基于Th1细胞正反馈正常化肿瘤新生血管重构免疫微环境的益气活血法治疗肝癌的机制研究

Blood stasis is one of the main pathological basis for the development of malignant tumor. However, there is still controversy in the clinical application of the method of activating blood and removing blood stasis. Studies have shown that large dose drugs of Invigorating Qi combined with the drugs of Activating Blood Circulation and removing blood stasis had the function of activating Th1 cells and anti-angiogenesis. Meanwhile the latest study indicated that there was a positive feedback loop between Th1 cells and tumor vascular normalization. Our precious result displayed that Buyanghuanwu decoction had effect of immunological enhancement and tumor vascular normalization on the prevention of hepatocellular carcinoma(HCC), Thereby we put forward the hypothesis that Buyanghuanwu decoction have the effect of activating Th1 to promote the tumor vascular normalization in positive feedback loop to reprogram immunomicroenvironment on HCC. To elucidate the mechanism of the effect of Buyanghuanwu decoction on HCC by activating Th1 to promote the tumor vascular normalization in positive feedback loop to reprogram immunomicroenvironment, the chemicals induced HCC mouse model, H22 subcutaneous transplantation mouse model, orthotopic liver transplantation for lung metastasis mouse model and co-culture system should be used in this study to compare the effect of Invigorating Qi component, activating blood components and whole components on HCC. By this way experimental support would be provided for the treatment of malignant tumor using drugs for promoting blood circulation and removing blood stasis combined with high dose drugs for Invigorating Qi, and also new experimental evidence would be proved for Blood stasis being caused by Qi deficiency due to the TCM theory of "Qi and blood".

瘀血阻滞是恶性肿瘤发生发展的主要病理基础之一，但活血化瘀治法在临床运用上尚存争议。已有研究表明大剂量益气扶正药物联合活血化瘀药物具有活化Th1细胞及抗肿瘤血管新生作用，结合Th1细胞与肿瘤血管正常化之间存在正反馈环路的最新进展，在前期研究发现增强细胞免疫抑制肿瘤新生血管可能是补阳还五汤防治肝癌重要途径的基础上，提出补阳还五汤作用于Th1细胞正反馈正常化肿瘤新生血管重构免疫微环境起到治疗肝癌作用的研究假说。本研究将利用化学诱导发生肝癌小鼠模型、小鼠皮下移植瘤模型和裸鼠原位移植肝转移瘤模型以及体外细胞共培养体系，对比观察补阳还五汤中益气组分、活血组分以及全方组分对肝癌的抑制作用，并从Th1细胞正反馈正常化肿瘤新生血管重构免疫抑制微环境的角度探讨益气活血法治疗肝癌的作用机制，为恶性肿瘤治疗使用活血化瘀药物需联合大剂量益气扶正药物提供实验支撑，同时为因虚致瘀的中医“气血”理论提供新的实验依据。

瘀血阻滞是恶性肿瘤发生发展的重要环节，但临床上能否使用和如何使用活血化瘀药物尚存争议。本研究从肿瘤血管新生与免疫细胞串话的角度探讨补阳还五汤重构免疫微环境治疗HCC的作用机制。.首先通过网络药理学探索补阳还五汤君药黄芪在肝癌治疗中的作用靶点，获得“药物—成分—疾病—靶点”作用关系，包括HIF-1、Ras信号通路等信号通路是补阳还五汤君药黄芪在肝癌治疗中的可能作用靶点。通过建立H22和Hep1-6细胞小鼠肝癌原位瘤模型，对比观察补阳还五汤合方及其拆方组分（益气组分和活血组分）对肿瘤生长的影响，并检测血管生成相关因子VEGF、ANG1、PDGF及其受体VEGFR、Tie-2、PDGF、炎症相关的CXCL10、缺氧相关的HIF-1α表达水平的变化，研究结果提示合方相较拆方组分，对H22肝癌原位荷瘤小鼠的肿瘤抑制作用明显，与VEGF和ANG等肿瘤血管新生因子相关。但在Hep1-6原位荷瘤小鼠中观察到补阳还五汤合方的促瘤效应，提示基因突变的差异影响补阳还五汤的治疗作用。结合网络药理学分析的结果，进一步建立Ras/c-myc和Ras/β-Catenin两种基因突变小鼠肝癌模型，在Ras/c-Myc基因突变的小鼠模型中，补阳还五汤全方及活血组分的促瘤作用明显，尤以活血组分为甚，益气组分具有一定的抑瘤作用；而在Ras/β-Catenin基因突变小鼠模型中，补阳还五汤合方以及拆方组分均具有抑瘤作用，同时益气组分和活血组分显示出协同效应，进一步机制研究表明补阳还五汤具有重构Ras/β-Catenin基因突变小鼠肝癌免疫抑制微环境的作用，其作用途径与Th1细胞正反馈环路正常化肿瘤新生血管相关。.本研究从活化TH1细胞正反馈正常化肿瘤新生血管重构肿瘤免疫微环境的角度阐明益气活血治法治疗恶性肿瘤的作用机制，为肿瘤治疗临床实践中活血化瘀药物应联合大剂量益气扶正药使用提供了实验依据。