扶正祛毒方通过影响DC－T淋巴细胞轴关键分子表达调节HBV免疫耐受机制研究

The chronic HBV carriers have two characteristics : HBV DNA copies in high level and Transaminase always in normal level， these characteristics usually lead to a high proportion developing into malignant diseases ,so it is necessary to treat the chronic HBV carriers. Modern medicine had learned that, immunological tolerance is the critical pathogenesis during the chronic HBV carrying process,and the deficiency of T cell-mediated immunity cross the DC-T axis plays the role in immunological tolerance mechanism,but so far there is no treatment. Traditional Chinese medicine has so many preponderances such as immunological regulation in both direction , treat in multi target ,but there is no uniform syndrome differentiation because of the absence of symptoms. Our group learned the key pathogenesis of chronic HBV carriers is "deficiency of qi and hidden toxin ",with the work we have done in the past,we discovered that Fuzhengqudu compound formula has the efficiency of strengthening genuine qi to eliminate toxin,it can inhabit the copies of HBV while protect the liver function , so we sets out from pathology and pathogenesis of traditional Chinese medicine and modern medicine mechanism convergence,using the regulation of immune cells and cytokines as the breakthrough point, puts forward this hypothesis :"Fuzhengqudu compound formula can break out the immunological tolerance,rebuild the immunological response by changing the express of critical molecular in DC-T-lymphocytes axis”,to verify the immunological regulation efficiency of Fuzhengqudu compound formula ,to study the immunological regulation mechanism of Fuzhengqudu compound formula,to reveal the convergence of mechanism in traditional Chinese medicine and modern medicine,supplements understanding of the pathogenesis of HBV with traditional Chinese medicine. And the research will employ the immune tolerance period after the HBV infection as treatment time to provide the basis for clinical practice.

慢性HBV携带者具有“HBV高水平复制”和“转氨酶正常或低水平”两大特点，结局高危需要治疗。西医认识到其核心机制为免疫耐受，以树突细胞（DC）－T淋巴细胞轴为主的细胞免疫功能缺陷是免疫耐受的关键，但无有效治疗手段。中医药具有双向调节、多靶点作用的优势，但本病常无症状限制辨证分型。课题组认识到本病“正虚毒伏”的病机关键，经过前期临床和实验研究，观察到具扶正祛毒功效的扶正祛毒方抑制HBV复制和保护肝细胞的作用，从中医病机理论和现代医学病理机制共通性出发，以对细胞免疫的调节为切入点，提出“扶正祛毒方通过影响DC－T淋巴细胞轴关键分子的表达，打破HBV免疫耐受，重建有效免疫应答”的假说，阐释扶正祛毒方对慢性HBV携带者免疫调节的药理学基础，探讨其作用机制，揭示中医“正虚毒伏”理论与慢性HBV携带者免疫耐受机制的共同性，补充中医学对慢性HBV携带者的认识。拟将免疫耐受期作为治疗时机，为临床提供依据。

免疫耐受是乙肝病毒（HBV）慢性携带的核心机制，慢性HBV携带者则是隐匿进展为肝纤维化、肝细胞癌的重要群体。本课题基于“正虚毒伏”理论与免疫耐受机制的共通性，利用HBV转基因小鼠模型，通过空白组对照、阳性对照、中药高、中、低剂量治疗的不同处理，以树突细胞（DC）-T淋巴细胞轴上关键分子表达的变化为目标，综合评价治疗药物扶正祛毒方抑制模型鼠HBV复制的作用，以及其对HBV诱导的慢性免疫耐受小鼠模型的免疫调节作用。. 实验结果表明，具有扶正祛毒功效的扶正祛毒方可以抑制HBV模型鼠病毒复制。并上调了DCs表面共刺激分子表达，增强了抗原提呈信号以及对T细胞的活化，调整了Th1/Th2比例，以促使其向Th1细胞分化，活化免疫反应。这可能是其发挥HBV抑制作用的关键所在。. 从不同浓度的治疗药物来看，综合不同剂量组HBVDNA的表达、肝细胞炎症反应、以及免疫应答的活化程度，考虑中药中剂量为最佳治疗浓度。从12周和24周不同时点来看，虽然不同时点的动物模型背景不完全一致，但24周肝细胞炎症反应较12周明显，符合疾病的进展规律；而在DCs表面共刺激分子表达、外周血CD4/CD8变化，以及Th1/Th2细胞因子比例等免疫应答层面，12周时实验药物对免疫耐受的调节优势优于24周时，可见，病程越长，干预效果越差，药物活化免疫应答方面的优势越不明显，但扶正祛毒方对免疫应答的调节作用仍较对照药物更持久。. 本实验从整体、组织、蛋白、基因水平，从免疫角度阐释了扶正祛毒方的作用及机制，为慢性病毒感染、免疫耐受的中医药治疗提供实验依据和理论借鉴，为临床无法进行辨证论治的病症提供病证结合的诊疗思路。