活血化瘀药物促进肿瘤新生血管"正常化"，改善肿瘤微环境，增加化疗药物靶向性的研究

Traditional Chinese medicine-huoxuehuayu has emerged as a potent strategy for cancer therapeutic, but the mechanisms are not well understood. the role which huoxuehuayu herb is promoting or suppress tumor angiogenesis is controversial. The first time, we propose that huoxuehuayu herb induce tumor vascular normalization, and thereby may improve tumor microenvironment. To establish the validity of this hypothesis, we investigated the therapeutic effect of Biejiajianwan(TCM compound) and dan-shen root in BALB/c mice bearing xenografted Hepatocellular Carcinoma and in vitro. A morphologically and functionally changes of vascular network which including microvessel density of tumor, pericyte coverage (α-smoothmuscle actin immunostaining), collagen-covered tumor vessels, and lymphatic vasculature, are determined. Tumors of interstitial fluid pressure (IFP) and tumor hypoxic fraction are measured. We also investigated therapeutic benefits and accumulation and subsequent deep diffusion of doxorubicin in mice bearing Hepatocellular Carcinoma which were treated with doxorubicin and huoxuehuayu herb(Biejiajianwan and dan-shen root).We propose that huoxuehuayu herb will promote normalization and remodeling of the tumor vasculature, and a resultant reduction in tumor hypoxia and interstitial fluid pressure. These in turn can lead to an improvement in the metabolic profile of the tumor microenvironment, the delivery and efficacy of exogenously administered chemotherapeutics. Thus, vascular normalization may contribute to the improved survival rates in tumor-bearing animals treated with huoxuehuayu herb in combination with cytotoxic therapies. The ?ndings will provide insights into the mechanisms underlying the effects of huoxuehuayu, which could lead to rational design of antitumor therapeutic strategies. The relationship between "Huoxuehuayu" and " morphological structure and function of vascular "may interpret Pathophysiologic mechanism of blood stasis syndrome.

活血化瘀药物抗肿瘤的作用机制尚不明确，对肿瘤血管生成的作用是促进或抑制，一直颇有争议。本课题首次从诱导肿瘤血管正常化出发，探讨活血化瘀药物的作用。本课题选择治疗肝癌的活血化瘀古方鳖甲煎丸及单药丹参进行动物实验和体外实验，观察其对原位移植肝癌的肿瘤内异常增生新生血管的形态，结构、功能，淋巴管结构，以及对肿瘤微环境（间质内液压（IFP）、乏氧状态）等的影响，并观察其对化疗药物在肿瘤内穿过性和浓度，和对化疗药物疗效的影响，通过研究活血化瘀药物对肝细胞癌异常增生血管的"正常化"、改善肿瘤微环境，降低肿瘤间质内液压的作用，探讨活血化瘀药治疗肿瘤、增加化疗药物疗效的新机制。为活血化瘀药物用于肿瘤治疗提供实验基础和依据。从"活血化瘀"与"血管形态结构"之间的关系 ，阐释血瘀证的病理机制。

为了研究丹参酮ⅡA和鳖甲煎丸对肿瘤新生血管结构及功能、肿瘤微环境的影响，以及其对阿霉素脂质体的增效作用。实验中建立小鼠肝癌细胞（Hepa1-6）皮下转移瘤模型，随机分为对照组，丹参酮ⅡA组，鳖甲煎丸组和贝伐单抗组（阳性对照组），与不同方式连续给药14天后处死小鼠，剥取肿瘤组织，分别进行免疫组织化学染色、免疫荧光染色、乏氧染色、液相色谱检测等项目，采集切片所得实验图片，并进行统计分析。另外，采用b-FGF诱导小鼠腹壁Matrige1种植体方法和体外培养大鼠腹主动脉环血管生成模型进行血管生成实验。研究发现丹参酮ⅡA和鳖甲煎丸均能够减少肿瘤血管新生，并且增加血管内皮细胞表面周细胞、基底膜的覆盖，使血管结构趋于正常。同时减少肿瘤内的淋巴管数量。另外，二者均能改善肿瘤的乏氧状态，增加阿霉素脂质体在瘤内的渗透距离及输送到肿瘤组织内部的量。由此推测，以丹参酮ⅡA和鳖甲煎丸为代表的活血化瘀药物能够促进肿瘤新生血管结构和功能趋向正常化，改善肿瘤微环境，与化疗药物联合使用能够起到增效作用。