The heart size was photographed using a digital camera (Olympus C5060WZ, Japan). The HW, LW, BW, and tibial length (TL) were measured. The LW/TL, HW/BW, and HW/TL ratios were calculated as an index of LV hypertrophy or HF. The heart was fixed in 4% paraformaldehyde solution for 24 hours and then embedded in paraffin. The sections (4 μm) were stained with hematoxylin and eosin (H&E) and Masson’s trichrome staining using the standard procedure (40, 44). To assess myocyte size, heart sections were stained with tetramethyl rhodamine isothiocyanate (TRITC)–labeled wheat germ agglutinin [WGA; 50 μg/ml in 1 × phosphate-buffered saline (PBS)] for 1 hour. The cell area was calculated by measuring at least 200 cells per slide. Digital photographs were taken at ×200 magnification of over 20 random fields from each heart and analyzed by Image Pro Plus 3.0 (Nikon, Japan) (40).

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