Dissecting apoptosis and intrinsic death pathways in the heart.

The limited regenerative capacity of postnatal ventricular myocytes coupled with their meager ability for genetic manipulation has presented a major technical obstacle for deciphering apoptosis initiation and execution signals in the heart. In this report, we describe the technical approaches used to study the intrinsic death pathways in postnatal ventricular myocytes during acute hypoxic injury. Discussed are methods for hypoxia, recombinant adenovirus-mediated gene transfer, cellular viability assays using the vital dyes calcein acetomethoxyester and ethidium homodimer-1, analysis of nuclear morphology by use of Hoechst dye 33258, and assessment of the state of the mitochondrial permeability transition pore.

Nuclear factor-kappaB-mediated cell survival involves transcriptional silencing of the mitochondrial death gene BNIP3 in ventricular myocytes.

Background: A survival role for the transcription factor nuclear factor-kappaB (NF-kappaB) in ventricular myocytes has been reported; however, the underlying mechanism is undefined. In this report we provide new mechanistic evidence that survival signals conferred by NF-kappaB impinge on the hypoxia-inducible death factor BNIP3.

Methods And Results: Activation of the NF-kappaB signaling pathway by IKKbeta in ventricular myocytes suppressed mitochondrial permeability transition pore (PTP) opening and cell death provoked by BNIP3.

Distinct pathways regulate proapoptotic Nix and BNip3 in cardiac stress.

Up-regulation of myocardial Nix and BNip3 is associated with apoptosis in cardiac hypertrophy and ischemia, respectively. To identify mechanisms of gene regulation for these critical cardiac apoptosis effectors, the determinants of Nix and BNip3 promoter activation were elucidated by luciferase reporter gene expression in neonatal rat cardiac myocytes. BNip3 transcription was increased by hypoxia but not by phenylephrine (10 microM), angiotensin II (100 nM), or isoproterenol (10 microM).

Apoptosis of ventricular myocytes: a means to an end.

One of the most compelling issues to impact on contemporary cardiology is arguably the phenomenon of programmed cell death or apoptosis. Studies in the nematode Caenorhabditis elegans provided the first indication that determinants of cell fate crucial for normal worm development were under genetic influences of the ced-3 and ced-9 genes, which promote or prevent cell death, respectively. Extrapolation of these seminal findings led to the discovery of the mammalian ced-3 and ced-9 homologs, which broadly encompass a family of cellular cysteine proteases known collectively as caspases and the Bcl-2 proteins.

Nuclear factor-kappaB represses hypoxia-induced mitochondrial defects and cell death of ventricular myocytes.

Background: Oxygen deprivation for prolonged periods of time provokes cardiac cell death and ventricular dysfunction. Preventing inappropriate cardiac cell death in patients with ischemic heart disease would be of significant therapeutic value as a means to improve ventricular performance. In the present study, we wished to ascertain whether activation of the cellular factor nuclear factor (NF)-kappaB suppresses mitochondrial defects and cell death of ventricular myocytes during hypoxic injury.

Therapeutic opportunities for cell cycle re-entry and cardiac regeneration.

Over the last two decades, considerable effort has been made to better understand putative regulators and molecular switches that govern the cell cycle in attempts to reactivate cell cycle progression of cardiac muscle. Rapid advancements on the field of stem cycle biology including evidence of cardiac progenitors within the adult myocardium itself and reports of cardiomyocyte DNA synthesis, which each suggest that the adult myocardium may in fact have the capacity for de novo myocyte regeneration. Augmenting cardiomyocyte number by targeting specific cell cycle regulatory genes or by stimulating cardiac progenitor cells to differentiate into cardiac muscle may be of therapeutic value in repopulating the adult myocardium with functionally active cells in patients with end-stage heart failure.

Inducible expression of BNIP3 provokes mitochondrial defects and hypoxia-mediated cell death of ventricular myocytes.

In this study, we provide evidence for the operation of BNIP3 as a key regulator of mitochondrial function and cell death of ventricular myocytes during hypoxia. In contrast to normoxic cells, a 5.6-fold increase (P<0.

IKK beta is required for Bcl-2-mediated NF-kappa B activation in ventricular myocytes.

The transcription factor nuclear factor kappa B (NF-kappa B) is regulated by cytoplasmic inhibitor I kappa B alpha. An integral step in the activation of NF-kappa B involves the phosphorylation and degradation of I kappa B alpha. We have previously reported that I kappa B alpha activity is diminished in ventricular myocytes expressing Bcl-2 (de Moissac, D.