以PLGA为药物载体的雷帕霉素洗脱MgCa合金食道支架的研究

Recently, magnesium alloys have attracted much attention as potential biodegradable implant materials due to their biodegradability in the bio-environment, inherent biocompatibility and their good mechanical properties. In January 2013 in Lancet a study reported the latest research results of magnesium alloy coronary drug-eluting stents, and in February "Nature Review Cardiology" remarked highly on this work, and claimed that "turning dreams of absorbable stent into reality", all these indicated a promising future for biodegradable Mg alloys in clinical applications. However, little is known about the interaction of esophageal stent and magnesium alloys. Our previous study showed good effects of 20% biodegradable magnesium alloy extracts on IEC-6, but their susceptibility to corrosion has disadvantage on cell cycle and apoptosis of IEC in vitro, indicating that the adjustment of the degradation rate by polymer coating is needed for using magnesium alloy as a promising surgical material. So, we put forward the hypothesis that sirolimus-eluting magnesium alloy stent coated by biodegradable polylactic acid-co-polyglycolic acid (PLGA) with providing adequate mechanical support, appropriate degradation rate, and physiological functions of preventing esophageal stricture would adjust the degradation rate in order to improve the biocompatibility and descend the restenosis rate between stent of esophagus and magnesium alloys. To test this hypothesis, the degradation characteristics and interfacial bioactivity of biodegradable PLGA carried sirolimus-eluting magnesium alloy stent was studied using human esophageal epithelial cells (HEEC) and animal models of pigs by atomic force microscopy(AFM), contact angle instrument and scanning electron microscopy after the interaction of HEEC and biodegradable PLGA carried sirolimus-eluting magnesium alloy stent. The biocompatibility and safety of biodegradable PLGA carried sirolimus-eluting magnesium alloy stent was evaluated from the molecular, cellular, tissue and whole animal levels through in vitro and in vivo experiments using Cell Counting Kit-8(CCK-8), flow cytometry, real-time PCR and Western blotting tests. This work will make a solid foundation of stent of alimentary tract of magnesium alloy for the clinical application.

镁合金因弹性模量低及独特的可降解性能，成为医用金属材料领域中的研究热点。2013年1月《柳叶刀》报道了镁合金冠脉支架的最新研究成果，同年2月《Nature Review Cardiology》对该研究给予了高度评价，未来临床应用前景看好。迄今镁合金在食道支架领域的研究尚无。我们前期实验发现肠上皮细胞（IEC）在20% 浓度镁合金浸提液中有很好的生物相容性，但镁合金的快速降解对IEC有不利影响，提示需要通过可降解高分子作为涂层进一步控制其降解速度，在本项目中我们提出研发以聚乳酸聚羟基乙酸共聚物(PLGA)为药物载体的雷帕霉素洗脱MgCa合金食道支架，兼具提供足够的力学支撑、合适降解速度及预防食道狭窄的功能；将通过人食管上皮细胞系HEEC和动物猪模型，采用扫描电镜、CCK-8等手段，研究该实验支架与食道细胞及组织相互作用后的表面性能与降解机理及生物相容性，为其成为新型食道支架提供理论依据。

目前在消化系统疾病中支架放置已经广泛应用于临床，但是，金属及塑料支架的不易取出、疼痛、穿孔、出血等不良反应使得支架植入术仍然面临着众多挑战；高分子生物可降解支架力学性能较差。镁合金不仅可以降解而且具有合适的物理力学性能，作为消化道支架的材料具有优势。本课题对镁合金进行了体外与体内两方面的实验。体外实验中，研究发现Mg-Ca合金浸提液在一定范围内的较高浓度时易诱导人结肠黏膜上皮NCM460细胞Caspase-3、Integrin 1 和E-cadherin的表达，而抑制Bcl-2的表达，且随浓度升高影响越大，可能与钙离子浓度有关。高浓度的镁钙合金浸提液对NCM460细胞中的MMP9及TIMP3基因表达有一定的影响，容易导致早期炎症反应的发生，其机制可能与浸提液中的钙离子浓度有关。但在体内实验时，镁钙合金的降解是一个逐渐的过程，钙离子的释放会比较缓慢，由其引起的早期炎症反应是比较轻微的。此外，研究还发现， PLLA/紫杉醇/镁锌钇钕合金在模拟体液中降解速度减缓；PLLA/紫杉醇/镁锌钇钕合金抑制人结肠黏膜上皮NCM460细胞的增殖。体内实验中，研究发现MAO/PLLA/雷帕霉素/镁锌钇钕合金支架没有增加血液中的镁离子浓度, 对肝肾功能也没有影响。将制成的MAO/镁锌钇钕合金支架、MAO/PLLA/镁锌钇钕合金支架及MAO/PLLA/雷帕霉素/镁锌钇钕合金支架植入动物肠道内进行观察。体内研究显示，由于肠道环境复杂，短期内MAO/PLLA/雷帕霉素/镁锌钇钕合金支架具有良好的力学特征，但长期观察，发现MAO/PLLA/雷帕霉素/镁锌钇钕合金支架仍然降解较快，仍需进一步研究以改善其力学特征。