Effect of Hypoxia on Cerebral Cortical Genomic DNA in the Guinea Pig Fetus at Term † 1895
| Autor: | Om P Mishra, Imran Qayyum, Alan Zubrow, Dennis McWeeney, Maria Delivoria-Papdopoulos |
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| Rok vydání: | 1998 |
| Předmět: | |
| Zdroj: | Pediatric Research. 43:323-323 |
| ISSN: | 1530-0447 0031-3998 |
| Popis: | Our previous studies have shown that hypoxia in fetal guinea pigs modifies cortical nuclear membranes resulting in increased high-affinity Ca++-ATPase activity and increased number (Bmax) of nuclear membrane IP3 receptors, resulting in calcium influx into the nucleus. The increased intranuclear Ca++ could activate nuclear Ca++-dependent endonuclease,which cuts genomic DNA at specific cleavage sites, causing specific DNA fragmentation. The present study tests the hypothesis that brain tissue hypoxia results in fragmentation of genomic DNA via endonuclease activity in the cerebral cortex of the guinea pig fetus. Guinea pig fetuses were obtained from anesthetized, normoxic (21% oxygen for 60 min, n=6) and hypoxic (7% oxygen for 60 min, n=6) mothers. Fetal brains were removed immediately after the experimental period. Brain tissue hypoxia was documented biochemically by decreased levels of ATP and phosphocreatine. Cerebral cortical nuclei were isolated and purified using a discontinuous sucrose gradient. DNA was isolated and purity was determined by the ratio of absorbance at 260/280nm. DNA samples were separated by gel electrophoresis on 1% agarose and the DNA bands stained with ethidium bromide. The density of the DNA bands was analyzed by imaging densitometery. In normoxic nuclei, only one DNA band was visualized, whereas DNA from hypoxic nuclei demonstrated six discrete peaks ranging in size from 100 to 2000 basepairs. DNA profile suggests that there is specific fragmentation of DNA in a “ladder” pattern in hypoxic nuclei at the internucleosomal linker regions. No random DNA cleavage (which would appear as a dense “smear” due to randomized fragments) was observed. The data demonstrate that there is fragmentation of nuclear DNA, a hallmark of programmed cell death, in cerebral cortex in the guinea pig fetus during acute hypoxia in utero. We speculate that the specific fragmentation of nuclear DNA in the brain cells of the guinea pig fetus is due to increased endonuclease activity as a result of altered nuclear membrane mechanisms of Ca++ influx, leading to hypoxia induced programmed cell death. |
| Databáze: | OpenAIRE |
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