Inhibition of HSP90 S‐nitrosylation Alleviates Cardiac Fibrosis via TGFβ/SMAD3 Signaling Pathway

Background and Purpose: Effective anti-fibrotic therapeutic solutions are unavailable so far. The heat shock protein 90 (HSP90) exerts deleterious effects in some fibrotic diseases. S-nitrosylation (SNO) of HSP90 affects its own function, however, little is known about its role in pathological stress. Here, we investigated the effect of SNO-HSP90 on cardiac fibrosis. Experimental Approach: SNO-HSP90 level was measured by biotin-switch. SNO sites were identified through mass spectrometry. S-nitrosylation site-mutated plasmids or adeno-associated virus, gene deletion and pharmacological antagonists were used to identify the contribution of SNO-HSP90 in myocardial fibrosis. Key Results: SNO-HSP90 level was positively correlated with fibrosis marker expression in hearts from patients and significantly higher in fibrotic hearts from spontaneously hypertensive rats and mice subjected to transverse aortic constriction, as well as in angiotensin II- or isoproterenol-treated neonatal rat cardiac fibroblasts. S-nitrosylated site of HSP90 at cysteine 589 was identified. Inhibition of SNO-HSP90 by Cys589 mutation reduced fibrosis in angiotensin II- or isoproterenol- treated cardiac fibroblasts. Administration of recombinant adeno-associated virus of Cys589 mutation improved heart function and alleviated fibrosis in transverse aortic constriction mice. Mechanismly, SNO-HSP90 stimulated transforming growth factor- β type II receptor (TGFβ RII) binding to HSP90 in response to fibrotic stimuli, subsequently increased phosphorylation and nuclear translocation of SMAD3. Additionally, inducible nitric oxide synthase (iNOS) deficiency or iNOS inhibitor, 1400W, reduced SNO-HSP90 level and the activation of TGFβ/SMAD3 signaling pathway. Conclusions and Implications: We demonstrate that genetic or pharmacological inhibition of SNO-HSP90 mitigates fibrosis through blocking TGFβ/SMAD3 signaling pathway, providing a potential therapy for cardiac remodeling.