基于IDO1生物靶标探讨"补肾健脾解毒方"抗慢性HBV感染的免疫调控机制

For the development of chronic hepatitis B virus (HBV) infection, mmune tolerance is an important mechanism. Thus, immunotherapy is expected to play a relieving and therapeutic role. Traditional Chinese medicine have a unique advantage in the fields of the immune regulation. Clinical trials confirmed that Bushen Jianpi Jiedu Prescription has the role of improving the immune tolerance status of patients with HBV infection. However, its efficacy targets and molecular mechanisms are still not fully understood. Based on the critical roles of indoleamine 2, 3-dioxygenase 1 (IDO1) in tryptophan metabolism to regulate the maturation and function of Treg cells and dendritic cells and on our previous studies, we hypothesize that Bushen Jianpi Jiedu Prescription can mediate tryptophan metabolism through intervention of IDO1 to regulate the maturation and function of Treg cells and dendritic cells, so as to produce an effective immune response to inhibit or eliminate HBV. In our study, the role of IDO1 in immune tolerance will be confirmed by in vivo and in vitro experiments. Moreover, CRISPR/cas9 and Lentiviral technologies will be used to supress IDO1 gene for identification and screening of the IDO1 signal pathway and the prescription active ingredients biological targets. Simultaneously, combined with IDO1 gene knockout mice, to further verification the detail endpoints of IDO1 and the resistant target of the prescription active ingredients in regulation of immune. Finally, the underlying mechanism of IDO1 and biological effects of Bushen Jianpi Jiedu Prescription in chronic HBV infection will be clearly elucidated. Our study is helpful to clear the inherent mechanism of Bushen Jianpi Jiedu Prescription against chronic HBV infection, and provide scientific evidences for its clinical application.

免疫耐受是慢性乙肝病毒（HBV）感染的重要因素，故免疫治疗有望对其起到缓解及治疗作用。中医药在调节免疫方面具有独特优势，临床试验证实中药复方"补肾健脾解毒方"能改善慢性HBV感染免疫耐受状态，但其药效靶点及分子机制尚不完全明确。基于吲哚胺2, 3-双加氧酶1（IDO1）干预色氨酸代谢以调控免疫的功能，结合前期研究我们提出："补肾健脾解毒方"可通过干预IDO1介导色氨酸代谢以调控Treg细胞及树突状细胞的功能，从而产生有效的免疫应答以抑制HBV。本研究拟通过体内外实验确定IDO1参与免疫耐受的作用，并运用CRISPR/cas9结合慢病毒技术修饰IDO1基因进行靶标调控机制探索及药物靶向分析，最后结合IDO1基因敲除小鼠模型深入验证，并明确"补肾健脾解毒方"干预HBV感染的免疫内涵及IDO1靶向调控机制。本研究将进一步揭示"补肾健脾解毒方"抗慢性HBV感染的内在机理，为其临床应用提供科学依据。

慢性乙型肝炎病毒（HBV）感染是一个全球性的公共问题。免疫耐受是慢性乙肝病毒（HBV）感染的重要因素，故免疫治疗有望对其起到缓解及治疗作用。中医药在调节免疫方面具有独特优势，临床试验证实中药复方“补肾健脾解毒方”能改善慢性HBV感染免疫耐受状态。前期已通过临床实验证实该方能够降低慢性HBV携带者血清TGF-β、IL-10的含量，下调CD4+CD25+Treg细胞，提高树突状细胞（DCs）表面分子表达率，促进DCs成熟，从而改善慢性HBV携带者的疾病进展。基于吲哚胺2, 3-双加氧酶1（IDO1）干预色氨酸代谢以调控免疫的功能，通过体内外实验确定IDO1基因的缺失使减轻机体的“色氨酸耗竭”状态，下调CD4+CD25+Treg细胞数量，调节T细胞比例，提高DCs成熟表面标记物的表达水平，增强DCs诱导混合淋巴细胞反应的功效。而高剂量的“补肾健脾解毒方”可能通过干预IDO1诱导的“色氨酸耗竭”状态以调控T细胞及DCs的功能和成熟，从而产生有效的免疫应答以达到抑制HBV作用，为“补肾健脾解毒方”的临床应用提供科学依据。