Acute lung injury (ALI) is a devastating syndrome characterized by diffuse alveolar damage, elevated airspace levels of pro-inflammatory cytokines, and flooding of the alveolar spaces with protein-rich edema fluid. Interleukin-1 (IL-1) is one of the most biologically active cytokines in the distal airspaces of patients with ALI. IL-1 has been shown to increase lung epithelial and endothelial permeability. In this study, we hypothesized that IL-1 would decrease vectorial ion and water transport across the distal lung epithelium. Therefore, we measured the effects of IL-1 on transepithelial current, resistance, and sodium transport in primary cultures of alveolar epithelial type II (ATII) cells. IL-1 significantly reduced the amiloride-sensitive fraction of the transepithelial current and sodium transport across rat ATII cell monolayers. Moreover, IL-1 decreased basal and dexamethasone-induced epithelial sodium channel ␣-subunit (␣ENaC) mRNA levels and total and cellsurface protein expression. The inhibitory effect of IL-1 on ␣ENaC expression was mediated by the activation of p38 MAPK in both rat and human ATII cells and was independent of the activation of ␣v6 integrin and transforming growth factor-. These results indicate that IL-1 may contribute to alveolar edema in ALI by reducing distal lung epithelial sodium absorption. This reduction in ion and water transport across the lung epithelium is in large part due to a decrease in ␣ENaC expression through p38 MAPK-dependent inhibition of ␣ENaC promoter activity and to an alteration in ENaC trafficking to the apical membrane of ATII cells. Acute lung injury (ALI)1 is a devastating clinical syndrome manifested by diffuse alveolar damage, capillary injury, and disruption of the alveolar epithelium. The acute phase of ALI is characterized by the influx of protein-rich edema fluid that impairs gas exchange, causing arterial hypoxemia and respiratory failure with an overall mortality rate of 30 -40% (1). Along with an increase in lung endothelial and epithelial permeability to protein, this syndrome is associated with abnormal surfactant production and decreased vectorial fluid transport across the lung epithelial barrier (2, 3). A number of inflammatory mediators have been found to be elevated in the alveolar space during the early phase of ALI, including interleukin (IL)-1, tumor necrosis factor-␣, IL-6, and IL-8 (4). IL-1 is one of the most biologically active cytokines in pulmonary edema and bronchoalveolar lavage fluids of patients with ALI (4 -6). Indeed, IL-1 increases microvascular lung epithelial permeability in in vitro and in vivo models of ALI (7). IL-1 also enhances alveolar epithelial repair by increasing cell spreading (8) and fibroblast activation and proliferation (5). In contrast, the role of IL-1 in distal lung epithelial ion transport remains unclear. In other epithelia such as the colonic epithelium, IL-1 inhibits aldosterone-induced electrogenic sodium absorption and attenuates aldosterone-induced up-regulation of -and ␥-subunit ...
Previous studies have shown that heat shock protein 72 (Hsp72) is found in the extracellular space (eHsp72) and that eHsp72 has potent immunomodulatory effects. However, whether eHsp72 is present in the distal air spaces and whether eHsp72 could modulate removal of alveolar edema is unknown. The first objective was to determine whether Hsp72 is released within air spaces and whether Hsp72 levels in pulmonary edema fluid would correlate with the capacity of the alveolar epithelium to remove alveolar edema fluid in patients with ALI/ARDS. Patients with hydrostatic edema served as controls. The second objective was to determine whether activation of the stress protein response (SPR) caused the release of Hsp72 into the extracellular space in vivo and in vitro and to determine whether SPR activation and/or eHsp72 itself would prevent the IL-1β-mediated inhibition of the vectorial fluid transport across alveolar type II cells. We found that eHsp72 was present in plasma and pulmonary edema fluid of ALI patients and that eHsp72 was significantly higher in pulmonary edema fluid from patients with preserved alveolar epithelial fluid clearance. Furthermore, SPR activation in vivo in mice and in vitro in lung endothelial, epithelial, and macrophage cells caused intracellular expression and extracellular release of Hsp72. Finally, SPR activation, but not eHsp72 itself, prevented the decrease in alveolar epithelial ion transport induced by exposure to IL-1β. Thus SPR may protect the alveolar epithelium against oxidative stress associated with experimental ALI, and eHsp72 may serve as a marker