| 摘要: | 背景 呼吸器引發的肺傷害 (ventilator-induced lung injury: VILI) 涵蓋了肺泡內皮及外皮細胞損傷、通透度增加、肺發炎介質增加、肺水腫形成以及肺部纖維質溶解逆轉並產生纖維質沉積於肺泡區間。肺部內皮組織通透度增加乃是經由一個與Rho訊息相關的傳導途徑 (Rho signal pathway) 來調節，活化Rho signal pathway可以使內皮細胞功能減弱，促進肌動纖維蛋白重組而造成內皮細胞通透度增加。VILI的另一機轉乃是經由血漿纖溶酶原啟動物抑制劑Plasminogen activator inhibitor (PAI-1) 的活化使得肺泡纖維質溶解反應逆轉，PAI-1是種抗纖維溶解的蛋白酶，負責降低纖溶酶plasmin活性和纖溶fibrinolysis潛力。昇高的肺部和全身性 PAI-1含量以及增加的肺泡纖維蛋白沉積曾出現在先前VILI 的動物模式的研究中。抗發炎藥物Ibuprofen曾以抑制Rho activation 的生體外作用機轉被使用在脊柱脊髓損傷及老年癡呆症的細胞模式中。但對於Ibuprofen 作用在VILI 所引發的Rho signal pathway生體內機轉尚未清楚。組織纖溶酶原啟動物Tissue plasminogen activator (tPA) 負責活化 plasminogen以造成fibrin分解，並已用於急性心肌梗塞患者溶栓治療，推論應可利用 tPA 纖溶原理以改善VILI 所造成的肺傷害。
目標 藉由動物模式，研究Ibuprofen 與tPA作用在高潮氣機械呼吸模式引發的肺傷害的機轉。由此發現影響 VILI 肺傷害的藥理途徑，進而產生臨床上急性呼吸窘迫症相關疾病的治療策略。
材料與方法 在探討Ibuprofen機轉研究中，成年雄性大鼠隨機接受兩種不同呼吸策略，其一為沒有介入呼吸器的對照組，另一為接受2小時高潮氣量與零-正呼氣末正壓 （high tidal volume zero positive-end-expiratory pressure：HVZP）呼吸策略且僅注射溶劑的傷害組與傷害前分別注射兩種不同劑量 ibuprofen 的處置組共四組。我們分析支氣管肺泡灌洗液（BALF）與肺組織的水腫情況、發炎反應與Rho signal pathway 的活化情形。Rho kinase活性藉由測定磷酸化的ezrin、radixin and moesin （p-ERM）與總 ERM 的比率。VILI 的特點是增高的肺洩漏蛋白、肺濕乾重量比、支氣管肺泡灌洗液 （BALF）細胞因子 (cytokines) 濃度和Rho guanine nucleotide exchange factor (GEFH1)、RhoA activity、p-ERM/total ERM 和 p-myosin light chain （p-MLC）的表現強度。在探討TPA機轉研究中，老鼠也被分為沒有介入呼吸器的對照組及接受2小時HVZP呼吸策略且僅注射溶劑的傷害組與傷害前先行靜脈注射tPA 的處置組共三組，並分析BALF、血液及肺組織標本，作生化和組織分析 PAI-1表現與組織 fibrin沉積。VILI 的特點是增高的肺洩漏蛋白、BALF cytokine濃度、PAI-1及D-dimer表現和組織 fibrin沉積。
結果 本研究發現Ibuprofen前處理明顯減少 HVZP 呼吸模式引發的動物肺水腫、 肺組織RhoA及Rho kinase pathway表現並呈現少許發炎抑制效果。TPA 前處理可顯著降低 VILI 引發的肺水腫及肺傷害、血漿 D-dimer表現與肺組織fibrin及血栓thrombi 沈積。但並不會影響肺部發炎反應。
結論 Ibuprofen抑制高潮氣量 VILI 引發的肺動脈內皮細胞高通透反應。這種保護效果與Ibuprofen的 RhoA/Rho kinase activity抑制作用相關聯 。而tPA藉由抑制PAI-1的作用，也呈現了減少VILI引發的肺水腫、肺損傷及肺纖維沈積的效果。
Background The spectrum of ventilator-induced lung injury (VILI) includes disruption of endothelial and epithelial cells, increase endothelial and epithelial permeability, increase pulmonary inflammatory mediators, formation of pulmonary edema, pulmonary fibrin turnover and the deposition of fibrin in the alveolar space. Pulmonary edema caused by increase endothelial permeability is regulated through Rho-dependent signaling pathway. Activation of Rho signaling pathway may contribute to impaired endothelial function, promote actin stress fiber formation, and increase endothelial permeability. The major mechanisms that contribute to increased alveolar fibrin turnover are caused by increased in plasminogen activator inhibitor (PAIs). PAI-1, a fibrinolytic antiprotease, is responsible for decreased plasmin activity and fibrinolytic potential. High pulmonary and systemic PAI-1 levels and increased alveolar fibrin deposition is a feature of animal model of VILI. Ibuprofen inhibits Rho activation in cell models of spinal-cord injury and Alzheimer’s disease. But the in vivo effects of ibuprofen on the Rho signaling pathway in VILI are not well understood. Tissue plasminogen activator (tPA) is responsible for fibrin degradation via the activation of plasminogen and has been used for thrombolysis in patients with acute myocardial infarction. Theoretical considerations suggest that fibrinolytic therapy with tPA might benefit animals with VILI.
