Absence of chromosome breakage in patients with retinoblastoma

Summary Mixed lymphocyte cultures were employed to assess the degree of spontaneous chromosome fragility in patients with retinoblastoma. There was no difference between the patients and their controls. If chromosome instability plays a role in the inherited tumour, more sensitive methods need be employed to elucidate it.     

The rate of chromosome breakage is age dependent in lymphocytes of adult controls

Summary Chromosome breaks and chromatid-type lesions from a prospective study of more than 1000 lymphocyte karyotypes from each of six controls were analysed. These lesions were more frequent in older (75 years old on average) than in younger (29 years old on average) controls, especially after 72h cultures. All controls were found to be carriers of fragile sites. The most frequent were 3p14.3 and 16q23, especially in older controls. At least one fra(X)(q27) mitosis was found in each control. Most deletions occurred after breakage in heterochromatin or in late-replicating euchromatin. As almost all radials were either mitotic chiasmata or triradials (branched chromosomes), it is concluded that chromatid exchanges between non-homologous segments are very rare, and indicate chromosomal instability syndrome or recent exposure to a mutagen.     

Folic acid and chromosome breakage III. Types and frequencies of spontaneous chromosome aberrations in proliferating lymphocytes

The types and frequencies of spontaneous chromosome aberrations were studied in human lymphocytes cultured for 96 h in minimal essential medium (MEM) or MEM without folic acid (MEM-FA). In both media, the most frequent aberrations were chromatid gaps, isochromatid gaps and chromatid breaks. Chromosome (isochromatid) breaks and dicentrics were seen less frequently. Neither of these less frequent aberrations was seen in 4000 cells from MEM, but both were seen in 4000 cells from MEM-FA.     

Folic acid and chromosome breakage. III. Types and frequencies of spontaneous chromosome aberrations in proliferating lymphocytes

The types and frequencies of spontaneous chromosome aberrations were studied in human lymphocytes cultured for 96 h in minimal essential medium (MEM) or MEM without folic acid (MEM-FA). In both media, the most frequent aberrations were chromatid gaps, isochromatid gaps and chromatid breaks. Chromosome (isochromatid) breaks and dicentrics were seen less frequently. Neither of these less frequent aberrations was seen in 4000 cells from MEM, but both were seen in 4000 cells from MEM-FA.     

Distribution of chromosome breakage point induced by arabinosylcytosine on in-vitro human lymphocytes]

The cytogenetic effects of arabinosyl cytosine on human leukocytes in vitro has been studied using continuous and discontinuous treatments. Optimum activity occurs at the end of the S period. Interchromosomal distribution of breaks is non-random. The hypothesis of a very late replication for some chromatid segments is discussed. The relations between banding and the position of breaks are discussed.     

The somatostatin analogue peptide TT-232 induces apoptosis and chromosome breakage in cultured human lymphocytes

Somatostatin receptors are supposed to be important in the regulation of apoptosis. In this study, we measured apoptosis occurring spontaneously, or induced by the synthetic somatostatin analogue, the peptide TT-232. We examined isolated human peripheral blood lymphocytes (PBL) from 32 nurses exposed bedside to cytostatic drugs, 12 chronic lymphoid leukaemia (CLL) patients prior to treatment, and 19 unexposed, healthy donors without anamnestic occupational exposure to genotoxic agents. Cells were stimulated by phytohaemagglutinin-P (PHA) and cultured for 69 h with or without 15 microg/ml TT-232, respectively. Cell kinetic parameters and apoptosis were determined by flow cytometry after staining with FITC-labeled anti-BrdU and propidium iodide (PI) and the results on spontaneous and peptide-induced apoptosis were compared with the obtained chromosome aberration frequencies (CA). The peptide TT-232 unexpectedly induced chromosome breakage in addition to apoptosis. The mean spontaneous apoptotic fractions were 6.65+/-0.89%, 6.46+/-0. 53%, and 3.07+/-0.57%, and the mean CA yields in the samples without TT-232 were 1.74+/-0.46%, 2.44+/-0.40%, and 4.50+/-1.05%, for healthy subjects, nurses, and CLL patients, respectively. A total of 15 microg/ml TT-232 treatment in healthy subjects increased the mean CA frequency (10.38+/-1.57%), as well as the apoptotic cell fraction (2.63+/-0.45 times higher than the corresponding untreated sample). In TT-232-treated PBLs of nurses, CA remained unchanged and the mean apoptotic cell fraction showed only a slight increase (1.24+/-0.11 times higher than the untreated). Among CLL patients, TT-232 treatment significantly increased both CA (up to 17.83+/-4.04%) and the ratio of apoptotic cells (21.78+/-11.00 times higher than the untreated). These results demonstrated significant differences in apoptosis sensitivity in controls, nurses and CLL donors, after 15 microg/ml TT-232 treatment. Data also indicate that the induced CA yields in CLL donors with high CA are in correlation with TT-232-induced apoptosis.     

