Abstract 1325: Morphological Characterization of Intervened Basal and Starvation-induced Autophagy in Cultured Adult Rat Cardiomyocytes
Autophagy is originally an important physiological process for cell survival but its pathophys-iological significance is largely unknown, although autophagic degenaration/death of cardio-myocytes has been recently reported to be associated with progression of heart failure. We induced autophagy in isolated adult rat ventricular cardiomyocytes (ARVC) by incubation with glucose-depleted/mannitol-supplemented medium for 72 hours. The control was ARVC incubated with glucose-containing medium. Intracellular vacuoles were apparent in the glucose-depleted ARVC under an electron microscope, which contained degenerated subcel-lular organelles such as mitochondria, indicating that the vacuoles were autophagic. Incidences of the ARVC immunopositive for LC3 were significantly greater in the glucose-depleted group than those in the glucose-supplemented group. Western blotting for LC3 confirmed this. Spontaneously occurring apoptosis was also observed in cultured ARVC, of which incidence was similar between the groups, and apoptotic ARVC were negative for LC3. The glucose-depleted ARVC showed significantly shorter survival intervals as assessed by Trypan blue dye exclusion, compared with controls (survival rate, 19+/−5.2% vs. 55+/−8.3% after 72 hour-incubation). Electron microscoic examination revealed that necrosis was the final form of death of autophagic ARVC. Next, ARVC were loaded with the inhibitors of autophagy: 3-methyladenine; bafilomycin A1; and leupeptin. All of the inhibitors were found to significantly reduce viability of the ARVC in a dose dependent manner in both glucose-supplemented and glucose-depleted groups. Thus, autophagy is likely to be protective for the ARVC survival. The present findings may imply a possibility that autophagic degeneration of cardiomyocytes reflects not a previous notice of programmed cell death but a compensatory response to stress for cell survival and that autophagic cardiomyocyte death may simply represent the unsuccessful outcome for survival, reaching necrosis.