Kempen Diederik H R, Lu Lichun, Hefferan Teresa E, Creemers Laura B, Maran Avudaiappan, Classic Kelly L, Dhert Wouter J A, Yaszemski Michael J
Tissue Engineering and Biomaterials Laboratory, Department of Orthopedic Surgery, Mayo Clinic College of Medicine, 200 First Street SW, Rochester, MN 55905, USA.
Biomaterials. 2008 Aug;29(22):3245-52. doi: 10.1016/j.biomaterials.2008.04.031. Epub 2008 May 9.
In this study, we investigated the in vitro and in vivo biological activities of bone morphogenetic protein 2 (BMP-2) released from four sustained delivery vehicles for bone regeneration. BMP-2 was incorporated into (1) a gelatin hydrogel, (2) poly(lactic-co-glycolic acid) (PLGA) microspheres embedded in a gelatin hydrogel, (3) microspheres embedded in a poly(propylene fumarate) (PPF) scaffold and (4) microspheres embedded in a PPF scaffold surrounded by a gelatin hydrogel. A fraction of the incorporated BMP-2 was radiolabeled with (125)I to determine its in vitro and in vivo release profiles. The release and bioactivity of BMP-2 were tested weekly over a period of 12 weeks in preosteoblast W20-17 cell line culture and in a rat subcutaneous implantation model. Outcome parameters for in vitro and in vivo bioactivities of the released BMP-2 were alkaline phosphatase (AP) induction and bone formation, respectively. The four implant types showed different in vitro release profiles over the 12-week period, which changed significantly upon implantation. The AP induction by BMP-2 released from gelatin implants showed a loss in bioactivity after 6 weeks in culture, while the BMP-2 released from the other implants continued to show bioactivity over the full 12-week period. Micro-CT and histological analysis of the delivery vehicles after 6 weeks of implantation showed significantly more bone in the microsphere/PPF scaffold composites (Implant 3, p<0.02). After 12 weeks, the amount of newly formed bone in the microsphere/PPF scaffolds remained significantly higher than that in the gelatin and microsphere/gelatin hydrogels (p<0.001), however, there was no statistical difference compared to the microsphere/PPF/gelatin composite. Overall, the results from this study show that BMP-2 could be incorporated into various bone tissue engineering composites for sustained release over a prolonged period of time with retention of bioactivity.
在本研究中，我们调查了从四种用于骨再生的缓释载体释放的骨形态发生蛋白2（BMP-2）的体外和体内生物活性。BMP-2被掺入（1）明胶水凝胶、（2）嵌入明胶水凝胶中的聚（乳酸-共-乙醇酸）（PLGA）微球、（3）嵌入聚富马酸丙二醇酯（PPF）支架中的微球以及（4）被明胶水凝胶包围的嵌入PPF支架中的微球。一部分掺入的BMP-2用（125）I进行放射性标记，以确定其体外和体内释放曲线。在原代成骨细胞W20-17细胞系培养和大鼠皮下植入模型中，在12周的时间内每周测试BMP-2的释放和生物活性。释放的BMP-2体外和体内生物活性的结果参数分别是碱性磷酸酶（AP）诱导和骨形成。在12周期间，四种植入物类型显示出不同的体外释放曲线，植入后发生了显著变化。从明胶植入物释放的BMP-2诱导的AP在培养6周后生物活性丧失，而从其他植入物释放的BMP-2在整个12周期间持续显示生物活性。植入6周后对载体进行的显微CT和组织学分析显示，微球/PPF支架复合材料（植入物3，p<0.02）中有明显更多的骨。12周后，微球/PPF支架中新形成的骨量仍显著高于明胶和微球/明胶水凝胶中的骨量（p<0.001），然而，与微球/PPF/明胶复合材料相比没有统计学差异。总体而言，本研究结果表明，BMP-2可掺入各种骨组织工程复合材料中，以长时间持续释放并保留生物活性。
