基于TGF-β/Smad信号通路的中药莲子心抗肺纤维化物质基础及作用机制研究

Idiopathic pulmonary fibrosis(IPF) is a progressive interstitial lung disease，which is characterized by a poor prognosis as well as a high lethality. Recent studies have proved that conventional treatment of glucocorticoid and acetylcysteine are inefficient, and there have not been effective drugs to cure the IPF presently. Pirfenidone(PFD), which acts on the TGF-β pathway, is the first drug approved to treat IPF. Our studies found that total alkaloid of Plumula nelumbinis possess a remarkable effect of anti-lung fibrosis. It can significantly reduce the proliferation of the lung fibroblasts as well as accumulation of extracellular matrix of mice. In addition, the high expression of TGF-βmRNA was inhibited as well. The effect of total alkaloid of plumula nelumbinis is comparable to that of PFD. As alkaloids of liensinine share the same Pyridine structure with PFD , it is promising to find new leading compounds which are structurally and functionally similar to PFD. In this project, we establish cell level high content screening to screen active fractions and active compounds that have the anti-lung fibrosis effect. RT-PCR and Western Blot are employed in the investigation of compounds impact on the pathway of TGF-β/smad, explaining their material basis and mechanisms and providing a foundation for the development of novel anti-lung fibrosis drugs.

特发性肺纤维化(IPF)是一种渐进性肺间质疾病，具有预后较差，病死率高的特点。近期研究证明传统用于治疗IPF的糖皮质激素及乙酰半胱氨酸无效，目前临床上缺乏有效的治疗药物。吡非尼酮（PFD）是全球首个获批治疗IPF的药物，其作用于TGF-β通路。我们研究发现中药莲子心总生物碱活性部位具有显著的抗IPF作用，可显著减少小鼠肺成纤维细胞增殖及胞外基质沉积，显著抑制小鼠TGF-βmRNA的表达；并且莲子心生物碱类成分具有与PFD相似的结构，因此有希望从莲子心获得与PFD结构及作用途径类似的新型抗IPF先导化合物。本项目建立基于荧光细胞成像技术的高内涵药物筛选系统，追踪莲子心抗IPF活性成分，从细胞-整体动物水平研究活性成分的抗IPF作用，利用RT-PCR、Western Blot等技术研究其对TGF-β/smad信号通路的影响，揭示莲子心抗IPF物质基础及作用机制，为研发新型抗IPF药物奠定基础。

特发性肺纤维化（IPF）是一种发病率高、致死率高的肺部疾病，是目前医学的治疗难题，临床上缺乏有效的治疗药物。本课题在前期研究基础上，对中药莲子心抗肺纤维化的物质基础及其调控TGF-β1/Smad 信号通路的作用机制进行研究。项目首先采用HPLC法建立了药材中有效成分生物碱类成分的含量测定方法，有效地控制药材的质量；运用现代分离技术对莲子心生物碱活性部位及其化学成分进行分离、纯化、富集，分离并鉴定了其中16个生物碱类成分。建立体内体外活性筛选方法用于提取物及化合物活性筛选，明确了总生物碱的活性部位。采用博来霉素气管内注入法成功复制了小鼠肺纤维化模型，评价莲心总碱对肺纤维化小鼠的保护作用，采用RT-PCR、Western-blot等方法，阐明了莲心总碱调控TGFβ1/Smad信号通路的作用机制。研究结果表明莲心总碱对博来霉素致肺纤维化小鼠具有明显的保护作用，其机制可能通过抑制炎症因子的生成，减轻炎症反应；下调TGFβ1/Smad信号通路中TGF-β1、Smad3及α-SMA基因的表达，进而降低其翻译产物TGF-β1、Smad3及其磷酸化蛋白p-Smad3和α-SMA蛋白的表达，降低肺组织中HYP含量；抑制小鼠氧化应激反应，清除氧自由基及抑制脂质过氧化物生成，进而有效干预IPF的进程。该研究为IPF防治提供科学依据，并为新药研制奠定基础。