Biomedical subjects

J B Little

Publications and source records attributed to J B Little.

At least 145 records · Page 8Linked to original sources

PubMed1989

Retinoic acid inhibits the fixation of initial transformational damage in X-irradiated Balb/3T3 mouse fibroblasts in vitro.

We have examined the effects of all-trans retinoic acid (RA) on confluent holding recovery (cell survival) and on the fixation of initial transformational damage expressed as the ultimate yield of transformed foci following X-irradiation of density-inhibited cultures of Balb/3T3 cells. Non-cytotoxic concentrations of RA suppressed both recovery of potentially lethal damage and neoplastic transformation in a dose-dependent manner when added for 24 h during post-irradiation confluent holding after a dose of 5 Gy. At 100 microM, RA inhibited the fixation of initial transformational damage by 80%. These findings are discussed in terms of the hypothesis that retinoids may allow a selective enhancement of the inactivation of certain irradiated tumor cells in vivo while reducing the risk of secondary malignancies in successfully treated patients.

Animals↗

PubMed1989

In vitro models of carcinogenesis: expression of recessive genes by chromosomal mutations.

There has been considerable recent interest in the mechanisms by which recessive mutations involving cancer genes may be expressed. We have developed an in vitro model to study this phenomenon in an endogenous autosomal gene in human cells. We have analyzed the molecular structural changes that lead to loss of heterozygosity at the thymidine kinase (tk) locus. The results indicate that expression of a recessive allele frequently occurs by loss of heterozygosity at that locus. Over 90% of spontaneous mutants at the tk locus arose by allele loss. The fraction of induced mutants that arose by this mechanism depended upon the inducing agent. Loss of the active tk allele was often accompanied by loss of linked genetic loci on the long arm of chromosome 17. These results suggest that large-scale chromosomal mutations resulting from events such as deletion or mitotic recombination may be an important mechanism for the expression of activated or mutated recessive genes in human cells. Such recessive mutations could involve oncogenes or other growth regulatory genes important in carcinogenesis.

Cell Division↗

PubMed1989

The molecular nature of mutants induced by X rays is altered by the presence of the radioprotector cysteamine.

The molecular structure of mutants induced in human lymphoblast cells by 500 cGy X rays in the presence of the radioprotector cysteamine (25 mM) has been compared with that induced by an equally mutagenic treatment of 150 cGy X rays alone. Sets of mutants at the hypoxanthine-guanine phosphoribosyl transferase locus were analyzed by Southern blot. Of 24 mutants induced by X rays in the presence of cysteamine, 67% exhibited no change in the restriction fragment pattern and thus were defined as point mutations; 8% appeared to be total gene deletions and 25% were partial deletions or rearrangements. In contrast, among 28 mutants induced by X rays alone (Liber et al., Mutat. Res. 178, 143-153 (1987)), 46% were point mutations, while 50% were total gene deletions and only 1 mutant (4%) was a partial deletion or rearrangement. Thus mutants isolated in the presence of cysteamine consisted of more point mutations and partial deletions/rearrangements, and considerably fewer total gene deletions. These results suggest that cysteamine may protect selectively against processes which lead to large-scale molecular changes.

Blotting, Southern↗

PubMed1989

Age-dependent modification by lactate of repair of potentially lethal damage in normal human diploid fibroblasts.

We investigated age-related changes in normal human diploid fibroblasts (strain AG 1522) followed throughout their lifespan in vitro. Radiosensitivity and the capacity to repair potentially lethal radiation damage (PLD) remained constant until senescence was reached. However, the effects of incubation with lactate on PLD repair were age dependent and complex. Low millimolar concentrations of lactate increased the capacity to repair PLD, whereas higher concentrations suppressed it in a concentration-dependent manner. With increasing in vitro age, the cells became less sensitive to the modifying effects of lactate; in presenescent cells, the modifying effects of lactate on PLD repair had completely vanished.

Cell Survival↗

PubMed1988

Persistently elevated sister chromatid exchanges in ethylene oxide-exposed primates: the role of a subpopulation of high frequency cells.

