Stem Cell and Regenerative Medicine

Abstract

High Glucose in Culture Media of Adipose Derived Mesenchymal Stem Cells – Gene Expression Alteration and Early Senescence

Barzelay Aya, Levy Ran, Katz Sebastian, Kohn Emmanuelle, Nitzan Anat, Meilik Benjamin, Loewesntein Anat, Krief Bryan and Adiel Barak

Subcutaneous adipose tissue derived mesenchymal stem cells (ASCs) are an attractive cell population for cell therapy applications. Cell culture conditions vary among reported protocols and can affect the quality of stem cells. The aim of this study was to determine the effect of glucose concentration on adipose tissue derived mesenchymal stem cells' culture. ASCs were isolated from subcutaneous fat of patients undergoing abdominoplasty. Cells were cultured with glucose concentration of 4.5 grams/liter or 1 gram/liter. The phenotypic characterization of the stem cells was evaluated by immunostaining and FACS analysis. Proliferation was assessed by division rate study.
Senescence ratio was evaluated by beta galactosidase assay at an advanced passage. Gene profiling for senescence was performed by real time PCR and western blot. Multipotency ability was evaluated by differentiation studies. Culturing adipose tissue derived mesenchymal stem cells with high glucose resulted in reduced proliferative capacity (DMEM high glucose 53 ± 7.5 h, DMEM low glucose 32 ± 4 h), and increased senescence ratio (DMEM high glucose 50% ± 3, DMEM low glucose 23.4% ± 9). Gene profiling revealed decreased Ki67 and PCNA expression and western blot analysis revealed decreased NOX-4 expression in stem cells cultured with high glucose. Moreover, ASCs cultured with high glucose exhibited poorer differentiation potential in comparison to cells cultured with low glucose. In conclusion, culturing ASCs with high glucose resulted in decreased division rate and proliferative capacity, early senescence, lower expression of NOX-4, and reduced multipotency.

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