Direct Aldosterone Action on Mouse Cardiomyocytes Detected with Atomic Force Microscopy

Abstract

There is growing evidence that aldosterone acts on heart where it causes cellular remodeling and hypertrophy. Since it is still unclear whether aldosterone directly acts on cardiomyocytes or indirectly, by an altered electrolyte balance in the organism, we applied atomic force microscopy (AFM) in primary cultures of neonatal mouse cardiomyocytes to measure hormone-induced changes in cell volume and plasma membrane surface. AFM measures cell volume and, at the same time, provides quantitative information on cell surface properties. Neonatal mouse cardiomyocytes were cultured for 28 hours in absence or presence of 100 nM aldosterone. Spironolactone was applied as a selective aldosterone receptor antagonist. At the microscopic level, single cell volume and single cell surface were found unchanged by aldosterone. However, nanoscopy of the cell surface, i.e. analysis of the plasma membrane at the nanometer level, revealed a specific increase in plasma membrane nano-enfoldings (roughness). This aldosterone-mediated increase in cell surface roughness was completely prevented by spironolactone. We conclude: (i) Aldosterone directly acts upon cardiomyocytes. (ii) At the microscopic level, no changes of cell volume and cell surface are detectable. (iii) At the nanoscopic level, aldosterone increases plasma membrane roughness. These nanometer changes, detectable only with AFM in cells scanned in fluid after fixation under physiological conditions, indicate plasma membrane remodeling of cardiomyocytes by mineralocorticoids.

Cardiac corticosteroid receptors mediate the enlargement of the ovine fetal heart induced by chronic increases in maternal cortisol

“…Other investigators have found that aldosterone directly stimulates myocyte surface area (Okoshi et al 2004) and remodeling of myocyte membrane (Kliche et al 2006) in cultures of neonatal myocytes, an effect presumed to be mediated by MR in the myocytes. Cortisol also increases expression of atrial natriuretic peptide in cultured neonatal myocytes, and both cortisol and aldosterone potentiate the effect of phenylephrine on hypertrophy in these cultures (Lister et al 2006), also indicating an intracardiac action at MR in these cultures.…”

DOCA and TGF-β Induce Early Growth Response Gene-1 (Egr-1) Expression

“…For western blot analysis, 2x10 6 cells were washed with ice-cold PBS and detached from flasks on ice using a cell scraper. After centrifugation (1400 rpm at 4°C), pelleted cells were dissolved in 60 µl lysis buffer (1% NP40; 150 mM NaCl; 50 mM Tris/HCL, pH 8 plus protease inhibitors Complete, EDTA-free, Roche, Germany) and incubated overnight at -70°C.…”

“…The underlying signaling pathways comprise mesangial and fibroblast activation as well as tubular epithelial-mesenchymal transition [1]. Stimulators of renal fibrosis include the mineralocorticoids aldosterone and deoxycorticosterone acetate (DOCA), hormones that decrease renal salt excretion [2], increase salt appetite [3,4], increase blood pressure [5], and lead to remodelling of the heart [6,7]. Activation of the renin-angiotensinaldosterone system (RAAS) stimulates transforming growth factor-β (TGF-β) expression in the kidney through various mechanisms and upregulates receptors for TGF-β.…”

Ultra-rapid non-invasive clinical nano-diagnostic of inflammatory diseases

“…Similarly, T-lymphocytes in thyroid associated ophtalmopathy show a higher diameter and roughness compared with control group 43 . For example, drugs as paclitaxel modifies the roughness of epithelia cells 44 , or aldosteron remodels the plasma membrane of cardiomyocytes 45 . Moreover, cell membrane is sensitive to change of environmental conditions such as salt, pH, ionic strength, etc.…”