基于治胆从肝理论干预胆石症的作用机制研究

According to the theory of Chinese medicine theory "biliary diseases treated by differentiating syndromes of the liver", the common key factors of cholelithiasis are liver dysfunction, gallbladder lost flowing and descending, and bile coagulation. "Dispersing liver and regulating qi, clearing heat and removing dampness" is the main principle for clinical treating of cholelithiasis. Lidantang is a recipe applied specially for treating cholelithiasis which is a well-treated disease in our department of surgery. It's used for treating cholelithiasis conservatively and preventing the recrudescence of gallstone after gallstone removal with the gallbladder preserved, resulting in a remarkable clinical effect. Experimental research reveals that Lidantang has an obvious function of dispersing stagnated liver qi for promoting bile flow, notably increasing the amount of bile secreted from the livers of rats, and the amount of bile acid in bile, decreasing the amount of cholesterol in bile secreted from liver. Based on the former clinical and experimental investigation, this study is proposed to establish animal model for cholesterol gallstone, observe the effect of Lidantang on ABCG5/ABCG8, SCP2, ABCB 11, APN, CCK, and CaMKⅡ, further clarify the molecular mechanism for intervening the secretion of cholesterol and bile acid from liver, cholesterol nucleation and the function of gallbladder, provide a theoretical basis to the clinical application of Lidantang for dispersing stagnated liver qi for promoting bile flow, and supply a choice of effective recipe to treating cholelithiasis and preventing gallstone formation after gallstone removal with the gallbladder reserved.

胆石症是临床最常见的疾患之一，根据 "治胆从肝"的中医理论，"肝失疏泄，胆失通降，胆液凝结"是胆石症共有的关键病机。胆石症的临床治疗主要是"疏肝理气、清热利湿"。利胆汤是我院外科多年用于胆石症的有效方剂，有良好的临床效果，且前期初步研究发现利胆汤可以显著增加大鼠胆汁分泌量，明显增加胆汁中的胆汁酸含量，降低胆汁中胆固醇的含量，但其作用机制有待深入研究。本项目拟在以往临床和实验研究的基础上，利用合作单位在药学领域的研究优势，采用药理学、生物化学、分子生物学，病理学等方法，建立胆固醇结石实验动物模型，观察利胆汤对ABCG5/ABCG8、SCP2，ABCB 11，APN和CCK、CaMKⅡ等的影响，从而阐明利胆汤干预肝脏胆固醇分泌、胆汁酸分泌，成核和胆囊功能的分子机制，为临床上使用利胆汤疏肝利胆治疗提供理论依据，为胆石症的治疗和保胆取石后预防结石的复发提供有效方剂，为研制我国特色创新中药奠定基础。

胆石症是临床最常见的疾患之一，根据 “治胆从肝”的中医理论，“肝失疏泄，胆失通降，胆液凝结”是胆石症共有的关键病机。胆石症的临床治疗主要是“疏肝理气、清热利湿”。利胆汤是我院外科多年用于胆石症的有效方剂，有良好的临床效果，但作用机理不清，限制了其临床使用。本项目通过四年的研究，已经按照计划完成申请书中的研究内容，主要发现如下：利胆汤可显著改善高脂饮食造成的胆结石形成，表现为减少结石数量，降低胆结石发病率。生化分析显示，利胆汤可显著降低胆结石小鼠血清中甘油三酯、胆固醇、低密度脂蛋白以及胆汁酸含量。胆汁检测中发现，利胆汤可显著降低胆汁中胆汁酸与胆固醇含量，降低胆汁中胆固醇饱和指数。同时，本研究发现利胆汤可显著改善高脂高胆固醇饮食造成的肝脏损伤，表现为利胆汤可显著降低血清中ALT, AST水平，减轻肝脏炎性细胞浸润现象。机制研究发现：一方面，利胆汤可显著降低胆结石小鼠模型肝脏中胆固醇转运蛋白的基因与蛋白表达（ABCG5， ABCG8， SCP2），但对成核蛋白APN表达无显著影响。另一方面，利胆汤可提升胆囊壁对CCK-8与Ach敏感性，促进胆囊收缩功能，增强胆汁排出量，减少胆汁中胆固醇的聚集，但是，体外研究发现，利胆汤对健康胆囊无直接引发收缩的功能，提示利胆汤发挥的胆囊收缩作用不是直接激活钙离子信号所产生。大量研究表明，过饱和胆汁酸可引发胆囊炎症，本研究中发现高脂高胆固醇饲料可引发，利胆汤可通过抑制胆囊壁炎症反应，减少炎性细胞浸润，抑制TNF-α/ Caspase8 / Caspase3级联诱导细胞凋亡信号通路，减轻胆囊壁中运动起搏细胞ICLCs细胞凋亡，从而改善胆囊运动功能。综上所述，利胆汤可从提高胆囊运动功能和降低胆囊内胆固醇沉积两方面，抑制胆结石的形成，降低胆结石数量与发病率，该研究成果为利胆汤合理用于胆结石的防治工作提供了坚实的理论基础和科学依据。