Abstract：
Tendons are frequently injured and have limited regenerative capacity. This motivates tissue engineering efforts aimed at restoring tendon function through strategies to direct functional tendon formation. Generation of a crosslinked collagen matrix is paramount to forming mechanically functional tendon. However, it is unknown how lysyl oxidase (LOX), the primary mediator of enzymatic collagen crosslinking, is regulated by stem cells. This study investigates how multiple factors previously identified to promote tendon formation and healing (transforming growth factor [TGF]β1 and TGFβ2, mechanical stimuli, and hypoxia-inducible factor [HIF]-1α) regulate LOX production in the murine C3H10T1/2 mesenchymal stem cell (MSC) line. We hypothesized that TGFβ signaling promotes LOX activity in C3H10T1/2 MSCs, which is regulated by both mechanical stimuli and HIF-1α activation. TGFβ1 and TGFβ2 increased LOX levels as a function of concentration and time. Inhibiting the TGFβ type I receptor (TGFβRI) decreased TGFβ2-induced LOX production by C3H10T1/2 MSCs. Low (5 mPa) and high (150 mPa) magnitudes of fluid shear stress were applied to test impacts of mechanical stimuli, but without TGFβ2, loading alone did not alter LOX levels. Low loading (5 mPa) with TGFβ2 increased LOX at 7 days greater than TGFβ2 treatment alone. Neither HIF-1α knockdown (siRNA) nor activation (CoCl2) affected LOX levels. Ultimately, results suggest that TGFβ2 and appropriate loading magnitudes contribute to LOX production by C3H10T1/2 MSCs. Potential application of these findings includes treatment with TGFβ2 and appropriate mechanical stimuli to modulate LOX production by stem cells to ultimately control collagen matrix stiffening and support functional tendon formation.
摘要：
肌腱经常受伤，再生能力有限。这激发了旨在通过指导功能性肌腱形成的策略来恢复肌腱功能的组织工程努力。交联的胶原基质的产生对于形成机械功能性肌腱是最重要的。然而,目前尚不清楚赖氨酰氧化酶（LOX），胶原蛋白酶交联的主要介质，受干细胞调控。这项研究调查了先前确定的促进肌腱形成和愈合的多种因素（转化生长因子（TGF）β1和TGFβ2，机械刺激和缺氧诱导因子（HIF）-1α）如何调节鼠C3H10T1/2间充质干细胞（MSC）系的LOX产生。我们假设TGFβ信号传导促进C3H10T1/2MSCs中的LOX活性，受机械刺激和HIF-1α激活的调节。TGFβ1和TGFβ2作为浓度和时间的函数增加LOX水平。抑制TGFβI型受体(TGFβRI)降低了TGFβ2诱导的C3H10T1/2MSC的LOX产生。低（5mPa）和高（150mPa）的流体剪切应力大小被用于测试机械刺激的影响，但没有TGFβ2，单独加载并不改变LOX水平。TGFβ2的低负荷(5mPa)在7天时比单独的TGFβ2处理增加LOX。HIF-1α敲低(siRNA)和激活(CoCl2)均不影响LOX水平。最终,结果表明，TGFβ2和适当的加载量有助于C3H10T1/2MSCs产生LOX。这些发现的潜在应用包括用TGFβ2和适当的机械刺激进行处理以调节干细胞的LOX产生,从而最终控制胶原基质硬化并支持功能性肌腱形成。
