Biomedical subjects

J B Little

Publications and source records attributed to J B Little.

At least 163 records · Page 9Linked to original sources

PubMed1987

Epidermal growth factor induces cytogenetic damage in mammalian cells.

It has been previously shown that potent tumor promoters, such as the phorbol ester 12-O-tetradecanoylphorbol-13-acetate (TPA), can induce both chromosomal aberrations and sister chromatid exchanges (SCE) in mammalian cells. We show here that epidermal growth factor (EGF) is also capable of inducing SCE. EGF and TPA did not enhance the frequency of SCE in an additive fashion, suggesting these agents do not act by completely independent mechanisms.

Cell Line↗

PubMed1987

Spontaneous transformation to anchorage-independent growth of a xeroderma pigmentosum fibroblast cell strain.

A skin fibroblast cell strain (GM2995) derived from a patient with xeroderma pigmentosum was received at low passage. As the cells were serially passaged (1:4 dilution), their size and growth characteristics changed. By passage 13, approximately 90% of the colonies produced by cells seeded at low density were composed of small, densely packed cells. Cells capable of anchorage-independent growth were observed after passage 7; they formed round, smooth-edged colonies in soft agar. The frequency of cells exhibiting anchorage-independent growth increased rapidly at subsequent passages, reaching 35-50% of the population by passage 20. This phenomenon was accompanied by the appearance of aneuploidy. These cells are still proliferating actively at passage 35. These late-passage GM2995 cells retain the extreme hypersensitivity to the cytotoxic effect of UV radiation characteristic of early-passage GM2995 cells.

Cell Division↗

PubMed1987

Differential effects of radical scavengers on X-ray-induced mutation and cytotoxicity in human cells.

The cytotoxic and mutagenic effects of X irradiation on a human lymphoblast cell line were examined in the presence of two radioprotective agents which modulate damage to DNA. The cells were treated with X rays alone or in the presence of either dimethyl sulfoxide or cysteamine. Surviving fraction and mutation to trifluorothymidine resistance (tk locus) and to 6-thioguanine resistance (hgprt locus) were measured. Survival was enhanced when the cells were irradiated in the presence of dimethyl sulfoxide; the D0 rose from 58 to 107 rad. However, at both genetic loci the induced mutant fractions were identical in the presence or absence of dimethyl sulfoxide. Survival was enhanced to a greater degree when the cells were irradiated in the presence of cysteamine; the D0 rose from 58 to 200 rad. Cysteamine also protected the cells from X-ray-induced mutation; the frequencies of X-ray-induced mutation at both the tk and hgprt loci were reduced by 50-75%. No protective effects were observed unless dimethyl sulfoxide or cysteamine was present during irradiation. These findings are discussed in terms of the hypothesis that, unlike for cell killing, radiation-induced mutagenesis in human lymphoblast cells is not mediated by the actions of aqueous free radicals, but rather by the direct effects of ionizing radiation.

Cell Line↗

PubMed1986

Chromosome 14 marker appearance in a human B lymphoblastoid cell line of nonmalignant origin.

A human B cell lymphoblastoid cell line of nonmalignant origin, derived 16 years ago, now carries a 14q+ marker chromosome in 100% of the cells analyzed. This marker was not apparent in karyotypes prepared 1 year after establishment of the line and, thus, presumably arose in culture. The cell line now grows more rapidly than typical B cell cultures and has an unusually high cloning efficiency (60%-90%). Using Southern blot hybridization, we have analyzed the arrangement of 13 onc genes that are often associated with human malignancy, including c-myc. None of these onc genes appear to be rearranged in this cell line.

B-Lymphocytes↗

PubMed1986

Toxicity and mutual interactions of cadmium and zinc ions in normal and carcinogen-transformed mouse cells.

The toxicity of chloride salts of physiological (zinc, manganese, nickel) and non-physiological (cadmium) bivalent metal ions was studied in normal or carcinogen-transformed mouse embryo fibroblast cells. The dose response curves for toxicity to both types of cells exhibited similar shapes. The transformed cells, however, were about twice as sensitive to zinc toxicity as normal cells. When normal and transformed cells were grown together and incubated for several hours with an appropriate concentration of zinc, the malignant cells were selectively killed. Cadmium was much more toxic than the three other metal ions in both types of cells. Its toxic effect was reversed by simultaneous addition of zinc at nontoxic concentrations.

Animals↗

PubMed1986

Somatic mutations at a heterozygous autosomal locus in human cells occur more frequently by allele loss than by intragenic structural alterations.

