六味地黄苷糖对睡眠剥夺小鼠所致焦虑抑郁样行为的作用及其机制研究

Effects and mechanisms of Liuwei Dihuang Active Fraction Combination on anxiety-and depression-like behavior induced by sleep deprivation in mice

目的观察六味地黄苷糖(LW-AFC)对睡眠剥夺小鼠所致焦虑抑郁样行为的作用及其机制。方法雄性C57小鼠依据小鼠体质量和自主活动性随机分为正常对照组、睡眠剥夺模型组、LW-AFC 0.8 g/kg、1.6 g/kg和3.2 g/kg组。应用72 h多平台水环境法建立失眠动物模型。给药组预防给药7 d后进行睡眠剥夺72 h,同时给予LW-AFC至建模后第14天。采用旷场实验评价小鼠的焦虑样行为,采用悬尾和强迫游泳实验评价小鼠的抑郁样行为,应用液相芯片分析技术检测小鼠海马组织中促炎因子IL-1β、IL-6、TNF-α以及抗炎因子IL-4的含量。采用Pearson相关性分析LW-AFC对睡眠剥夺小鼠细胞因子的影响与焦虑抑郁样行为的相关性。采用Western印迹法检测小鼠海马星形胶质细胞激活标志物胶质原纤维酸性蛋白(GFAP)的变化。结果与正常对照组相比,睡眠剥夺模型组小鼠在旷场中央的探索时间显著缩短(P<0.05),悬尾和强迫游泳的不动时间显著延长(P<0.05),小鼠海马组织中GFAP表达显著升高(P<0.01),IL-1β、IL-6和TNF-α含量均显著增加(P<0.01),IL-4含量显著降低(P<0.01);与睡眠剥夺模型组相比,LW-AFC 1.6和3.2 g/kg组小鼠旷场实验的中央探索时间显著延长(P<0.05),GFAP表达量显著降低(P<0.05)。LW-AFC 3个剂量组小鼠悬尾和强迫游泳的不动时间显著缩短(P<0.05,P<0.01),海马组织中IL-1β和IL-6含量均显著降低(P<0.05,P<0.01);LW-AFC 1.6和3.2 g/kg组小鼠海马组织中TNF-α含量均显著降低(P<0.01),IL-4含量显著增加(P<0.05,P<0.01)。Pearson相关性分析结果提示,LW-AFC改善小鼠的焦虑抑郁样行为与促炎细胞因子(IL-1β、TNF-α)密切相关(P<0.05,P<0.01)。结论 LW-AFC能改善睡眠剥夺导致的小鼠焦虑和抑郁样行为改变,其机制可能与其调节细胞因子的失衡以及抑制星形胶质细胞活化有关。

Objecive To investigate the effects of Liuwei Dihuang Active Fraction Combination(LW-AFC)on anxiety-and depression-like behavior induced by sleep deprivation(SD)in mice and its mechanism.Methods The male C57 mice were randomly divided into normal control group,SD model group and LW-AFC 0.8,1.6 and 3.2 g/kg groups according to their body weight and autonomic activity.SD animal model was established by 72-hour multi-platform water environment method.After 7 days of preventive administration of LW-AFC,mice in the LW-AFC groups were deprived of sleep for 72 hours,and LW-AFC was given to mice until the14 th day of the end of SD.Anxiety-like behavior of mice was evaluated by open field test,depression-like behavior was evaluated by the tail suspension and forced swimming tests;and the levels of pro-inflammatory factors(IL-1β,IL-6,and TNF-alpha)and anti-inflammatory factor(IL-4)in hippocampus of mice were detected by Luminex protein array system.The Pearson correlation analysis was used to analyze the correlation between the effect of LW-AFC on cytokines and anxiety-depression-like behavior in SD mice.The expression of glial fibrillary acidic protein(GFAP),an activation marker of astrocyte in hippocampus of mice was examined by Western blot.Results Compared with the normal control group,the time of central exploration in the open field of the SD model group was significantly shortened(P<0.05),and the immobility time in the tail suspension and forced swimming tests was significantly prolonged(P<0.05),the expression of GFAP was significantly inereased(P<0.01),the level of IL-1β,IL-6 and TNF-αin the SD model group was significantly increased(P<0.01),while the level of IL-4 was significantly decreased(P<0.01).Compared with the SD model group,the central exploration time was significantly prolonged in the LW-AFC 1.6 and 3.2 g/kg groups(P<0.05),and the immobility time in the tail suspension and forced swimming tests were significantly shortened in the LW-AFC 0.8,1.6 and 3.2 g/kg groups(P<0.05,P<0.01),and the expression of GFAP was significantly decreased in 1.6 and 3.2 g/kg groups(P<0.05).Meanwhile,the level of IL-1βand IL-6 was significantly decreased in the LW-AFC 0.8,1.6 and 3.2 g/kg groups(P<0.05,P<0.01)and the level of TNF-αwas significantly decreased in the LW-AFC 1.6 and 3.2 g/kg groups(P<0.01),while the level of IL-4 was significantly increased(P<0.05,P<0.01)compared with the SD model group.The Pearson correlation analysis suggested that LW-AFC improved the anxiety-and depression-like behavior in mice,which was closely related to the proinflammatory cytokines(IL-1βand TNF-α)(P<0.05,P<0.01).Conclusion LW-AFC could ameliorate the anxiety-and depression-like behavior induced by SD in mice.The mechanism was most likely related to the regulation of imbalanced cytokines and inhibition of the astrocyte activation.