金黄虎头蜂蜂毒抑制类风湿性关节炎炎症和血管生成的作用机制及物质基础研究

Vespa mandarinia belongs to Vespa genus, family Vespidae. Possessing the natures of large body, aggressiveness, and producing a large amount of venom, this species has been traditionally utilized as concocted venom wine prepared by soaking the insects into alcohol by Jingpo minority at Yunnan Dehong region for the treatment of rheumatoid arthritis (RA) and paralysis. In the preliminary work of this project, we disclosed that the vespa venom demonstrated the inhibitory effect against angiogenesis in chick chorioallantoic membrane, and therapeutic effect on the rat collagen-induced arthritis, along with the analgesic effect, on the basis of solving the problems of batch culture of the insect, the improvement on the technology of constantly collecting venom of high purity. The major targets of this research project are to adopt the model of collagen-induced arthritis in rats to further investigate the influence of the V. mandarinia venom on the distribution pattern of the inflammatory cell subsets, on the expression of the cytokines, and on the expression levels of endothelial growth factor and its receptor 2, in the peripheral blood and synovial fluid at the startup phase of synovitis. Furthermore, the study on the bioactivities of vespa venom on in vitro cultured fibroblast-like synovial cell proliferation and the expression of apoptotic factor and vascular endothelial growth factor will also be accomplished in this proposal. Moreover, parsing of the structures of the active components existed in vespa venom will be also thoroughly performed, by means of cluster of modern proteomic analytic techniques such as ultrafiltration, electrophoresis, and mass spectrometry. The aim of this project is to, through these studies, elaborate the acting mechanism of action of the vespa venom upon intervening the RA local inflammation and the regulatory effect of inflammatory cytokines in synovial fluid and angiogenic inhibition. Then to elucidate the active substances of this vespa species, it will help to enrich the research background of the V. mandarinia, which is expected to provide a scientific basis for better developing this Yunnan's special insect resource.

金黄虎头蜂是云南德宏景颇族群众传统应用炮制蜂毒酒用于治疗类风湿关节炎等疾病的一种胡蜂科虎头蜂属昆虫，具有个头大，攻击性强、毒量大的特点。前期在解决了金黄虎头蜂批量养殖的难题，完善取毒技术并能获得高纯度蜂毒的基础上；摸索出蜂毒具有抑制鸡胚尿囊膜血管生成、治疗大鼠胶原诱导性关节炎和镇痛的作用。本项目拟进一步研究金黄虎头蜂蜂毒对胶原诱导性关节炎大鼠模型滑膜炎症期的外周血和关节滑液中炎性细胞亚群分布格局及炎症因子表达的影响、对血管内皮生长因子及其受体2表达水平的影响；以及蜂毒对体外培养的成纤维样滑膜细胞增殖、凋亡因子和血管内皮生长因子表达的影响。并用超滤、电泳和质谱等现代分离分析技术对蜂毒物质基础进行解析。通过这些研究，阐述金黄虎头蜂蜂毒干预RA局部炎症，调节炎症因子群落及抗血管生成作用的作用机制，了解其物质基础，丰富金黄虎头蜂作为传统药用昆虫的研究资料，为更好的开发云南特色资源提供科学依据。

本项目以金黄虎头蜂蜂毒抑制类风湿关节炎炎症和血管生成的作用机制及物质基础为研究目标：研究内容包括金黄虎头蜂蜂毒对胶原诱导性关节炎大鼠模型滑膜炎症及滑膜细胞生长的影响、对血管内皮生长因子表达水平的影响以及对各类炎症因子表达的影响。结果显示金黄虎头蜂蜂毒对CIA大鼠有显著的疗效作用，主要表现在可以减轻CIA大鼠关节肿胀度及关节周长，并减轻CIA大鼠间质细胞激活、炎性细胞浸润、内膜细胞增生程度，能抑制促炎因子和血管内皮生长因子的表达水平，能调控各类免疫细胞的分布格局。并用生化分离分析技术对蜂毒物质分离纯化21个蛋白质和小肽分子。.另外，项目根据金黄虎头蜂蜂毒有较强的纤溶活性增强和抗凝活性的实验结果，开展了胡蜂蜂毒制剂对小鼠耐缺氧和抗心肌缺血作用研究初探，以及对大鼠脑缺血再灌注损伤的保护作用的开展了初步研究。结果显示结果发现胡蜂蜂毒能减少脑梗塞面积，减轻神经元的坏死，其机制可能与抗炎相关。.本项目的实验结果丰富了金黄虎头蜂蜂毒的研究资料，丰富了胡蜂作为传统药用昆虫的研究资料，并为更好的开发云南特色资源提供科学依据。