Effect of shear stress and substrate on endothelial DAPK expression, caspase activity, and apoptosis.
BMC Res Notes
; 6: 10, 2013 Jan 10.
Article
em En
| MEDLINE
| ID: mdl-23305096
ABSTRACT
BACKGROUND:
In the vasculature, misdirected apoptosis in endothelial cells leads to pathological conditions such as inflammation. Along with biochemical and molecular signals, the hemodynamic forces that the cells experience are also important regulators of endothelial functions such as proliferation and apoptosis. Laminar shear stress inhibits apoptosis induced by serum depletion, oxidative stress, and tumor necrosis factor α (TNFα). Death associated protein kinase (DAPK) is a positive regulator of TNFα induced apoptotic pathway. Here we investigate the effect of shear stress on DAPK in endothelial cells on glass or silicone membrane substrate. We have already shown a link between shear stress and DAPK expression and apoptosis in cells on glass. Here we transition our study to endothelial cells on non-glass substrates, such as flexible silicone membrane used for cyclic strain studies.RESULTS:
We modified the classic parallel plate flow chamber to accommodate silicone membrane as substrate for cells, and validated the chamber for cell viability in shear stress experiments. We found that adding shear stress significantly suppressed TNFα induced apoptosis in cells; while shearing cells alone also increased apoptosis on either substrate. We also found that shearing cells at 12 dynes/cm2 for 6 hours resulted in increased apoptosis on both substrates. This shear-induced apoptosis correlated with increased caspase 3/7 activities and DAPK expression and activation via dephosphorylation of serine 308.CONCLUSION:
These data suggest that shear stress induced apoptosis in endothelial cells via increased DAPK expression and activation as well as caspase-3/7 activity. Most in vitro shear stress studies utilize the conventional parallel plate flow chamber where cells are cultured on glass, which is much stiffer than what cells encounter in vivo. Other mechanotransduction studies have utilized the flexible silicone membrane as substrate, for example, in cyclic stretch studies. Thus, this study bridges the gap between shear stress studies on cells plated on glass to studies on different stiffness of substrates or mechanical stimulation such as cyclic strain. We continue to explore the mechanotransduction role of DAPK in endothelial apoptosis, by using substrates of physiological stiffness for shear stress studies, and by using silicone substrate in cyclic stretch devices.
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Estresse Fisiológico
/
Endotélio Vascular
/
Apoptose
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Proteínas Quinases Dependentes de Cálcio-Calmodulina
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Proteínas Reguladoras de Apoptose
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Caspase 3
/
Caspase 7
Limite:
Animals
Idioma:
En
Revista:
BMC Res Notes
Ano de publicação:
2013
Tipo de documento:
Article
País de afiliação:
Estados Unidos