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J R Savage

Publications and source records attributed to J R Savage.

At least 19 recordsLinked to original sources
PubMed1992

The effect of X-irradiation on cell cycle progression and chromatid aberrations in stimulated human lymphocytes using cohort analysis studies.

Stage sensitivity for the production of chromatid-type aberrations and mitotic delay has been investigated in a stimulated human lymphocyte population, following an absorbed dose of 1.5 Gy 250 kVp X-rays. BrdU replication banding was used to obtain a fine analysis of the cell cycle and to permit cohort analysis. Fluctuations in yield with sample time were found for all aberration categories, but these could not be related simply to either the developmental stage of the cells at time of exposure, or to the time-to-run to metaphase. In general G2 and late S cells had higher aberration yields than early S and pre S cell populations. Mitotic delay and perturbation at this dose extends to all sub-phases of S and is as great, if not greater, in the earliest S cells as it is in G2.

Cells, Cultured↗
PubMed1991

A BrdU pulse double-labelling method for studying adaptive response.

We describe a pulse BrdU double-labelling method which we have used for a cohort-analysis study of a conventional adaptive response experiment in stimulated human lymphocytes. This method allows us to identify the cycle position occupied by each scored metaphase at the time of primer and at the time of challenge. Responses in cells at defined developmental stages can then be compared irrespective of mitotic perturbation. Using a primer X-ray dose of 1.0 cGy, and a challenge dose of 1.5 Gy 6 h later, we found a transitory depression in frequency of total aberrations present in cells sampled at 6 h but not at 9 h after challenge. When defined cohorts were summed for both sample times, no significant adaptive response was observed. A repeat experiment where terminal BrdU labelling was used after the challenge failed to provide any positive response at either 6 or 9 h. The technique presented not only ensures that similar cell mixtures from a heterogeneous, asynchronous population can be scored and compared, but it serves also to demonstrate the kinetic complexity of stimulated lymphocyte cultures.

Adaptation, Physiological↗
PubMed1990

BrdU pulse/reverse staining protocols for investigating chromosome replication.

By using a reverse Giemsa staining procedure (TT chromatin pale, TB chromatin dark) it is possible to detect replication in metaphase chromosomes with short (approximately 10 min) 5-bromodeoxyuridine (BrdU) pulses. A pulse protocol allows us to consider the question "What is replicating at this point in time?" and we have investigated replication patterns during cycle transit in stimulated human female lymphocytes. A clear-cut demarcation between R-zone early and G-zone late was not found. Instead, whilst replication commences (with a very staggered start) in R-zones, activity soon appears to transgress band boundaries and gives rise to cells with unclassifiable patterns where chromosomes take on a mottled or reticulate appearance. Replication in R-zones dies out leaving a clear G-zone pattern persisting for the remainder of S which terminates with a very staggered finish. When pulse duration is increased (approximately 1 h) the frequency of unclassifiable cells falls and occasional "mixed-pattern" cells appear which have, within the same cell, typical R- and G-zone regions. The existence of such cells indicates that if a mid-S replication pause exists (and the absence of any mid-S wave of pale stained cells suggests that it does not) it does not make exclusive separation between dark R- and G-band zones.

Bromodeoxyuridine↗
PubMed1990

Two additional cases of t dic(9:12) in acute lymphocytic leukemia (ALL): prognosis in ALL with dic(9:12).

We report two occurrences of dic(9;12) in acute lymphoblastic leukemia and review previous cases. Cases of dic(9;12) share common features with cases of 9p and 12p rearrangements, but prognosis seems particularly good in cases of dic(9;12). The persistence of a specific dicentric in stable clones is remarkable and points to unusual centromeric behavior and/or marked selective advantage of the anomaly.

Adult↗
PubMed1989

The production of chromosome structural changes by radiation.

This paper attempts an update and comment upon some of the topics of chromosome aberration formation which Lea raised in Chapter VI of his classic work 'Actions of Radiations on living Cells'. Only the first nine sections of this chapter are covered, which deal primarily with the qualitative aspects of aberrations, their formation, classification and interrelationships. In commenting upon these topics, pertinent references are made to work with mammalian and human cells. Increased knowledge of the importance of DNA as a fundamental target and the integral part it plays in the complex structure of the chromosome, coupled with cellular techniques not available to these earlier workers necessitate some revision and modification of early ideas. However, inspite of the enormous accumulation of data and ideas since the original work was published in 1946, the foundation that these early workers laid is still very solid. Surprisingly, we are still puzzled by many of the problems that perplexed them.

Animals↗
PubMed1989

Production of chromosome aberrations, micronuclei, and sister-chromatid exchanges by 24-keV epithermal neutrons in human G0 lymphocytes.

The induction of chromosome aberrations, micronuclei and sister-chromatid exchanges in human G0 lymphocytes by 24-keV epithermal neutrons has been measured. Positive linear dose responses were obtained for the 3 end points, with a tendency to saturation at higher dose for SCE production. In all cases, the responses to 24-keV neutrons were characteristic of high-LET radiations.

Chromosome Aberrations↗
PubMed1989

A pulse BrdU method for SCE.

Using 'reverse' harlequin staining (bromouracil-substituted chromatin staining dark), it is possible to detect at metaphase a pulse of bromodeoxyuridine (BrdU) incorporated during S-phase. Provided that this pulse is of reasonable duration, fairly uniform staining along the chromatids of some S-cells is achieved, and no difficulty is encountered in observing and scoring SCE in such cells at second division. Thus, it is possible to define within an asynchronous population a narrow cohort of target cells and recover these for SCE scoring at second division irrespective of treatment induced perturbation. This serves to reduce the heterogeneity found in the usual terminal BrdU SCE protocols for such populations and should lead to more reliable and repeatable quantitative results. The method is illustrated for mitomycin C given to dividing human blood lymphocytes using both simultaneous and delayed pulse modes.

