Measurement of aldosterone/cortisol concentration

H295R cells were plated to 60% confluence in 24‐multiwell plates. Thereafter, they were exposed to either 100 mg·dL⁻¹ d‐glu, 450 mg·dL⁻¹ d‐glu, 100 mg·dL⁻¹ d‐glu plus 100 nmol·L⁻¹ AII (for aldosterone), or 450 mg·dL⁻¹ d‐glu plus 100 nmol·L⁻¹ AII (for aldosterone) for 72 h. The aldosterone and cortisol concentrations of the media were thereafter measured by Aldosterone EIA Kit and Cortisol EIA Kit (Cayman Chemical), respectively, after their extraction with dichloromethane according to the manufacturer's instructions. The obtained data were normalized by the protein concentrations measured by Protein Assay Kit (Bio‐Rad).