Liang Haixiang, Li Xudong, Shimer Adam L, Balian Gary, Shen Francis H
Department of Orthopaedic Surgery, Orthopaedic Research Laboratories, University of Virginia School of Medicine, Cobb Hall, Room B057, Box 800159, Charlottesville, VA 22908, USA.
Department of Orthopaedic Surgery, Orthopaedic Research Laboratories, University of Virginia School of Medicine, Cobb Hall, Room B057, Box 800159, Charlottesville, VA 22908, USA; Department of Biochemistry and Molecular Genetics, University of Virginia School of Medicine, Jordan Hall 6007, 1340 Jefferson Park Ave, Charlottesville, VA 22908, USA.
Spine J. 2014 Mar 1;14(3):445-54. doi: 10.1016/j.spinee.2013.09.045. Epub 2013 Oct 25.
Although the use of mesenchymal stem cells (MSC) with scaffolds for bone repair has been considered an effective method, the interactions between implanted materials and bone tissues have not been fully elucidated. At some specific sites, such as the vertebral body (VB) of the spine, the process of bone repair with implanted biomaterials is rarely reported. Recently, adipose tissue was found to be an alternative source of MSC besides bone marrow. However, the strategy of using adipose-derived stromal (ADS) cells with bioactive scaffold for the repair of spinal bone defects has seldom been studied.
To use a sintered poly(lactide-co-glycolide) acid (PLGA) microspheres scaffold seeded with induced rat ADS cells to repair a bone defect of the VB in a rat model.
Basic science and laboratory study.
A sintered porous microspheres scaffold was manufactured by PLGA. ADS cells were isolated from Fischer 344 rats and then induced by osteogenic medium with growth and differentiation factor 5 (GDF5) in vitro. Before implantation, cells were cultured with inductive media for 2 weeks as a monolayer situation and 1 more week on a PLGA scaffold as a three-dimensional structure. These assembled bioactive scaffolds then were implanted in lumbar VB bone defects in Fischer 344 rats. The ex vivo differentiation of the cells was confirmed by von Kossa staining and real-time polymerase chain reaction. The performance of cells on the scaffold was detected by scanning electron microscopy and (3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium) assay. In vivo bone formation was quantitatively measured by computed tomography study. And the effect of tissue repair was also evaluated by histological studies.
Proliferation and differentiation of cells were confirmed before in vivo implantation. Quantification of bone formation in vivo through serial three-dimensional computed tomography images revealed that the VB implanted with GDF5-induced cells demonstrated more bone formation than the control groups. Besides the bone formation period that occurred between 2 and 4 weeks in all groups, a second bone formation period was found to occur only in the groups that received cells with previous induction in vitro. This second period of significant bone formation happened simultaneously with collapsing of the scaffolds. It was then demonstrated histologically that vascularization early in the process and cooperation between host bone and implanted cells accompanied by collapse of the scaffold may be the factors that influence bone formation. This study not only provides a therapeutic strategy of using biomaterial for bone repair in the spine, but also may lead to a technological method for studying the relationship between implanted stem cells and host tissue.
Adipose-derived stromal cells maintained in culture on a scaffold and treated with osteogenic induction with growth factor ex vivo could be used to enhance bone repair in vivo.
尽管使用间充质干细胞（MSC）与支架进行骨修复被认为是一种有效的方法，但植入材料与骨组织之间的相互作用尚未完全阐明。在一些特定部位，如脊柱椎体（VB），使用植入生物材料进行骨修复的过程鲜有报道。最近，脂肪组织被发现是除骨髓之外MSC的另一个来源。然而，使用脂肪来源的基质（ADS）细胞与生物活性支架修复脊柱骨缺损的策略鲜有研究。
使用接种诱导大鼠ADS细胞的聚（丙交酯-共-乙交酯）酸（PLGA）微球烧结支架修复大鼠模型中的VB骨缺损。
基础科学与实验室研究。
用PLGA制造烧结多孔微球支架。从Fischer 344大鼠中分离出ADS细胞，然后在体外用人骨生长与分化因子5（GDF5）的成骨培养基诱导。植入前，将细胞在诱导培养基中单层培养2周，然后在PLGA支架上作为三维结构再培养1周。然后将这些组装好的生物活性支架植入Fischer 344大鼠的腰椎VB骨缺损处。通过von Kossa染色和实时聚合酶链反应确认细胞的体外分化。通过扫描电子显微镜和（3-（4,5-二甲基噻唑-2-基）-5-（3-羧甲氧基苯基）-2-（4-磺基苯基）-2H-四唑）测定法检测细胞在支架上的性能。通过计算机断层扫描研究定量测量体内骨形成。并通过组织学研究评估组织修复效果。
在体内植入前确认了细胞的增殖和分化。通过连续三维计算机断层扫描图像对体内骨形成进行定量分析，结果显示植入GDF5诱导细胞的VB比对照组显示出更多的骨形成。除了所有组在2至4周之间出现骨形成期外，还发现仅在体外预先诱导细胞的组中出现第二个骨形成期。这个显著的骨形成的第二阶段与支架的塌陷同时发生。然后组织学证明，该过程早期的血管化以及宿主骨与植入细胞之间的协同作用以及支架的塌陷可能是影响骨形成的因素。本研究不仅提供了一种使用生物材料修复脊柱骨的治疗策略，还可能导致一种研究植入干细胞与宿主组织之间关系的技术方法。
在支架上培养并经体外生长因子成骨诱导处理的脂肪来源基质细胞可用于增强体内骨修复。