Bromodeoxyuridine↗
PubMed1988

A method for comparing DNA replication programmes at the level of the chromosome bands.

During DNA synthesis, each pair of homologous chromosomes replicates its bands in a precise order and at a specific time. When using asynchronous cell populations, this replication programme has to be reconstructed from a series of "stills"--serial samples taken at intervals through S-transit. Obviously, the result obtained is dependent upon the kinetic progression of cells through the cycle, and any perturbation of the cycle. A difficulty arises when we wish to compare the replication programme of a chromosome in two different cultures (e.g. cells from different origins, or after different treatments). Kinetic differences between cultures make it almost impossible to obtain two samples for analysis containing the same "mixture" of cells. Thus, a false programme difference could be introduced, or a real one masked. In this paper, we present a method of comparison that overcomes this problem. It is based upon the observation that with serial sampling of steady-state cell populations through S-phase, band appearance curves are sigmoidal and are very well approximated by cumulative Normal distributions with very similar standard deviations. If a family of such curves, closely spaced in time, is sampled twice, the two observed frequencies for each curve are related, their probits all lying on a single straight line. This line has a slope of 45 degrees and its displacement from the origin is a function of the time interval between the two samples. Given two identical families of such curves, exactly the same relationship will hold if one sample is drawn from each. If, however, the two families differ (in order, spacing, standard deviation etc.), the probit/probit plot will deviate in various ways from a straight line with 45 degrees slope. Any two subsets of chromosome replication-band frequencies can be regarded as derived from a family of cumulative Normal curves and probit/probit comparisons used to test the similarity of their replication programmes.

Ataxia Telangiectasia↗
PubMed1988

Replication kinetics of X chromosomes in fibroblasts and lymphocytes.

The kinetics of replication for early and late replicating X chromosomes in karyotypically normal fibroblasts and lymphocytes was studied using terminal bromodeoxyuridine (BrdU) treatment followed by Hoechst/light/Giemsa staining. Although the order of band appearance differs between the two tissues, the programme (order and interval between band appearances) for early replicating bands (dark R-bands) is identical in the two homologues. This is probably also the case for later replicating bands (dark G-bands) though the criteria for determining mean band appearance times are less reliable for these bands when terminal BrdU treatment is used. This means that the late X has a delayed start but thereafter proceeds at the same pace as its early counterpart.

Bromodeoxyuridine↗
PubMed1988

The behaviour of fragile X and other aberrations during recovery from low folate conditions.

Whole blood from two mentally retarded fra-X brothers was grown in low folate medium where fra-X expression was enhanced. Bromodeoxyuridine was added to mitigate the low folate conditions and metaphases were sampled sequentially, and stained for replication banding, through one cell cycle of recovery. The replication bands allowed detailed analysis of the cell cycle and the allocation of individual cells to precise sub-phases. Various classes of fra-X and all other types of chromosomal aberrations were scored in these classified cells. The fra-X does not conform in morphology to any of the known simple chromatid intrachange types, which were often present within the same cells, but the subsequent fall in frequency once bromodeoxyuridine was added closely paralleled that of the conventional aberrations. Normal folate level frequencies of fra-X are restored by the time early S-phase cells (sub-phase SkI) reach metaphase. When sub-phased cells are rearranged in true chronological sequence, there is a suggestion of a sudden fall in frequency between SkII-III (about 70% of the transit of S). This suggests that the critical point for low folate enhancement occurs in this region of the S-phase. This is somewhat earlier than the band-appearance distribution curve for Xq27 which lies within sub-phase SkIV.

Bromodeoxyuridine↗
PubMed1988

Generalized blocking in S phase by methotrexate.

An attempt was made to enhance the frequency of prometaphase cells for high-resolution-banding studies in untransformed Syrian hamster fibroblasts using a typical methotrexate (MTX) block/bromodeoxyuridine release schedule. The recovery 'wave' was serially sampled and detailed sub-phase analysis made using the replication bands resulting from bromodeoxyuridine uptake. Of the 3 batches of MTX used, one (Sigma greater than 2 years old) was found to have decayed to a non-toxic compound which produced almost no measurable perturbation of the cell cycle at any concentration used. The other 2 (new Sigma and new Lederle), whilst producing mitotic index fluctuations which could be construed to indicate blocking and "synchrony", gave absolutely no evidence of any specific blocking site, but rather a general stoppage (or slowing down) during MTX treatment and continuation in exactly the same order as untreated controls upon release.

Animals↗
PubMed1988

Induction of sister chromatid exchanges (SCE) in G0 lymphocytes by plutonium-238 alpha-particles.

Irradiation of human G0 lymphocytes with plutonium-238 alpha-particles and X-rays was performed to investigate the production of sister chromatid exchanges (SCE). Alpha-particles produce a significant increase in SCE and this elevation is more significant when separated lymphocytes are irradiated. X-ray irradiation did not induce any significant increase in SCE. Therefore the relative biological effectiveness (RBE) for the induction of SCE by alpha-particles in this system is undefined and effectively infinite.

Alpha Particles↗
PubMed1988

Karyotype evolution in the bone marrow of a patient with Fanconi anemia: breakpoints in clonal anomalies of this disease.

A 21-year-old Fanconi anemia patient developed refractory anemia. Laboratory studies revealed a transitory increased platelet count and a typical del(5q). Bone marrow karyotyping showed a -6, +der(6)t(1;6)(q12;p25) rearrangement and, two years later, a mosaic -6, +der(6),t(1:6)(q12;p25)/-2, +der 2), t(1;2)(q12;q37) constitution. The chromosome mechanism operating in this patient is discussed.

Adult↗