Human Lymphocyte Chromosomes Research Articles

Early and later replication patterns of increased resolution in human lymphocyte chromosomes.

High resolution patterns of DNA replications in human lymphocyte chromosomes during early and late S-phases were studied by means of the BrdU-Hoechst-Giemsa technique. The late replicating bands were found to be identical with highly detailed G-bands. Between early replicating bands and R-bands subtile differences were observed. A possible correlation between a replication band seen on the chromosomal level and a replication cluster observed after fiber autoradiography is discussed.

Heterogeneity of human chromosome 9 constitutive heterochromatin as revealed by sequential distamycin A/DAPI staining and C-banding.

Distamycin A/DAPI staining and sequential C-banding of human lymphocyte chromosomes reveals the regular occurrence of differentially staining subfractions of chromosome 9 constitutive heterochromatin. These subfractions are regionally organized as two subsegments: a distal one, which fluoresces brightly with DAPI after preincubation with distamycin A and a proximal one, which stains intensely with Giemsa after sequential C-banding. Observations are presented that indicate an occasionally independent genetic behavior of these heterochromatin subfractions.

Combined action of isoniazid and para-aminosalicylic acid in vivo on human chromosomes in lymphocyte cultures

Two antitubercular drugs, viz., isoniazid (INH) and para-aminosalicylic acid (PAS), in combination, were evaluated for their in vivo clastogenic effects of human lymphocyte chromosomes. Lymphocyte cultures from tuberculosis patients taking a therapeutic dose of INH and PAS for a period of not less than 3 months and from two sets of controls were used: (1) newly diagnosed tuberculosis patients who were not yet under therapy and (2) healthy individuals from the general population. Chromosome aberration frequency was very significantly increased in the patients exposed to combined INH and PAS therapy as compared with controls. The most frequently observed aberrations were chromatid breaks and gaps. Isoniazid, the major antituberculosis drug, has been reported not to be clastogenic by itself. However, we observed that the INH--PAS combination commonly used in therapy was clastogenic. From this observation it may be concluded that INH and PAS act synergistically in producing chromosomal aberrations.

Binding of lectins to mitotic chromosomes and interphase nuclear substructures

We have investigated binding of the lectins wheat germ agglutin (WGA) and concanavalin A (Con-A) to mitotic chromosomes and interphase nuclear substructures using fluorescein and rhodamineconjugated lectins. Standard cytogenetic preparations of human lymphocyte chromosomes were incubated with WGA. Mitotic chromosomes and interphase nuclei are brightly fluorescent. Con-A did not detectably bind chromosomes prepared in this way. Mitotic chromosomes display a distinctive banding pattern with WGA similar to quinacrine banding. We also studied binding to interphase “nuclear scaffolds”, which are prepared by treating isolated Hela cell nuclei with DNAse l and high salt. WGA binds nuclear scaffolds internally while Con-A binds to the periphery. The specific binding of lectins to chromosomal structures derived from these two different methods indicates that glycosides are true chromosomal components and that glycoproteins may have a role in chromosome organization.

Recovery kinetics of radiation-induced chromosome aberrations in human G0 lymphocytes.

Declining yields of radiation-induced dicentric chromosomes in human G0 lymphocytes were observed in split-dose experiments with time intervals varied up to 8 h. In agreement with microdosimetric intratrack-intertrack interaction models, only the dose-squared yield component was reduced and approached an asymptotic value equal to one half of the corresponding single exposure yield. For 150 kV X-rays and 13 MeV electrons, at total doses up to 6 Gy, the time constant tau of the approximately exponential decline was practically dose- and quality-independent within a range of 100-180 min. For 10 kV X-rays, in the presence of a dominant linear yield component, only a small split-dose effect, but with a consistent tau-value, was observed for a total dose of 5 Gy. Since tau can be interpreted as the mean life time of "primary lesions" in chromatin fibres, its independence from absorbed dose and radiation quality means that radiation damage of the split-dose recovery mechanism can be excluded for doses up to 6 Gy. By correlating the observed split-dose reduction of the acentric fragment yield to the reduction of the dicentric yield, (1.64 +/- 0.03) acentrics/dicentric for 150 kV X-rays and (1.51 +/- 0.11) acentrics/dicentric for 13 MeV electrons were obtained. Acentrics formed in the course of dicentric formation as well as in other binary interactions of "primary lesions" are represented in these ratios. Post-irradiation recovery during time intervals between irradiation and cell stimulation up to 24 h did not occur. The relations to comparable results in cell lethality experiments are discussed, and a hypothesis of "fast" and "slow" binary interactions of "primary lesions" is put forward.

Effects of two cytostatics (degranol and bleomycin) on the chromosomes of human lymphocytes in culture

Effects of two cytostatics (degranol and bleomycin) on the chromosomes of human lymphocytes in culture

Involvement of sulfhydryl groups of chromosomal proteins in sister chromatid differentiation.

