非常规层层自组装构建川芎嗪/葛根素配伍洗脱涂层促进血管再生

Delayed endothelial healing caused by drug-eluting stents implantation may result in high incidence of late thrombosis. Using new drugs with both anti-proliferative effect on smooth muscle cells (SMC) and promotive effect on re-endothelialization instead of traditional anti-proliferative drugs is important to accelerating vascular regeneration and preventing late thrombosis. In this research, the ligustrazine/puerarin combination will be chosen as the drug because of the enhanced therapeutic efficacy by the cooperation of active constituents from Chinese traditional medicine. Biocompatible and degradable coatings which can promote endothelialization will be fabricated via unconventional layer-by-layer self assembly, and the cooperation of biofunctional coatings and effective drug combination will be achieved. Ligustrazine and puerarin will be introduced into the multilayered films based on Pluronic polymer mixed micelle, chitosan oligosaccharide and heparin via the preassembly of building blocks. The relationship among the nanostructure of building blocks, the structure of multilayered films and the functions of multilayered films will be explored. The influence of ligustrazine/puerarin combination-eluting coatings on the thrombin-induced proliferation of SMC and endothelial cell injury will be demonstrated via the in vitro cell culture experiments. The effect of ligustrazine/puerarin combination-eluting stents on reducing the rate of in-stent restenosis and accelerating vascular regeneration will be researched in a hypercholesterolemic rabbit iliac artery stent model. The cooperation of multiple effects is designed to improve the security and effectiveness of drug-eluting stents in clinical application. The results from this original research will provide a new approach to the fabrication of functional drug-eluting coatings and the development of Chinese traditional medicine-eluting stents with independent intellectual property.

血管内皮损伤难以修复会增加药物洗脱支架植入后晚期血栓形成的风险。以阻抗平滑肌增殖与促内皮化协同作用的药物替代传统抗增殖药物有助于促进血管再生、降低晚期血栓形成风险。本项目以药效协同增强的中药有效成分川芎嗪/葛根素配伍为涂层药物，利用非常规的静电层层自组装技术构筑具有促内化功能的生物相容性可降解涂层，期望获得涂层生物功能与药物疗效的协同作用。本项目利用预组装体的形成，将川芎嗪与葛根素载入基于Pluronic聚合物混合胶束、壳寡糖与肝素的层状多层膜；研究多层膜组装过程，探索组装基元结构-多层膜结构-多层膜性能间的内在关联；通过凝血酶干预的体外细胞实验与家兔髂动脉支架植入实验，评价药物洗脱涂层阻抗平滑肌增殖与保护内皮功能、促进血管内皮修复的作用。本项目通过多途径协同作用提高药物洗脱支架临床应用的安全性与有效性，为药物洗脱涂层功能化提供新方法，并为具有我国自主知识产权的中药洗脱支架研究提供新思路。

本项目利用静电层层自组装技术构筑Pluronic/多糖复合的药物洗脱涂层，评价Pluronic/多糖复合膜作为药物洗脱支架涂层材料的可行性。研究制备了端基磺酸化的Pluronic P123，其与Pluronic F127在水溶液中自组装形成电负性混合胶束，并与壳寡糖通过静电作用形成稳定、分散、纳米球形的预组装体。调节预组装体组成，可获得表面呈正电性的纳米单元。预组装体与肝素可在基材表面实现交替沉积，且预组装体在表面沉积过程中保持稳定。将中药活性组分葛根素载入复合膜，肝素/壳寡糖/胶束复合膜的载药量低，且抗菌性与血液相容性均不理想；相同组装条件下构筑肝素/壳聚糖/胶束复合膜，膜内载药量显著提高，膜表面抑菌性和血液相容性也得到明显改善，但仍未满足药物洗脱支架涂层的要求。本研究结果揭示了Pluronic聚合物/多糖复合膜的性能与预组装体组装基元的结构密切相关，这类非常规静电层层自组装过程具有其特殊性，有助于探索组装基元结构-多层膜性能间的内在关联。本项目进一步研究壳寡糖/胶束预组装体作为葛根素口服给药载体的可行性。体外释药实验结果表明，混合胶束与壳寡糖作用形成预组装体，葛根素的释放速率与释放量降低。Caco-2细胞单层模型中的跨膜转运实验结果显示，将葛根素载入预组装体可促进其跨膜转运，跨膜转运量随着预组装体中壳寡糖含量的增加而降低。以小鼠为模型，载药预组装体溶液灌胃给药10 min左右，血药浓度达到最高。阿尔茨海默病模型小鼠实施载药预组装体溶液灌胃治疗30天后，Morris水迷宫实验与脑组织MDA含量、AchE活力及GSH-PX酶活力测定的结果显示，预组装体具有提高葛根素口服给药药效的作用。鉴于壳寡糖/胶束预组装体的性能可由体系组成调控，其在口服给药载体方面的应用具有一定的潜力。