Ethylene oxide (EtO) is a potent DNA-alkylating agent which has been shown to induce sister chromatid exchanges (SCE) in the peripheral blood lymphocytes of exposed workers. To study further the persistence of EtO-induced SCE, we have examined lymphocytes from a group of cynomolgus monkeys exposed to EtO in control, 50-ppm, and 100-ppm concentrations for 7 h/day, 5 days/week over the years 1979-1981. The data collected in 1987 were compared with those generated immediately prior to the cessation of exposure in 1981. EtO-induced SCE persisted at levels significantly above those of the nonexposed controls. Comparison of the distributions of SCE between 1979 and 1987 shows that, although mean SCE decreased from 1981 to 1987, the mean SCE in the top 10% of the distribution has not diminished over time. Consequently, the increased level of SCE is entirely attributable to a subpopulation of cells with high frequencies of SCE. These findings suggest that long-lived lymphocytes may inefficiently repair EtO-induced lesions which produce SCE. The results also have important implications for the proper use of SCE analytical techniques in the epidemiological study of cytogenetic damage after chronic exposure to DNA-alkylating agents.

Animals↗

PubMed1988

Radiosensitivities of ten apparently normal human diploid fibroblast strains to cell killing, G2-phase chromosomal aberrations, and cell cycle delay.

We have examined the sensitivity to X-irradiation of ten apparently normal human fibroblast cell strains received as coded samples. Three different end points were examined: cytotoxicity; induction of chromosomal aberrations after irradiation in G2; and G1-phase block after irradiation and release from confluent growth. Three of these ten strains showed a moderate degree of hypersensitivity to X-rays by all three assays, falling within the range previously reported for ataxia telangiectasia heterozygotes. This variability in the "normal" response may render difficult the use of these techniques in the detection of gene carriers for such disorders in the general population. Furthermore, it indicates that care should be taken in the selection of reference control strains in studies of the sensitivity of cells from various genetic disorders to radiation.

Cell Cycle↗

PubMed1988

Effects of cigarette smoking and solvent exposure on sister chromatid exchange frequency in painters.

A cross-sectional study of sister chromatid exchange frequency (SCE) in peripheral blood lymphocytes from 117 members of the International Brotherhood of Painters and Allied Tradesman was conducted in union locals in two major U.S. cities. Chronic solvent exposure intensity and duration were estimated from interviewer-administered-questionnaire data. SCE for all of the workers in the study were scored by one reader. A second reader determined the SCE frequency from a random sample of 30 workers. No significant difference in SCE frequency was associated with reader or time of reading. Age, coffee and alcohol intake and chronic solvent exposure (both intensity and duration, estimated over the working lifetime and over the year prior to study for each worker) did not significantly elevate SCE. The effect of smoking on SCE frequency, assessed by analysis of variance controlling for other possible confounding factors, showed that smoking increased SCE frequency (P less than .0001). The SCE frequency in the smoking, solvent-exposed (estimated as lifetime exposure) painters was 6.75 SCE/cell; in the non-smoking, solvent-exposed workers the SCE frequency was 5.73 SCE/cell; the controls had SCE levels of 6.84 SCE/cell (smokers) and 5.90 SCE/cell (non-smokers), respectively. These observations are consistent with other work suggesting that chronic solvent exposure in the paint trade is not associated with an elevation in SCE in peripheral blood lymphocytes. However, further work is necessary to address adequately the question of the genotoxicity of acute solvent exposure in these workers.

Art↗

PubMed1988

Multiple abnormalities in the ultraviolet light response of cultured fibroblasts derived from patients with the basal cell nevus syndrome.

Basal Cell Nevus Syndrome (BCNS) is a rare autosomal-dominant inherited disorder associated with a marked hypersusceptibility to spontaneous and radiation-induced skin cancer. We examined the changes in cell survival, unscheduled DNA synthesis (UDS) and the frequency of sister chromatid exchanges (SCE) induced by ultraviolet light (UVL) in confluent normal and BCNS fibroblasts. BCNS cells appeared slightly hypersensitive to the cytotoxic effects of UVL. The rate of UDS induced by UVL exposure in normal cell strains increased linearly following doses up to 30 J/m2, whereas in BCNS cells UDS became saturated at doses of 10 J/m2 showing no further increase with doses up to 30 J/m2. UDS activity persisted for longer periods after UVL exposure in BCNS as compared with normal cells. The dose-response relationship for UVL-induced SCE was similar in normal and BCNS fibroblasts. However, the frequencies of UVL-induced SCE declined to near background levels in normal cells following 12-24 hr of confluent holding prior to subculture whereas they remained elevated in BCNS cells with holding times up to 24 hr after UVL exposure. Overall, these results suggest that BCNS fibroblasts may have a diminished capacity for the repair of some type of DNA damage as compared with normal fibroblasts.

Adult↗

PubMed1988

Effect of duration of exposure to benzo(a)pyrene diol-epoxide on neoplastic transformation, mutagenesis, cytotoxicity, and total covalent binding to DNA of rodent cells.

