Abstract Body (Do not enter title and authors here): Introduction. The Hippo pathway is an intracellular pathway that negatively regulates cell survival. Experimental evidence demonstrated that the activation of mammalian sterile 20-like kinase 1 (MST1), the core component of the Hippo Pathway, leads to dilated cardiomyopathy and heart failure. However, the role of MST1 in endothelial dysfunction caused by cardiovascular risk factors has not yet been investigated.
Hypothesis. We tested the hypothesis that MST1 activation in response to metabolic stress may exert detrimental endothelial effects.
Methods. We studied MST1 activity in endothelial cells (HUVECs) treated with metabolic stress (high glucose, HG; oxidized low-density lipoproteins, oxLDL). Adenoviruses overexpressing either a wild type (AD-MST1) or a dominant negative form of MST1 (AD-DN-MST1) were used for overexpression and inhibition studies, respectively. Angiogenesis, apoptosis, oxidative stress, nitric oxide (NO), RAC1/NOX2 activity were assessed. We studied endothelial function in mesenteric arteries isolated from mice with endothelial-specific deletion of MST1 (MST1eKO) subjected to HG or oxLDL treatment. We also analyzed MST1 levels in peripheral blood mononuclear cells (PBMCs) from patients with coronary artery disease (CAD).
Results. Endogenous levels of MST1 increase in response to HG or ox-LDL (p<0.05). Forced overexpression of MST1 by AD-MST1 induces apoptosis (p<0.001), impairs angiogenesis (p<0.001) and NO metabolism (p<0.001) in HUVECs. Inhibition of MST1 rescues the deleterious endothelial effects of both HG and oxLDL. Mechanistically, we demonstrated that AD-MST1 promotes RAC1/NOX2-induced oxidative stress (p<0.01). Pharmacological or genetic RAC1/NOX2 inhibition rescues endothelial dysfunction induced by MST1 overexpression. We also observed that endothelial-dependent relaxation, oxidative stress and NO production are preserved in MST1eKO mice undergoing metabolic stress (p<0.001). Finally, we found that MST1, NOX2 and RAC1 levels were significantly higher in patients with CAD compared to those without, and MST1 correlated with CAD severity, evaluated by the SYNTAX Score (rho=0.209, p=0.028).
Conclusion. This study provides novel evidence that MST1 activation plays a pivotal role in endothelial/vascular dysfunction induced by cardiovascular risk factors. Inhibition of MST1 may be considered a potential strategy for the prevention of cardiovascular diseases.