Engineering a Mechanoactive Fibrous Substrate with Enhanced Efficiency in Regulating Stem Cell Tenodifferentiation.

Electrospun-aligned fibers in ultrathin fineness have previously demonstrated a limited capacity in driving stem cells to differentiate into tendon-like cells. In view of the tendon's mechanoactive nature, endowing such aligned fibrous structure with mechanoactivity to exert mechanical stimulus by itself, namely, without any forces externally applied, is likely to potentiate its efficiency of tenogenic induction. To test this hypothesis, in this study, a shape-memory-capable poly(l-lactide--caprolactone) (PLCL) copolymer was electrospun into aligned fibrous form followed by a "stretching-recovery" shape-programming procedure to impart shape memory capability. Thereafter, in the absence of tenogenic supplements, human adipose-derived stem cells (ADSCs) were cultured on the programmed fibrous substrates for a duration of 7 days, and the effects of constrained recovery resultant stress-stiffening on cell morphology, proliferation, and tenogenic differentiation were examined. The results indicate that the enacted mechanical stimulus due to shape memory effect (SME) did not have adverse influence on cell viability and proliferation, but significantly promoted cellular elongation along the direction of fiber alignment. Moreover, it revealed that tendon-specific protein markers such as tenomodulin (TNMD) and tenascin-C (TNC) and gene expression of scleraxis (), , , and collagen I () were significantly upregulated on the mechanoactive fibrous substrate with higher recovery stress compared to the counterparts. Mechanistically, the Rho/ROCK signaling pathway was identified to be involved in the substrate self-actuation-induced enhancement in tenodifferentiation. Together, these results suggest that constrained shape recovery stress may be employed as an innovative loading modality to regulate the stem cell tenodifferentiation by presenting the fibrous substrate with an aligned tendon-like topographical cue and an additional mechanoactivity. This newly demonstrated paradigm in modulating stem cell tenodifferentiation may improve the efficacy of tendon tissue engineering strategy for tendon healing and regeneration.