Activation of the stress protein response inhibits the STAT1 signalling pathway and iNOS function in alveolar macrophages: role of Hsp90 and Hsp70

• Published in: Thorax Vol. 65; no. 4; pp. 346 - 353
• Main Authors: Howard, Marybeth, Roux, Jérémie, Lee, Hyon, Miyazawa, Byron, Lee, Jae-Woo, Gartland, Brandi, Howard, Amanda J, Matthay, Michael A, Carles, Michel, Pittet, Jean-François
• Format: Journal Article
• Language: English
• Published: London BMJ Publishing Group Ltd and British Thoracic Society 01.04.2010; BMJ Publishing Group; BMJ Publishing Group LTD
• Subjects: Adenoviruses; Animals; Biological and medical sciences; Cardiology. Vascular system; Cell Line; Chaperones; Gene Expression Regulation - drug effects; heat shock; Hot Temperature; HSP70 Heat-Shock Proteins - physiology; HSP90 Heat-Shock Proteins - physiology; Humans; innate immunity; Interferon-gamma - pharmacology; lung; macrophage biology; macrophages; Macrophages, Alveolar - drug effects; Macrophages, Alveolar - metabolism; Medical sciences; Mice; Nitric oxide; Nitric Oxide - biosynthesis; Nitric Oxide Synthase Type II - genetics; Nitric Oxide Synthase Type II - physiology; Phosphorylation; Pneumology; Protein expression; Proteins; RNA, Messenger - genetics; signal transduction; Signal Transduction - physiology; STAT1 Transcription Factor - genetics; STAT1 Transcription Factor - physiology; STAT1–Hsp90 interaction; Stress, Physiological - drug effects; Stress, Physiological - physiology; United States > US; California; Response; Anesthesia; Activation; Circulatory system; Cardiology; Protein; Stress; Macrophage
• ISSN: 0040-6376; 1468-3296
• DOI: 10.1136/thx.2008.101139

Background and aimAlveolar fluid clearance is impaired by inducible nitric oxide synthase (iNOS)/nitric oxide (NO)-dependent mechanisms in acute lung injury (ALI)/acute respiratory distress syndrome (ARDS). The activation of the stress protein response (SPR) in alveolar macrophages on iNOS-dependent NO production in response to interferon γ (IFNγ), a major cytokine present in the airspace of patients with ALI, was investigated.MethodsThe SPR was activated in murine and primary human alveolar macrophages prior to analysis of signal transducer and activator of transcription factor 1 (STAT1) activation, iNOS mRNA and protein synthesis, and NO production.ResultsSPR activation resulted in inhibition of IFNγ-mediated NO production (p=0.001) with >95% detergent insolubilisation of the STAT1 protein. Its subsequent proteasomal degradation was partially reversed with pretreatment of cells with the chemical chaperone glycerol. This early effect of the SPR was caused by the complete disruption of heat shock protein 90 (Hsp90)–STAT1 binding, as shown by immunoprecipitation. Recovery of STAT1 activation and recovery of iNOS synthesis occurred within 12 h after SPR activation (p=0.02). NO production (as compared with non-SPR controls) did not occur until 48 h later (p=0.02). SPR-induced Hsp70 (Hsp70i) expression caused a late inhibition of NO production (p=0.02). Inhibiting >50% Hsp70i expression recovered NO production to control levels whereas overexpressing Hsp70i in the absence of the SPR inhibited NO production (p=0.02).ConclusionEarly inhibition of STAT1 following its dissociation from Hsp90, and later inhibition of iNOS activity by Hsp70i, represent novel mechanisms by which SPR activation modulates the IFNγ signalling in alveolar macrophages. These results highlight a potential clinical application for Hsp90 inhibitors in modulating NO signalling during the early phase of acute lung injury.