lec15_rein_blas

lec15_rein_blas - Widespread programmed cell death in...

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Widespread programmed cell death in proliferative and postmitotic regions of the fetal cerebral cortex Anne J. Blaschke, Kristina Staley, and Jerold Chun, 1996 Presented by Jonathan Reinharth April 5, 2005
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Programmed Cell Death Apoptosis Fragments DNA Nucleosomal ladders Found in post-mitotic cells Assumed not to occur in embryonic cortical development (specifically, in proliferative areas)
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Experimental Goal Characterize the distribution and rates of PCD in the embryonic cerebral cortex (in mice) during embryonic days 10-18
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In Situ End Labeling Identifies fragmented nuclear DNA in dying cells Attaches labeled nucleotides to free DNA ends, which can then be visualized in tissue sections Past studies with ISEL Identifies PCD specifically (Gavrieli et al, 1992) ISEL+: Section incubated with mix containing terminal- deoxynucleotidal-transferase (TdT) and either a) digoxigenin-11-dUTP incubated with blocking solution 1hr incubated with anti-digoxigenin Fab fragments overnight; or b) [ 32 P]dCTP incubated at 37 o C for 1hr incubated at 65 o C for 2hr; washed
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In-Situ End Labeling + Does it identify cells undergoing PCD? Does it only identify these cells, or others as well? Does it do this in many areas of the nervous system?
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The Effectiveness of Isel+ Labeling ISEL+ in the thymus ISEL+ in the retina ISEL+ marking in vitro apoptosis ISEL+ under heat inactivation Only PCD fragments? RNase DNase Okazaki fragments
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ISEL+ in the Thymus Dexamethasone is known to increase PCD amongst immature T-cells ISEL+ detected this effect (compared with control) Primarily in thymic cortex (97% death) DNA fragments undergoing apoptosis (nucleosomal ladders in LMPCR) ISEL+ labeling of dying cells: a) Normal thymus b) Dexamethasone treated thymic section Source: Blaschke, A. J., and K. Staley, et al. "Widespread Programmed Cell Death in Proliferative and Postmitotic Regions of
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This note was uploaded on 11/11/2011 for the course BIO 7.344 taught by Professor Bobsauer during the Spring '08 term at MIT.

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lec15_rein_blas - Widespread programmed cell death in...

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