Huang Shuo, Xu Liangliang, Zhang Yifeng, Sun Yuxin, Li Gang
Department of Orthopaedics and Traumatology, Li Ka Shing Institute of Health Sciences, The Chinese University of Hong Kong, Prince of Wales Hospital, Shatin, Hong Kong, PR China.
Cell Transplant. 2015;24(12):2643-55. doi: 10.3727/096368915X687219. Epub 2015 Feb 2.
Mesenchymal stem cells (MSCs) are immune privileged and a cell source for tissue repair. Previous studies showed that there is systemic mobilization of osteoblastic precursors to the fracture site. We hypothesized that both systemic and local administration of allogeneic MSCs may promote fracture healing. Bone marrow-derived MSCs and skin fibroblasts were isolated from GFP Sprague-Dawley rats, cultured, and characterized. Closed transverse femoral fracture with internal fixation was established in 48 adult male Sprague-Dawley rats, which were randomly assigned into four groups receiving PBS injection, MSC systemic injection, fibroblast systemic injection, and MSC fracture site injection; 2 × 10(6) cells were injected at 4 days after fracture. All animals were sacrificed at 5 weeks after fracture; examinations included weekly radiograph, micro-CT, mechanical testing, histology, immunohistochemistry, and double immunofluorescence. The callus size of MSC injection groups was significantly larger among all the groups. Radiographs and 3D reconstruction images showed that the fracture gaps united in the MSC injected groups, while gaps were still seen in the fibroblast and PBS injection groups. The mechanical properties were significantly higher in the MSC injection groups than those in the fibroblast and PBS groups, but no difference was found between the MSC local and systemic injection groups. Immunohistochemistry and double immunofluorescence demonstrated that GFP-positive MSCs were present in the callus in the MSC injection groups at 5 weeks after fracture, and some differentiated into osteoblasts. Quantitative analysis revealed the number of GFP-positive cells in the callus in the MSC systemic injection group was significantly lower than that of the MSC local injection group. The proportion of GFP osteoblasts in GFP-positive cells in the MSC systemic injection group was significantly lower than that of the MSC local injection group. These findings provide critical insight for developing MSC-based therapies, and systemic injection of allogeneic MSCs may be a novel treatment method for promoting fracture repair.
间充质干细胞（MSCs）具有免疫特权，是组织修复的细胞来源。先前的研究表明，成骨前体细胞会向骨折部位进行全身动员。我们推测，异体间充质干细胞的全身给药和局部给药都可能促进骨折愈合。从绿色荧光蛋白（GFP）转基因的斯普拉格-道利大鼠中分离出骨髓来源的间充质干细胞和皮肤成纤维细胞，进行培养并鉴定。对48只成年雄性斯普拉格-道利大鼠造成闭合性股骨横行骨折并进行内固定，将其随机分为四组，分别接受磷酸盐缓冲液（PBS）注射、间充质干细胞全身注射、成纤维细胞全身注射以及间充质干细胞骨折部位注射；在骨折后4天注射2×10⁶个细胞。所有动物在骨折后5周处死；检查包括每周的X线片、显微CT、力学测试、组织学、免疫组织化学以及双重免疫荧光。在所有组中，间充质干细胞注射组的骨痂大小显著更大。X线片和三维重建图像显示，间充质干细胞注射组的骨折间隙已愈合，而成纤维细胞和PBS注射组仍可见间隙。间充质干细胞注射组的力学性能显著高于成纤维细胞和PBS组，但间充质干细胞局部注射组和全身注射组之间未发现差异。免疫组织化学和双重免疫荧光显示，骨折后5周，间充质干细胞注射组的骨痂中存在绿色荧光蛋白阳性的间充质干细胞，且部分分化为成骨细胞。定量分析显示，间充质干细胞全身注射组骨痂中绿色荧光蛋白阳性细胞的数量显著低于间充质干细胞局部注射组。间充质干细胞全身注射组中绿色荧光蛋白阳性细胞中成骨细胞的比例显著低于间充质干细胞局部注射组。这些发现为开发基于间充质干细胞的治疗方法提供了关键见解，异体间充质干细胞的全身注射可能是促进骨折修复的一种新的治疗方法。
