肠道菌移位介导的人参联合环磷酰胺抗癌增效机制研究

Recent study show that the anti-cancer activity was enhanced by the combined application of ginseng and cyclophosphamide, however the mechanisms involved need to be further elucidated. The latest researches had proved that cyclophosphamide could increase the intestinal permeability and induce the translocation of lactobacillus and enterococcus genera intestinal bacteria into secondary lymphoid organs, and subsequently stimulate the expression of anti-cancer related factors in vivo and finally help shape the anticancer response. Our previous studies found that ginseng could induce the propagation of lactobacillus and enterococcus genera probiotics in intestine. Therefore, a hypothesis of the mechanisms of the anti-cancer enhancement by the combined application of ginseng and cyclophosphamide is proposed: ginseng will promote the propagation of gut bacteria in intestine, while cyclophosphamide will increase the intestinal permeability, and then increase the translocation of gut bacteria into secondary lymphoid organs, which trigger the expression of much more anti-cancer related factors, and finally enhance the anti-cancer activity. The aim of this project is to illuminate the mechanisms of anti-cancer enhancement by the combined application of ginseng and cyclophosphamide, which can provide scientific basis for the combination of ginseng and cyclophosphamide for the treatment of cancer, and novel idea for anti-cancer drug development through combination of medicinal herbs and chemicals.

阐明中药与西药协同增效机制对中西医结合事业的发展具有重要意义。最新研究证明，环磷酰胺可通过增大肠壁通透性、触发乳酸菌和肠球菌等肠道益生菌移位至次级淋巴器官、激活抗癌相关因子表达，产生抗癌效应。我们前期研究发现人参能促进肠道益生菌如乳酸菌和肠球菌等增殖，人参联合环磷酰胺应用有抗癌增效作用，但机制有待深入研究。本课题拟利用乳腺癌动物模型，通过对联合应用前后肠壁通透性、肠道和次级淋巴器官中可移位菌在的表征及活性和抗癌效应等方面进行研究，建立人参-环磷酰胺-肠道菌-抗癌效应的关联性，验证如下科学假说：人参联合环磷酰胺应用时，人参促进乳酸菌和肠球菌等肠道益生菌增殖，环磷酰胺触发更多的可移位肠道菌移位至次级淋巴器官，继而诱导机体大量表达抗癌相关因子，从而提高抗癌效能。通过该假说的验证，揭示肠道菌移位介导的人参联合环磷酰胺抗癌增效的机制，为人参与环磷酰胺联合抗癌提供科学依据，也为抗癌药物研究提供新思路。

临床上常将环磷酰胺与人参联合应用，起到提高环磷酰胺抗癌效能的目的，但机制不明。本项目针对这一临床需求开展一系列研究，研究结果如下：（1）应用UPLC-QTOF-MS/MS技术创建了30种人参皂苷同时定性定量方法，为实验过程中人参的质量控制提供了有效监控手段；（2）在动物水平证实人参能够提高环磷酰胺的抗乳腺癌效能，同时可以降低环磷酰胺的毒性，然而抗癌效率随着人参剂量的增大而降低，但减毒效能随着人参剂量的增大而增大，综合考虑人参与环磷酰胺联合应用时抗癌效能和毒性，中剂量的人参与环磷酰胺联合应用最为合适。此外还发现，当同时给予万古霉素后，中剂量人参几无提高环磷酰胺抗乳腺癌效能的作用，表明人参与环磷酰胺联用抗癌增效作用与肠道菌密切相关；（3）长期使用环磷酰胺导致肠道菌紊乱，其中降低Lactobacillus johnsonii和Lactobacillus murinus等乳杆菌的水平，而这些乳杆菌与环磷酰胺的抗癌效能密切相关。人参可回调环磷酰胺所致的肠道菌紊乱，其中可提高环磷酰胺所致的Lactobacillus降低和Escherichia、Enterobacteraceae升高，最值得注意的是，人参可提高肠道中Lactobacillus johnsonii和Lactobacillus murinus等乳杆菌水平。回调肠道菌是人参提高环磷酰胺抗乳腺癌效能的机制之一。（4）肠黏膜炎在环磷酰胺抗癌作用中是一把双刃剑，适当的肠道损伤可触发Lactobacillus johnsonii和Lactobacillus murinus等乳杆菌移位，提高环磷酰胺的免疫抗癌效能，但严重的肠道损伤，会造成大量致病菌的移位，从而造成机体感染，甚至发生脓毒血症等。研究发现人参可以通过调控NFκB等通路缓解环磷酰胺所致的肠黏膜损伤，减少肠道内病原微生物通过肠粘膜进入机体的程度，从而降低二次感染发生的可能性。降低肠黏膜炎损伤程度，减少致病菌移位及其所致的感染，是人参提高环磷酰胺抗乳腺癌效能的另一机制。总之，人参通过多种途径提高环磷酰胺的抗乳腺癌效能，一方面通过调节环磷酰胺所致的肠道菌紊乱，另一方面通过改善肠黏膜炎，促使适量的肠道菌转移至体内产生免疫抗癌效能。该研究为人参与环磷酰胺联合应用提供理论依据和实验依据，同时也为中西医结合抗肿瘤创新研究提供新思路。