Objectives First, we investigated ibuprofen effects on Rho signaling pathway and pulmonary edema in high tidal volume association VILI rats. Second, we tested the hypothesis that the tPA may counteract the inhibitory effect of PAI and attenuate lung injury in a rat model of VILI.
Materials and methods In the investigation of ibuprofen, adult male Sprague-Dawley rats were randomized to receive a ventilation strategy with three different interventions for 2 h: (1) a high-volume zero-positive-end-expiratory pressure (PEEP) (HVZP) group; (2) an HVZP + ibuprofen 15 mg/kg group; and (3) an HVZP + ibuprofen 30 mg/kg group. A fourth group without ventilation served as the control group. Rho-kinase activity was determined by ratio of phosphorylated ezrin, radixin, and moesin (p-ERM), substrates of Rho-kinase, to total ERM. VILI was characterized by increased pulmonary protein leak, wet-to-dry weight ratio, cytokines level, and Rho guanine nucleotide exchange factor (GEF-H1), RhoA activity, p-ERM/total ERM, and p-myosin light chain (MLC) protein expression. In the investigation of tPA, the rats were randomized to receive HVZP with two different interventions for 2 h: (1) HVZP group receiving HVZP + vehicle injection; (2) HVZP + tPA group receiving tPA (1.25 mg/kg, iv) 15 min before ventilation. A third group without ventilation served as the control group. The BALF, blood and lungs were collected for biochemical and histological analyses for PAI-1 and fibrin deposition. VILI was characterized by increased pulmonary protein leak, enhanced serum PAI-1 level, lung tissue PAI-1 expression and fibrin deposition.
Results Ibuprofen pretreatment significantly reduced the HVZP-induced increase in pulmonary edema, pulmonary endothelial permeability and RhoA/Rho kinase activity. Ibuprofen pretreatment partially reduced HVZP-induced inflammatory reaction.. TPA pretreatment significantly decreased the HVZP-induced increased plasma D-dimer levels, and lung vascular fibrin thrombi expression. TPA treatment showed no effect on HVZP-induced lung inflammation..
Conclusion Ibuprofen attenuated high tidal volume VILI-induced pulmonary endothelial hyperpermeability. This protective effect was associated with a reduced Rho-kinase activity. TPA attenuated VILI by decreasing capillary-alveolar protein leakage as well as local and systemic coagulation as shown by decreased lung vascular fibrin and PAI-1 deposition, and plasma D-dimers. |