A human B-cell lymphoblastoid cell line heterozygous at the thymidine kinase (TK) locus (i.e., carrying one functional and one nonfunctional thymidine kinase allele) was used to study the molecular nature of mutations leading to loss of TK activity. A total of 113 mutant clones, both spontaneous and induced, were examined by restriction enzyme mapping and by the use of a restriction fragment length polymorphism (RFLP) at the TK locus. A majority (71%) of all mutant clones examined had lost the entire functional TK allele, becoming either homozygous or hemizygous for the nonfunctional allele. The remaining mutants had either no detectable changes (26%) or had obvious structural alterations (less than 5%) in the active TK gene. These results emphasize the importance of allele loss, presumably by mitotic chromosomal mechanisms, in mutagenesis at autosomal loci, and suggest that in vitro models for recessive somatic mutation which are based at hemizygous loci may ignore a large category of genetically significant events.

B-Lymphocytes↗

PubMed1986

Hypersensitivity of cells from a new chromosomal-breakage syndrome to DNA-damaging agents.

Cells derived from a patient with severe chromosomal breakage, immunodeficiency, and growth retardation were found to resemble those from individuals with ataxia telangiectasia (A-T) in terms of their sensitivity to cell killing and the induction of cytogenetic abnormalities by X-rays. Their response to other DNA-damaging agents, including 254-nm UV light, mitomycin C, MNNG, and bleomycin was also A-T-like. In contrast to classical A-T, however, X-irradiated cells exhibited a G1 block after release from density inhibition of growth that was not significantly different from that of normal controls.

Adult↗

PubMed1986

X-rays mutate human lymphoblast cells at genetic loci that should respond only to point mutagens.

We have demonstrated that X-rays induce mutations at 4 of 5 genetic loci. 2 of these loci, which code for a mRNA synthesis factor (resistance to 5,6-dichlororibofuranosylbenzimidazole) and tubulin (resistance to podophyllotoxin), are "small-marker" loci, in that they theoretically respond only to mutations which eliminate a toxin-binding site while leaving the major function of the protein intact. Thus mutations induced by X-rays in these two loci are most likely due to base-pair substitution-type alterations. X-Rays did not induce mutations in the Na+/K+ ATPase (resistance to ouabain), another small-marker locus. Two other loci, hypoxanthine guanine phosphoribosyl transferase (resistance to 6-thioguanine) and thymidine kinase (resistance to trifluorothymidine), are "whole-gene" targets in that they theoretically respond to a wide variety of mutagenic changes. X-Rays induced dose-dependent increases in mutant fraction at both of these loci. Ethyl methanesulfonate (EMS), an agent thought to produce mutations primarily through a base-pair substitution mechanism, induced mutations at all genetic loci tested. The pattern of mutations at the small-marker loci induced by EMS was different than that induced by X-rays, suggesting that the specificities of the mutagens and/or of the loci are different.

Dichlororibofuranosylbenzimidazole↗

PubMed1986

Characteristics of radiation-induced neoplastic transformation in vitro.

Data are presented to support the hypothesis that the initial step in the morphologic transformation of irradiated rodent cells is a frequent cellular event involving a large fraction of the irradiated population. This process appears to involve DNA damage, but not to represent a targeted mutation in specific structural gene(s). Morphologic transformation and immortalization appear to be distinct steps in the overall process of transformation. In contradistinction to rodent cells, immortalization is a very rare event in human diploid cells which is induced at extremely low frequencies. The hypothesis is presented that immortality develops among clones of cells bearing stable chromosomal rearrangements which emerge during the proliferation of a population of radiation damaged cells.

Animals↗

PubMed1986

The in vitro genetic effects of fibrous erionite and crocidolite asbestos.

Epidemiologic evidence has recently identified an association between an endemic outbreak of pleural and peritoneal mesothelioma in the Urgup region of Turkey and exposure to zeolite fibres. This malignancy is usually associated with exposure to asbestos dusts whose mineralogical characteristics differ from those of zeolites. The present study further defines the in vitro biologic activity of erionite, a common zeolite fibre found in the Urgup region of Turkey. Both erionite and crocidolite asbestos fibres were clastogenic in synchronous Chinese hamster ovary (CHO) fibroblasts. Both fibres also altered CHO ploidy. Erionite, unlike crocidolite or Min-U-Sil quartz, caused a slight increase in sister chromatid exchanges in synchronous CHO cells. Neither erionite nor crocidolite was mutagenic in a human lymphoblastoid cell line. Erionite fibres thus produced in vitro cytogenic changes similar to those caused by asbestiform mineral dusts and, like asbestos fibres, did not induce mutations in human lymphoblastoid cells.

Aluminum Silicates↗

PubMed1986

Effect of aliphatic amides on oncogenic transformation, sister chromatid exchanges, and mutations induced by cyclopenta[cd]-pyrene and benzo[a]pyrene.

