ROS-磁性双重靶向智能纳米粒的构建及促创面愈合的应用研究

Stromal cell derived factor 1 (SDF-1) can promote the mobilization, chemotaxis, homing and targeted differentiation of bone marrow mesenchymal stem cells (BMSCs) in the process of wound healing to participate in the regeneration and repair of blood vessel. In view of high level of reactive oxygen species (ROS) in wound tissues, we developed a kind of ROS-responsive nanoparticles carrying SDF-1. After its intravenous administration, SDF-1 could be released targeting to wound and accelerated vascularization of wound could be seen. But there existed three problems, including low drug-loading capacity due to strong hydrophobicity of nano polymer, high level of static SDF-1 release in vivo due to its hypersensitive response to ROS, false positive SDF-1 release caused by high level of ROS located in other lesion region besides wound. This project aims to prepare smart nanoparticles with dual ROS-magnetic targeting to solve all these problems. Through modification of side chain of the polymer with polyethylene glycol (PEG), high drug-loading capacity will be obtained by increased hydrophilicity, and decreased ROS-responsive sensitivity will also be done by steric hindrance effect. Magnetic Fe3O4 nanoparticles will be constructed as the magnetic nucleus of new smart nanoparticles. By combination of magnetic with ROS targeting, false positive SDF-1 release will be ameliorated so that more accurately targeted release of SDF-1 to wound, higher biological utilization and higher safety will be realized. The characteristics of these nanoparticles will be investigated. After intravenous administration of these nanoparticles in diabetic rat with chronic skin wound, the wound-targeted release of SDF-1, and its biological effect on BMSCs homing to wound, as well as the speed of wound healing will be observed. The distribution and outlet of polymers, Fe3O4 and SDF-1 in vivo will also be followed by intact fluorescence imaging and MRI imaging methods.

创面愈合中基质细胞衍生因子1（SDF-1）动员和趋化骨髓间充质干细胞（BMSCs）归巢和定向分化，参与血管的再生和修复。利用创面的高活性氧（ROS）特性，课题组研制了携带SDF-1的ROS响应性纳米粒，静脉给药后具有创面靶向释药性和促血管化作用。存在的问题：纳米粒强疏水性致SDF-1载药量低；ROS过敏感致体内静态释药量大；机体内存在其他高ROS区域致SDF-1假阳性释放。.本项目拟制备ROS-磁性双重靶向智能纳米粒，通过聚合物侧链亲水性PEG修饰以提高SDF-1载药量，及空间阻碍效应适度钝化ROS敏感性；构建Fe3O4纳米粒子作为磁核，复合磁靶向减少假阳性释放，以实现更精准的创面靶向释药、更高的生物利用度和安全性。对其进行表征和特性鉴定。对具慢性创面的糖尿病大鼠输注纳米粒，从聚合物片段、核磁和蛋白三个环节活体示踪药物的转归，观察SDF-1创面靶向释放、BMSCs定向归巢效应及创面治疗效果。

创面愈合中基质细胞衍生因子1（SDF-1）动员和趋化骨髓间充质干细胞（BMSCs）归巢和定向分化，参与血管的再生和修复。前期研究中课题组利用创面的高活性氧（ROS）特性，研制了携带SDF-1的ROS响应性纳米粒，静脉给药后具有创面靶向释药性和促血管化作用。但是由于纳米粒强疏水性致SDF-1载药量低，而ROS过敏感致体内静态释药量大，另外机体内存在其他高ROS区域致SDF-1假阳性释放。.为了解决现有的问题，本项目采用PEG 修饰了PPADT，合成了新型ROS敏感纳米粒PPADT-PEG，其具有半亲水半疏水特性，能够提供更好的载药性能， ROS敏感性也更适宜的。为了减少假阳性释放对局部组织的影响，项目组制备Fe3O4 磁性纳米粒子，验证了其具有适宜的磁反应强度和较高的生物安全性。使用该Fe304磁性纳米粒子，赋予PPADT-PEG纳米粒磁性靶向功能，并通过磁场过滤法筛除未包裹磁核及磁性强度不足的纳米粒，合成和筛选了ROS-磁性双重靶向智能纳米粒。采用该纳米粒包裹SDF-1，形成(PPADT-PEG)-Fe3O4-SDF-1纳米粒，所合成纳米粒大小约139±46nm, 载药量约11.21%，包封率为35.89%。体外实验和大鼠在体实验验证其靶向性良好、静态释放率适宜、生物相容性优异。.项目组将(PPADT-PEG)-Fe3O4-SDF-1纳米粒应用于裸鼠全层皮肤缺损模型，自制创面外用的磁性敷料，于创面局部形成磁场。静脉输注(PPADT-PEG)-Fe3O4-SDF-1 纳米粒联合外源性GFP-BMSCs 移植，通过体内示踪进行创面靶向特性研究，证实其靶向性明显优于PPADT-SDF-1α纳米粒，能有效诱导BMSCs定向归巢。同时，创面愈合加速，作用机制主要为促进创面血管化。.本课题针对前期研究中携带SDF-1的ROS响应性纳米粒所存在的问题开展进一步研究，通过对纳米粒的改性，增加载药量，减少静释量，并通过增加磁性靶向，减少假阳性，按照课题计划完成了项目工作。这为创面修复，乃至组织修复与再生领域的治疗提供了新的思路和方法。但本项目仍需进行进一步研究，一方面需要提高操作在临床上的可行性，并进行进一步的安全性和有效性检测，为临床试验的开展打好基础，另一方面还需继续提高精准性，并对作用机制进行更深入的研究，为其他药物的精准治疗提供依据。