Normal red blood cells partially decrease diepoxybutane-induced chromosome breakage in cultured lymphocytes from Fanconi anaemia patients

Objectives: Fanconi anaemia (FA) is a cancer‐prone chromosome instability syndrome characterized by hypersensitivity to DNA cross‐linking agents, such as diepoxybutane (DEB). Previous studies have shown that normal red blood cells (RBC) can protect cultured lymphocytes against chromosomal breaks induced by DEB. The present study was designed to analyse influence of RBCs from normal individuals on frequency of DEB‐induced chromosome breaks in lymphocyte cultures from FA patients. Materials and methods: A comparative study was performed between DEB‐induced chromosome breaks in cultures of FA lymphocytes with either autologous or heterologous RBCs. A further comparative study was carried out between whole blood cultures from FA patients performed on two occasions, before and 1 week after transfusion of RBCs. Results: It was observed that normal RBCs compared to FA RBCs, partially reduced chromosome breaks in cultured FA lymphocytes. A significant reduction in DEB‐induced breaks was also observed in FA cultured lymphocytes obtained 1 week after transfusion of RBCs, in comparison to those observed in the same patients before RBC transfusion. Conclusions: This study shows that DEB‐induced chromosome instability in FA lymphocytes is partially reduced by normal RBCs. This effect may have some clinical relevance in vivo, whenever FA patients receive a RBC transfusion.     

The fragile breakage versus random breakage models of chromosome evolution

For many years, studies of chromosome evolution were dominated by the random breakage theory, which implies that there are no rearrangement hot spots in the human genome. In 2003, Pevzner and Tesler argued against the random breakage model and proposed an alternative “fragile breakage” model of chromosome evolution. In 2004, Sankoff and Trinh argued against the fragile breakage model and raised doubts that Pevzner and Tesler provided any evidence of rearrangement hot spots. We investigate whether Sankoff and Trinh indeed revealed a flaw in the arguments of Pevzner and Tesler. We show that Sankoff and Trinh's synteny block identification algorithm makes erroneous identifications even in small toy examples and that their parameters do not reflect the realities of the comparative genomic architecture of human and mouse. We further argue that if Sankoff and Trinh had fixed these problems, their arguments in support of the random breakage model would disappear. Finally, we study the link between rearrangements and regulatory regions and argue that long regulatory regions and inhomogeneity of gene distribution in mammalian genomes may be responsible for the breakpoint reuse phenomenon.     

The cellular response to chromosome breakage

DNA double-strand breaks (DSBs) are among the most deleterious types of damage that can occur in the genome of eukaryotic cells because failure to repair them can lead to loss of genetic information and chromosome rearrangements. DSBs can arise by failures in DNA replication and by exposure to environmental factors, such as ionizing radiations and radiomimetic chemicals. Moreover, they might arise when telomeres undergo extensive erosion, leading to the activation of the DNA damage response pathways and the onset of apoptosis and/or senescence. Importantly, DSBs can also form in a programmed manner during development. For example, meiotic recombination and rearrangement of the immunoglobulin genes in lymphocytes require the generation of site- or region-specific DSBs through the action of specific endonucleases. Efficient DSB repair is crucial in safeguarding genome integrity, whose maintenance in the face of DSBs involves branched signalling networks that switch on DNA damage checkpoints, activate DNA repair, induce chromatin reorganization and modulate numerous cellular processes. Not surprisingly, defects in these networks result in a variety of diseases ranging from severe genetic disorders to cancer predisposition and accelerated ageing.     

The controlling sequence for site-specific chromosome breakage in Tetrahymena

Site-specific chromosome breakage occurs in many ciliated protozoa during nuclear differentiation. We have determined the cis-acting sequence that controls this process in Tetrahymena thermophila. The Tetrahymena ribosomal RNA gene is bounded by two breakage sites. Injection of this gene into developing macronuclei leads to breakage at these sites. Deletion analysis has localized the sequences essential for breakage to a 28 bp region that includes a 15 bp sequence (Cbs) known to be present in other breakage sites. Insertions of Cbs allow breakage to occur at new sites, which is accompanied by elimination of surrounding DNAs and formation of telomeric sequences, as it is at natural sites. Thus, Cbs is the necessary and sufficient sequence signal for chromosome breakage in Tetrahymena.