The aberrant differentiation of valvular interstitial cells (VICs) to osteogenic lineages promotes calcified aortic valves disease (CAVD), partly activated by potentially destructive hemodynamic forces. These involve Rho A/ROCK1 signaling, a mechano-sensing pathway. However, how Rho A/ROCK1 signaling transduces mechanical signals into cellular responses and disrupts normal VIC homeostasis remain unclear. We examined Rho A/ROCK1 signaling in human aortic valves, and further detected how Rho A/ROCK1 signaling regulates mineralization in human VICs. Aortic valves (CAVD n = 22, normal control (NC) n = 12) from patients undergoing valve replacement were investigated. Immunostaining and western blotting analysis indicated that Rho A/ROCK1 signaling, as well as key transporters and enzymes involved in the Warburg effect, were markedly upregulated in human calcified aortic valves compared with those in the controls. In vitro, Rho A/ROCK1-induced calcification was confirmed as AMPK-dependent, via a mechanism involving metabolic reprogramming of human VICs to Warburg effect. Y-27632, a selective ROCK1 inhibitor, suppressed the Warburg effect, rescued AMPK activity and subsequently increased RUNX2 ubiquitin-proteasome degradation, leading to decreased RUNX2 protein accumulation in human VICs under pathological osteogenic stimulus. Rho A/ROCK1 signaling, which is elevated in human calcified aortic valves, plays a positive role in valvular calcification, partially through its ability to drive metabolic switching of VICs to the Warburg effect, leading to altered AMPK activity and RUNX2 protein accumulation. Thus, Rho A/ROCK1 signaling could be an important and unrecognized hub of destructive hemodynamics and cellular aerobic glycolysis that is essential to promote the CAVD process.

瓣膜间质细胞 (VIC) 向成骨谱系的异常分化促进钙化主动脉瓣疾病 (CAVD)，部分由潜在的破坏性血流动力学力量激活。这些涉及 Rho A/ROCK1 信号，这是一种机械感应通路。然而，Rho A/ROCK1 信号如何将机械信号转变成细胞反应并破坏正常的 VIC 稳态仍不清楚。我们检查了人主动脉瓣中的 Rho A/ROCK1 信号，并进一步检测了 Rho A/ROCK1 信号如何调节人 VIC 中的矿化。主动脉瓣 (CAVD n  = 22, 正常控制 (NC) n = 12) 来自接受瓣膜置换术的患者进行了调查。免疫染色和蛋白质印迹分析表明，与对照组相比，Rho A/ROCK1 信号以及参与 Warburg 效应的关键转运蛋白和酶在人钙化主动脉瓣中显着上调。在体外，Rho A/ROCK1 诱导的钙化被证实为 AMPK 依赖性，通过涉及人类 VIC 代谢重编程至 Warburg 效应的机制。Y-27632 是一种选择性 ROCK1 抑​​制剂，可抑制 Warburg 效应，挽救 AMPK 活性并随后增加 RUNX2 泛素-蛋白酶体降解，导致病理性成骨刺激下人类 VIC 中 RUNX2 蛋白积累减少。Rho A/ROCK1 信号，在人类钙化主动脉瓣中升高，在瓣膜钙化中发挥积极作用，部分是通过其驱动 VIC 代谢转换为 Warburg 效应的能力，从而导致 AMPK 活性改变和 RUNX2 蛋白积累。因此，Rho A/ROCK1 信号可能是破坏性血液动力学和细胞有氧糖酵解的重要且未被认可的枢纽，这对于促进 CAVD 过程至关重要。