【作者】 易世宏；

【导师】 陈宝田；

【作者基本信息】 南方医科大学 ， 中西医结合临床， 2008， 博士

【摘要】 睡眠约占正常人生命三分之一的时间,是维持人体正常生命活动的一个重要的生理过程。健康的睡眠不但可以帮助人们消除疲劳,恢复体力和精力,提高学习、工作的效率,而且还能增强机体的免疫力,有益于抵抗疾病的发生和帮助疾病的恢复。近年来,随着社会的发展,生活节奏的加快,工作学习的紧张,竞争意识的增强,失眠的发生率也不断上升。失眠症是指睡眠的始发和睡眠维持发生障碍,致使睡眠质量不能满足个体需要而明显影响患者白天活动(如精神萎靡不振、注意力下降、反应迟钝等)的一种睡眠障碍综合征。中医学将该病属于“不寐”“不得眠”“不得卧”等范畴。现代医学对失眠的研究主要基于对睡眠机制的探索,从睡眠的相关神经结构到目前研究热点——睡眠物质,运用尖端的仪器设备,先进的分子生物技术,从器官、组织到细胞、分子、基因等水平进行深入的研究。由于睡眠的机制尚未完全清楚,因此镇静催眠就成为失眠治疗的权宜之计。由于现代医学用还原分析的方法研究睡眠,对于整体的重视程度相对薄弱,因而在研究睡眠机制和治疗睡眠相关疾病时,忽略了人身的整体性,比如用于治疗失眠的药物,往往直接针对中枢神经系统,对中枢神经系统有一定的抑制作用,同时还会使患者不同程度地产生依赖性、戒断症状和“宿醉”现象,对于失眠的康复不利,病人也难以接受。因此,人们积极地在中医学领域寻找和探索治疗失眠症的理论和方法。名中医陈宝田教授在研读《内经》的基础上,综合中医各家对失眠的认识,基于中医学从功能角度、从整体角度、从变化角度把握生命基本规律的特征,提出了“平阴阳、调五神、和营卫”的整体治疗大法,在经方柴胡桂枝汤的基础上创立了安眠方,临床上治疗失眠症取得了很好的疗效。本文旨在系统地观察安眠方治疗失眠症的临床疗效和通过动物实验探讨安眠方治疗失眠症的药效学作用基础及其机理的推测,为安眠方的开发提供可靠的理论和临床依据。目的:学习名中医的宝贵经验,挖掘中医药治疗失眠症的潜力。通过临床观察探讨安眠方治疗失眠症的临床效果并对“平阴阳、调五神、和营卫”整体治疗大法进行检验;通过动物实验探讨安眠方治疗失眠症的药效学基础及其治疗失眠症的神经-内分泌-免疫网络调节机制,为安眠方的开发提供可靠的理论和实验依据。方法:1、临床疗效观察:符合纳入标准的失眠患者80例,按照就诊的时间随机分为安眠方治疗组和艾司唑仑对照组。采用国际通用的匹兹堡睡眠质量指数量表(PSQI)、中医证候量表和阳性对照药物艾司唑仑进行疗效比较,进行治疗前后的自身对照和治疗后组间比较,观察安眠方对失眠患者的夜间睡眠质量和白天机能状态的改善情况,从而对安眠方治疗失眠症疗效做出科学、客观的评价。2、实验研究:1)药效学基础:用药理生理实验多用仪观察安眠方对昆明小鼠自主活动的影响,48只小鼠随机分为空白对照组、艾司唑仑对照组和安眠方组,每组16只,分别给予蒸馏水、艾司唑仑水溶液、安眠方水煎液灌胃,观察安眠方对小鼠自主活动的影响。24只昆明小鼠随机分为空白对照组、艾司唑仑对照组和安眠方组,每组8只。正常对照组给予蒸馏水,安眠方组给予安眠方煎液,艾司唑仑组给予艾司唑仑水溶液,灌胃体积及剂量同上,连续7天。于末次给药60min后,小鼠腹腔注射戊巴比妥钠30mg/kg,观察安眠方对小鼠的睡眠潜伏期及睡眠持续时间的影响。60只小鼠随机分为空白对照组、艾司唑仑对照组和安眠方组,每组20只,腹腔注射戊巴比妥钠后,分别给予蒸馏水、艾司唑仑水溶液、安眠方水煎液灌胃,观察安眠方对戊巴比妥钠阈下剂量小鼠睡眠率的影响。2)药理学机制:Wistar大鼠随机分为正常组、模型组、安眠方组和艾司唑仑对照组四组。用腹腔注射对氯苯丙氨酸(PCPA)的方法复制大鼠失眠模型,用荧光分光光度法、放射免疫、免疫组化、半定量RT-PCR等方法观察安眠方对大鼠脑组织五羟色胺(5-HT)、五羟吲哚乙酸(5-HIAA)、去甲肾上腺素(NE)、γ-氨基丁酸(GABA)、白介素-1β(IL-1β)、食欲素(Orexin)和神经肽Y(NPY)的影响。结果:1临床观察:两组在性别、年龄段、病程段等方面比较无统计学意义(P均＞0.05),具有可比性。匹兹堡睡眠质量指数调查,安眠方组和艾司唑仑组各自治疗前后比较均有统计学意义(P均＜0.01),安眠方和艾司唑仑均能改善患者的睡眠时间、入睡时间、睡眠效率、睡眠紊乱及总分。治疗后,安眠方组与艾司唑仑组睡眠时间和睡眠效率项目比较有统计学意义(P均＜0.01),安眠方在改善患者睡眠时间和睡眠效率方面优于艾司唑仑;而入睡时间、睡眠紊乱、日间功能和总分等项目比较均无统计学意义(P均＞0.05),两者疗效相当。安眠方组和艾司唑仑组改善中医症状各自治疗前后比较均有统计学意义(P均＜0.01),安眠方和艾司唑仑均能改善中医各症状。治疗后,烦躁郁闷症状比较有统计学意义(P=0.031),安眠方改善烦躁郁闷疗效优于艾司唑仑。入睡困难、惊悸易醒、晨起困倦、精神萎靡比较无统计学意义(P均＞0.05),两者疗效相当。两组临床疗效比较无显著意义,(P=0.107),安眠方疗效与艾司唑仑相当。两组中医症状疗效比较有显著意义(P=0.008),安眠方在改善中医症状疗效优于艾司唑仑。安眠方对失眠症各型的疗效比较无统计学意义(P=0.775),安眠方对各证型的疗效相当。艾司唑仑对失眠症各型的疗效比较无统计学意义(P=0.830),艾司唑仑对各证型的疗效相当。治疗后,安眠方组患者各实验室指标检测无异常,艾司唑仑组患者有2例肝功能异常,安眠方的临床安全性较高。2、实验研究:安眠方和艾司唑仑均能减少小鼠的自主活动次数,艾司唑仑效果优于安眠方。安眠方和艾司唑仑均能减少小鼠的睡眠潜伏期,两者效果相当。安眠方和艾司唑仑均能增加小鼠的睡眠持续时间,艾司唑仑优于安眠方(P=0.000)。安眠方和艾司唑仑均能提高小鼠的入睡率,两者效果相当。安眠方和艾司唑仑均能增加大鼠脑内5-HT的含量,安眠方效果优于艾司唑仑(P=0.000)。安眠方组能增加大鼠脑内的5-HIAA,与艾司唑仑组比较有统计学意义(P=0.000)。安眠方组和艾司唑仑组均能增加大鼠脑内的GABA含量,安眠方组与艾司唑仑组比较有统计学意义(P=0.000),安眠方效果优于艾司唑仑。安眠方组能减少大鼠脑内NE的含量,与艾司唑仑组比较有统计学意义(P=0.000),效果优于艾司唑仑。安眠方组能增加大鼠脑内IL-1β的含量,与艾司唑仑组比较有统计学意义(P=0.000),效果优于艾司唑仑。安眠方能抑制orexin在大鼠下丘脑的表达,与艾司唑仑组比较有统计学意义(P=0.000)。安眠方能增加NPYmRNA在下丘脑区域的表达,与艾司唑仑组比较有统计学意义(P=0.000),效果优于艾司唑仑。结论:1、临床研究:安眠方不仅能提高患者晚间的睡眠质量,而且能够改善患者白昼的机能状态,从而提高患者白昼的学习、工作效率。安眠方能够调整患者的阴阳失衡状况,调和营卫,使五脏功能协调,气血调和,五神各有所藏,从根本上治疗失眠症。“平阴阳、调五神、和营卫”治疗大法对失眠症行之有效。2、实验研究:安眠方能够使小白鼠自主活动次数减少,协同戊巴比妥钠增加小鼠的入睡率,减少睡眠潜伏期,增加睡眠持续时间。说明安眠方具有肯定的镇静促眠作用。安眠方可使PCPA化失眠大鼠脑内降低的5-HT,5-HIAA含量明显升高,达到正常水平,并可降低失眠大鼠脑内NE含量,提示该方改善失眠大鼠睡眠的作用是增加其脑内5-HT合成与代谢、降低NE含量,恢复5-HT通路与NE通路之间的相互平衡和制约,进而恢复正常的睡眠—觉醒节律,纠正失眠。安眠方可增加PCPA化失眠大鼠脑中IL-1β的含量,使之达到或超过正常水平,说明安眠方调节睡眠的机制与调节免疫功能、增加具有促眠作用的因子有关。安眠方能增加PCPA化失眠大鼠脑中抑制性氨基酸GABA含量,增强其抑制作用,从而改善PCPA化失眠大鼠的睡眠状况。安眠方能抑制或减少Orexin在大鼠下丘脑的表达;增加NPY mRNA在PCPA化失眠大鼠脑下丘脑区域的表达,改善大鼠的失眠状况。安眠方能够从多靶点、多层次作用于神经-内分泌-免疫网络,整体调节机体的内环境,改善失眠的内稳态,从根本上治疗失眠症,比化学合成药物更有优势,值得临床开发应用。更多还原

