Role of inflammatory response in aortic valve lesions

摘要 外科瓣膜置换术是钙化性主动脉瓣狭窄的唯一有效治疗方法。 尽管慢性炎症被认为在疾病的发生和发展中起主要作用 启动和维持炎症过程的信号机制 在瓣膜细胞中的作用仍不清楚。对炎性和成骨反应的了解 瓣膜细胞对于预防主动脉瓣病变及其进展具有重要意义。我们展示了 内毒素刺激Toll样受体4上调骨组织的表达 人主动脉瓣肌成纤维细胞中BMP-2的表达及其意义 在TLR4诱导的成骨反应中起主要作用，以Runx2表达为特征， 碱性磷酸酶(ALP)和骨桥蛋白。此外，我们发现TLR4的慢性刺激 促进老年小鼠主动脉瓣钙化和增厚。初步研究表明，TLR4 信号转导上调HAVMF中Notch1胞内结构域(NICD1)的水平。中和 Inf？是一种受TRIF调节的1型干扰素，它消除了细胞内NICD1的增加，并减少了 刺激TLR4后BMP-2水平的变化。此外，抑制Notch1也会减少BMP-2 TLR4刺激后表达。综上所述，这些新的发现表明，INF？激活 NOTCH1和NICD1增强TLR4诱导的BMP-2表达。有趣的是，高水平的 在TLR4刺激后24 h，人AVMF中同时存在lNICD 1和BMP-2。很可能是 NICD1不仅在早期阶段调节BMP-2的表达，而且在 调制BMP-2信号。事实上，初步研究表明，Notch1的激活增强了 BMP-2诱导Runx2的表达。因此，TLR4信号可诱导大鼠成骨反应 其机制涉及BMP-2和Notch1信号通路之间的相互作用。 我们制定了三个相互关联的指标来确定TLR4诱导的瓣膜机制 成骨反应：1)验证TLR4信号诱导主动脉瓣成骨的假说 通过BMP-2介导的Runx2上调的反应，2)检验NICD1的假设 通过调节核因子B的活性来增强BMP-2的表达，以及3)检验以下假设 NICD1通过与Smad1/5/8的相互作用增强BMP-2信号。这些研究将提供 对促炎信号介导的分子机制的重要见解 AVMF成骨反应和主动脉瓣病变。

Summary Surgical valve replacement is the only available therapy for calcific aortic valve stenosis. Although chronic Inflammation is believed to play a major role in the pathogenesis and progression of aortic valve lesions, the signaling mechanisms that initiate and sustain the inflammatory process in valvular cells remain unclear. Understanding of the inflammatory and osteogenic responses in valvular cells is important for prevention aortic valve lesions and their progression. We demonstrate that stimulating Toll-like receptor 4 (TLR4) with endotoxin up-regulates the expression of bone morphogenetic protein 2 (BMP-2) in human aortic valve myofibroblasts (HAVMFs) and that BMP-2 plays a major role in TLR4-induced osteogenic responses, characterized by expression of Runx2, alkaline phosphatase (ALP) and osteopontin. Further, we found that chronic stimulation of TLR4 promotes aortic valve calcification and thickening in old mice. Preliminary studies show that TLR4 signaling up-regulates the levels of Notch1 intracellular domain (NICD1) in HAVMFs. Neutralizing INF¿, a TRIF-regulated type 1 interferon, abrogates the increase in cellular NICD1 and reduces BMP-2 levels following stimulation of TLR4. Further, inhibition of Notch1 also reduces BMP-2 expression following TLR4 stimulation. Together, these novel findings indicate that INF¿ activates Notch1 and that NICD1 enhances TLR4-induced BMP-2 expression. Interestingly, elevated levels of lNICD1 and BMP-2 coexist in human AVMFs at 24 h following TLR4 stimulation. It is likely that NICD1 plays a role not only in regulating BMP-2 expression in the early phase but also in modulating BMP-2 signaling. Indeed, preliminary studies show that activation of Notch1 enhances BMP-2-induced expression of Runx2. Thus, TLR4 signaling induces the osteogenic responses in human AVMFs, and mechanisms involve the interplay between the BMP-2 and Notch1 pathways. We formulated three interrelated aims to determine the mechanisms of TLR4-induced valvular osteogenic responses: 1) to test the hypothesis that TLR4 signaling induces aortic valve osteogenic responses through BMP-2-mediated up-regulation of Runx2, 2) to test the hypothesis that NICD1 enhances BMP-2 expression through modulation of NF-¿B activity and 3) to test the hypothesis that NICD1 enhances BMP-2 signaling via interaction with Smad1/5/8. These studies will provide important insights into the molecular mechanisms by which pro-inflammatory signaling mediates AVMF osteogenic responses and aortic valve lesions.