Evaluation of the effects of ascorbic acid on metabolism of human mesenchymal stem cells

Background: Mesenchymal stem cells (MSCs) are multipotent cells holding much promise for applications in regenerative medicine. However, with problems such as aging, increases in heteroploid cells, genomic instability, and reduced maintenance of stemness, more stable culturing methods and the production of MSCs with an improved therapeutic effect are desired. Ascorbic acid (AsA), which is a cofactor for a variety of enzymes and has an antioxidant effect, cannot be synthesized by certain animals, including humans. Nevertheless, little attention has been paid to AsA when culturing MSCs. Methods: We analyzed the effect of adding AsA to the culture medium on the proliferation and metabolism of human MSCs by serial analysis of gene expression and metabolome analysis. Results: We found that AsA promotes MSC proliferation, and is particularly useful when expanding MSCs isolated from bone marrow. Serial analysis of gene expression and metabolome analysis suggested that, due to HIF1 alpha accumulation caused by decreased activity of the enzymes that use AsA as a coenzyme in cultures without AsA, genes downstream of HIF1 alpha are expressed and there is a conversion to a hypoxia-mimetic metabolism. AsA promotes HIF1 alpha breakdown and activates mitochondria, affecting cell proliferation and metabolism. Comprehensive evaluation of the effects of AsA on various metabolic products in MSCs revealed that AsA increases HIF1 alpha hydroxylase activity, suppressing HIF1 alpha transcription and leading to mitochondrial activation. Conclusions: Adding AsA during MSC expansion leads to more efficient preparation of cells. These are expected to be important findings for the future application of MSCs in regenerative medicine.