Massive cell apoptosis distinguishes cryoinjury from ventricular resection. (A and C) AFOG (Acid Fuchsin Orange-G) staining labels healthy muscle cells (light orange), damaged cells with plasma (dark orange), and collagen (blue). (B and D) Immunostaining with Tropomyosin (TPM) for cardiac muscle (blue), and nuclear MEF-2 as a marker for healthy cardiomyocytes (red). Apoptotic cells are detected by the TUNEL assay (green). (B' and D') Higher magnifications of the framed area in the left panels. (A-B) Adjacent longitudinal sections of the heart one day after ventricular resection. (A) The ventricle (v) comprises a narrow stripe of damaged cardiomyocytes (dark orange) just above the amputation plane (dashed line). Underneath the amputation plane a blood clot (bc) seals the wound; a, atrium; ba, bulbus arteriosus. (B-B') A narrow layer of damaged TPM-positive myocardium above the amputation plane (dashed line) display enhanced apoptosis and reduced MEF-2 expression (arrowheads). (C-D) Adjacent cross sections of the heart one day after cryoinjury. (C) The ventricle encompasses a large disk-shaped damage (encircled with dashed line). (D-D') This region contains abundant apoptotic TPM-positive cardiomyocytes that downregulate MEF-2 expression. Scale bars in (A-B'), 300 μm.