Fig. 3.

Mineralocorticoid receptor (MR) is the major receptor for aldosterone (Aldo) in muscle cells. (A) Interaction between Aldo and MR in mouse C2C12 myoblasts (MBs) using binding enzyme-linked immunosorbent assay (ELISA). Varying amounts of cell lysates were incubated in Aldo- or bovine serum albumin (BSA)-coated wells. Thereafter, the level of MR was determined by ELISA (n=4). (B) Western blot analysis to determine the MR expression in mouse C2C12 cells without or with 2% horse serum for 2 days (n=3). (C, D) Mouse C2C12 MBs were differentiated into myotubes (MTs) with 2% horse serum in the presence or absence of 100 nM Aldo and/or 1 μM eplerenone (Eple) for 3 days. (C) Intracellular reactive oxygen species levels were measured using a fluorescent probe, chloromethyl derivative of 2′,7′-dichlorofluorescein diacetate (CM-H₂DCFDA; n=3). (D) MTs were stained with anti-MyHC antibody while nuclei were counterstained with 4,6-diamidino-2-phenyindole (DAPI). Quantitative results per field are presented (n=3). Scale bars: 100 μm (C) and 100 μm (D). OD, optical density. ^aP<0.05 vs. control; ^bP<0.05 vs. 100 nM Aldo.