Department of Orthopaedic Surgery, Yong Loo Lin School of Medicine, National University of Singapore, Singapore, Singapore.
Tissue Eng Part A. 2013 Feb;19(3-4):350-9. doi: 10.1089/ten.TEA.2012.0274. Epub 2012 Nov 7.
In this study, we tested the hypothesis that a surface functionalization delivery platform incorporating heparin onto strontium alginate microbeads surfaces would convert this "naive carriers" into "mini-reservoirs" for localized in vivo delivery of recombinant human bone morphogenetic protein-2 (rhBMP-2) that will induce functional bone regeneration. In vitro evaluation confirmed that (1) heparin incorporation could immobilize and prolong rhBMP-2 release for approximately 3 weeks; (2) a significant decrease (p<0.01) in rhBMP-2 burst release is attainable depending on initial protein load; and (3) rhBMP-2 released from surface functionalized microbeads retained bioactivity and stimulated higher alkaline phosphatase activity in cultured C(2)C(12) cells when compared with daily administration of fresh bolus rhBMP-2. Subsequently, surface functionalized microbeads were used for in vivo delivery of rhBMP-2 at local sites of posterolateral spinal fusion surgery in rats. The microbeads were loaded into the pores of medical-grade polyepsilone caprolactone-tricalcium phosphate scaffolds before implantation. Results revealed robust bone formation and a biomechanically solid fusion after 6 weeks. When compared with a control group consisting of an equivalent amount of rhBMP-2 that was directly adsorbed onto bare-surfaced microbeads with no heparin, a 5.3-fold increase in bone volume fraction and a 2.6-fold increase in bending stiffness (flexion/extension) were observed. When compared with collagen sponge carriers of rhBMP-2, a 1.5-fold and a 1.3-fold increase in bone volume fraction and bending stiffness were observed, respectively. More importantly, 3D micro-computed tomography images enabled the visualization of a well-contained newly formed bone at ipsilateral implant sites with surface functionalized rhBMP-2 delivery. This was absent with collagen sponge carriers where newly formed bone tissue was poorly contained and crossed over the posterior midline to contralateral implants. These findings are important because of complications with current rhBMP-2 delivery method, including excessive, uncontrolled bone formation.
在这项研究中，我们测试了一个假设，即通过将肝素结合到锶藻酸盐微珠表面上的表面功能化递药平台，将这些“原始载体”转化为局部体内递送重组人骨形态发生蛋白-2（rhBMP-2）的“迷你储库”，从而诱导功能性骨再生。体外评估证实：（1）肝素的掺入可以固定并延长 rhBMP-2 的释放约 3 周；（2）根据初始蛋白质负载，可以实现 rhBMP-2 突释的显著减少（p<0.01）；（3）与每日给予新鲜的 rhBMP-2 相比，从表面功能化微珠释放的 rhBMP-2 保持生物活性，并刺激培养的 C(2)C(12)细胞中碱性磷酸酶活性更高。随后，在大鼠后路脊柱融合手术的局部部位，使用表面功能化微珠进行 rhBMP-2 的体内递送。在植入前，将微珠装入医用级聚己内酯-磷酸三钙支架的孔中。结果显示，在 6 周后出现了强大的骨形成和生物力学上坚固的融合。与由等量的 rhBMP-2 组成的对照组相比，对照组由等量的 rhBMP-2 直接吸附在没有肝素的裸露表面微珠上，骨体积分数增加了 5.3 倍，弯曲刚度（弯曲/伸展）增加了 2.6 倍。与 rhBMP-2 的胶原海绵载体相比，骨体积分数和弯曲刚度分别增加了 1.5 倍和 1.3 倍。更重要的是，3D 微计算机断层扫描图像能够可视化具有表面功能化 rhBMP-2 递送的同侧植入部位的新形成骨。而使用胶原海绵载体时，新形成的骨组织无法很好地包裹，并且跨越中线到达对侧植入物。这些发现很重要，因为当前 rhBMP-2 递送方法存在并发症，包括过度、不受控制的骨形成。
