Differential contribution of necrosis and apoptosis in myocardial ischemia-reperfusion injury

Abstract

Necrosis and apoptosis differentially contribute to myocardial injury. Determination of the contribution of these processes in ischemia-reperfusion injury would allow for the preservation of myocardial tissue. Necrosis and apoptosis were investigated in Langendorff-perfused rabbit hearts (n = 47) subjected to 0 (Control group), 5 (GI-5), 10 (GI-10), 15 (GI-15), 20 (GI-20), 25 (GI-25), and 30 min (GI-30) of global ischemia (GI) and 120 min of reperfusion. Myocardial injury was determined by triphenyltetrazolium chloride (TTC) staining, terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL), bax, bcl2, poly(ADP)ribose polymerase (PARP) cleavage, caspase-3, -8, and -9 cleavage and activity, Fas ligand (FasL), and Fas-activated death domain (FADD). The contribution of apoptosis was determined separately (n = 42) using irreversible caspase-3, -8, and -9 inhibitors. Left ventricular peak developed pressure (LVPDP) and systolic shortening (SS) were significantly decreased and infarct size and TUNEL-positive cells were significantly increased (P < 0.05 vs. Control group) at GI-20, GI-25, and GI-30. Proapoptotic bax, PARP cleavage, and caspase-3 and -9 cleavage and activity were apparent at GI-5 to GI-30. Fas, FADD, and caspase-8 cleavage and activity were unaltered. Irreversible inhibition of caspase-3 and -9 activity significantly decreased (P < 0.05) infarct size at GI-25 and GI-30 but had no effect on LVPDP or SS. Myocardial injury results from a significant increase in both necrosis and apoptosis (P < 0.05 vs. Control group) evident by TUNEL, TTC staining, and caspase activity at GI-20. Intrinsic proapoptotic activation is evident early during ischemia but does not significantly contribute to infarct size before GI-25. The contribution of necrosis to infarct size at GI-20, GI-25, and GI-30 is significantly greater than that of apoptosis. Apoptosis is significantly decreased by caspase inhibition during early reperfusion, but this protection does not improve immediate postischemic functional recovery.

Mitochondrial network in the heart

“…Necrosis, rather than apoptosis is the major cause of cell death during ischemia (McCully et al, 2004). Cardiomyocyte apoptosis is evident only at the long term of ischemia (more than 25 minutes), mainly exists adjacent to the ischemia necrotic zone, and inhibition of apoptosis fails to rescue the function of the ischemic cardiomyocyte (Zhao et al, 2000).…”

Molecular mechanism and therapy application of necrosis during myocardial injury

“…Inhibition of apoptosis has been applied to the protection of cardiomyocytes in heart diseases 64, 65. But the inhibition of apoptosis by caspase inhibitors during early reperfusion did not improve the post‐ischaemic heart function 66. During the heart I/R injury, both apoptosis and necrosis would happen, but Leist and his coworkers showed that loss of intracellular ATP would switch the apoptosis to necrosis.…”

Cardioplegia and Diazoxide Modulate STAT3 Activation and DNA Binding

“…Nuclear protein extracts were isolated from pooled heart tissue (n = 6 for each group of hearts) as previously described [9][10][11]. Protein concentration was determined by BCA (bicinchoninic acid) protein assay reagent (Pierce, Rockford, IL).…”

“…Protein concentration was determined by BCA (bicinchoninic acid) protein assay reagent (Pierce, Rockford, IL). Nuclear protein (20 μg) was separated by 6% SDS-polyacrylamide gel electrophoresis and transferred to nitrocellulose membranes as previously described [11]. Efficiency of protein transfer was confirmed by Ponceau S staining [9,11].…”

“…Nuclear protein (20 μg) was separated by 6% SDS-polyacrylamide gel electrophoresis and transferred to nitrocellulose membranes as previously described [11]. Efficiency of protein transfer was confirmed by Ponceau S staining [9,11]. Blots were incubated with appropriate primary antibodies (1:200 dilution), washed and incubated with appropriate secondary antibodies (1:2000), and detected using ECL Plus detection system (Amersham Pharmacia Biotech, Piscataway, NJ), as previously described [9,11].…”