The fluorescence of human lymphocyte chromosomes stained with sulfhydryl group-specific fluorochromes is markedly enhanced by a mild near-ultraviolet irradiation pretreatment, indicating breakage of protein disulfide bonds. When metaphase preparations of cells cultured in the presence of BrdU during two cell cycles are irradiated and subsequently stained with the sulfhydryl group-specific fluorescent reagents used in this study, a differential fluorescence of sister chromatids is observed. After staining with the DNA-specific fluorochrome DAPI an opposite pattern of lateral differentiation appears. It can be concluded that the chromatid containing bifilarly BrdU-substituted DNA has a higher content of sulfhydryl groups than the chromatid containing unifilarly BrdU-substituted DNA. This implies a more pronounced effect of breakage of disulfide bonds in the chromatid with the higher degree of BrdU-substitution. BrdU-containing chromosomes pretreated with the mild near-ultraviolet irradiation procedure used by us, do not show any differentiation of sister chromatids after Feulgen staining. Using sulfhydryl group-specific reagents, differential fluorescence of sister chromatids could still be induced by irradiation with near-ultraviolet light after the complete removal of DNA from the chromosomes by incubation with DNase I. Thus, the protein effect of irradiation of BrdU-containing chromosomes takes place independently of what occurs to DNA.

A method for determining the statistical significance of mutation frequencies

AbstractA method is devised for determining the statistical significance of the difference between a mutation frequency observed after treatment with a potential mutagen and its control frequency. This procedure, however, can be used only when the respective control value is based on large and reliable material (“stable” controls), a requirement that seems to be met in at least part of the systems used for mutagenesis screening. The procedure proposed has some advantages when compared with the 2 and the KASTENBAUM‐BOWMAN test, the most important one being that it can be employed when these two tests cannot (because of expected frequencies <5 in the 2 test and limited tabulations of the N and p values in the KASTENBAUM‐BOWMAN test). After explaining the procedure, an instruction how to carry it out is presented and illustrated by two examples (recessive sex‐linked lethal mutations in Drosophila melanogaster and dicentric chromosomes in human lymphocytes).

Nonrandom distribution of chromosomal aberrations induced by three chemicals

The G-band locations of 3244 breakpoints induced by cis-platinum (II) diamminedichloride (PDD), 1460 breakpoints induced by cytosine arabinoside (ara-C), and 1257 breakpoints induced by triethylenemelamine (TEM) in human lymphocyte chromosomes were identified. The breakpoints induced by each of these chemicals demonstrated a significantly nonrandom distribution within the human karyotype. The overall pattern of the interarm distribution was dependent upon the chemical used, but certain chromosome arms exhibited similar responses to all 3 chemicals. Comparison of the frequencies of breakpoints within individual G-bands indicated that (1) certain bands were susceptible to damage induced by all 3 chemicals; (2) certain bands were resistant to damage by all 3 chemicals; (3) certain bands demonstrated variable susceptibility to induced damage dependent upon the chemical agent; and (4) other bands demonstrated near expected frequencies of damage (by length) to all 3 agents.

Simultaneous staining of sister chromatid exchanges and Q-bands in human chromosomes after treatment with methyl methane sulphonate, quinacrine mustard, and quinacrine.

Human peripheral lymphocyte chromosomes were stained simultaneously for sister chromatid exchanges (SCEs) and Q-banding. No effect of treatment with MMS, QM, and Q on the distribution of SCEs in chromosomes was found compared with controls. The SCEs were distributed between chromosomes roughly according to metaphase length, with the shorter chromosomes underrepresented. The majority of SCEs were located to pale bands, while a few occurred in bright bands and at interfaces between pale and bright bands. A greater frequency than expected of SCEs had occurred at identical sites in homologous chromosomes. This frequency was significantly increased after treatment with MMS.

Sensitivity of Human Peripheral Lymphocyte Chromosomes to Various X-ray Doses and Subsequent Storage in Plexiglass or Glass Containers

Sensitivity of Human Peripheral Lymphocyte Chromosomes to Various X-ray Doses and Subsequent Storage in Plexiglass or Glass Containers

Bleomycin and sister-chromatid exchange in human lymphocyte chromosomes

Bleomycin and sister-chromatid exchange in human lymphocyte chromosomes

Sister-chromatid exchange induced by X-ray of human lymphocytes and the effect of l-cysteine

A staining technique that detects sister-chromatid exchanges (SCEs) has been used to examine the response of human lymphocyte chromosomes to various dosages of X-irradiation. The SCE frequency was markedly increased following irradiation. However, the increase was of a significantly smaller magnitude when irradiation occurred in the presence of an antimutagenic agent. Scoring SCEs may provide a useful technique for assaying the mutagenic effects of environmental carcinogens as well as the protective effects of antimutagenic agents.

