gms | German Medical Science

Objectives: Does the local application of hUCMSC-EVs enhance the osseointegration, bone regeneration and fracture stability in an osteoporotic metaphyseal fracture?

Methods: Estrogen dysregulation combined with a dietary treatment was achieved with a ventral ovariectomy performed on 52 female Sprague Dawley rats at 12 weeks. The osteoporotic effect was enhanced in all ovariectomized (OVX) rats by a Phosphate-, Vitamin D3-, Soja, and Phytoestrogen reduced diet for 6 weeks post-surgery. OVX rats were analyzed by peripheral quantitative computed tomography pQCT) at biweekly time points to confirm reduced bone mineral density and content compared to controls (CTRL) over 14 weeks. After 8 weeks, a 1mm fracture of the right distal metaphyseal area of the femur was internally stabilized with a Y-shaped mini plate. OVX rats were divided into three subgroups: OVX without vehicle (n=9), OVX with alginate scaffold carrier (n=21), and OVX with hUCMSC-EVs on alginate scaffold (2.0 x 109 hUCMSC-EVs) (n=21). The corresponding treatments were administered directly into the fracture gap and monitored with weekly X-rays.

Results and Conclusion: pQCT analysis of the proximal tibia metaphysis showed a significant difference in the volumetric cancellous bone mineral density (Trabecular density/TRAB_DEN) between OVX and CTRL rats (P<0.0001). TRAB_DEN for OVX rats revealed a mean of 176.0mg/cm3±57.1 and 284.9mg/cm3±31.5 for CTRL rats. A significant difference was also observed in the volumetric total bone mineral density (Total density /TOT_DEN) with a mean of 518.2mg/cm3±31.1 for OVX and 637.0mg/cm3±11.1 for CTRL rats (P<0.0001). Fracture consolidation on X-ray was observed in all three groups. After 6 weeks biomechanical testing for bending stiffness and torsion will be carried out. Micro-computed tomography will provide measurements of bone volume, trabecular bone number and thickness in all groups. All results will be confirmed by histology at 2-, 4-, and 6-weeks post-surgery on formalin-fixed paraffin-embedded femoral sections. Movat Pentachrome and Mallory's Aniline Blue stains will be used, and osteoclast activity will be determined by TRAP staining and RANKL immunohistochemical analysis. We expect that osteoporotic fracture healing in rats will be enhanced by the direct application of hUCMSC-EVs using alginate as a carrier into the fracture to improve osseointegration, increase fracture stability and promote bone regeneration.