清燥救肺汤分解剂干预MP感染LAMPs模式识别及其调控通路的机制研究

Mycoplasma pneumonia (M. pneumoniae) is one of the most common causes of pediatric pneumonia, with widespread and high pathogenicity. The adhesion and invasion was mainly mediated by M. pneumonia derived lipid-associated membrane proteins (LAMPs), which induced the apoptosis and pulmonary injury. Qingzao Jiufei (QZJF) decoction had an obvious effect in clinical practice and was confirmed that it was also effective on inhibiting inflammation spread in several experimental researches. Therefore, we proposed the hypothesis that there was correlation between dryness-heat injuring lung-yin and M. pneumonia derived LAMPs patten recognization and its regular pathway. QZJF decoction was tonification and purgation in combination, including four rules of clearing, ascending, desending and moistening. Begaining with active substance research, we combined the techniques of monoclonal antibodies with HPLS-MS pharmacokinetic study to explore material basics and target organ. Study was a factorial design. Balb/c mice infected by M. pneumonia were divided into exogenous dryness group and non-exogenous dryness group to observe LAMPs pattern recognization and dynamic change of components through NF-κB dependent-signal pathway with techniques of Western-blot, RT-PCR and so on. For providing research and theory basis, our study attempted to explore the content of dryness and lung fluid through modern biological indicators, to compare effectiveness of QZJF decoction and different decomposer and to discuss the mechanism of integrated regulation how QZJF decoction improved the immune pulmonary injury caused by M. pneumonia.

肺炎支原体(MP)具有广泛致病性，是儿童肺炎重要病原之一。主要由MP脂质相关膜蛋白(LAMPs)介导黏附入侵，损伤细胞，诱导凋亡而导致肺损伤。清燥救肺汤临床疗效确切，实验研究发现能抑制肺部炎症，因而提出"燥热灼伤肺津与MP感染LAMPs模式识别及其通路调控存在一定相关性"假说。清燥救肺汤攻补兼施，内含清宣降润四法，以其有效物质群为切入点，采用单克隆抗体技术，结合HPLC-MS进行全方和分解剂药动学研究，探寻物质基础及作用靶器官。通过析因设计，以Balb/c小鼠MPP模型（有无外燥环境）为对象，采用Western-blot、RT-PCR等方法，观察LAMPs模式识别、NF-κB通路"元件"动态变化，阐明"燥热"、"肺津"现代生物指标内涵，探讨清燥救肺汤分解剂药能效果差异性，揭示改善MPP免疫炎性损伤的整合调节机制，为临床应用提供实验依据与理论支撑。

肺炎支原体(MP)具有广泛致病性，是儿童肺炎重要病原之一。MP感染具有燥热致病的特点，治燥名方-清燥救肺汤干预MP感染取得了较好疗效。该方清燥润肺，攻补兼施，驱邪-“清-宣-降”法，扶正-“润-补”法，可依“法”拆方研究。本课题通过分析MPP所致的“燥热灼伤肺津”的病机特点，创新性的提出“燥热”、“肺津”并与MPP的生物学指标关联，研究清燥救肺汤及其分解剂干预靶器官、靶向效应机制。采用高效液相波长切换法结合单克隆抗体检测技术对清燥救肺汤分解剂活性成分定量测定，筛选最佳提取工艺，并进行质量控制研究。以高效液相色谱法进行清燥救肺汤血清指纹图谱研究，分析其特征峰和共有峰。建立MP镜下观察分级标准，并与测定的CCU和实时荧光定量PCR测定结果进行关联，呈正相关。模拟温燥环境，并与MPP模型比较，发现肺部炎症、SPA、AQP5等指标变化呈一致性，温燥伤肺与MPP炎症具有一定关联性。清燥救肺汤能够减轻肺部炎症、清除毒素、升高AQP5表达、增加SPA含量而体现“润燥”作用。清燥救肺汤控制肺部炎症、减少促炎因子、减少毒素、调节免疫、抑制凋亡、调控TLR2/NF-κB信号通路，分解剂I起主要作用；清燥救肺汤上调免疫、增加抑炎因子，分解剂Ⅱ起主要作用。本研究较系统地揭示清燥救肺汤及其分解剂对改善MPP免疫炎性损伤的整合调节机制，为“从燥论治MP感染”提供理论支持和实验依据。

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