Date of Graduation
1997
Document Type
Thesis
Abstract
Redoxyendonuclease activity was detected in extracts of human neuroblastoma cells using a base-release assay specific for thymine glycol in DNA. The level of redoxyendonuclease activity was more than 2-fold higher in dividing cells compared to quiescent cells, suggesting that quiescent cells may have a reduced capacity to repair oxidative DNA base damages. Cells were synchronized by serum deprivation and then stimulated to enter the cell cycle by the addition of serum to determine enzyme activity at different stages of the cell cycle. The redoxyendonuclease activity was regulated in a biphasic manner with a peak in early G1 and a peak in S phase. This suggests that at specific times during the cell cycle actively growing cells may be more resistant to oxidative DNA damage due to increased repair capacity. The repair capacity of neuroblastoma cells was quantified as the decrease in enzyme-sensitive sites determined by alkaline sucrose density gradient centrifugation following treatment with the oxidant osmium tetroxide. Actively dividing cells repaired the oxidative damage in approximately 24 hours, while the quiescent cells failed to excise the damaged sites and subsequently died. These results indicate that non-dividing cells do not effectively repair oxidative DNA damage, as compared to the dividing cells. Similarly, quiescent cells, treated with osmium tetroxide and fed a serum-enriched media, failed to re-enter the cell cycle and did not repair the oxidative damage. The data indicate that non-dividing cells, such as neurons, do not have the capacity to repair excess oxidative damage and may suffer the biological consequences of DNA damage accumulation, including cellular death, mutagenesis or carcinogenesis. When synchronized cells were damaged with osmium tetroxide, there were differential DNA repair rates, depending on the stage of the cell cycle. These DNA repair rates coordinated with the redoxyendonuclease activity profile. The results of the studies described contribute to a further understanding of the DNA repair pathways which are interlinked with complex processes of cell cycle regulation.
Recommended Citation
Walpita, Deepika, "Cell cycle regulation of redoxyendonuclease activity in human neuroblastoma cells." (1997). Graduate Theses, Dissertations, and Problem Reports. 9961.
https://researchrepository.wvu.edu/etd/9961