SPECT/CT imaging for the evaluation of the differentiation efficacy of canine adipose tissue derived mesenchymal stem cells (cAD-MSC) into osteogenic lineage

The purpose of our study was to isolate, amplify and characterize adipose-derived MSC from Beagle dogs, with a particular analysis of their osteogenic differentiation potential. SPECT/CT imaging was performed to observe the ability of cAD-MSC to promote ectopic bone formation on a three-dimensional scaffold. cAD-MSCs were harvested from subcutaneous and visceral fat of Beagle dogs. Histological and phenotypic characterization was made before in vivo studies. Cells from young donors with the highest proliferative and osteogenic potential were chosen to evaluate in-vivo behavior of the cAD-MSC towards osteogenic differentiation. Each applied chitosan gelatin scaffold cross-linked with glutaraldehyde contained 2x105 cAD-MSC. After 2 days proliferation in regular medium and one week in osteomedium, scaffolds were implanted in 10 nude mice, 5-6 weeks old divided in 2 groups used for imaging studies. Ali mice received scaffold containing osteoinduced cells on the right side. The 5 mice from the second group were implanted also with an empty scaffold on the left side. The SPECT/CT whole body images were performed after two months. 80 Bq (100 µI) Tc-99m-MDP were injected into the tail vein of the mice. Two hours later imaging was made by laboratory SPECT/CT hybrid camera (nanoSPECT/CT, Mediso Ltd, Hungary). SPECT/CT analysis of the dorsal region of mice containing an osteoinduced cAD-MSC-seeded scaffold showed Tc-99m-MDP uptake without exceptions. Bone-like structure was evidenced in the CT slices. On the contrary Tc-99m-MDP accumulation was not present in the empty scaffolds in the control mice. Histological analysis by von Kossa staining showed high mineral concentration in the osteoinduced cAD-MSC scaffolds. Isolated cAD-MSC have osteogenic potential and have a possible application for the treatment of bone diseases in veterinary medicine. Our results demonstrate that direct in vivo imaging with Tc-99m-MDP is a valid noninvasive method for the evaluation of new bone formation in tissue engineering as it is adsorbed preferentially by newly formed minerals and for clinical studies to determine the bone formation dynamics.