PAI-1 调节血管平滑肌细胞的细胞骨架和固有硬度。
PAI-1 Regulates the Cytoskeleton and Intrinsic Stiffness of Vascular Smooth Muscle Cells.
机构信息
Departments of Medicine (H.B.K., M.V., F.I.R.-P., M.M.-Q., Y.J., L.A.M.-L., W.P.F.), University of Missouri, Columbia.
Medical Pharmacology and Physiology (A.S., C.A.F., S.-Y.C., M.A.H., L.A.M.-L., W.P.F.), University of Missouri, Columbia.
出版信息
Arterioscler Thromb Vasc Biol. 2024 Oct;44(10):2191-2203. doi: 10.1161/ATVBAHA.124.320938. Epub 2024 Jun 13.
BACKGROUND
Plasma concentration of PAI-1 (plasminogen activator inhibitor-1) correlates with arterial stiffness. Vascular smooth muscle cells (SMCs) express PAI-1, and the intrinsic stiffness of SMCs is a major determinant of total arterial stiffness. We hypothesized that PAI-1 promotes SMC stiffness by regulating the cytoskeleton and that pharmacological inhibition of PAI-1 decreases SMC and aortic stiffness.
METHODS
PAI-039, a specific inhibitor of PAI-1, and small interfering RNA were used to inhibit PAI-1 expression in cultured human SMCs. Effects of PAI-1 inhibition on SMC stiffness, F-actin (filamentous actin) content, and cytoskeleton-modulating enzymes were assessed. WT (wild-type) and PAI-1-deficient murine SMCs were used to determine PAI-039 specificity. RNA sequencing was performed to determine the effects of PAI-039 on SMC gene expression. In vivo effects of PAI-039 were assessed by aortic pulse wave velocity.
RESULTS
PAI-039 significantly reduced intrinsic stiffness of human SMCs, which was accompanied by a significant decrease in cytoplasmic F-actin content. PAI-1 gene knockdown also decreased cytoplasmic F-actin. PAI-1 inhibition significantly increased the activity of cofilin, an F-actin depolymerase, in WT murine SMCs, but not in PAI-1-deficient SMCs. RNA-sequencing analysis suggested that PAI-039 upregulates AMPK (AMP-activated protein kinase) signaling in SMCs, which was confirmed by Western blotting. Inhibition of AMPK prevented activation of cofilin by PAI-039. In mice, PAI-039 significantly decreased aortic stiffness and tunica media F-actin content without altering the elastin or collagen content.
CONCLUSIONS
PAI-039 decreases intrinsic SMC stiffness and cytoplasmic stress fiber content. These effects are mediated by AMPK-dependent activation of cofilin. PAI-039 also decreases aortic stiffness in vivo. These findings suggest that PAI-1 is an important regulator of the SMC cytoskeleton and that pharmacological inhibition of PAI-1 has the potential to prevent and treat cardiovascular diseases involving arterial stiffening.
背景
血浆 PAI-1（纤溶酶原激活物抑制剂-1）浓度与动脉僵硬度相关。血管平滑肌细胞（VSMCs）表达 PAI-1，而 VSMCs 的固有僵硬度是总动脉僵硬度的主要决定因素。我们假设 PAI-1 通过调节细胞骨架促进 VSMCs 僵硬度，并且 PAI-1 的药理学抑制可降低 VSMCs 和主动脉僵硬度。
方法
使用 PAI-039（一种 PAI-1 的特异性抑制剂）和小干扰 RNA 抑制培养的人 VSMCs 中的 PAI-1 表达。评估 PAI-1 抑制对 VSMCs 僵硬度、F-肌动蛋白（丝状肌动蛋白）含量和细胞骨架调节酶的影响。使用 WT（野生型）和 PAI-1 缺陷型小鼠 VSMCs 来确定 PAI-039 的特异性。进行 RNA 测序以确定 PAI-039 对 VSMC 基因表达的影响。通过主动脉脉搏波速度评估 PAI-039 的体内作用。
结果
PAI-039 显著降低人 VSMCs 的固有僵硬度，同时细胞质 F-肌动蛋白含量显著降低。PAI-1 基因敲低也降低了细胞质 F-肌动蛋白。PAI-1 抑制显著增加了 WT 小鼠 VSMCs 中的 F-肌动蛋白解聚酶 cofilin 的活性，但在 PAI-1 缺陷型 VSMCs 中没有增加。RNA 测序分析表明，PAI-039 在 VSMCs 中上调 AMPK（AMP 激活的蛋白激酶）信号，Western 印迹也证实了这一点。AMPK 的抑制阻止了 PAI-039 对 cofilin 的激活。在小鼠中，PAI-039 显著降低主动脉僵硬度和中膜 F-肌动蛋白含量，而不改变弹性蛋白或胶原蛋白含量。
结论
PAI-039 降低了 VSMCs 的固有僵硬度和细胞质应激纤维含量。这些作用是通过 AMPK 依赖性 cofilin 的激活介导的。PAI-039 还在体内降低了主动脉僵硬度。这些发现表明 PAI-1 是 VSMCs 细胞骨架的重要调节剂，并且 PAI-1 的药理学抑制有可能预防和治疗涉及动脉僵硬度的心血管疾病。
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