Abstract
Tissue engineering, especially cell sheets-based engineering, offers a promising approach to tendon regeneration; however, obtaining a sufficient source of cells for tissue engineering applications is challenging. Adipose-derived stem cells (ASCs) are essential sources for tissue regeneration and have been shown to have the potential for tenogenic differentiation in vitro via induction by growth differentiation factor 5 (GDF-5). In this study, we explored the feasibility of ASCs cell sheets stimulated by GDF-5 for engineered tendon repair. As shown by quantitative polymerase chain reaction and western blotting, tenogenesis-related markers (Col I&III, TNMD, biglycan, and tenascin C) were significantly increased in GDF-5-induced ASCs cell sheets compared with the uninduced. Moreover, the levels of SMAD2/3 proteins and phospho-SMAD1/5/9 were significantly enhanced, demonstrating that GDF-5 may exert its functions through phosphorylation of SMAD1/5/9. Furthermore, the cell sheets were combined with P(LLA-CL)/Silk fibroin nanoyarn scaffolds to form constructs for tendon tissue engineering. Terminal deoxynucleotidyl transferase dUTP nick end labeling and immunofluorescence assays demonstrated favorable cell viability and tenogenesis-related marker expression in GDF-5-induced constructs. In addition, the constructs showed the potential for tendon repair in rabbit models, as demonstrated by histological, immunohistochemical, and biomechanical analyses. In our study, we successfully produced a new tissue-engineered tendon by the combination of GDF-5-induced ASCs cell sheets and nanoyarn scaffold which is valuable for tendon regeneration.
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