0.05,P<0.01）；皮层H₂O₂和MDA水平明显下降（P<0.05），下丘脑中SOD活力明显升高（P<0.05）,MDA水平明显下降（P<0.05）；第1、4天的血清NEFA、TC水平显著降低（P<0.05,P<0.01），第4天的血糖水平显著下降（P<0.01）；血清中IL-1β、IL-6水平明显降低（P<0.05）,IL-10水平明显升高（P<0.05）；下丘脑组织中BMAL1蛋白表达明显上调（P<0.05）,PER2蛋白表达显著下调（P<0.01）。结论 BXDJJ以镇静催眠作用为基础，可能通过调节昼夜节律基因BMAL1、PER2的表达，从而改善睡眠剥夺导致的大鼠昼夜节律异常、氧化应激、糖脂代谢紊乱及炎症反应。更多还原

【Abstract】 Objective To investigate the effect and mechanism of modified Tianwang Buxin Dan （BXDJJ） in improving sleep deprivation injury in rats.Methods （1） Twenty-four SD male rats were randomly divided into Diazepam group (3 mg·kg^-1) and BXDJJ low-,medium-and high-dose groups (4.8,9.6 and 19.2 g·kg^-1),and24-hour baseline sleep was used as control for single gavage administration to investigate the sedative-hypnotic effects of the drug in each group from the EEG level.（2） Another 32 SD rats were randomly divided into normal control group,model group,Diazepam group (3 mg·kg^-1) and BXDJJ group (9.6 g·kg^-1) and administered by gavage once daily for 7 consecutive days.The multi-platform water environment sleep deprivation rat model was replicated with 20 hours of sleep deprivation per day for 7 consecutive days.During the experiment,the general status of the rats was observed and serum levels of superoxide dismutase （SOD）,hydrogen peroxide （H₂O₂）,malondialdehyde （MDA）,free fatty acids（NEFA）,total cholesterol （TC） and blood glucose were measured on days 1,4 and 7 of sleep deprivation.On the seventh day of sleep deprivation,forelimb grip strength test and 24-hour spontaneous activity monitoring were performed in rats.The levels of interleukin-1β （IL-1β）,interleukin-6 （IL-6） and interleukin-10 （IL-10） in serum were measured by ELISA method,the levels of SOD,MDA and H₂O₂in prefrontal cortex and hypothalamus were detected by biochemical reaction kit,and the protein expression levels of circadian genes BMAL1 and PER2 in hypothalamus were detected by Western Blot.Results （1） Compared with baseline,the percentage of total sleep in the rats in the BXDJJ medium-and high-dose groups was significantly increased （P<0.05）.BXDJJ had a sedativehypnotic effect and the medium-dose of BXDJJ (9.6 g·kg^-1) was selected for subsequent experiments.（2） Compared with the normal control group,the forelimb grasping strength of rats in the model group was significantly reduced （P<0.05）,the spontaneous activity distance at 2∶00 was significantly reduced （P<0.01） and the dark cycle activity was significantly decreased （P<0.05）;serum SOD activity was significantly decreased （P<0.01） and H₂O₂and MDA levels were significantly increased （P<0.05,P<0.01） on day 1,both serum H₂O₂and MDA levels were significantly increased （P<0.05,P<0.01） on day 4,and serum H₂O₂level was significantly increased （P<0.01）on day 7;levels of both cortical H₂O₂and MDA were significantly increased （P<0.01,P<0.05）,SOD activity in hypothalamus was significantly decreased （P<0.05） and MDA level was significantly increased （P<0.01）;serum NEFA and TC levels on days 1 and 4 were significantly increased （P<0.01）,and blood glucose level on day 4 was significantly increased （P<0.01）;serum IL-1β and IL-6 levels were significantly increased,and IL-10 level was significantly decreased （P<0.05）.The protein expression of BMAL1 was significantly down-regulated （P<0.01）and that of PER2 protein expression was significantly up-regulated （P<0.01） in hypothalamus tissue.Compared with the model group,the forelimb grasp strength of rats in the BXDJJ group was significantly increased （P<0.05）,and the total distance of spontaneous activity at 2∶00 was significantly increased （P<0.05）;serum SOD activity was significantly increased （P<0.05） and both H₂O₂and MDA levels were significantly reduced on day 1 （P<0.05,P<0.01） and on day 4 （P<0.05,P<0.01）;levels of cortical H₂O₂and MDA were significantly reduced （P<0.05）,SOD activity in the hypothalamus was significantly increased （P<0.05） and MDA level was significantly reduced（P<0.05）;serum NEFA and TC levels on days 1 and 4 were significantly reduced （P<0.05,P<0.01）,and the blood glucose level was significantly decreased on day 4 （P<0.01）;serum IL-1β and IL-6 levels were significantly decreased （P<0.05） and IL-10 level was significantly increased （P<0.05）;BMAL1 protein expression was significantly up-regulated （P<0.05） and PER2 protein expression was significantly down-regulated （P<0.01） in hypothalamic tissue.Conclusion BXDJJ is based on sedative-hypnotic effects and may improve abnormal circadian rhythms,oxidative stress,disrupted glucolipid metabolism and inflammatory responses caused by sleep deprivation in rats by regulating the expressions of circadian genes BMAL1 and PER2.更多还原