Protective Effect of Inhaled Rho-Kinase Inhibitor on Lung Ischemia-Reperfusion Injury

摘要

Background Rho-kinase, an intracellular serine/threonine kinase, is a key regulator of cytoskeletal dynamics. Recent studies have demonstrated that Rho-kinase is involved in the ischemia-reperfusion injury (IRI) pathogenesis of many organs; however, its involvement with lung IRI remains unclear. This study assessed the association of Rho-kinase with lung IRI and evaluated the protective effect of inhaled Rho-kinase inhibitors in lung IRI.;Methods The study included isolated rat lung perfusion models, divided into three groups: sham, Rho-kinase inhibitor, and warm ischemia (n?= 6 each). The lungs were exposed to 60 minutes of warm ischemia by perfusion cessation. At the onset of ischemia, nebulized fasudil, a novel Rho-kinase inhibitor, and saline were inhaled in the Rho-kinase inhibitor and warm ischemia groups, respectively. Perfusion was restarted after the ischemic period, and physiologic data were collected for 90 minutes. Lungs in the sham group were continuously perfused without ischemia or drug administrations. Protein expression in tissue specimens related to the Rho-kinase pathway was evaluated by Western blotting.;Results Warm ischemia and subsequent reperfusion enhanced Rho-kinase activity, and this was suppressed by fasudil inhalation. Fasudil inhalation significantly attenuated IRI pathophysiology, including pulmonary vascular contraction, dynamic compliance, lung edema, and oxygenation. Molecular analysis showed that Rho-kinase suppressed myosin phosphatase and endothelial nitric oxide synthase activities, suggesting these are downstream targets of Rho-kinase during lung IRI pathogenesis.;Conclusions The present study suggests that Rho-kinase activation is involved in lung IRI pathogenesis and that inhaled Rho-kinase inhibitors may attenuate this pathogenesis.;Despite significant advances in organ preservation, ischemia-reperfusion injury (IRI) remains one of the major causes of primary graft dysfunction after lung transplantation [1, 2, 3]. IRI pathogenesis is multifaceted, and the exact molecular mechanisms remain unclear [4, 5]. Previous reports have shown that xanthine oxidase [6][6], nicotinamide adenine dinucleotide phosphate oxidase [7][7], nitric oxide synthases [8][8], nuclear factor-κB [9][9], Toll-like receptor 4 [10][10], and proinflammatory cytokines [11][11] are involved in the development of lung?IRI. Therefore, a thorough understanding of the physiologic, cellular, and molecular changes in lung IRI?is crucial for a good prognosis in lung transplant recipients.Rho-kinase is an intracellular serine/threonine kinase that was identified as one of the effectors of the small guanosine triphosphate-binding protein Rho [12, 13]. It is ubiquitously present in mammalian cells and forms a molecular switch in the cells for regulating cytoskeletal dynamics. Rho-kinase plays a significant role in the development of various vascular diseases, such as coronary hyperconstriction [14][14], vasospasm after subarachnoid hematoma [15][15], systemic hypertension [16][16], and pulmonary hypertension [17][17]. Ischemia and subsequent reperfusion also induce pathophysiologic changes to the vasculature of the lungs, including pulmonary vascular contraction and increased vascular permeability [18][18]. Studies have reported that Rho-kinase is related to IRI pathogenesis of several organs, including the heart [19][19], liver [20][20], and kidney [21][21]; however, the role of Rho-kinase in lung IRI pathogenesis is still not determined.We hypothesized that Rho-kinase plays a key role in the development of lung IRI and that inhibition of Rho-kinase would have a protective effect against IRI. To test this hypothesis, we examined the effect of Rho-kinase and its inhibition on lungs by using an isolated rat lung perfusion model.Jump to SectionMaterial and MethodsAnimalsSet-up of Isolated Lung Perfusion SystemProtocol After Set UpDrug AdministrationMeasurementsPhysiologic DataWet/dry lung weight ratio and histologic evaluationRho-