We examined the effects of the aliphatic amides isopropyl-valeramide (IVA) and allylisopropylacetamide (AIA) on oncogenic transformation and sister chromatid exchanges (SCE) induced by cyclopenta[cd]pyrene (CPP) and benzo[a]pyrene (B[a]P) in C3H/10T1/2 cells and on B[a]Pdiol-epoxide (BPDE)-induced mutation at the HGPRT locus in Chinese hamster ovary (CHO) cells. IVA and AIA significantly suppressed B[a]P and CPP transformation in vitro. Both amides were effective when given just prior to, simultaneously with, or 24 h after carcinogen exposure. On the other hand, IVA and AIA did not affect cytotoxicity, the frequencies of SCE induced by CPP or B[a]P, nor BPDE-induced mutations in CHO cells. These and previous results suggest that the mechanism of inhibition of transformation by IVA or AIA may be very specific and probably not related to the early initiation event in oncogenic transformation in vitro.

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

PubMed1986

Fibroblast radiosensitivity and intraocular fibrovascular proliferation following radiotherapy for bilateral retinoblastoma.

A 5-day-old female patient was found to have large hereditary retinoblastomas in the posterior pole of each eye. The patient received radiation treatment over a 39-day period, with each retina receiving 4600 rad. Two weeks after the complete treatment the tumours had regressed to approximately one-quarter of their original size. By 14 weeks following completion of radiotherapy the patient had developed in each eye extensive iris neovascularisation with progressive closure of the filtration angles, secondary glaucoma, and retinal detachments resulting from fibrovascular proliferation on the retinal surface. Radiosensitivity studies were from separate conjunctival biopsies obtained before and after radiation. These showed a D0 (calculated survival curve parameters, defined in the Methods section) in the exponential growth phase of 110 prior to radiation and a postirradiation exponential growth phase D0 of 70. Karotype studies showed several chromosomal abnormalities following radiotherapy. The clinical course and pathology findings are thought to represent an unusually severe orbital and ocular response to radiation therapy. These findings are consistent with our hypothesis that some patients with hereditary retinoblastoma may have a defect in the accumulation repair of x-irradiation induced DNA damage.

Cell Survival↗

PubMed1986

Site-specific chromosomal rearrangements induced in human diploid cells by x-irradiation.

A total of 130 stable, two-break reciprocal translocations were scored in G-banded karyotypes prepared from 375 metaphase spreads from a strain of human diploid fibroblasts irradiated with 400 or 600 rads and analyzed 1-20 mean population doublings later. The chromosomal location of each of the 260 breakpoints was mapped. The sites of 121 chromosomal breaks and deletions in the first postirradiation mitosis were also scored. Unlike the random distribution of these latter events, the translocation breakpoints showed not only a nonrandom distribution among chromosomes but also the existence of specific sites within chromosomes that were more frequently involved in translocations. The most notable finding was a marked excess of translocations involving the short arm of chromosome 1, in particular, band 1p22. The specific types of translocations were random, although the breakpoints were not. Eight of the 12 most frequently involved chromosomal sites were regions in which fragile sites have been mapped in human lymphocytes.

Cells, Cultured↗

PubMed1986

Enhancement of benzo[a]pyrene-induced sister chromatid exchanges in lymphocytes from cigarette smokers occupationally exposed to asbestos.

The frequencies of base-line and benzo[a]pyrene [(BP) CAS 50-38-8]-induced sister chromatid exchanges (SCE) were measured in peripheral blood lymphocytes from 22 male asbestos-exposed workers and 10 nonexposed workers of comparable age. A clear association between cigarette smoking and asbestos exposure in the sensitivity of lymphocytes to BP was observed. Among asbestos-exposed workers, lymphocytes from those who smoked cigarettes were significantly more susceptible to the induction of SCE by in vitro exposure to BP (P = .01) than were lymphocytes from nonsmokers. Active smoking elevated the base-line SCE frequency in both asbestos-exposed and nonexposed workers (P = .001), and an interaction between smoking and asbestos in the production of base-line SCE was suggested (P = .07). Asbestos exposure alone was not associated with an enhanced susceptibility to the induction of SCE by BP or with an elevation of base-line SCE. Increased age was associated with an increase in SCE inducibility by BP (P = .01), and a history of smoking was marginally associated with SCE inducibility by BP (P = .07). These findings support the hypothesis that an increased susceptibility of asbestos-exposed individuals to polyaromatic hydrocarbon-induced cancer results from an enhanced sensitivity to the induction of genetic damage rather than to an asbestos-induced differential cellular metabolic capacity.

Adult↗

PubMed1986

Induction of neoplastic transformation by low-dose-rate exposure to tritiated water.

