In Vitro Induction of Tendon-Specific Markers in Tendon Cells, Adipose- and Bone Marrow-Derived Stem Cells is Dependent on TGFβ3, BMP-12 and Ascorbic Acid Stimulation

Abstract

Mesenchymal Stem Cells (MSCs) and tissue-specific progenitors have been proposed as useful tools for regenerative medicine approaches in bone, cartilage and tendon-related pathologies. The differentiation of cells towards the desired, target tissue-specific lineage has demonstrated advantages in the application of cell therapies and tissue engineering. Unlike osteogenic and chondrogenic differentiation, there is no consensus on the best tenogenic induction protocol. Many growth factors have been proposed for this purpose, including BMP-12, b-FGF, TGF-β3, CTGF, IGF-1 and ascorbic acid (AA). In this study, different combinations of these growth factors have been tested in the context of a two-step differentiation protocol, in order to define their contribution to the induction and maintenance of tendon marker expression in adipose tissue and bone marrow derived MSCs and tendon cells (TCs), respectively. Our results demonstrate that TGF-β3 is the main inducer of scleraxis, an early expressed tendon marker, while at the same time inhibiting tendon markers normally expressed later, such as decorin. In contrast, we find that decorin is induced by BMP-12, b-FGF and AA. Our results provide new insights into the effect of different factors on the tenogenic induction of MSCs and TCs, highlighting the importance of differential timing in TGF-β3 stimulation.

Supporting Cell-Based Tendon Therapy: Effect of PDGF-BB and Ascorbic Acid on Rabbit Achilles Tenocytes In Vitro

“…From biochemical supplementation, stimulation by different growth factors, including transforming growth factor-beta (TGF-β) [8], bone morphogenetic protein-12 (BMP-12) [26], connective tissue growth factor (CTGF) [27], and platelet-derived growth factor-BB (PDGF-BB) [28] has been explored. Small bioactive molecules like ascorbic acid (AA), i.e., Vitamin C [29][30][31] or glutamine [12] have also been investigated. Interestingly, bioactive molecules are often investigated concerning their effect on stem cell tenogenic differentiation under serum+ conditions [18,[32][33][34] and serum-free conditions [35,36], while effects on tenocytes have not been investigated in-depth so far, and only few reports for human or animal tenocytes exist [37][38][39].…”

“…On the other hand, AA is known to promote collagen biosynthesis [44,45], and it has also been suggested as a molecule that could help tendons to heal [46]. While its effects on matrix synthesis are known to some extent [30], what changes its supplementation might bring to the phenotype of tenocytes is less known.…”

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

“…The incremental response of scleraxis expression to TGF-β has been suggested to be mediated by the canonical Smad signaling pathway (Czubryt, 2012). Moreover, TGF-β3 as inducer of scleraxis (Perucca Orfei et al, 2019), acts as a potential biomolecular trigger of tenogenesis and can inhibit later markers (Perucca Orfei et al, 2019).…”

“…Disruption of TGF-β signalling in TGF-β2 −/− and TGF-β3 −/− double mutant embryos resulted in the loss of tendons and ligaments in limbs, trunk, tail and head (Pryce et al, 2009). Furthermore, TGF-β3 is the main inducer of scleraxis, an early expressed tendon marker (Perucca Orfei et al, 2019), acting as a potential trigger of tenogenesis. TGF-β3 has also been described to decrease tendon adhesion and promote tendon healing (Jiang et al, 2016), highlighting TFG-β3 as a molecule of interest in tendon biology and regenerative mechanisms.…”

Engineering the Tenogenic Niche In Vitro with Microenvironmental Tools

“…Low glucose dose has also been shown to promote tenogenic phenotype, as evidenced by upregulated tenogenic marker expression and enhanced collagen expression in tenocyte culture . However, these supplements are not sufficiently tenogenic when used in isolation and are often used in addition to other microenvironmental cues …”