Pulmonary Hypertension Research Group, Quebec Heart and Lung Institute Research Center, Laval University, Quebec City, Canada.
Am J Respir Crit Care Med. 2014 Aug 1;190(3):318-28. doi: 10.1164/rccm.201402-0383OC.
Pulmonary arterial hypertension (PAH) is characterized by significant exercise intolerance, which is multifactorial and involves skeletal muscle alterations. There is growing evidence that microRNAs (miRs) are involved in PAH pathogenesis.
We hypothesized that miR-126, an endothelial-specific, proangiogenic miR, is down-regulated in the peripheral muscles of patients with PAH, which would account for skeletal muscle microcirculation loss and exercise intolerance.
Patients with PAH displayed decreases in exercise capacity ([Formula: see text]o2max) and microcirculation loss on quadriceps muscle biopsy (in CD31(+) immunofluorescence experiments) compared to control subjects. Exercise capacity correlated with muscle capillarity (r = 0.84, P < 0.01). At the cellular level, vascular endothelial growth factor (VEGF) and VEGF receptor 2 expression were similar in both groups. Conversely, PAH was associated with a 60% decrease in miR-126 expression in a quantitative reverse transcriptase polymerase chain reaction experiment (P < 0.01), resulting in up-regulation of its targeted protein, Sprouty-related, EVH1 domain-containing protein 1 (SPRED-1), and a marked decrease in the downstream effectors of the VEGF pathway, p-Raf/Raf and p-ERK/ERK, as determined by immunoblot analysis. Using freshly isolated CD31(+) cells from human quadriceps biopsies, we found that the down-regulation of miR-126 in PAH triggered the activation of SPRED-1, impairing the angiogenic response (Matrigel assay). These abnormalities were reversed by treating the PAH cells with miR-126 mimic, whereas inhibition of miR-126 (antagomir) in healthy CD31(+) cells fully mimicked the PAH phenotype. Finally, miR-126 down-regulation in skeletal muscle of healthy rats decreased muscle capillarity in immunofluorescence assays (P < 0.05) and exercise tolerance in treadmill tests (P < 0.05), whereas miR-126 up-regulation increased them in monocrotaline PAH rats.
We demonstrate for the first time that exercise intolerance in PAH is associated with skeletal muscle microcirculation loss and impaired angiogenesis secondary to miR-126 down-regulation.
肺动脉高压（PAH）的特征是显著的运动不耐受，这是多因素的，并涉及骨骼肌改变。越来越多的证据表明 microRNAs（miRs）参与了 PAH 的发病机制。
我们假设内皮特异性促血管生成 miR-126 在 PAH 患者的外周肌肉中下调，这将导致骨骼肌微循环丧失和运动不耐受。
与对照组相比，PAH 患者的运动能力（[Formula: see text]o2max）和股四头肌活检中的微循环丧失（在 CD31（+）免疫荧光实验中）降低。运动能力与肌肉毛细血管（r = 0.84，P < 0.01）相关。在细胞水平上，两组的血管内皮生长因子（VEGF）和 VEGF 受体 2 表达相似。相反，PAH 与定量逆转录聚合酶链反应实验中 miR-126 表达降低 60%（P < 0.01）相关，导致其靶向蛋白 Sprouty 相关 EVH1 结构域蛋白 1（SPRED-1）上调，以及 VEGF 通路下游效应物 p-Raf/Raf 和 p-ERK/ERK 的显著减少，通过免疫印迹分析确定。使用从人类股四头肌活检中新鲜分离的 CD31（+）细胞，我们发现 miR-126 在 PAH 中的下调触发了 SPRED-1 的激活，从而损害了血管生成反应（Matrigel 测定）。通过用 miR-126 模拟物处理 PAH 细胞，这些异常得到了逆转，而在健康的 CD31（+）细胞中抑制 miR-126（antagomir）则完全模拟了 PAH 表型。最后，健康大鼠骨骼肌中 miR-126 的下调降低了免疫荧光测定中的肌肉毛细血管（P < 0.05）和跑步机测试中的运动耐量（P < 0.05），而 miR-126 的上调增加了这些在野百合碱 PAH 大鼠中的作用。
我们首次证明，PAH 中的运动不耐受与骨骼肌微循环丧失和血管生成受损有关，这是由于 miR-126 的下调。
