Abstract
[3H]-thymidine is commonly used to analyze the accumulation of [3H]-labeled chromatin fragments in cells undergoing apoptosis. This study shows that [3H]-thymidine incorporation within DNA is sufficient per se to inhibit growth and to induce apoptosis in canine kidney epithelial cells and porcine aorta endothelial cells. Despite high-level [3H]-thymidine-DNA labeling, rat vascular smooth muscle cells (VSMC) showed only modest inhibition of growth and induction of apoptosis compared to other cell types. Similarly to serum deprivation, apoptosis triggered by [3H]-thymidine labeling was sharply potentiated by VSMC transfection with a functional analogue of c-myc, E1A-adenoviral protein (VSMC-E1A), and was suppressed by stimulation of cAMP signaling with forskolin as well as by and Na/K pump inhibition with ouabain. Both apoptosis induction and growth suppression seen in [3H]-thymidine-treated VSMC-E1A were reduced by the pan-caspase inhibitor z-VAD.fmk. Thus, our results show that the differential efficiency of the apoptotic machinery determines cell type-specific attenuation of growth in cells with [3H]-thymidine-labeled DNA. They also demonstrate that [3H]-thymidine-treated and serum-deprived VSMC employ common intermediates of the apoptotic machinery, including steps that are potentiated by E1A-adenoviral protein and inhibited by activation of cAMP signaling as well as by inversion of the intracellular [Na+] i /[K+] i ratio.
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Orlov, S.N., Pchejetski, D.V., Der Sarkissian, S. et al. [3H]-thymidine labelling of DNA triggers apoptosis potentiated by E1A-adenoviral protein. Apoptosis 8, 199–208 (2003). https://doi.org/10.1023/A:1022931028235
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DOI: https://doi.org/10.1023/A:1022931028235