针药结合对抑郁症模型大鼠海马神经元信号转导机制的影响及临床研究
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摘要
目的:
     1.明确针刺结合抗抑郁药治疗抑郁症的疗效;分析、总结针药结合治疗抑郁症的优势。2.探索针(电针)、药(氟西汀)、电针结合氟西汀对抑郁症模型大鼠海马神经元保护和促进神经元发生的信号转导机制影响;3.探求中医学对抑郁症发病机制的认识,明确抑郁症的病位,明确抑郁症辨证论治的目标。分析抑郁症辨证的正确方法,为针药结合治疗抑郁症提供辨证理论基础。
     方法:
     目的1:(1)文献综述针药结合、针(电针)治疗抑郁症的临床与实验研究。(2)临床研究:将45例患者根据单盲、随机原则分为观察组和对照组。对照组单纯采用SSRIs类药物进行治疗;观察组采用针药结合治疗:在SSRIs类药物治疗基础上,加以督脉经穴为主、导气针法的针刺治疗。共治疗6周。分别于治疗前及治疗后1、2、4、6周对HAMD总分、因子分进行评价,对两组得分进行t检验,判断组间差异;根据HAMD量表的减分率,于治疗6周后分别对两组的临床疗效进行评定,卡方检验分析其疗效差异。目的2:(1)文献综述抑郁症海马神经元损伤和神经元发生障碍的神经生物学机制,说明信号转导对海马神经损伤和神经元发生障碍的作用和重要性。分析抗抑郁药对受体后信号转导机制的作用环节,寻找针药结合对海马信号转导机制影响研究的作用位点。(2)实验研究:采用分养和中等强度不可预见应激制备抑郁症大鼠模型,根据行为学得分随机分为电针组、药物(氟西汀)组、针药组,和正常组,以观察电针、药物(氟西汀)、针药结合(电针结合氟西汀)对模型大鼠体重、open-field运动得分、24小时糖水消耗量及海马内cAMP浓度、cAMP反应元件结合蛋白(CREB)、脑源性神经生长因子(BDNF)在治疗7天、21天时的变化。放射免疫法检测cAMP浓度,免疫组织化学法检测CREB、BDNF的表达,计算机图象分析系统半定量分析CREB、BDNF的在海马CA3区、齿状回的平均光密度。检测结果在各治疗组之间进行对照(F检验),并分别与正常组进行对照(t检验)。目的3:(1)对抑郁症中医辨证文献进行证型证候因子频次分析,寻找抑郁症脏腑辨证的规律和基础病机、明确病理性质。(2)总结抑郁症的辨证方法。(3)利用脏腑升降理论从整体性论述脑与脏腑关系。
     结果:
     目的1:(1)针药结合文献综述表明:针药结合治疗可以①有效地改善虑/躯体化、体重、睡眠因子症状优于单纯抗抑郁药物治疗;②提高疗效,对于临床抑郁症的治疗难点,如难治性抑郁症、症状中的失眠等;③减少抗抑郁药用量;④体现在弥补抗抑郁药延迟方面。电针治疗抑郁症的临床研究表明:单纯电针治疗抗抑郁效应等同于抗抑郁药(阿米替林、麦普替林,氟西汀);电针抗抑郁效应产生的机理研究表明:电针可以调整抑郁症模型大鼠中枢及外周单胺递类质水平、受体的数量和功能、亢进的HPA轴功能,可以提高抑郁症模型大鼠皮质和海马p-CREB表达水平、保护海马BDNF、抑制慢性应激造成的大鼠海马神经元凋亡。(2)临床研究结果:从开始治疗第1周起,与对照组相比,观察组HAMD总评分、焦虑/躯体化、体重、阻滞、睡眠障碍因子出现了明显的改善,这种效应持续至治疗第4周,而焦虑/躯体化一直延续到第6周。观察组体重因子从第2周始停止下降,而对照组体重的增长到第6周时与观察组持平。只有认知因子改善与对照组无明显差异(P>0.05);根据HAMD量表的减分率,结果经卡方检验,P<0.05,表明两组治疗6周后在疗效方面存在显著性差异,观察组优于对照组。
     目的2:(1)文献研究:抑郁症存在海马神经元损伤和神经元发生障碍,BDNF的表达下调是海马神经元损伤和发生障碍的关键环节。抗抑郁药(氟西汀)的药理作用在于突触间隙单胺类递质浓度增加后引起的受体后信号转导机制的适应性调节。其中cAMP-CREB通路是介导BDNF表达增加的关键性环节。(2)电针、氟西汀、电针结合氟西汀都可以改善应激造成的抑郁症模型大鼠的体重、24小时糖水消耗量、及水平和垂直运动积分下降;针药组增长幅度最高,但与单纯电针和药物治疗相比差异无显著性(P>0.05)。