Guo Xuran, Wang Xianliu, Tang Han, Ren Yajuan, Li Donghong, Yi Bingcheng, Zhang Yanzhong
College of Chemistry, Chemical Engineering and Biotechnology, Donghua University, Shanghai 201620, China.
Shanghai Engineering Research Center of Nano-Biomaterials and Regenerative Medicine, Donghua University, Shanghai 201620, China.
ACS Appl Mater Interfaces. 2022 May 11. doi: 10.1021/acsami.2c04294.
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.
先前已证明，超细的电纺排列纤维在驱动干细胞分化为肌腱样细胞方面能力有限。鉴于肌腱的机械活性本质，赋予这种排列的纤维结构机械活性，使其能够自行施加机械刺激，即在没有外部施加任何力的情况下，可能会增强其成腱诱导效率。为了验证这一假设，在本研究中，将具有形状记忆功能的聚（L-丙交酯-ε-己内酯）（PLCL）共聚物静电纺丝成排列纤维形式，然后通过“拉伸-恢复”形状编程程序赋予其形状记忆能力。此后，在没有成腱补充剂的情况下，将人脂肪来源干细胞（ADSCs）在编程的纤维基质上培养7天，并研究受限恢复产生的应力硬化对细胞形态、增殖和成腱分化的影响。结果表明，由于形状记忆效应（SME）产生的机械刺激对细胞活力和增殖没有不利影响，但显著促进了细胞沿纤维排列方向的伸长。此外，研究发现，与对照相比，在具有较高恢复应力的机械活性纤维基质上，肌腱特异性蛋白标记物如肌腱调节蛋白（TNMD）和腱生蛋白-C（TNC）以及硬骨素（Scx）、Ⅱ型胶原蛋白（Col2）、Ⅰ型胶原蛋白（Col1）的基因表达显著上调。从机制上讲，Rho/ROCK信号通路被确定参与了底物自驱动诱导的腱分化增强。总之，这些结果表明，受限形状恢复应力可作为一种创新的加载方式，通过为纤维基质提供排列的肌腱样拓扑线索和额外的机械活性来调节干细胞腱分化。这种新证明的调节干细胞腱分化的模式可能会提高肌腱组织工程策略对肌腱愈合和再生的疗效。
