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Introduction Mesenchymal stem/ stromal cells (MSCs) represent one of the most extensively investigated adult stem cell populations due to their pro-regenerative activity observed following their application into injured tissues. Diabetes mellitus type 2 (T2D) is a serious civilization disease occurring frequently in western societies. The prolonged high glucose level in blood may cause damage to many organs, including bone and joint disorders e.g., osteoarthritis. It has been shown that the physiological and clinical status of human cell donors, as well as the type of source material for MSCs isolation, could affect biological properties of these cells. Thus, this study aimed to examine the biological properties of adipose tissue-derived MSCs (AT-MSCs) harvested from healthy and T2D donors.
Methodology Human AT-MSCs obtained from healthy donors and patients suffering from T2D were purchased from Lonza and underwent comparable biological features characterisation including assessment of: morphology, viability in various conditions, and antigenic profile, proliferation rate as well as trilineage differentiation potential in vitro. In our study, three types of expansion cell culture media (CM) were used: i) control – standard CM, ii) standard CM containing high glucose concentration (to mimic diabetic conditions), or iii) standard CM containing high glucose concentration and addition of insulin (to mimic insulin-treated diabetic conditions). Appropriate media (StemPro Differentiation Kits) were used to stimulate cell differentiation into adipocytes, chondrocytes, and osteocytes and were supplemented with high glucose concentration or high glucose concentration and addition of insulin accordingly to the initial expansion conditions.
Results In our preliminary study, we observed no significant differences in morphology, viability, proliferation rate as well as expression of positive (CD90, CD105) and negative (CD19, CD45) MSCs markers between AT-MSCs derived from healthy and T2D donors in the tested media. In the case of osteogenic differentiation, we also did not observe any significant differences in expression of genes related to this differentiation pathway at different time points (3, 7, 14, or 21 days of differentiation) between the groups, which was confirmed by Anilazine Red histochemical staining. However, AT-MSCs derived from T2D donors exhibited a significant decrease in adipogenic genes expression along with decreased aggregation of lipid droplets observed after histochemical staining, when compared to healthy donors. In case of chondrogenic differentiation, we observed a statistically significant impact of glucose or glucose and insulin supplementation on expression of selected genes activated during chondrogenesis, which indicates important role of culture conditions on healthy and diabetic MSCs.
Conclusions To summarise, AT-MSCs obtained from healthy and T2D donors exhibited similar biological properties including proliferation, viability, morphological and antigenic stability in the tested conditions as well as mesodermal differentiation capacity in vitro, and as such may be considered for future use in autologous applications in human patients. However, the appropriate concentration of glucose or glucose and insulin in the expansion medium may affect the expression of some genes critical for differentiation of AT-MSCs, which requires future optimization, especially for clinical applications.
Acknowledgements: This study was funded by NCBR grant STRATEGMED3 (Strategmed3/ 303570/5/NCBR/2017) to EZS and NCN grant MAESTRO 11 (2019/34/A/NZ3/00134) to EZS.
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