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Kinetic analysis of apoptosis induction in human cell lines by UVA and 8-MOP
B R Vowels1, E K Yoo, F P Gasparro
1Department of Dermatology, School of Medicine, University of Pennsylvania, Philidelphia 19104-6142, USA. vowelsb@mail.med.upenn.edu
Abstract:
Whereas previous studies have indicated that DNA damage as a result of 8-methoxypsoralen (8-MOP) and UVA treatment leads to cell death, this study establishes the minimum concentrations of 8-MOP and UVA necessary to induce apoptosis in human T-lymphocytic and monocytic cell lines. In order to asses apoptosis, we used fluorescent microscopy to examine changes in light scattering as well as internucleosomal DNA fragmentation. Generation of a dose response curve showed that the minimum combination of UVA and 8-MOP that was necessary to induce greater than background levels of apoptosis within 24 h of treatment was 0.5 J/cm2 UVA and 12.5 ng/mL of 8-MOP. A striking observation was that UVA alone at doses > or = 1.0 J/cm2, but not 8-MOP alone (0-300 ng/mL), induced significant apoptosis in the Sup-T1 cells induced by UVA alone was not as great as that of 8-MOP and UVA in combination, a highly significant correlation between the product of the concentration of 8-MOP (ng/mL) times the dose of UVA (J/cm2) and the percentage of apoptotic cells was observed. This correlation provides an important tool for studying the relationship of UVA-induced DNA damage to apoptosis induction. moreover, it will provide a means by which early events in the apoptotic pathway can be dissected.
Insights
This study identifies the minimum 8-methoxypsoralen (8-MOP) and UVA levels needed to trigger apoptosis in human T-cells. A combined 0.5 J/cm2 UVA and 12.5 ng/mL 8-MOP effectively induced programmed cell death.
Area of Science:
- Cell Biology
- Photobiology
- Molecular Biology
Background:
- Previous research linked 8-methoxypsoralen (8-MOP) and UVA treatment to cell death.
- Apoptosis induction mechanisms by combined 8-MOP and UVA require precise quantification.
Purpose of the Study:
- To determine the minimum effective concentrations of 8-MOP and UVA for inducing apoptosis in human T-lymphocytic and monocytic cell lines.
- To establish a quantitative correlation between 8-MOP/UVA exposure and apoptosis levels.
Main Methods:
- Fluorescent microscopy was used to assess apoptosis via light scattering and DNA fragmentation.
- Dose-response curves were generated to identify minimum effective treatment levels.
- Statistical correlation analysis was performed between treatment product and apoptosis percentage.
Main Results:
- The minimum effective combination for inducing apoptosis was 0.5 J/cm2 UVA and 12.5 ng/mL 8-MOP.
- UVA alone (≥1.0 J/cm2) induced significant apoptosis in Sup-T1 cells, whereas 8-MOP alone did not.
- A strong correlation was observed between the product of 8-MOP concentration and UVA dose, and the percentage of apoptotic cells.
Conclusions:
- A precise threshold of 8-MOP and UVA is identified for effective apoptosis induction.
- This quantitative relationship aids in studying UVA-induced DNA damage and apoptosis.
- The findings provide a tool for dissecting early events in the apoptotic pathway.