除药物组外,电针组、针药组均可以在7天的治疗后显著升高cAMP浓度至正常组水平;治疗21天后,三种治疗均可以升高cAMP浓度、cAMP/cGMP比值,组间比较无差异(P>0.05)。抑郁症模型大鼠存在海马CA3区、齿状回CREB、BDNF平均光密度下降(P<0.01)。治疗7天后,电针组、针药组在CA3区、齿状回的CREB和齿状回BDNF平均光密度与正常组水平相当(P>0.05),药物组低于正常组(P<0.05);在CA3区BDNF平均光密度,电针组、药物组和针药组与正常组相当(P>0.05)。治疗21天后,CREB平均光密度只有针药结合治疗可以同时提高CA3和齿状回至正常水平;而电针组均低于正常组(P<0.01);药物组在CA3区低于正常组(P<0.01),在齿状回与正常组相当(P>0.05)。对于BDNF平均光密度,只有针药组在CA3区和齿状回能同时提高至正常组水平;药物组在CA3区可以,电针组在齿状回可以。
     目的3:(1)证候因子频次分析:抑郁症涉及脏腑众多;肝郁及其相关证型是抑郁症脏腑辨证中惟一的实证因子,频次最高;而其它脏腑证候因子都为虚证。(2)气机紊乱是抑郁症发生基本病机。(3)抑郁症辨证方法有二:以证型归纳症状和症状-证候因子-证型(4)抑郁瘟病位在脑,与脏腑密切相关。抑郁症辨治目标应围绕脑和脏腑的功能失调、脑与脏腑功能的联系失调三个方面进行。(5)抑郁症是属于升降失调的病证。
     结论:
     1.单纯电针、氟西汀、电针结合氟西汀治疗均对抑郁症模型大鼠海马信号转导机制具有调整作用,以电针结合氟西汀治疗疗效最为显著。
     2.抑郁症中医辨证目前主要针对脏腑病变进行,气机紊乱是基本病机,病理性质虚实夹杂。阳虚可能是抑郁症脑部病变机制的主要方面。
     3.针药结合有效且必要。针灸治疗抑郁症具有多靶点、多途径的特点,与治疗靶点明确的抗抑郁药物联合运用,可以有效地弥补抗抑郁药物的不足,实现对抑郁症治疗的整体、全面和系统化。
Keywords: depression; electric-acupuncture; fluoxetine; cAMP; CREB
     [Purpose]: 1. Demonstrate the curative effect of the combination of acupuncture treatmentin combination with antidepressant and analyze and summarize the benefits of the treatmentmethod 2. Explore the signal transduction mechanism of the hippocampal neuronsprotection and the neurogenesis promotion of depressive model rats when treated byelectric-acupuncture, fluoxetine, and combination of both treatment approaches. 3. Explorethe pathogenetic knowledge of depression within the area of traditional Chinese medicine:based in the review of the differentiation of the symptoms and signs according to thepathological patterns of the disorders of depression by identify the location of the disorderand target the cause of depression, continue by analyzing which method will be theappropriate one to apply and propose the rationale explanation behind the choice of thecombination therapeutic regiment for the treatment of depression.
