The Matricellular Protein Cyr61 Signaling Axis in Arterial Restenosis

血管平滑肌细胞（SMC）迁移是再狭窄时动脉重塑的重要环节 血管成形术和支架植入术后尽管最近药物洗脱技术取得了进展， 仍然是一个具有挑战性的并发症。为了开发创新的治疗方法，发现新的 需要再狭窄的调节分子和机制。大量证据表明，血小板- 衍生生长因子（PDGF）在血管损伤后SMC迁移到内膜中起着重要作用。 我们最近的出版物揭示，从头基质细胞蛋白Cyr 61（CCN 1）是介导 PDGF通过与整合素α6β1相互作用，通过“由外向内”的信号通路诱导SMC迁移 和αvβ3，其导致细胞内粘着斑激酶（FAK）活化。这表明Cyr 61信号 动脉再狭窄轴。有趣的是，我们最近也观察到小鼠中Cyr 61的高水平表达， 血管成形术导丝诱导的股动脉病变。为了追求的作用和新的信号成分， Cyr 61轴在动脉再狭窄中的作用，我们探索了一系列蛋白激酶激活在极化的主导作用中的作用。 边缘的SMC使用一种新的pseudopodium隔离方法。令人兴奋的是，我们的初步数据显示， 以下发现：PDGF诱导新的伪足富集的非典型激酶1（PEAK 1）的活化， 其定位于SMC的极化前缘;敲低Cyr 61阻断PDGF诱导的PEAK 1 PEAK 1的缺失阻断了PDGF诱导的SMC迁移; PEAK 1（p-PEAK 1）在SMC中与p-FAK相互作用。重要的是，我们观察到Cyr 61水平和PEAK 1 激活在导丝诱导的小鼠股动脉损伤中被稳健地诱导。此外，Cyr 61 p-PEAK 1定位于病变SMC而不定位于图尼卡中膜SMC。这些数据有力地表明Cyr 61 而新的伪足激酶PEAK 1在损伤诱导的动脉再狭窄中起关键作用。基于 根据这些新的观察结果，我们推测Cyr 61-α6β1/αvβ3-p-PEAK 1-p-FAK介导了损伤诱导的 SMC迁移和Cyr 61和p-PEAK 1控制动脉再狭窄中的血管重塑。我们的假设 将在以下具体目标中进行测试。目的1：确定PDGF中p-PEAK 1如何与p-FAK相互作用 途径，确定作为治疗靶点的抑制肽，并检查Cyr 61对 使用来自我们最近创建的SMC特异性的新Cyr 61无效SMC激活细胞内级联反应， Cyr 61 KO小鼠。目的2：探讨PDGF诱导的人肝癌细胞凋亡中α6β1和αvβ3之间可能形成的复合物 Cyr 61通路，并使用来源于 来自创新的Cyr 61 dm/dm、Cyr 61 D125 A和SMC特异性Cyr 61-/-小鼠。目的3：确定Cyr 61的作用 和新的酪氨酸激酶PEAK 1在血管成形术诱导的血管重塑中使用三种创新的Cyr 61 遗传模型Cyr 61 dm/dm、Cyr 61 D125 A和SMC Cyr 61 KO小鼠。拟议的研究预计将确定 预防和治疗动脉再狭窄的新靶点。

Vascular smooth muscle cell (SMC) migration is a critical event in the arterial remodeling in restenosis following angioplasty and stenting. Despite recent advances in drug-eluting technology, arterial restenosis remains a challenging complication. In order to develop innovative therapeutic approaches, discovery of novel regulatory molecules and mechanisms of restenosis is needed. Substantial evidence has shown that platelet- derived growth factor (PDGF) plays a prominent role in SMC migration into the intima following vascular injury. Our recent publication revealed that de novo matricellular protein Cyr61 (CCN1) is the key molecule mediating the PDGF-induced SMC migration via an “outside-in” signaling route through its interaction with integrins α6β1 and αvβ3, which leads to intracellular focal adhesion kinase (FAK) activation. This suggests a Cyr61 signaling axis in arterial restenosis. Interestingly, we also recently observed high levels of Cyr61 expression in mouse angioplasty guidewire-induced femoral arterial lesions. To pursue the role and novel signaling components of the Cyr61 axis in arterial restenosis, we explored an array of protein kinase activation in the polarized leading edge of SMCs using a novel pseudopodium isolation approach. Excitingly, our preliminary data revealed the following findings: PDGF induces activation of a novel pseudopodium-enriched atypical kinase 1 (PEAK1), which is localized in the polarized leading edge of SMCs; knockdown of Cyr61 blocks PDGF-induced PEAK1 activation; depletion of PEAK1 blocks PDGF-induced SMC migration; and PDGF-induced phosphorylated PEAK1 (p-PEAK1) interacts with p-FAK in SMCs. Importantly, we observed that Cyr61 levels and PEAK1 activation are robustly induced in guidewire-induced mouse femoral arterial lesions. Furthermore, both Cyr61 and p-PEAK1 localize in lesion SMCs but not in tunica media SMCs. These data strongly suggest that Cyr61 and the novel pseudopodium kinase PEAK1 play crucial roles in injury-induced arterial restenosis. Based on these new observations, we hypothesize that the Cyr61-α6β1/αvβ3-p-PEAK1-p-FAK mediates injury-induced SMC migration and that Cyr61 and p-PEAK1 control vascular remodeling in arterial restenosis. Our hypothesis will be tested in the following specific aims. Aim 1: Determine how p-PEAK1 interacts with p-FAK in the PDGF pathway, identify the inhibitory peptides serving as therapeutic targets, and examine the key role of Cyr61 on the activation of intracellular cascades using novel Cyr61 null SMCs from our recently created SMC-specific Cyr61 KO mice. Aim 2: Explore the possible complex formation between α6β1 and αvβ3 in PDGF-induced Cyr61 pathway and determine the consequent activation of the intracellular molecules using SMCs derived from innovative Cyr61dm/dm, Cyr61D125A, and SMC-specific Cyr61-/- mice. Aim 3: Determine the role of Cyr61 and novel tyrosine kinase PEAK1 in angioplasty-induced vascular remodeling using three innovative Cyr61 genetic models Cyr61dm/dm, Cyr61D125A, and SMC Cyr61 KO mice. The proposed studies are expected to identify novel targets for prevention and treatment of arterial restenosis.