【Abstract】 Sleep, occupying one-third time of human lives, is an important physiological process to let people keep fit. Reports indicate that good-sleep not only can help people to eliminate fatigue, recover physical fitness and vigour, raise the efficiency of work and study, but also can enhance immunity of body, counteract and recover disease.The incidence rate of insomnia becomes higher and higher because of development of society , fast living, stress of working and studying and high competition in recent years. Insomnia is a syndrome of sleep-disorder somnipathy, which dosen’t work well during begining and keeping to make people feel tired in daytime. Insomnia is called "Bu mei""Bu de mian ""Bu de wo"etc in traditional Chinese medicine. Modern medical science focus on the mechanism of sleep. It begins from nerve fibric to sleeping substance with modern apparatus and advanced technique. The method of sedative and hypnosis is makeshift for insomnia because of being unaware of the mechanism of sleep nowadays. Modern medicine science pays attention to the method of reduction and ignores the concept of entirety body in the course of researching sleep and treating sleep-disorder disease. Such as drugs for insomnia, it aims at central nervous system and inhibits its function. It will make patients depend on drugs, occur abstinence syndrome and hangover. It is adverse to recover disease and the patients can’t accept it. So people seek after new theory and method about insomnia in traditional Chinese medicine. Professor Chen is a famous doctor in TCM, who is goood at treating sleep-disorder disease. Based on his research on one of TCM classical works ’NeiJing’, the treatment method of "Pingyinyang, Tiaowushen, Heyingwei"was put forward. It accords with the nature of TCM: concept of function, concept of entirety body, concept of change. Professor Chen also founded AnMianfang and got remarkably clinical curative effect. This paper aims at observing the clinical curative effect of treating insomnoia with AnMianfang. And explore the mechanism of treating insomnoia with AnMianfang through animal experiment. It will privide clinical and academic proof for developing AnMianfang reliably.Objective: To observe the effect of AnMianfang in treating insomnia and test the method of "Pingyinyang, Tiaowushen, Heyingwei". And to explore the mechanism of treating rats of insomnoia model with AnMianfang.Methods: Clinical observation: 80 cases are selected and devided into two groups at random: AnMianfang group and Estazolam group. Comparing the effcet before and after the treatment interclass to know the situation of sleeping at night and stutas during the daytime by the estimate tables of PSQI and TCM symptom schedule.Experiment: 1) pharmacodynamics: observing the effect of KunMing rats’ autonomic activities with pharmacology and physiology instrument. 