纳米颗粒复合可注射多功能双相水凝胶微球系统修复椎间盘损伤的实验研究

The goal of this project is the development of a novel strategy using nanoparticle-conjugated dual-phase multifunctional injectable hydrogel microsphere system of gelatin-hydroxyphenyl propionic acid (Gtn-HPA) to facilitate the reparative response in the intervertebral disc (IVD) after discectomy. A liquid formulation of Gtn-HPA will be used as a carrier for the injection of the preformed Gtn-HPA microspheres into the site of the discectomy. The novel aspect of the liquid formulation of the gel is that the degree and rate of cross-linking can be controlled independently by the concentrations of peroxidase and peroxide that are added to the Gtn-HPA during the injection process. The two-phase system of the preformed microspheres and carrier gel enable broader control over the mechanical and degradative properties, and release of multiple regulatory molecules and, if necessary in the future, exogenous cells. Therefore, for the treatment of IVD after discectomy, it would be useful to develop an injectable two-phase gel system: one phase is the performed microspheres of stiffer gel with higher mechanical strength but slower degradative speed, which can contribute to the early mechanical stabilization of the violated disc immediately after being injected into the lesion; the other phase is a liquid formulation of Gtn-HPA, which would form a softer gel between the microspheres and the host tissue after injection, to facilitate the migration of endogenous cells into the defect for a reparative response. The preformed microsphere can also serve as a vehicle for the controlled release of therapeutic agents. In this study, the PDGF-BB will be incorporated in the microspheres to facilitate cell recruiting and contained in nanoparticles to control its release. The objective of this project is: 1) to evaluate, in vitro, the biological and mechanical behavior of the novel injectable dual-phase microsphere system in order to determine the most promising formulation to evaluate histomorphometrically the tissue response in a standardized defect in the IVD in an animal model; 2) using a goat IVD model to evaluate the repairing tissue response of the dual-phase microsphere system histologically.

椎间盘组织由于自身修复能力差，目前尚无满意的再生和修复间盘损伤的治疗方法。本项目将利用新型可注射共价交联水凝胶—羟苯基丙酸/明胶（Gtn-HPA）构建纳米颗粒复合可注射多功能双相微球水凝胶系统修复椎间盘损伤：用较软的低浓度Gtn-HPA作为基质相，有助于细胞迁移和增殖的；用硬度高浓度Gtn-HPA作为微球相，防止水凝胶被过快过早吸收，有助于维持受损间盘的早期稳定性；同时，将载有血小板源性生长因子（PDGF-BB）的纳米颗粒与高浓度Gtn-HPA微球相进行复合，超长时持续释放PDGF-BB，诱导内源性干细胞和血管前期细胞迁移入间盘缺损（水凝胶系统）内，促进修复再生进程。在本项目研究中，我们将先通过体外实验对该新型水凝胶系统的物理化学性质、生物亲和性、内源性细胞诱导迁移能力等进行评和研究，优化和确定体系参数；再用山羊作为腰间盘损伤大动物实验模型，在组织学上评价修复效果，研究间盘损伤修复机制。