We examined the effect of different durations of exposure (20 sec to 24 hr) to (+/-) 7-beta,8 alpha-dihydroxy-9 alpha, 10 alpha -epoxy-7,8,9,10-tetrahydrobenzo(a)pyrene (BPDE I) on the induction of transformation in C3H/10T 1/2 cells and of mutations to 6-thioguanine resistance in Chinese hamster ovary cells (CHO), as well as on BPDE I-DNA binding in these two cell lines. A 20-sec exposure of the cells to BPDE I was sufficient to induce mutations and morphological transformation in vitro. However, the transformation frequency in CH3 mouse-embryo-derived 10T 1/2 cells increased twofold and the frequency of mutations in CHO cells sixfold when the exposure time to BPDE I was increased from 20 sec to 8 h. Cytotoxicity increased under similar conditions. A large number of BPDE I-DNA adducts were formed in both cell lines within the first 15-min of exposure of the cells to this ultimate carcinogen. The total covalent binding did not increase with longer than 15-min incubation times. These results suggest that in addition to its covalent binding to DNA, BPDE I may influence other cellular mechanism(s) that are responsible for the initiation of transformation and mutagenesis.

7,8-Dihydro-7,8-dihydroxybenzo(a)pyrene 9,10-oxide↗

PubMed1988

Studies of mutagenesis and neoplastic transformation by bivalent metal ions and ionizing radiation.

We examined the influence of nontoxic concentrations of each of two essential (Zn++ and Mn++) and one nonessential (Ni++) bivalent metal ions on spontaneous and radiation-induced neoplastic transformation and specific gene mutations in mammalian cells. All three metals induced low levels of transformation in mouse BALB/3T3 cells but exerted no mutagenic effect in CHO cells (hprt locus) over a broad range of concentrations. Continuous incubation for 8 or 15 days with each of the metal ions did not enhance the frequency of cell killing, transformation, or mutations induced by acute exposure to x-rays. Zn++, however, had a small but consistent protective effect on the induction of all three endpoints by x-irradiation.

Animals↗

PubMed1988

Recovery of mitomycin C-treated mouse 10T1/2 cells during confluent holding.

The relationship between cytotoxicity, sister-chromatid exchanges (SCE) and the repair of DNA crosslinks was studied in mouse 10T1/2 cells during confluent holding following either acute or protracted MMC treatment. No cytotoxic effects were observed with increasing doses of MMC until SCE frequencies 1.8 times background levels were induced. Protracted MMC treatments were less cytotoxic than acute MMC exposure at doses which yielded similar frequencies of SCE. The kinetics of recovery during confluent holding in acute MMC-treated cells were similar for cytotoxicity and the repair of DNA interstrand crosslinks. These results suggest that a type of non-lethal DNA damage which causes SCE may persist for long periods of time in MMC-treated cells. This non-lethal damage may accumulate during protracted MMC exposure while damage leading to cell killing is repaired.

Animals↗

PubMed1988

Studies of ionizing radiation as a promoter of neoplastic transformation in vitro.

The induction of malignant transformation was examined in a standard promotion protocol in which BALB/3T3 cells were incubated continuously with tritiated water (3HOH) following acute treatment with various doses of either X-rays or benzo(a)pyrene (BP). In no case was there any evidence that protracted exposure to ionizing radiation from 3HOH enhanced the yield of transformants induced by the primary carcinogen over that predicted if the effects of the two agents were additive.

Animals↗

PubMed1988

Survival of human diploid skin fibroblasts from normal individuals after X-irradiation.

The cytotoxic effect of X-rays was measured by a colony formation assay in multiple experiments with fibroblast cell strains derived from 24 presumably normal individuals, received as 65 different coded and blinded samples. Each strain was received on two or more occasions at different times and bearing different codes. The means and standard deviations of the survival curve parameters for the 24 strains were: D0 = 123 +/- 23; D10 = 273 +/- 42 cGy. The D0 ranged from 89 to 175 and the D10 from 196 to 372 cGy. The degree of interexperimental variation, though generally minimal, differed considerably among cell strains. There was no systematic effect of passage level, cloning efficiency, serum lot, age or sex of the donor on X-ray survival. These results confirm that the intrinsic radiosensitivity varies significantly among skin fibroblasts isolated from clinically normal individuals, apparently owing to as yet unidentified genetic factors.

Adolescent↗

PubMed1987

Normal cytotoxic response of skin fibroblasts from patients with Li-Fraumeni familial cancer syndrome to DNA-damaging agents in vitro.

