Spontaneous Apoptosis of Thymocytes Is Uncoupled with Progression through the Cell Cycle |
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Authors: | C. Pellicciari M. G. Bottone V. Schaack S. Barni A. A. Manfredi |
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Affiliation: | aDipartimento di Biologia Animale and Centro di Studio per l'Istochimica del CNR, Piazza Botta 10, I-27100, Pavia, Italy;bLaboratorio di Immunoterapia Adottiva, H. S. Raffaele, Via Olgettina, 69, I-20133 Milan, Italy |
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Abstract: | Most developing lymphocytes spontaneously die in the thymus during positive and negative selection of the T cell repertoire. By evaluating the expression of the proliferation antigens Ki-67 and PCNA, we demonstrated here that more than 95% of thymocytes are potentially proliferating. The coincidence within the same cell population of death and proliferation is thus apparent in developing thymocytes. Using dual-parameter cytometric techniques to evaluate in single cells the amount of DNA versus light-scattering values, we found that spontaneous thymocyte apoptosis occurs with similar frequency in all the cycle phases, whereas apoptosis induced by the anti-topoisomerase-II, etoposide (which is the consequence of irreversible DNA damage), takes place with higher frequency in S and G2phases (i.e., in those cycle phases in which DNA is subjected to torsional constraints). The capability of thymocytes to enter apoptosis was also monitored by digesting DNAin situwith DNase I (a nuclease that cleaves DNA mimicking the nuclear damage common to most apoptotic suicides). We also show that endonuclease-mediated DNA digestion occurs to a similar extent in cells with different DNA contents, i.e., in cycle phases in which the superstructural organization of chromatin is markedly different. |
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