Evaluation of tenogenic differentiation potential of selected subpopulations of human adipose‐derived stem cells

Identification of a suitable cell source and bioactive agents guiding cell differentiation towards tenogenic phenotype represents a prerequisite for advancement of cell-based therapies for tendon repair. Human adipose-derived stem cells (hASCs) are a promising, yet intrinsically heterogenous population with diversified differentiation capacities. In this work, we investigated antigenically-defined subsets of hASCs expressing markers related to tendon phenotype or associated with pluripotency that might be more prone to tenogenic differentiation, when compared to unsorted hASCs. Subpopulations positive for tenomodulin (TNMD+ hASCs) and stage specific early antigen 4 (SSEA-4+ hASCs), as well as unsorted ASCs were cultured up to 21 days in basic medium or media supplemented with TGF-β3 (10 ng/ml), or GDF-5 (50 ng/ml). Cell response was evaluated by analysis of expression of tendon-related markers at gene level and protein level by real time RT-PCR, western blot, and immunocytochemistry. A significant upregulation of scleraxis was observed for both subpopulations and unsorted hASCs in the presence of TGF-β3. More prominent alterations in gene expression profile in response to TGF-β3 were observed for TNMD+ hASCs. Subpopulations evidenced an increased collagen III and TNC deposition in basal medium conditions in comparison with unsorted hASCs. In the particular case of TNMD+ hASCs, GDF-5 seems to influence more the deposition of TNC. Within hASCs populations, discrete subsets could be distinguished offering varied sensitivity to specific biochemical stimulation leading to differential expression of tenogenic components suggesting that cell subsets may have distinctive roles in the complex biological responses leading to tenogenic commitment to be further explored in cell based strategies for tendon tissues.