     [Methods]: Purposel: (1) Summarize recent clinical and experimental studies andliteratures about the treatment of electric-acupuncture and the combinationelectric-acupuncture and anti -depressants. (2) Clinical research involves 45 patientsdivided into an observation group and control group unilaterally blind randomly. Thecontrol group was only treated by SSRIs anti-depressants and the observation groupreceived acupuncture as well as anti-depressants. The choice of points was mainly from theDu channel and the manipulations were based on fortifying method. Both groups weretreated for 6 weeks. The evaluations were done by the HAMD score at the onset of thetreatment as well as the first, second, fourth and sixth weeks. The differences wereevaluated by T-test the scores of two groups. The clinical therapeutic effects were evaluated on the reduction of the basis of chi square rating scale of the HAMD score. Purpose 2: (1)The overview of medical literatures explaining the function and the importance of signaltransduction in the neurobiological mechanisms of hippocampus neurons injury andneurogenesis condition during depression. Analyze the effects of antidepressant (fluoxetine)in the transduction of the post-acceptor signal and observe the situs in the signal oftransduction of the treatment in the combination treatment of acupuncture and anti--depressant. (2) Experimental research: After replicated the model, and divided thedepressed rat into the electric-acupuncture group and fluoxetine group. Evaluate thecombination group and fluoxenie group according to ethological scores by observing theweight of model rats, open-field behavior scores, sacchar consumption within 24hours, anddensity of cAMP, CREB, BDNF in hippocampus when treated by different therapy at day 7and day 21. Analyze the data within each therapy groups through F-test and compared withnormal group. Purpose 3: (1) Statistical analysis were done on the type of syndromefactor's frequency in all literatures about depression differentiation of symptoms and signsand rules of entrails differentiation of symptoms and signs about depression and elementarypathogenesis were evaluated to remove the pathological nature. (2) Summarize the methodsof differentiation of symptoms and signs about depression. (3) Analyze the theory of the"brain in relation to the mind". (4) Discuss the relationship between the brain and internalorgans from the view of the theory of ascend and descend.
     [Result]: Purpose 1: (1) The literature overview indicated the treatment of combinationacupuncture and antidepressant could have three purposes:①improve the effect ofsymptoms such as insomnia and weight-loss better than the single antidepressant treatment;②reduce the dosage of antidepressant.③redeem the delay of the effect of theantidepressant since electro-acupuncture also has an anti-depression curative effect. (2)Clinical research result: After the first week of treatment, HAMD total scores, and anxiety,body weight, emotional and intellectual blockage, insomnia factors of the observationgroup improved faster than the control group and increased during the following 4 weeks.Only the cognition factor had no obvious difference when compared with the control group(P>0.05). According to the HAMD score, there was an obvious difference between the twogroups. The observation group had better results according to the HAMD score whencompared with the control group (P<0.05). Purpose 2: (1) Hippocampal neurons damagedand neurogenesis blocked occur in depressive disorder. Down-regulation BDNF expressionis the key point. The pharmaceutical effects of antidepressant contribute to the post-receptor signal transduction mechanism adaptability regulation after the increase of density ofmonoamine transmitter in synaptic cleft. Path of cAMP-CREB is a critical link which madethe mediate BDNG expressions increased. (2) The weight of model rats, open-fieldbehavior scores, sacchar consumption within 24 hours all increased in the combinationtreatment of electro-acupuncture with antidepressant and the controlled group with nosignificant difference (P>0.05).The density of cAMP within 7days which have nodifference with controlled group (P>0.05 ) however, after 21 days of treatment, it increasedin the three groups with no difference existing among them (P>0.05). Average optical ofCREB and BDNF descended in hippocampal CA3 and dentatus gyrus(DG) area ofdepressive model rats. Only the average optical of CREB and BDNF of the CA3 or DGarea of the combination group ascended to the normal level when treated in 7days to 21 days.Purpose 3: (1) Statistical analysis was effectuated on the syndrome factor's frequencyinvolved in the differentiation of symptoms and signs in the depression syndrome.Stagnation of liver-Qi and correlated type of syndrome is the only excessive syndrome witha high frequency and occurs in the early period. Other types of syndrome are all deficiencysyndrome. The disorders of qi are based on the pathogenesis of depression. (2) Basicpathological nature of depression is asthenia syndrome. (3) The location of the syndromeof depression is in brain and is closely related with the internal organs. The differentiationof the signs and symptoms of depressions and the treatment should affect three aspects:functional disorder of the brain and internal organs and the disharmony between the brainand internal organs. (4) Yang deficient is the principle aspect of the differentiation of thesyndrome of depression.
     [Conclusion]: 1. The combination of electro-acupuncture and fluoxetine group andcontrolled groups can both regulate the hippocampal signal transduction pathway ofdepressive model rats. However, the combination treatment is the most effective one.2. InChinese medicine: The disorders of qi are the basis pathogenesis of depression. The basicpathological nature of depression is asthenia syndrome. Yang deficient is the principalaspect of the differentiation of the depressive symptoms and signs of the brain.3. It'seffective and necessary when depression is treated by the combination of acupuncture andwestern medicine. Acupuncture treatment carries out a multitarget multipath clinical effect.It can redeem antidepressant efficiently, treat depression entirety, generically, andsystematically.
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