of SPR activation in the distal air spaces of patients with ALI.Keywords pulmonary edema; alveolar fluid clearance; heat shock response; respiratory distress syndrome; heat shock protein 70 ACUTE LUNG INJURY (ALI) is a common cause of acute respiratory failure in critically ill patients. The early phase of ALI is characterized by the accumulation and activation of inflammatory cells (neutrophils and macrophages) within the distal air spaces that release high levels of oxidant species (46). Alveolar epithelial and lung endothelial injury leads to increased permeability, Copyright © 2006 the American Physiological SocietyAddress for reprint requests and other correspondence: M. T. Ganter, Dept. of Anesthesia and Perioperative Care, San Francisco General Hospital, 1001 Potrero Ave., Rm. San Francisco, CA 94110 (mt.ganter@gmail.com).. NIH Public AccessAuthor Manuscript Am J Physiol Lung Cell Mol Physiol. Author manuscript; available in PMC 2009 October 21. NIH-PA Author ManuscriptNIH-PA Author Manuscript NIH-PA Author Manuscript pulmonary edema, and acute respiratory failure. An important mechanism to prevent alveolar flooding is the maintenance or upregulation of lung alveolar fluid clearance (AFC), requiring an intact alveolar epithelium (47). However, AFC has been shown to be impaired in the majority of ALI patients, and an impaired AFC was associated with worse clinical outcomes (47).Heat shock or stress proteins (Hsp) are a ...
Activation of the stress protein response (SPR) inhibits iNOS-dependent release of NO from alveolar macrophages by blocking the activation of the STAT1 pathway in response to IFN-γ, a major cytokine present in the airspace of patients with acute lung injury. Inhibition of STAT1 phosphorylation after heat stress was associated with detergent insolubilization of the STAT1 protein and its proteasomal degradation which was reversed with the pretreatment of cells with glycerol, a chemical chaperone that reduces the extent of heat-induced protein denaturation. This early effect of SPR is the result of the disruption of the Hsp90 binding to the STAT1 protein. Our results also demonstrated that a late effect of SPR activation involves the regulation of iNOS function by inducible Hsp70 (Hsp70i). The STAT1 signaling pathway recovered function within 12 hours post-SPR activation and synthesis of the iNOS protein, however, NO production did not occur until 72 hours later. Inhibiting Hsp70i expression after heat stress recovered iNOS function whereas overexpressing Hsp70i in the absence of heat stress inhibited iNOS function. In summary, heat stress-induced transient inhibition of STAT1 following its dissociation from Hsp90, and the later transient inhibition of iNOS activity by inducible Hsp70, represent novel mechanisms by which the activation of the stress protein response inhibits the IFN-γ signaling pathway in alveolar macrophages. Since Hsp90 inhibitors have been shown to be safe in humans, these results also highlight a potential clinical application for this class of drugs in modulating NO signaling during the early phase of acute lung injury.
Alveolar fluid clearance is impaired by iNOS/NO‐dependent mechanisms in ALI/ARDS. We investigated whether activation of the stress protein response (SPR) by heat, or using an Hsp90 inhibitor inhibits the STAT1 signaling pathway, which is activated in ALI/ARDS, and thus inhibits iNOS/NO production. Inhibition of STAT1 activation after heat stress results in detergent insolubilization of the STAT1 protein and its proteasomal degradation which is reversed with the pretreatment of cells with glycerol, a chemical chaperone that reduces the extent of heat‐induced protein denaturation. This early effect of SPR is the result of the disruption of the Hsp90 binding to the STAT1 protein and confirmed using an Hsp90 inhibitor. Recovery of STAT1 activation and iNOS synthesis occurs by 12 hours post‐SPR induction, however, NO production did not occur until 48 hours later. These results demonstrate a late effect of SPR activation involving the regulation of iNOS function by inducible Hsp70 (Hsp70i). Inhibiting Hsp70i expression after heat stress recovers iNOS function whereas overexpressing Hsp70i in the absence of heat stress inhibits iNOS function. These results represent novel mechanisms by which the activation of SPR inhibits STAT1 signaling in alveolar macrophages and highlight a potential clinical application for Hsp90 inhibitors in modulating NO signaling during the early phase of acute lung injury. [Research support: UCSF academic senate grant (MH); NIH GM62188 (JFP)]
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