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| Thanks to the following for materials & support |
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Apoptosis & Cell Death
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Apoptosis differs from necrosis in that physiological extracellular stimuli initiate cell death via numerous signaling cascades. These cascades involve targeting of mitochondria and increase in mitochondrial permeability. This results in release of mitochondrial proteins and activation of proteases called caspases which target many organelles, including the nucleus and DNA fragmentation (Fig. 1).
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Fig 1. Staurosporine-induced apoptosis is identified in cultured myocytes by laddering of DNA
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This process is vital to the development of tissues and organs and to the removal of damaged or diseased cells. However, defective apoptotic pathways can lead to promotion of cell death, leading to heart disease, Type 1 diabetes, neurodegeneration and numerous other pathologies.
Downregulation leads to uncontrolled proliferation of cells and cancer. The BRC is currently investigating the bioenergetic basis of apoptosis by assessing metabolic parameters during and after ischemic interventions, in both neurons and cardiac myocytes. Such interventions include hypoxia-reperfusion treatment and oxidative stress. In collaboration with the Jonas Laboratory at the Yale School of Medicine, the BRC is studying the mechanism by which BCL-xL attenuates apoptosis and cell death in both hippocampal neurons and cardiac myocytes (Fig 2). Staining with terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL; shown in yellow) detects DNA fragmentation as a result of apoptotic signal cascades. Hypoxia (24h) followed by reoxygenation leads to an increase in percentage of TUNEL-positive nuclei (Fig 2). The percentage of TUNEL-positive nuclei, relative to total nuclei (DAPI; shown in blue), is reduced in neonatal cardiac myocytes that overexpress BCL-xL.
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Fig 2. |
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| • Anion release in response to hypotonic challenge >> |
• Flux studies from mammalian emobryos as indicators of
viability & cell death >> |
| • Mitochondrial channel activity within a living synapse >> |
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| Garber, S.S., Messerli, M.A., Hubert, M., Lewis, R., Hammar, K., Indyk, E. and Smith, P.J.S. 2005. Monitoring Cl- movement in single cells exposed to hypotonic solution. Journal of Membrane Biology, 203:101-110. |
| Trimarchi, J.R., Liu, L., Smith, P.J.S. and Keefe, D.L. 2002. Apoptosis recruits two-pore domain potassium channels used for homeostatic volume regulation. American Journal of Physiology. Cell Physiology. 282: C588-C594. |
| Trimarchi, J.R., Liu, L., Smith, P.J.S. and Keefe, D.L. 2000. Non-invasive measurement of potassium efflux as an early indicator of cell death in mouse embryos. Biology of Reprodution, 63: 851-857. |
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Equipment