The effects of busulphan on the chromosomes of normal human lymphocytes

In vitro exposure of human lymphocytes to busulphan (BUS) produced an increase in chromosome aberrations and in sister-chromatid exchange (SCE) frequency. The distribution of chromosome breaks throughout the karyotype was non-random and they occurred mainly in the G-negative bands. Certain bands had a marked susceptibility to BUS and comparisons with the human chromosome-break distributions reported for a number of drugs revealed that some of these bands were equally susceptible to other alkylating agents. Both the number of chromosome gaps and breaks and the SCE frequency increased with BUS concentration, but only the SCE dose-response was a clearly defined linear relationship. Therefore a standard SCE dose-response curve was constructed for future comparison with the results of similar investigations of patients on BUS therapy.

Localization of γ-rays induced chromatid breaks using a three consecutive staining-technique

Three staining techniques (Giemsa, Q-banding and R-banding) are used consectively to localize the breakage points in chromosomes of human lymphocytes, irradiated during G 2-phase with γ-rays, at doses ranging from 50 to 200 rad. The large majority, about 85% of the breaks, occurs at the interbands, between R- and Q-bands. The discrepancy of this result, with regard to previously reported ones, is attributed to the strong bias of analysis when only one staining technique is used.

The effects of busulphan on the chromosomes of normal human lymphocytes

In vitro exposure of human lymphocytes to busulphan (BUS) produced an increase in chromosome aberrations and in sister-chromatid exchange (SCE) frequency. The distribution of chromosome breaks throughout the karyotype was non-random and they occurred mainly in the G-negative bands. Certain bands had a marked susceptibility to BUS and comparisons with the human chromosome-break distributions reported for a number of drugs revealed that some of these bands were equally susceptible to other alkylating agents. Both the number of chromosome gaps and breaks and the SCE frequency increased with BUS concentration, but only the SCE dose-response was a clearly defined linear relationship. Therefore a standard SCE dose-response curve was constructed for future comparison with the results of similar investigations of patients on BUS therapy.

Cytogenetic effects of styrene and styrene oxide on human lymphocytes and Allium cepa.

Styrene and styrene oxide induce cytogenetic effects already at very low concentrations (0.01% v/v or even less); the effects are similar in both in vitro human lymphocytes and in vivo onion root tip cells (Allium cepa L.). It is characteristic that styrene treatment is more potent in causing chromosome breakage in both systems. In Allium styrene induced inhibition of mitotic spindle action as revealed by a strong c-mitotic effect. Also the number of micronuclei and nuclear bridges increased in both test systems, especially after styrene oxide treatment. Furthermore, the metaphase chromosome morphology in the cells treated with styrene oxide was strongly affected. In both systems, chromosome destruction was observed, or else the chromosome material was decondensed and resulted in a characteristic fuzzy appearance of Allium chromosomes or a banded appearance of human lymphocyte chromosomes. A specific effect of styrene oxide on the chromosomal proteins is thus suggested. The data obtained from the autoradiographic studies with Allium support the idea that [7--3H] styrene oxide binds irreversibly to the cytoplasmic and nuclear macromolecules.

Quantitative distribution of satellite associations with different numbers of chromosomes in human lymphocytes

Statistical analysis of the quantitative distribution of associations formed by 2, 3, 4, 5, and 6 acrocentrics (associations differing in numerical composition) revealed good agreement between the empirical frequencies and the theoretical values calculated by the exponential law of distribution to base 2. The high agreement in all donors tested points to the strict regularity of the quantitative distribution of associations with different numerical values and their special role in human lymphocytes.

The RBE of 30 kV X-rays for the induction of dicentric chromosomes in human lymphocytes.

The frequency of dicentric chromosomes induced by the irradiation of human lymphocytes in Go phase was determined with hard (150 kV) and soft (30 kV) X-rays. When a linear-quadratic dose-effect relation was used to fit the experimental data, a significant linear contribution was found for 30 kV X-rays. The RBE of 30 kV compared with 150 kV X-rays approaches the value 3 at a 30 kV X-rays dose of 20 rad and decreases with increasing dose. From the results it may be concluded that chromatids with primary breaks, undergoing second order reactions and thus forming dicentric chromosomes, are produced by traversals of low-energy electrons through chromatin material.

Marijuana exposure in vivo and human lymphocyte chromosomes

Sequential chromosome examinations of peripheral lymphocyte cultures were carried out on 21 adult male volunteers who smoked natural blend marijuana cigarettes containing about 1%, 2%, or no Δ 9-THC. For a limited number of subjects, blood samples from a single venipuncture were cultured independently in two cytogenetic laboratories, and later the slides were exchanged for re-analysis. There were significant differences between laboratories in the absolute break frequencies recorded. These inter-laboratory differences were demonstrated for both techniques of cell culture and metaphase analysis. Neither laboratory found a statistically significant increase in break frequencies associated with marijuana smoking. The present study, therefore, failed to detect a measurable effect of marijuana smoking on chromosomal aberrations in subjects experienced in the use of the drug.