Lack of Apaf-1 expression confers resistance to cytochrome c-driven apoptosis in cardiomyocytes

Cardiac fibroblasts play an essential role in the physiology of the heart. These produce extracellular matrix proteins and synthesize angiogenic and cardioprotective factors. Although fibroblasts of cardiac origin are known to be resistant to apoptosis and to remain metabolically active in situations compromising cell survival, the underlying mechanisms are unknown. Here, we report that cardiac fibroblasts were more resistant than dermal or pulmonary fibroblasts to mitochondriadependent cell death. Cytochrome c release was blocked in cardiac fibroblasts but not in dermal fibroblasts treated with staurosporine, etoposide, serum deprivation, or simulated ischemia, precluding caspase-3 activation and DNA fragmentation. Resistance to apoptosis of cardiac fibroblasts correlated with the expression of the anti-apoptotic protein Bcl-2, whereas skin and lung fibroblasts did not express detectable levels of this protein. Bcl-x L, Bax, and Bak were expressed at similar levels in cardiac, dermal, and lung fibroblasts. In addition, the death of cardiac fibroblasts during hypoxia was not associated with the cleavage of Bid but rather with Bcl-2 disappearance, suggesting the requirement of the mitochondrial apoptotic machinery to execute death receptor-induced programmed cell death. Knockdown of bcl-2 expression by siRNA in cardiac fibroblasts increased their apoptotic response to staurosporine, serum, and glucose deprivation and to simulated ischemia. Moreover, dermal fibroblasts overexpressing Bcl-2 achieved a similar level of resistance to these stimuli as cardiac fibroblasts. Thus, our data demonstrate that Bcl-2 is an important effector of heart fibroblast resistance to apoptosis and highlight a probable mechanism for promoting survival advantage in fibroblasts of cardiac origin.

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“…of three independent experiments. DNA fragmentation was assessed by conventional agarose gel electrophoresis as previously reported (14).…”

Section: Methodsmentioning

confidence: 99%

“…However, the exact role that mitochondria play in cardiac apoptosis is still a matter of debate (13). In this regard, we have recently reported that cardiac myocytes are resistant to apoptosome-driven apoptosis by a mechanism involving the silencing of Apaf-1 expression (14).…”

mentioning

confidence: 99%

Bcl-2 Is a Key Factor for Cardiac Fibroblast Resistance to Programmed Cell Death

Mayorga

Bahí

Ballester

et al. 2004

Journal of Biological Chemistry

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“…The mechanisms of cell death in terminally differentiated, nondividing cells such as cardiomyocytes and neurons are less well defined. Recent studies have shown that Apaf-1 levels in post-mitotic cells such as cardiomyocytes and sympathetic neurons are markedly reduced (as compared to mitotic cells) [108,109,116], resulting in a significant decrease in apoptosome activity. The reduced activity of E2F1 [30], which is a transcriptional regulator of Apaf-1 [93] in terminally differentiated cardiomyocytes, is likely to contribute to the marked reduction of Apaf-1 in these cells.…”

Section: Apoptosismentioning

confidence: 99%

Rho kinase in the regulation of cell death and survival

Shi

Wei

2007

Arch. Immunol. Ther. Exp.

219

183

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SummaryRho kinase (ROCK) belongs to a family of serine/threonine kinases that are activated via interaction with Rho GTPases. ROCK is involved in a wide range of fundamental cellular functions such as contraction, adhesion, migration, and proliferation. Recent studies have shown that ROCK plays an important role in the regulation of apoptosis in various cell types and animal disease models. Two ROCK isoforms, ROCK1 and ROCK2, are assumed to be function redundant, based largely on kinase construct overexpression, and chemical inhibitors (Y27632 and fasudil), which inhibit both ROCK1 and ROCK2. Gene targeting and RNA interference approaches allow further dissection of distinct cellular, physiological and patho-physiological functions of the two ROCK isoforms. This review, based on recent molecular, cellular and animal studies, focuses on the current understanding of ROCK signaling in the regulation of apoptosis and highlights the new findings from recently generated ROCK-deficient mice.

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“…Protein expression was detected in protein extracts diluted in Tris-buffered 2% SDS solution at pH 6.8 as reported earlier. 30 Antibody data can be found in the Supplementary Table 2.…”

Section: Methodsmentioning

confidence: 99%

“…Executioner caspase enzymatic activity was measured in cell extracts as reported earlier 30 using the fluorogenic substrate Z-DEVD-AFC (Caspase-3 Substrate IV; Merck-Calbiochem) at 50 mM in 96-well plates. Fluorescence was detected with a Biotek Instruments Inc. FL Â 800 Fluorescence Reader (Izasa, Spain) with excitation filter set at 360 nm and emission filter at 530 nm.…”

Section: Methodsmentioning

confidence: 99%

Polypyrimidine tract binding proteins (PTB) regulate the expression of apoptotic genes and susceptibility to caspase-dependent apoptosis in differentiating cardiomyocytes

et al. 2009

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Cardiac morphologic abnormalities in mice deficient for key regulators of the caspase-dependent signaling underscored its role in heart development. However, the mechanisms regulating apoptotic gene expression in the developing heart are unknown. As polypyrimidine tract binding proteins (PTB) determine gene isoform expression during myoblast differentiation and contribute to Apaf-1 translation in cell lines, we investigated whether PTB regulate apoptotic gene expression in differentiating cardiomyocytes. Our results show that PTB are expressed in the embryonic heart and are silenced during development, coinciding with a reduction in the expression of apoptotic genes. Overexpression of PTB in postnatal cardiomyocytes, which express low levels of PTB and apoptotic genes, induced an increase in the amount of pro-apoptotic proteins without affecting abundance of their respective transcripts. Translation of the reporter gene Firefly Luciferase preceded by the 5 0 -untranslated region of Apaf-1 or Caspase-3 was enhanced by PTB in cardiomyocytes. PTB silencing in fibroblasts induced a decrease of apoptotic protein levels. PTB overexpression in cardiomyocytes induced caspase activity and caspase-dependent DNA fragmentation during ischemia, which is otherwise caspase-independent in differentiated cardiomyocytes. Our results show that PTB contribute to apoptotic gene expression and modulate the susceptibility to caspase activation in differentiating rat cardiomyocytes.

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Lack of Apaf-1 expression confers resistance to cytochrome c-driven apoptosis in cardiomyocytes

Cited by 66 publications

References 42 publications

Bcl-2 Is a Key Factor for Cardiac Fibroblast Resistance to Programmed Cell Death

Bcl-2 Is a Key Factor for Cardiac Fibroblast Resistance to Programmed Cell Death

Rho kinase in the regulation of cell death and survival

Polypyrimidine tract binding proteins (PTB) regulate the expression of apoptotic genes and susceptibility to caspase-dependent apoptosis in differentiating cardiomyocytes

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