Understanding the effects of substrate stiffness on rejuvenation of aging MSCs via single cell analysis.

骨髓间充质干细胞(MSCs)因其良好的治疗特性而被广泛应用于临床。然而，MSCs在组织培养皿(TC)上的体外扩增会导致衰老(复制性衰老)，数量和质量下降，机制不明。第一年的报道表明，在水凝胶上培养的早期传代MSCs表现出反向衰老表型。在这份第二年的报告中，为了确认水凝胶的再生能力，对已经经历细胞周期停滞的晚期传代MSCs进行了评估。与年轻MSCs的结果一致，凝胶上也观察到衰老标志物的下调。用CSK张力抑制剂处理TC上的MSCs，下调了这些标志物的表达，强调了CSK张力对细胞衰老的重要性。为了深入了解机械刺激对氧化还原动态平衡的影响，用过氧化氢诱导细胞提前衰老，并评估细胞的应激反应。在TC上，处理细胞增加了ROS、抗氧化基因和衰老标记物。该水凝胶可以减弱过氧化氢诱导的氧化应激和抑制衰老的作用，表明在水凝胶上培养的MSCs具有更强的清除自由基的能力，能够更好地维持氧化还原动态平衡。击倒氧化还原调节剂MASTER可促进细胞衰老，提示机械信号、氧化还原动态平衡和细胞衰老之间存在相互联系。

Mesenchymal stem cells (MSCs) from bone marrow are widely used in clinical applications due to their therapeutic properties. However, in vitro expansion of MSCs on tissue culture dish (TC) induce aging (replicative senescence), which reduce their quantities and qualities with undefined mechanism.The first-year report demonstrated that the early passage MSCs cultured on hydrogels exhibited the reverse senescence phenotypes. In this second-year report, to confirm the rejuvenation capacity of the hydrogels, the late passage MSCs that already undergone cell cycle arrest were evaluated. In agreement with the results obtained from the young MSCs, down-regulation of senescence markers was also observed on the gels. Treating the MSCs on TC with CSK tension inhibitors downregulated these markers, emphasizing the importance of the CSK tension on cellular senescence. To gain insight into the effect of mechanical stimuli on redox homeostasis, premature senescence was induced using hydrogen peroxide and the cellular stress response was evaluated. On TC, the treated cells increased ROS, antioxidant genes and senescence markers. The hydrogels could attenuate the effects of hydrogen peroxide induced oxidative stress and suppressed senescence, indicating that the MSCs cultured on the hydrogels possess higher radical scavenging capacity and could better maintain redox homeostasis. Knocking down the master of redox regulator enhanced the senescence, suggesting the interconnection between mechanical signaling, redox homeostasis and cellular senescence.