1st Department of Critical Care and Pulmonary Medicine and "Marianthi Simou" Applied Biomedical Research and Training Center, Medical School, University of Athens, 45-47 Ispilandou str, 10676, Athens, Greece.
Department of Pathology, Medical School, University of Athens, Athens, Greece.
Respir Res. 2017 Dec 13;18(1):209. doi: 10.1186/s12931-017-0694-7.
Inspiratory resistive breathing (IRB), a hallmark of obstructive airway diseases, is associated with large negative intrathoracic pressures, due to strenuous contractions of the inspiratory muscles. IRB is shown to induce lung injury in previously healthy animals. Src is a multifunctional kinase that is activated in the lung by mechanical stress. ERK1/2 kinase is a downstream target of Src. We hypothesized that Src is activated in the lung during IRB, mediates ERK1/2 activation and IRB-induced lung injury.
Anaesthetized, tracheostomized adult rats breathed spontaneously through a 2-way non-rebreathing valve. Resistance was added to the inspiratory port to provide a peak tidal inspiratory pressure of 50% of maximum (inspiratory resistive breathing). Activation of Src and ERK1/2 in the lung was estimated during IRB. Following 6 h of IRB, respiratory system mechanics were measured by the forced oscillation technique and bronchoalveolar lavage (BAL) was performed to measure total and differential cell count and total protein levels. IL-1b and MIP-2a protein levels were measured in lung tissue samples. Wet lung weight to total body weight was measured and Evans blue dye extravasation was estimated to measure lung permeability. Lung injury was evaluated by histology. The Src inhibitor, PP-2 or the inhibitor of ERK1/2 activation, PD98059 was administrated 30 min prior to IRB.
Src kinase was activated 30 min after the initiation of IRB. Src inhibition ameliorated the increase in BAL cellularity after 6 h IRB, but not the increase of IL-1β and MIP-2a in the lung. The increase in BAL total protein and lung injury score were not affected. The increase in tissue elasticity was partly inhibited. Src inhibition blocked ERK1/2 activation at 3 but not at 6 h of IRB. ERK1/2 inhibition ameliorated the increase in BAL cellularity after 6 h of IRB, blocked the increase of IL-1β and returned Evans blue extravasation and wet lung weight to control values. BAL total protein and the increase in elasticity were partially affected. ERK1/2 inhibition did not significantly change total lung injury score compared to 6 h IRB.
Src and ERK1/2 are activated in the lung following IRB and participate in IRB-induced lung injury.
吸气抵抗性呼吸（IRB）是气道阻塞性疾病的一个标志，由于吸气肌的剧烈收缩，导致胸腔内产生很大的负压。IRB 被证明会导致先前健康的动物肺部损伤。Src 是一种多功能激酶，机械应激可使其在肺部激活。ERK1/2 激酶是 Src 的下游靶标。我们假设在 IRB 过程中 Src 在肺部被激活，介导 ERK1/2 的激活和 IRB 诱导的肺损伤。
麻醉并气管切开的成年大鼠通过双向非再呼吸阀自主呼吸。在吸气口增加阻力，以提供 50%最大潮气量吸气峰压（吸气抵抗性呼吸）。在 IRB 过程中估计肺部 Src 和 ERK1/2 的激活情况。IRB 持续 6 小时后，通过强迫振荡技术测量呼吸系统力学，并进行支气管肺泡灌洗（BAL）以测量总细胞计数和分类细胞计数以及总蛋白水平。测量肺组织样本中的 IL-1b 和 MIP-2a 蛋白水平。测量湿肺重与体重的比值，并通过 Evans 蓝染料渗出评估肺通透性。通过组织学评估肺损伤。在 IRB 之前 30 分钟给予 Src 抑制剂 PP-2 或 ERK1/2 激活抑制剂 PD98059。
IRB 开始后 30 分钟，Src 激酶被激活。Src 抑制可改善 6 小时 IRB 后 BAL 细胞计数的增加，但不能改善肺内 IL-1β 和 MIP-2a 的增加。BAL 总蛋白和肺损伤评分的增加不受影响。组织弹性的增加部分受到抑制。Src 抑制在 3 小时而非 6 小时 IRB 时阻断 ERK1/2 的激活。ERK1/2 抑制可改善 6 小时 IRB 后 BAL 细胞计数的增加，阻断 IL-1β 的增加，并使 Evans 蓝染料渗出和湿肺重恢复至对照值。BAL 总蛋白和弹性的增加部分受到影响。与 6 小时 IRB 相比，ERK1/2 抑制对总肺损伤评分无明显影响。
IRB 后肺部 Src 和 ERK1/2 被激活，并参与 IRB 诱导的肺损伤。
