Effects of Dimethyloxaloglycine-Preconditioning on Osteogenesis and Vascularization in Adipose Derived Mesenchymal Stromal Cells Using a Rat Spinal Fusion Model

Introduction: Adipose-derived mesenchymal stromal cells (ASCs) offer a promising alternative to bone marrow-derived mesenchymal stromal cells (BMSCs) for bone regeneration. However, low cell survival in these cells due to in vivo hypoxic environment remains challenging. Hypoxia Inducible Factor-1α (HIF-1α) aids cellular adaptation to hypoxia while Dimethyloxaloylglycine (DMOG) has been shown to stabilize HIF-1α and enhance osteogenic and angiogenic capacity. We aimed to demonstrate that DMOG and hypoxia preconditioning of ASCs enhance osteogenesis and vascularization thus improving spinal fusion in a rat model.

Methods: ASCs were isolated from the inguinal fat pads of syngeneic 6–8-week-old Lewis rats, culture-expanded, and preconditioned at passage 1 (P1) (80% confluency) with 1ng of DMOG for 24 hours. Passage 2 (P2) ASCs (2x106) were seeded onto Vitoss scaffolds for transplantation. Lewis rats underwent L4-L5 posterolateral spinal fusion and were assigned to one of two groups: [1]Vitoss+DMOG-preconditioned ASCP2s; and, [2]Vitoss+non-preconditioned ASCP2s. Fusion was assessed eight weeks post-surgery via manual palpation (scored as 0=non-fused, 1=partial fusion, and, 2=fused), micro-computed tomography (µCT) imaging (0=non-fused; 1=unilateral fusion; 2=bilateral fusion), and histology. Histology was used to assess bone quality and neovascularization within the fusion masses.

Results: µCT imaging data suggested that DMOG-preconditioned ASCP2 group yielded significantly higher fusion mass volumes than non-preconditioned ASCP2 group (23.49 mm3 vs. 15.39 mm3, p=.001). DMOG-preconditioned ASCP2 group showed a trend towards higher rates of µCT fusion and manual palpation scores compared to the non-preconditioned ASCP2 group (1.16 vs. 0.50, p=0.06; and 1.25 vs. 0.66, p>0.05). On histology, the preconditioned group displayed greater osteoid deposition, denser trabeculae, greater osteoblast density, and greater vascularization.

Conclusion : In our rat posterolateral fusion model, hypoxia preconditioned culture-expanded ASCs exhibited significantly increased fusion mass volumes compared to non-preconditioned ASCs. Additionally, µCT and manual palpation scores were higher with greater bone formation and vascularity on histology in the group pretreated with DMOG.