Liu Zunpeng, Zhu Yue, Zhu Haitao, He Xiaoning, Liu Xinchun
Department of Orthopedics, First Affiliated Hospital, China Medical University, Shenyang, China Department of Orthopedics, Fourth Affiliated Hospital, China Medical University, Shenyang, China.
Department of Orthopedics, First Affiliated Hospital, China Medical University, Shenyang, China
J Biomater Appl. 2016 Oct;31(4):477-487. doi: 10.1177/0885328216643854. Epub 2016 Apr 7.
Mesenchymal stem cells have shown great potential for accelerating bone healing. In the present study, we evaluate the efficacy of fibrin glue/mesenchymal stem cells/recombinant human bone morphogenetic protein-2 composite for posterolateral spinal fusion in a rabbit model. Forty adult rabbits underwent posterolateral intertransverse fusion at the L5-L6 level. The animals were randomly divided into four groups based on the implant material: fibrin glue, fibrin glue/mesenchymal stem cells composite, fibrin glue-recombinant human bone morphogenetic protein-2 (fibrin glue/recombinant human bone morphogenetic protein-2) composite, and fibrin glue/mesenchymal stem cells/recombinant human bone morphogenetic protein-2 composite. After six weeks, the rabbits were euthanized for manual palpation, radiographic examination, biomechanical testing, and histology. Manual palpation results showed that the fusion rate for fibrin glue, fibrin glue/mesenchymal stem cells, fibrin glue/recombinant human bone morphogenetic protein-2, and fibrin glue/mesenchymal stem cells/recombinant human bone morphogenetic protein-2 was 0, 0, 40%, and 70%, respectively. Moreover, fusion rate determined by radiographic examination for fibrin glue, fibrin glue/mesenchymal stem cells, fibrin glue/recombinant human bone morphogenetic protein-2, and fibrin glue/mesenchymal stem cells/recombinant human bone morphogenetic protein-2 was 0, 0, 40%, and 80%, respectively. Gray analysis showed that fibrin glue/recombinant human bone morphogenetic protein-2 group had higher ossification area and density than fibrin glue group; and fibrin glue/mesenchymal stem cells/recombinant human bone morphogenetic protein-2 group had higher ossification area and density than fibrin glue/recombinant human bone morphogenetic protein-2 group. Formation of continuous bone masses between L5 and L6 level in mesenchymal stem cells/recombinant human bone morphogenetic protein-2/fibrin glue group was further confirmed by computed tomography scanning and three-dimensional reconstruction. Biomechanical testing demonstrated that the fusion strength (flexion, extension, lateral bending, and axial rotation) in fibrin glue/mesenchymal stem cells/recombinant human bone morphogenetic protein-2 group is significantly higher than that in fibrin glue/recombinant human bone morphogenetic protein-2 group. The formation of mature bone tissues between transverse processes of the fused specimens from both fibrin glue/recombinant human bone morphogenetic protein-2, and fibrin glue/mesenchymal stem cells/recombinant human bone morphogenetic protein-2 groups was confirmed by HE staining, and quantitative real-time polymerase chain reaction results showed the upregulation of CD31, type I collagen, osteocalcin, and osteonectin in the fibrin glue/mesenchymal stem cells/recombinant human bone morphogenetic protein-2 group. In conclusion, our findings show that mesenchymal stem cells delivered with recombinant human bone morphogenetic protein-2 using fibrin glue as carrier are more effective in enhancing spine fusion than recombinant human bone morphogenetic protein-2 without mesenchymal stem cells in the rabbit model.
间充质干细胞在加速骨愈合方面已显示出巨大潜力。在本研究中，我们在兔模型中评估了纤维蛋白胶/间充质干细胞/重组人骨形态发生蛋白-2复合物用于腰椎后路椎间融合的疗效。40只成年兔在L5-L6水平进行后路横突间融合。根据植入材料将动物随机分为四组：纤维蛋白胶组、纤维蛋白胶/间充质干细胞复合物组、纤维蛋白胶-重组人骨形态发生蛋白-2（纤维蛋白胶/重组人骨形态发生蛋白-2）复合物组和纤维蛋白胶/间充质干细胞/重组人骨形态发生蛋白-2复合物组。六周后，对兔子实施安乐死以进行手动触诊、影像学检查、生物力学测试和组织学检查。手动触诊结果显示，纤维蛋白胶组、纤维蛋白胶/间充质干细胞组、纤维蛋白胶/重组人骨形态发生蛋白-2组和纤维蛋白胶/间充质干细胞/重组人骨形态发生蛋白-2组的融合率分别为0%、0%、40%和70%。此外，影像学检查确定的纤维蛋白胶组、纤维蛋白胶/间充质干细胞组、纤维蛋白胶/重组人骨形态发生蛋白-2组和纤维蛋白胶/间充质干细胞/重组人骨形态发生蛋白-2组的融合率分别为0%、0%、40%和80%。灰度分析显示，纤维蛋白胶/重组人骨形态发生蛋白-2组的骨化面积和密度高于纤维蛋白胶组；纤维蛋白胶/间充质干细胞/重组人骨形态发生蛋白-2组的骨化面积和密度高于纤维蛋白胶/重组人骨形态发生蛋白-2组。计算机断层扫描和三维重建进一步证实了间充质干细胞/重组人骨形态发生蛋白-2/纤维蛋白胶组在L5和L6水平之间形成了连续的骨块。生物力学测试表明，纤维蛋白胶/间充质干细胞/重组人骨形态发生蛋白-2组的融合强度（前屈、后伸、侧弯和轴向旋转）显著高于纤维蛋白胶/重组人骨形态发生蛋白-2组。HE染色证实了纤维蛋白胶/重组人骨形态发生蛋白-2组和纤维蛋白胶/间充质干细胞/重组人骨形态发生蛋白-2组融合标本横突之间成熟骨组织的形成，定量实时聚合酶链反应结果显示纤维蛋白胶/间充质干细胞/重组人骨形态发生蛋白-2组中CD31、I型胶原、骨钙素和骨连接素上调。总之，我们的研究结果表明，在兔模型中，以纤维蛋白胶为载体递送重组人骨形态发生蛋白-2的间充质干细胞在增强脊柱融合方面比不含间充质干细胞的重组人骨形态发生蛋白-2更有效。
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