48 rats were devided into three groups at random: balnk group, estazolam group and AnMianfang group, 16 in each group. Rats were filled with distilled water, estazolam water-solution, AnMianfang decoction respectively to observe the effect of KunMing rats’ sleeping time. 60 rats were devided into three groups at random: balnk group, estazolam group and AnMianfang group, 20 in each group. Rats were filled with distilled water, estazolam water-solution, AnMianfang decoction respectively to observe the effect of KunMing rats’ rate of falling asleep after injecting Pentobarbital into abdominal cavity. 2) pharmacology: 80 Wistar rats were devided into four groups at random: normal group, model group, AnMianfang group and estazolam group, 20 in each group. Duplicate insomnia model rats by means of PCPA abdominal injection. Observe the express of neurotransmitter, cytokine, neuropeptide and etc in rats’ brains. The observing contents conclude 5-HT, 5-HIAA, NE, GABA, IL-1β, orexin and NPY. The methods of detection conclude radioimmunoassay, immunohistochemistry, reverse transcriptase PCR and etc. Results: 1、Clinical observation: The difference among sex, age and course ofdisease is not significant between AnMianfang group and Estazolam group. The twogroups are comparable (P>0.05). The difference of the items of PSQI are significantbefore and after treating with AnMianfang or Estazolam between AnMianfang groupand Estazolam group (P<0.01) . AnMianfang and Estazolam can improve thesleeping time, the falling sleep time, the sleep efficiency, the sleep disorder and thetotal sleep marks. The difference of sleeping time and sleep efficiency is significantbetween AnMianfang group and Estazolam group after treating (P<0.01) .AnMianfang can improve sleeping time and sleep efficiency better than Estazolam.The other items of PSQI are not significant between AnMianfang group andEstazolam group after treating (P>0.05) . AnMianfang and Estazolam both canimprove symptom of TCM remakbaly before and after treating (P<0.01) . Thesymptom of restlessness and morosity can be lessened visibly after treating (P =0.031) . AnMianfang can lessen restlessness and morosity better than Estazolam.The other symptom of TCM are not significant between AnMianfang group andEstazolam group after treating (P>0.05) . The clinical curative effect betweenAnMianfang group and Estazolam group is comparable (P>0.05) . The clinicalcurative effect of TCM between AnMianfang group and Estazolam group issignificant (P<0.01) . AnMianfang can improve symptom of TCM better thanEstazolam. The difference of the items of polysomnogram are significant before andafter treating with AnMianfang or Estazolam between AnMianfang group andEstazolam group (P<0.01) . AnMianfang and Estazolam can improve the totalsleeping time, REM sleep time, the sleep incubation period and the awareness time.After treating, the items of polysomnogram are comparable between AnMianfanggroup and Estazolam group (P>0.05) . The clinical curative effect of every patternof syndrome of insomnia is comparable treated by AnMianfang (P=0.775) . Theclinical curative effect of every pattern of syndrome of insomnia is comparabletreated by Estazolam (P = 0.830) . The indexs of laboratory for patients in AnMianfang group are normal after treating, but two patients whose hepatic function are abnormal in Estazolam group. AnMianfang is safty for insomnia patients.2 Experimental research: AnMianfang and Estazolam can reduce the frequency of rats’ autonomic activities, the effect of Estazolam