Analysis by FISH of the spectrum of chromosome aberrations induced by X-rays in G0 human lymphocytes and their fate through mitotic divisions in culture

Abstract

The induction, distribution, and persistence of chromosome aberrations in human lymphocytes exposed to X‐rays in G0 were analyzed in 48‐, 70‐, and 94‐hr cultures by conventional metaphase analysis and painting of chromosomes 1, 2, and 4 by FISH. All cells that had been scored by FISH were relocated to determine by differential staining of chromatids whether they had passed through 1, 2, or ≥3 divisions. FISH revealed a dose‐dependent induction of stable and unstable aberrations, while chromatid labeling showed mitotic lag caused by irradiation in G0. Relative to their DNA contents, there was a small but significant overrepresentation of chromosome 4 and underrepresentation of chromosome 2 among the aberrations involving chromosomes 1, 2, and 4. FISH slightly underestimated the genomic frequency of unstable aberrations measured by conventional metaphase analysis. There was a slight excess of translocations relative to dicentrics, but the data are compatible with the 1:1 ratio expected from cytogenetic theory. Many of the translocations were apparently incomplete (i.e., nonreciprocal). Incomplete translocations were more frequent at higher X‐ray dose and in first division, suggesting that they may be associated with complex damage and are more apt to be lost in mitosis than complete translocations. Among the incomplete translocations, t(Ab) outnumbered t(Ba) — a difference ascribable to the FISH technique. Aberration frequencies declined as the cells divided in culture. The overall decline in the frequency of aberrant cells (≈29% per cell generation) reflects a rapid decline in dicentrics and fragments (≈60% per cell generation) and the relative stability of translocations. The frequency of translocation‐bearing cells underwent a modest decline in culture (≈13% per cell generation). Environ. Mol. Mutagen. 33:94–110, 1999 © 1999 Wiley‐Liss, Inc.

Transmission of Induced Chromosomal Aberrations through Successive Mitotic Divisions in Human Lymphocytes after In Vitro and In Vivo Radiation

“…Using TC staining and extended lymphocyte culture times, we demonstrate, for the first time, that the yield of DCs decreased 50.12% between M1 and M2, much closer to the theoretical rate (50%) 13, 31 than found in all other studies 15, 17, 20, 32 . Extended lymphocyte culture times permitted to resolve of, not only the mitotic lag of exposed lymphocytes observed at 50 h, but also the difference in the proliferation indices between the three donors.…”

“…The scoring of CA for the estimation of the dose received, an important parameter for estimating the associated risk, is a powerful approach, in particular in the absence of or the impossibility to perform physical dosimetry 10 . The transmission of these aberrations has been previously investigated using conventional cytogenetic techniques (FPG) 13–17, 19, 30 and FISH 14, 20, 21 . However, investigation of the transmission of ACs was performed using conventional cytogenetic techniques (uniform staining) without any distinction between the different types of ACs, and their transmission remains unclear.…”

FISH-based analysis of 10- and 25-kV soft X-ray–induced DNA damage in 184A1 human mammary epithelial cells

“…In addition, Virsik-Peuckert et al (1997) investigated human fibroblasts and demonstrated a translocation to dicentric ratio different from 1. Several reasons given by the different authors are disparities of the repair processes (Virsik-Peuckert et al 1997), misscoring of dicentrics without the application of centromere-specific markers (Stephan and Pressl 1997) or the influence of scoring criteria (Hoffmann et al 1999). Additionally, the method of three colour FISH, as applied in the present work, might lead to the misscoring of complex chromosome aberrations as pseudo-simple exchanges, which might affect the exchange ratio.…”

“…For other cell lines, the published data are not consistent showing both verifications (Hoffmann et al 1999;Stephan and Pressl 1997;Braselmann et al 2003) and deviations (Knehr et al 1996;Virsik-Peuckert et al 1997;Barquinero et al 1999) from the expected ratio for human lymphocytes. In addition, Virsik-Peuckert et al (1997) investigated human fibroblasts and demonstrated a translocation to dicentric ratio different from 1.…”

Induction and Persistence of Multicentric Chromosomes in Cultured Human Peripheral Blood Lymphocytes Following High-Dose Gamma Irradiation

“…The intricate effects of processes associated with interphase cell death and cell-cycle progression to mitosis pose difficulties in examining metaphase cells for accurate dose assessment (Belloni et al 2008). Higher dicentric frequencies observed in cells within the same post-irradiation cell division derived from an extended culture time indicate either a delay in cell cycle progression of aberrant cells or the presence of different lymphocyte subpopulations with different radiosensitivities (Lloyd et al 1977, Boei et al 1996, Hoffman et al 1999, Pala et al 2001, Pujol et al 2012. To overcome technical difficulties related to cell proliferation kinetics, a recent study (Pujol et al 2012) has shown a method that yields a high proportion of mitotic cells at 48 h which was realized by blocking the G2/M cell-cycle checkpoint by cotreatment with caffeine and colcemid.…”