■肺内皮是导致肺移植后原发性移植物功能障碍的肺缺血再灌注损伤的主要靶标。我们假设在离体肺灌注(EVLP)期间通过短暂的热应激治疗受损的大鼠肺,以引起肺热休克反应,可以保护内皮免受严重的再灌注损伤。
■在37°C的恒温(T°)(EVLP37°C组)下,在EVLP平台上再灌注1h热缺血损伤的大鼠肺长达6h,或在41.5°C下从EVLP(EVLPHS组)1到1.5h的瞬时热应力(HS)之后。添加仅暴露于1小时EVLP的一组肺(预热条件)作为对照(基线组)。在第一个协议中,我们在选定的时间点(每个时间n=5/组)测量了肺热袜蛋白表达(HSP70,HSP27和Hsc70).在第二个协议中,我们确定(n=5/组)肺体重增加(水肿),肺顺应性,氧合能力,肺动脉压(PAP)和血管阻力(PVR),肺组织中PECAM-1(CD31)的表达以及Src激酶和VE-cadherin的磷酸化状态,以及细胞因子在灌注液中的释放(TNFα,IL-1β)和内皮生物标志物(sPECAM,vonWillebrand因子-vWF-,sE-选择素和sICAM-1)。组织学和免疫荧光研究评估了CD31肺内皮中血管周围水肿和3-硝基酪氨酸(过氧亚硝酸盐的标志物)的形成。
■HS诱导了HSP70和HSP27的早期(3小时)和持续性表达,而不影响Hsc70。EVLP37°C组的肺出现大量水肿,低顺应性和氧合,PAP和PVR升高,TNFα的大量释放,IL-1β,s-PECAM,vWF,E-选择素和s-ICAM,以及显著的Src激酶激活,VE-钙粘蛋白磷酸化,内皮3-NT形成和减少CD31表达。与此形成鲜明对比的是,所有这些改变要么被HS治疗废除,要么被显著减轻.
■在受损大鼠肺的EVLP过程中,短暂的热应激的治疗性应用降低了内皮通透性,减弱肺血管收缩,防止src激酶激活和VE-钙粘蛋白磷酸化,同时减少内皮过氧化亚硝酸盐的产生以及细胞因子和内皮生物标志物的释放。总的来说,这些数据表明,治疗性热应激可能是保护肺内皮免受严重再灌注损伤的有希望的策略。
UNASSIGNED: The pulmonary endothelium is the primary target of lung ischemia-reperfusion injury leading to primary graft dysfunction after lung transplantation. We hypothesized that treating damaged rat lungs by a transient heat stress during ex-vivo lung perfusion (EVLP) to elicit a pulmonary heat shock response could protect the endothelium from severe reperfusion injury.
UNASSIGNED: Rat lungs damaged by 1h warm ischemia were reperfused on an EVLP platform for up to 6h at a constant temperature (T°) of 37°C (EVLP37°C group), or following a transient heat stress (HS) at 41.5°C from 1 to 1.5h of EVLP (EVLPHS group). A group of lungs exposed to 1h EVLP only (pre-heating conditions) was added as control (Baseline group). In a first protocol, we measured lung heat sock protein expression (HSP70, HSP27 and Hsc70) at selected time-points (n=5/group at each time). In a second protocol, we determined (n=5/group) lung weight gain (edema), pulmonary compliance, oxygenation capacity, pulmonary artery pressure (PAP) and vascular resistance (PVR), the expression of PECAM-1 (CD31) and phosphorylation status of Src-kinase and VE-cadherin in lung tissue, as well as the release in perfusate of cytokines (TNFα, IL-1β) and endothelial biomarkers (sPECAM, von Willebrand Factor -vWF-, sE-selectin and sICAM-1). Histological and immunofluorescent studies assessed perivascular edema and formation of 3-nitrotyrosine (a marker of peroxinitrite) in CD31 lung endothelium.
UNASSIGNED: HS induced an early (3h) and persisting expression of HSP70 and HSP27, without influencing Hsc70. Lungs from the EVLP37°C group developed massive edema, low compliance and oxygenation, elevated PAP and PVR, substantial release of TNFα, IL-1β, s-PECAM, vWF, E-selectin and s-ICAM, as well as significant Src-kinase activation, VE-cadherin phosphorylation, endothelial 3-NT formation and reduced CD31 expression. In marked contrast, all these alterations were either abrogated or significantly attenuated by HS treatment.
UNASSIGNED: The therapeutic application of a transient heat stress during EVLP of damaged rat lungs reduces endothelial permeability, attenuates pulmonary vasoconstriction, prevents src-kinase activation and VE-cadherin phosphorylation, while reducing endothelial peroxinitrite generation and the release of cytokines and endothelial biomarkers. Collectively, these data demonstrate that therapeutic heat stress may represent a promising strategy to protect the lung endothelium from severe reperfusion injury.