Mechanobiological Strategies to Enhance Ovine (Ovis aries) Adipose-Derived Stem Cells Tendon Plasticity for Regenerative Medicine and Tissue Engineering Applications
Author:
Haidar-Montes Arlette A.1ORCID, Mauro Annunziata1ORCID, El Khatib Mohammad1ORCID, Prencipe Giuseppe1ORCID, Pierdomenico Laura2, Tosi Umberto1ORCID, Wouters Guy3, Cerveró-Varona Adrián1ORCID, Berardinelli Paolo1, Russo Valentina1ORCID, Barboni Barbara1ORCID
Affiliation:
1. Unit of Basic and Applied Biosciences, Faculty of Bioscience and Agro-Food and Environmental Technology, University of Teramo, 64100 Teramo, Italy 2. Center for Advanced Studies and Technology (CAST), University “G. d’Annunzio” of Chieti-Pescara, 66100 Chieti, Italy 3. FAT STEM Company, Erembodegem, 9300 Aalst, Belgium
Abstract
Adipose-derived stem cells (ADSCs) hold promise for tendon repair, even if their tenogenic plasticity and underlying mechanisms remain only partially understood, particularly in cells derived from the ovine animal model. This study aimed to characterize oADSCs during in vitro expansion to validate their phenotypic properties pre-transplantation. Moreover, their tenogenic potential was assessed using two in vitro-validated approaches: (1) teno-inductive conditioned media (CM) derived from a co-culture between ovine amniotic stem cells and fetal tendon explants, and (2) short- (48 h) and long-term (14 days) seeding on highly aligned PLGA (ha-PLGA) electrospun scaffold. Our findings indicate that oADSCs can be expanded without senescence and can maintain the expression of stemness (Sox2, Oct4, Nanog) and mesenchymal (CD29, CD166, CD44, CD90) markers while remaining negative for hematopoietic (CD31, CD45) and MHC-II antigens. Of note, oADSCs’ tendon differentiation potential greatly depended on the in vitro strategy. oADSCs exposed to CM significantly upregulated tendon-related genes (COL1, TNMD, THBS4) but failed to accumulate TNMD protein at 14 days of culture. Conversely, oADSCs seeded on ha-PLGA fleeces quickly upregulated the tendon-related genes (48 h) and in 14 days accumulated high levels of the TNMD protein into the cytoplasm of ADSCs, displaying a tenocyte-like morphology. This mechano-sensing cellular response involved a complete SOX9 downregulation accompanied by YAP activation, highlighting the efficacy of biophysical stimuli in promoting tenogenic differentiation. These findings underscore oADSCs’ long-term self-renewal and tendon differentiative potential, thus opening their use in a preclinical setting to develop innovative stem cell-based and tissue engineering protocols for tendon regeneration, applied to the veterinary field.
Funder
European Union’s Horizon 2020 research and innovation program Italian Ministry of University and Research (MUR) National Innovation Ecosystem
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