基于抑郁症和气虚证应激共易感性的病证结合动物模型研究

There is inherent difficulty in revealing the intrinsic association between disease and syndrome based on the model by simply adding the individual models for a disease and TCM syndrome. In contrast, the combination of disease and syndrome model based on the common vulnerability to disease and syndrome under genetic influence has potential advantages. Previous studies found that under the same chronic stress, depressive behaviors and syndromes differed from various strains of mice: Balb/c and 129S1 were both vulnerable to depression, while Balb/c also showed Qi deficiency vulnerability. This project plans to, with 129S1 strains as a control, use following different methods to establish co-vulnerability to depression and Qi deficiency of Balb/c strains for the combination of disease and syndrome model: 1. to confirm co-vulnerability to depression and Qi deficiency under different stress protocols; 2. to validate the model by comparing the efficetiveness of depression and Qi-deficiency following treatment with different Qi tonifying and non-Qi tonifying TCM formula ; 3. to analyze the evolution of depression and syndrome with stress progression， and test effects of syndrome-specific Chinese medicine at different time of disease stages; 4. to associate of the depression and syndromes, curative effects of traditional Chinese medicine with the transcriptome in hippocampus (depression control center), hypothalamus (emotion and homeostasis control center) and intestine (Qi related organ), as well as intestinal microflora, to obtain the specific genes and systems biology evidence of the disease and syndrome combination model. This project will provide a new and stable model for the in-depth research and development of the combination of depression and Qi deficiency syndrome.

将疾病和证候分别造模相加的模型在揭示病证内在关系上存在固有困难，对此，利用遗传影响下的疾病与证候共易感性建立的病证结合模型则有潜在优势。前期研究发现相同的慢性应激条件下，不同品系小鼠抑郁样行为与证候表现各异：其中Balb/c与129S1品系都抑郁易感，而Balb/c还表现为气虚易感。本项目计划以129S1品系为对照，采用多种方法确立Balb/c品系抑郁气虚应激共易感病证结合模型：1.不同应激造模方法下确认其抑郁气虚应激共易感性；2.比较不同益气与非益气中药病证疗效差异，反证病证模型有效性；3.解析病证随应激进程惯序演变规律及不同时间对证中药干预作用；4.将抑郁症证候及对证中药疗效与海马（抑郁调控中枢），下丘脑（情绪与躯体稳态调控中枢）和肠（气虚相关器官）基因表达组及肠道菌群相联系，获得该病证结合模型的特异性基因及系统生物学证据。此项目为深入的抑郁气虚病证结合研究开发提供新的稳定的模型基础。

将疾病和证候分别造模相加的模型在揭示病证内在关系上存在固有困难，对此，利用遗传影响下的疾病与证候共易感性建立的病证结合模型则有潜在优势。通过品系对照的差异性来揭示证候的特异性，在不同应激模型下，Balb/c 小鼠都表现出抑郁样行为与气虚证候，因此该品系具有抑郁症与气虚证的应激共易感性。在此基础上，方药反证结合分子生物学方法显示补气经典方药四君子汤而不是理气方越鞠丸可以明显改善 Balb/c 模型动物表现出抓力下降、负重游泳时间减少、皮毛评分增加的气虚样表型，也改善了糖偏好下降等抑郁样行为。而越鞠丸对 Balb/c 小鼠与129/S1 小鼠抑郁症均有一定治疗作用，可以改善其抑郁症行为表现，但对 Balb/c 小鼠的治疗作用不如四君子汤，并且越鞠丸对 Balb/c 小鼠虚证证候表现没有明显的治疗作用。这样的结果说明，针对疾病具体证候的治疗，效果可能优于单纯针对疾病本身的治疗。不同模型的抑郁气虚发生发展的时程变化上看，CORT模型中，Babl/c小鼠上先出现气虚表型，进一步发展形成抑郁表型；而129S2/SV小鼠上先出现抑郁表型，进一步发展形成气虚表型。Balb/c 小鼠在 CUMS 下的气虚抑郁病证类型。表明Balb/c小鼠的气虚证候发生在抑郁表现之前，客观揭示了疾病与证候出现及应激共易感的时序差异。四君子汤可逆转脑和肌肉 PKA-CREB 信号的下调。动物行为学和肠道菌群测序结果支持 Balb/c 小鼠是建立抑郁气虚应激共易感性的良好模型动物，而慢性习得性无助模型是建立抑郁伴气虚症状的稳定模型。归脾汤能长期改善模型小鼠的抑郁和气虚症状，其作用可能通过改善小鼠肠道菌群来体现。CUMS 造模后，给健脾解郁方治疗后，乳酸菌丰度恢复至正常水平，说明药物治疗有助于维护肠道益生菌的稳定。黄芪太子参可以改善抑郁所致的小鼠盲肠内容物菌群的紊乱，且在改善肠道菌群方面优于氟西汀。总之，我们通过病证的整体特性，选取调控抑郁症，气虚证候及整合脑-体稳态进行系统生物学的检测，通过比较气虚与非气虚，益气与理气的药物处理等，多角度，多水平揭示抑郁症气虚证候的特异性分子及其网络指标。

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