Skin fibroblasts from patients with the Li-Fraumeni familial cancer syndrome have been reported to show abnormalities in their response to X-irradiation. We have examined the response of fibroblasts from affected and nonaffected individuals in three families to treatment with four DNA-damaging agents: X-rays, UV light, N-methyl-N'-nitro-N-nitrosoguanidine, and mitomycin C. Test cells along with additional cell bank control strains were received coded and blinded. The same strains were studied on 2 or 3 separate occasions; each of these groups was coded differently. The cytotoxic effects of the four agents were examined by a colony formation assay. Sensitivity to the induction of chromosomal aberrations by X-rays was also measured. In all cases, the response of cells from affected individuals did not differ significantly from that of cells from unaffected (not a risk) family members nor of cell bank controls. The response of somatic cells from members of Li-Fraumeni cancer families to DNA-damaging agents does not appear to be a fruitful approach to the detection of at-risk individuals.

Cells, Cultured↗

PubMed1987

Interrelationships among X-ray-induced anchorage independence, mutagenesis and chromosomal rearrangements in human diploid fibroblasts.

The changes in the frequencies of X-ray-induced mutations to 6-thioguanine resistance, chromosomal abnormalities and transformation to anchorage-independent growth were examined during confluent holding recovery in density-inhibited cultures of human diploid fibroblasts. Chromosomal abnormalities studied included non-stable aberrations, deletions, and reciprocal translocations measured up to 20 mean population doublings post-irradiation. Complex recovery kinetics were observed. A comparison of these kinetics suggests that reciprocal translocations are associated with the induction of both transformation and mutations. Mutagenesis correlated with deletions and translocations, whereas transformation correlated with translocations only.

Cell Adhesion↗

PubMed1987

Detection of ataxia telangiectasia heterozygous cell lines by postirradiation cumulative labeling index: measurements with coded samples.

Coded skin fibroblast cell strains from ataxia telangiectasia (AT) families and apparently normal individuals were obtained from two different sources. AT homozygous strains were clearly identified on the basis of marked hypersensitivity to cell killing by X-irradiation. AT heterozygotes were intermediate in their cytotoxic response between AT homozygotes and five normal reference cell strains. When density-inhibited cultures were X-irradiated and immediately subcultured to low density, a large fraction of AT heterozygous cells were irreversibly blocked in G1 as determined by cumulative labeling indices following incubation with [3H]thymidine. No such block occurred in four reference normal or AT homozygous strains. Three coded cell strains from apparently normal individuals resembled AT heterozygotes in their response; two of these strains were heterozygous for lysosomal storage disease. Thus, although the phenotype associated with the cellular response of AT heterozygous cells to X-irradiation is not specific to this disorder, the cumulative labeling indices assay may be a useful method for the detection of AT heterozygotes in kindreds with known AT.

Ataxia Telangiectasia↗

PubMed1987

Molecular and biochemical analyses of spontaneous and X-ray-induced mutants in human lymphoblastoid cells.

We have isolated a series of 14 spontaneously arising and 28 X-ray-induced mutants at the hypoxanthine-guanine phosphoribosyltransferase (hgprt) locus in human lymphoblastoid cells. Among the spontaneous mutants, 5/14 (36%) had detectable alterations in their restriction fragment pattern after hybridization with a human cDNA probe for hgprt. Of the 10 remaining mutants, 4 had partial HGPRT enzyme activity, which suggested that they contained point mutations. Among the 28 mutants induced by 150 rad of X-rays, 15 (54%) had deletions of part or all of the hgprt gene. 5 of the remaining 13 (18% overall) had partial HGPRT enzyme activity, which suggested that they contained point mutations. These data imply that in this human cell system, X-rays induce both point mutants which have residual enzyme activity as well as mutations involving relatively large deletions of DNA.

DNA Restriction Enzymes↗

PubMed1987

Radiation sensitivity of fibroblast strains from patients with Usher's syndrome, Duchenne muscular dystrophy, and Huntington's disease.

The colony-forming ability of 10 normal human fibroblast cell strains and of 10 strains representing 3 degenerative diseases of either nerve or muscle cells was determined after exposure of the cells to X-rays or beta-particles from tritiated water. Both methods of irradiation yielded similar comparative results. The fibroblast strains from the 5 Usher's syndrome patients and from 1 of the 2 Huntington's disease patients were hypersensitive to radiation, while those from the 3 Duchenne muscular dystrophy patients and the second Huntington's disease patient had normal sensitivity to radiation. These results indicate both disease-specific and strain-specific differences in the survival of fibroblasts after exposure to ionizing radiation.

Cell Survival↗