is better than that of AnMianfang. AnMianfang and Estazolam can decrease the sleeping incubation period, the effect is comparable between AnMianfang and Estazolam (P>0.05) . AnMianfang and Estazolam can increase the sleep duration, the effect of Estazolam is better than that of AnMianfang (P = 0.000) . AnMianfang and Estazolam can raise the rate of falling sleep, the effect is comparable between AnMianfang and Estazolam. AnMianfang and Estazolam can increase the content of 5-HT in rat brain, the effect of AnMianfang is better than that of Estazolam (P = 0.000) . AnMianfang can enhance the content of 5-HIAA obviously comparing with Estazolam group (P = 0.000) . AnMianfang and Estazolam can increase the content of GABA in rat brain, the effect of AnMianfang is better than that of Estazolam (P=0.000) . AnMianfang can decrease the content of NE in rat brain, the effect of AnMianfang is better than that of Estazolam (P=0.000). AnMianfang can increase the content of IL-1βin rat brain, the effect of AnMianfang is better than that of Estazolam (P=0.000). AnMianfang can increase the express oof NPY mRNA in area of hypothalamus and hippocampus, the effect of AnMianfang is better than that of Estazolam (P=0.000) .Conclusions: 1 Clinical observation: AnMianfang can not only raise the quality of sleep at night but also can improve the state of body in daytime. It is benificial for patients to raise the efficiency of studying and working in daytime. It also can improve the state of lost of Ying and Yang, harmonize Ying Wei, WuZang and QiXue. It can cllect WuSheng for their owns and cure insomnia at all.2 Experimental research: AnMianfang can decrease the frequency of rats’ autonomic activities, increase the rate of falling sleep, decrease the incubation period of sleeping, and increase the time of remaining sleeping. It can prove that AnMianfang has the function of sedative and hypnosis.AnMianfang can reduce the level of 5-HT in model rats’ brains, improve the content of 5-HIAA obviously, or even than that of normal rats, debase the content of NE in rats’ brains. From above, we can coclude that the mechanism of AnMianfang is enhancing the synthesis and metabolism of 5-HT, debasing the content of NE, rebounding the iter balance between 5-HT and NE, and restoring the normal rhythm between awareness and sleeping.AnMianfang can increase the content of IL-1βobviously in rats’ brains, or even than that of normal rats. It shows that there is something with adjusting immune function and improving the cytokine of hypnosis for the mechanism of AnMianfang.AnMianfang can increase the content of GABA in rats’ brains, enhancing the role of inhibition to improve the state of insomnia rat with injected PCPA.AnMianfang can increase the express of Orexin and NPY mRNA in the area of hippocampus and hypothalamus of rats’ brains to improve the state of insomnia.The data indicates that AnMianfang possesses the property to facilitate the neroendocrine-immune network by multiple ways to exact its sedation and hypnosis and the treatment for insomnia. It also can adjust the internal environment of body and rebound the homeostasis of insomnia. It is less side effect and better curative effect than those of synthetic chemicals. It will be worthy of developping in clinic.更多还原