Dimethyloxalylglycine,a small molecule,synergistically increases the homing and angiogenic properties of human mesenchymal stromal cells when cultured as 3D spheroids

Strategies aiming at increasing the survival and paracrine activity of human mesenchymal stromal cells (MSCs) are of utmost importance to achieve the full therapeutic potential of these cells. Herein, we propose both physical and biochemical strategies to enhance the survival, homing, angiogenic, and immunomodulatory properties of MSCs in vitro. To that purpose, we compared the effect of exposing either 2D monolayer or 3D spheroids of MSCs to (i) hypoxia (2% O₂) or to (ii) a hypoxic-mimetic small molecule, dimethyloxalylglycine (DMOG), with cells cultured at 21% O₂. 3D-cultured MSC spheroids evidenced higher survival upon exposure to oxidative stress and expressed higher levels of factors involved in tissue repair processes, namely tumor necrosis factor-stimulated gene-6, matrix metalloproteinase-2, and vascular endothelial growth factor. MSCs cultured as 3D spheroids and further exposed to hypoxia or hypoxic-mimetic conditions provided by DMOG synergistically favored the expression of the cell surface marker C-X-C chemokine receptor type-4, involved in homing processes to injured tissues, and adhesion to extracellular matrix components as fibronectin. These results highlight the role of ex vivo preconditioning approaches, presenting a novel strategy that combine biochemical stimuli with 3D spheroid organization of MSCs to maximize their tissue regeneration potential.