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Abstract-We demonstrated recently that chronic administration of aldosterone to rats induces glomerular mesangial injury and activates mitogen-activated protein kinases including extracellular signal-regulated kinases 1/2 (ERK1/2). We also observed that the aldosterone-induced mesangial injury and ERK1/2 activation were prevented by treatment with a selective mineralocorticoid receptor (MR) antagonist, eplerenone, suggesting that the glomerular mesangium is a potential target for injuries induced by aldosterone via activation of MR. In the present study, we investigated whether MR is expressed in cultured rat mesangial cells (RMCs) and involved in aldosterone-induced RMC injury. MR expression and localization were evaluated by Western blotting analysis and fluorolabeling methods. Cell proliferation and micromechanical properties were determined by [ 3 H]-thymidine uptake measurements and a nanoindentation technique using an atomic force microscope cantilever, respectively. ERK1/2 activity was measured by Western blotting analysis with an anti-phospho-ERK1/2 antibody. Protein expression and immunostaining revealed that MR was abundant in the cytoplasm of RMCs. Aldosterone (1 to 100 nmol/L) dose-dependently activated ERK1/2 in RMCs with a peak at 10 minutes. Pretreatment with eplerenone (10 mol/L) significantly attenuated aldosterone-induced ERK1/2 phosphorylation. Aldosterone (100 nmol/L) treatment for 30 hours increased Key Words: mineralocorticoids Ⅲ aldosterone T he utility of mineralocorticoid receptor (MR) antagonists in renal injury has been suggested in preclinical and clinical studies. 1-12 MR blockade had no effect on systemic blood pressure but markedly ameliorated glomerular injury in stroke-prone spontaneously hypertensive rats 3 and rats treated with angiotensin II (Ang II) and an NO synthase inhibitor, 4 cyclosporine A 5 or radiation. 6 In patients with chronic renal failure 7 and early diabetic nephropathy, 8 addition of a nonselective MR antagonist, spironolactone, to angiotensinconverting enzyme (ACE) inhibitors had no hemodynamic effects but markedly reduced the urinary protein excretion rate (U protein V). For hypertensive patients, it has also been indicated that monotherapy with spironolactone 9 or a selective MR antagonist, eplerenone, 10 is more effective than ACE inhibitors in reducing U protein V. Furthermore, White et al 11 showed that in hypertensive patients, eplerenone has a similar blood pressure-lowering effect to a calcium antagonist, amlodipine, but reduced the urinary albumin-to-creatinine ratio to a greater extent than amlodipine. Thus, these observations support the notion that MR blockade has renoprotective effects through mechanisms that cannot be simply explained by hemodynamic changes.We demonstrated recently that chronic administration of aldosterone to rats induced glomerular injury characterized by mesangial matrix expansion and cell overgrowth. 12 We also observed that the aldosterone-induced glomerular injury was prevented by treatment with eplerenone. These results in...

Accumulating evidence implicates the significance of the physical properties of the niche in influencing the behavior, growth and differentiation of stem cells. Among the physical properties, extracellular stiffness has been shown to have direct effects on fate determination in several cell types in vitro. However, little evidence exists concerning whether shifts in stiffness occur in vivo during tissue development. To address this question, we present a systematic strategy to evaluate the shift in stiffness in a developing tissue using the mouse embryonic cerebral cortex as an experimental model. We combined atomic force microscopy measurements of tissue and cellular stiffness with immunostaining of specific markers of neural differentiation to correlate the value of stiffness with the characteristic features of tissues and cells in the developing brain. We found that the stiffness of the ventricular and subventricular zones increases gradually during development. Furthermore, a peak in tissue stiffness appeared in the intermediate zone at E16.5. The stiffness of the cortical plate showed an initial increase but decreased at E18.5, although the cellular stiffness of neurons monotonically increased in association with the maturation of the microtubule cytoskeleton. These results indicate that tissue stiffness cannot be solely determined by the stiffness of the cells that constitute the tissue. Taken together, our method profiles the stiffness of living tissue and cells with defined characteristics and can therefore be utilized to further understand the role of stiffness as a physical factor that determines cell fate during the formation of the cerebral cortex and other tissues.