Ex vivo antioxidant preconditioning improves the survival rate of bone marrow stem cells in the presence of wound fluid

Abstract The advancement of autologous mesenchymal stem cell (MSC) therapy for the treatment of non‐healing diabetic wounds is hampered by endogenous MSC dysfunction and limited viability of cells post‐transplantation into the pathological wound environment. The development of effective strategies to restore the functional capabilities of these impaired MSCs prior to transplantation may be a key to their ultimate success as wound repair mediators. The current study therefore investigated whether antioxidant preconditioning [7.5 mM N‐acetylcysteine (NAC) + 0.6 mM ascorbic 2‐phosphate (AAP)] could restore the growth rate, migration ability and viability of impaired MSCs and whether this restored state is maintained in the presence of diabetic wound fluid (DWF). Healthy control (source: wild type, C57BL/6J mice) (n = 12) and impaired/diabetic MSCs (source: obese prediabetic, B6.Cg‐Lepob/J mice) (n = 12) were isolated from the bone marrow of mice. Treatment groups post‐isolation were as follow: (a) No treatment ( baseline phenotype ): MSCs expanded in standard growth media (SGM) (±8 days) and only exposed to growth media. (b) DWF ( baseline response ): MSCs expanded in SGM (±8 days) followed by exposure to DWF (24 hours, 48 hours, 96 hours). (c) Antioxidant preconditioning ( preconditioned phenotype ): MSCs expanded in the presence of NAC/AAP (±8 days). (d) Antioxidant preconditioning + DWF ( preconditioned response ): MSCs expanded in the presence of NAC/AAP (±8 days) followed by exposure to DWF (24 hours, 48 hours, 96 hours). The results demonstrated that expansion of MSCs (both healthy control and impaired diabetic) in the presence of combined NAC/AAP treatment improved ex vivo MSC viability and protected MSCs in the presence of DWF. Despite improved viability, AAP/NAC could however not rescue the reduced proliferation and migration capacity of impaired diabetic MSCs. The protective effect of NAC/AAP preconditioning against the toxicity of DWF could however be a potential strategy to improve cell number post‐transplantation.