Systemic and local administration of allogeneic bone marrow derived mesenchymal stem cells promotes fracture healing in rats

Abstract

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 the GFP-Sprague-Dawley rats, cultured and characterized. Closed transverse femoral fracture with internal fixation was established in 48 adult male Sprague-Dawley rats, whom were randomly assigned into 4 groups receiving: PBS injection; MSCs systemic injection; Fibroblasts systemic injection and MSCs fracture site injection. 2x10⁶ cells were injected at 4 days after fracture. All animals were terminated at 5 weeks after fracture; examinations included weekly radiograph; Micro-CT; mechanical testing; histology, immunohistochemistry and double immunofluorescence. The callus size of MSCs injection groups were significant larger among all the groups. Radiographs and 3D-reconstruction images showed that the fracture gaps united in the MSCs injected groups, while gaps were still seen in the fibroblast and PBS injection groups. The mechanical properties were significantly higher in the MSCs injection groups than those in the fibroblast and PBS groups, but no difference was found between the MSCs local and systemic injection groups. Immunohistochemistry and double immunofluorescence demonstrated that GFP-positive MSCs were present in the callus in the MSCs injection groups at 5 weeks after fracture, and some have differentiated into osteoblasts. Quantitative analysis revealed the number of GFP-positive cells in the callus in the MSCs systemic injection group was significantly lower than that of the MSCs local injection group. The proportion of GFP-osteoblasts in GFP-positive cells in the MSCs systemic injection group was significantly lower than that of the MSCs 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.