生长因子次第控释系统对组织工程骨血管化的调控研究

Rapid and effective vascular reconstruction is a key to the success of tissue engineering bone graft, and regulation of growth factors on angiogenesis is crucial. Previous studies have shown that the inductive mechanism, effective dose and timing, synergistic effects of various growth factors are different. However, the optimal induced model of growth factors synergistic regulation is still unclear and the delivery system that could release multiple growth factors sequentially is rare. According to inductive properties and mechanism of growth factors and the advantage of supercritical fluid technology (SFT) in keeping proteins activity, our study attempts to construct a delivery system that could sequentially release multiple growth factors following the expected design using SFT. Through studying the differences of induction, effective dose and timing, and synergy combinations of various growth factors, we will establish the delivery model which could release different growth factors at the specific time and space, encapsulate growth factors into microspheres and construct multiple growth factors sequential released system combining with scaffolds by using SFT, and investigate effects of this delivery system on angiogenesis in vitro and in vivo. Through above researches, it expect to construct a multiple growth factors sequential released system simulating physiological repair process, realize effective vascularization of tissue engineering bone, and provide theoretical and experimental basis for promoting the practical application of tissue engineering bone.

迅速有效的血管重建是组织工程骨移植成功的关键，生长因子的调控对血管生成至关重要。我们前期研究证明，不同生长因子诱导血管生成的机理、有效剂量和时序、协同效应均不同，但各种因子协同调控血管生成的最佳诱导模式及机理尚不清楚，且缺乏一种能次第释放多种生长因子的载体系统。本项目根据不同生长因子对血管生成的诱导特性和机理，结合超临界流体技术利于保持蛋白药物活性的优势，拟构建一种能按照预期设计次第释放多种生长因子的载体系统。通过研究生长因子调控血管生成的诱导差异性、有效剂量和时序、协同作用组合，建立不同因子特定的时间和空间释药模式，采用超临界流体技术将不同生长因子分别载入各自微球中，与支架复合构建多因子次第控释系统，考察其在体内外诱导血管生成的效能。期望通过上述研究，构建一种模拟生理修复的过程次第释放多种生长因子的缓释系统，有效地实现组织工程骨的血管化，为推动组织工程骨的实际应用提供理论和实验依据。

生长因子的调控对血管生成至关重要。本项目根据PDGF、FGF-2和VEGF三种生长因子对血管生成的诱导特性和机理，构建了一种能次第释放多种生长因子的载体系统。首先通过PDGF、FGF-2和VEGF对血管内皮细胞增殖和血管分化的作用研究了三种生长因子调控血管生成的诱导差异性、有效剂量和时序、协同作用组合，建立了模拟生理修复的时间和空间释药模式；筛选制备微球和支架的制备材料和工艺参数，采用乳化法制备载PDGF的聚乳酸微球PDGF-PLLAms，微球呈规则完整球形，表面光滑，平均粒径约5μm，包封率约72.5%，二氯甲烷残留量为0.0025%；采用超临界二氧化碳发泡技术制备复合PDGF-PLLAms、VEGF和FGF-2的PLGA多孔支架，孔隙率为71.92%，孔径为50-100μm，抗压强度5.48Mpa，有效地实现了释放前期以VEGF和FGF-2为主，中后期以PDGF为主的释药模式。最后，分别考察了复合支架在体内外对血管生成的作用，结果证明复合支架能够有效地诱导体内外血管生成。综上所述，本研究为药物次第释放载体系统的制备提供了技术方法，并为组织工程骨快速有效的血管形成的奠定了坚实的工作基础。项目资助发表SCI论文4篇，EI论文1篇，CSCD论文1篇，申请发明专利1项，待发表论文2篇。