Exponentially growing mouse BALB/3T3 cells seeded at low density were incubated with various concentrations of tritiated water (3HOH). A dose-dependent increase in the yield of transformants occurred in cells incubated for 100 h with 25 to 500 micrograms/ml of 3HOH. The dose-response curve rose linearly at low doses, with no evidence of a threshold or quadratic component. The transformation frequencies were similar when the same total dose of radiation was given over periods of 5 to 168 h, although survival was slightly lower with 5 h exposure compared with longer intervals. Acute X-ray exposure appeared to be more efficient in inducing transformation than protracted exposure to 3HOH at doses of 200-400 rad, but slightly less effective in the 25-100 rad range. When normalized for survival, 3HOH was more effective than X rays even at the higher doses. This result is very similar to previous findings with [3H]thymidine. A comparison of [3H]thymidine and 3HOH results suggest that the transmutational effect of tritium decay for radioactivity incorporated into DNA may be a significant factor in cytotoxicity but not in transformation; the enhanced effectiveness of both types of tritium exposure for transformation and mutagenesis may result from a higher relative biological effectiveness for the tritium beta particle for these end points.

Animals↗

PubMed1985

Mechanisms of human cell neoplastic transformation: relationship of specific abnormal clone formation to prolonged lifespan in X-irradiated human diploid fibroblasts.

A total of 9 control and 46 X-irradiated human fibroblast cultures were followed throughout their lifespan in vitro; G-banded karyotypes were examined at regular intervals. The lifespan (mean population doublings) of irradiated cultures was slightly but significantly prolonged over that of controls. None of the cultures developed any changes in cell morphology characteristic of neoplastic transformation. A number of abnormal clones containing marker chromosomes emerged in the irradiated cultures. Most of these senesced early, but 2 clones were associated with a considerably increased lifespan. One of these had a deletion in the short arm of chromosome I (p22, p32), and the other had 2 specific translocations involving chromosome 22, t(1;22)(q25,q12) and t(6;22)(p22,q11). We hypothesize that the emergence of an abnormal clone with translocations in the vicinity of critical oncogenes may be associated with prolongation of lifespan and the induction of immortalization in human diploid cells, an event independent of the acquisition of other characteristics of the transformed phenotype.

Cell Survival↗

PubMed1985

Confluent holding leads to a transient enhancement in mutagenesis in UV-light-irradiated xeroderma pigmentosum, Gardner's syndrome and normal human diploid fibroblasts.

The influence of confluent holding periods of 0-24 h of UV-light-induced mutagenesis has been investigated in several human cell strains including xeroderma pigmentosum complementation group A (XPA), Gardner's syndrome (GS) and normal human diploid fibroblasts (NHDF). These cells strains were chosen for the variety of their responses in cytotoxicity experiments. Confluent cultures of NHDF exposed to UV light exhibited a time-dependent increase in survival when subculture was delayed up to 24 h after irradiation. GS and XPA fibroblasts showed no such increase. In dose-response experiments, GS cells from 3 kindreds were moderately hypersensitive to cell killing by UV light whereas XPA cells were strongly hypersensitive. Confluent cultures of GS cells were slightly and XPA markedly hypermutable to 6-thioguanine resistance (6-TGR) when the cells were subcultured immediately after UV exposure. When allowed confluent holding periods of 1.5-24 h, GS, XPA and NHDF all exhibited a transient enhancement of mutagenesis such that a 5-10-fold increase in mutation frequency was observed in cells subcultured at 6-9 h after irradiation as compared to cells subcultured at 3-6 h. A decline in mutation frequency prior to the mutagenesis peak was observed in GS and normal cells but not in XPA. After 24 h of confluent holding, the mutation frequency in irradiated GS and NHDF had returned to near background levels although XPA mutation frequencies remain similar to those observed in immediately subcultured cells. A model to explain these overall results is discussed.

Cell Survival↗

PubMed1985

Toxicity and mutagenicity of low dose rates of ionizing radiation from tritiated water in human lymphoblastoid cells.

In order to study the toxic and mutagenic effects of low-dose-rate exposure to ionizing radiation, human lymphoblast cells were grown continuously in tritiated water (3H2O) for up to 8 days. Dose rates ranged from 0.0054 to 0.064 rad/min. Mutation to trifluorothymidine resistance (TK locus) and 6-thioguanine resistance (HGPRT locus) was measured; comparable results were observed at both loci. The mutant fraction as a function of total absorbed dose was independent of dose rate over the range studied. At the lower doses, the dose-response curve was linear, with no indication of a threshold. Overall, it appeared to be slightly biphasic with a diminished slope at higher total doses. These data are discussed in relation to earlier studies utilizing high-dose-rate X-irradiation and incorporated [3H]TdR; 3H2O and [3H]TdR were more mutagenic per rad than X-rays, but 3H2O was less cytotoxic than either X-rays or [3H]TdR.

Cell Line↗