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Biomedical subjects

A R Collins

Publications and source records attributed to A R Collins.

At least 127 records · Page 7Linked to original sources
PubMed1981

Chinese hamster ovary mutant UV-1 is hypomutable and defective in a postreplication recovery process.

CHO-UV-1 is a mutant of the Chinese hamster cell CHO-K1 hypersensitive to killing by ultraviolet light but with normal resistance to X-ray. It is also hypersensitive to killing by ethyl methane sulfonate. Hybrid clones formed bu fusing UV-1 and Chinese hamster lung cells display the normal ultraviolet resistance of the latter. The sensitive phenotype behaves, therefore, in a genetically recessive manner. Ultraviolet sensitivity of UV-1 is not associated with a deficiency in excision repair. Alkaline sucrose gradient sedimentation analysis of nascent DNA from ultraviolet-irradiated cells reveals that UV-1 is, however, markedly deficient in postreplication recovery. Furthermore, UV-1 has a lower rate of induced mutation to 6-thioguanine resistance than does the parental cell when treated with ultraviolet light or ethyl methane sulfonate. These results suggest that the phenotype of UV-1 is due to a mutation in a form of postreplication recovery which in normal cells is error prone.

Animals↗
PubMed1981

Correlations of DNA damage and repair with nuclear and chromosomal damage in HeLa cells caused by methylnitrosamides.

N-Methyl-N-nitrosourethan induces breaks or alkali-labile sites in cellular DNA, many if not all of which are repaired rapidly. Other DNA lesions are repaired by an excision process. Hydroxyurea and 1-beta-D-arabinofuranosylcytosine cause an accumulation of DNA breaks after N-methyl-N-nitrosourethan treatment, probably by inhibiting the DNA-synthetic (but not the nucleolytic) stage of excision repair. Chromosome damage (fragmentation or attenuation of interphase chromosomes and decondensation and radial rearrangement of metaphase chromosomes) is present soon after treatment with N-methyl-N-nitrosourethan and is not reversed during further incubation. It is apparently associated with the longer-lived DNA lesions, probably those which are removed by excision, and is enhanced by incubation with hydroxyurea and 1-beta-D-arabinofuranosylcytosine. N-Methyl-N-nitrosourethan also inhibits cellular protein and the loss of nucleolar structure. N-Methyl-N-nitrosourea is less potent than N-methyl-N-nitrosourethan in causing DNA or chromosome damage and is less cytotoxic.

Cell Cycle↗
PubMed1980

Cell cycle-related variations in UV damage and repair capacity in Chinese hamster (CHO-K1) cells.

UV damage to CHO cell DNA, measured by formation of thymine-containing dimers, increases from mitosis to early S phase. Computer simulation of UV absorption by the DNA of an idealized CHO cell at different stages in the cell cycle resembles the cycle dependence of UV damage. Incision at UV damage sites, measured by the accumulation of breaks in preexisting DNA during 30 minutes' post-irradiation incubation with the DNA synthesis inhibitors 1-beta-D arabinofuranosylcytosine and hydroxyurea, increases from mitosis to interphase. Analysis of the dose dependence of DNA break accumulation indicates that both the affinity of the endonuclease for dimer sites and the maximum enzyme activity at saturating levels of dimers are significantly lower in mitosis than in interphase. The killing of CHO cells by UV is enhanced if repair is temporarily inhibited by are C. The DNA gyrase inhibitor novobiocin prevents UV-induced incision.

Animals↗
PubMed1979

Repair and survival after UV in quiescent and proliferating Microtus agrestis cells: different rates of incision and different dependence on DNA precursor supply.

Cultured cells of Microtus agrestis, the common field vole, perform unscheduled DNA synthesis after UV irradiation. They respond to incubation with a DNA synthesis inhibitor (1-beta-D-arabinofuranosylcytosine) following UV in ways typical of cells capable of excision repair, with reduced survival and an accumulation of breaks in pre-existing DNA. Microtus cells irradiated with UV in a quiescent pre-S-phase state are more sensitive to UV than are proliferating cells, in terms of survival. Adding DNA precursors (deoxyribonucleosides), and--in case of proliferating cells--growing in complete rather than dialysed serum, enhance UV survival. Quiescent cells show a higher rate of endonucleolytic incision of DNA after UV than do proliferating cells. The balance between incision (producing single-strand DNA breaks) and repair DNA synthesis (leading to rejoining of breaks) is shifted by the addition of deoxyribonucleosides, which suggests that DNA precursor supply is a rate-limiting factor in repair. The lower survival of quiescent cells (in the absence of added deoxyribonucleosides) may be due to insufficient precursor supply to meet the demands of the high incision rate.

Animals↗
PubMed1979

DNA damage in synchronized HeLa cells irradiated with ultraviolet.

The lethal effect of UV radiation of HeLa cells is least in mitosis and greatest in late G1-early S. Photochemical damage to HeLa DNA, as measured by thymine-containing dimer formation and by alkaline sucrose sedimentation, also increases from mitosis towards early S phase. Computer simulations of UV absorption by an idealized HeLa cell at different stages of the cell cycle indicate that changes in damage could be due solely to changes in chromatin geometry. But survival is not exclusively a function of damage.

Cell Division↗
PubMed1978

Recovery of a Sendai virus variant with temperature-sensitive hemolytic activity from persistently infected cells from mouse brain.

A persistently infected cell line designated MB/Senas was established by cultivation of mouse brain cells from four-day-old C3H mice infected intracerebrally at birth with 10(6) PFU of Sendai virus, strain 52. After 5 passages, 0.16 per cent of Sendai52 antiserum (containing two 50 per cent plaque reducing doses/ml of serum) was introduced into the culture medium. The addition of antiserum was accompanied by a rise in cell-associated viral antigen from a level of 5 per cent antigen positive cells to 100 per cent demonstrable by both intracellular and membrane immunofluorescence. A variant of Sendai52 virus, designated Sendaias, was recovered from MB/Senas by inoculation of supernatant medium into chick embryos. Infection of chick embryos at 37 degrees C was abortive. Fifty per cent or less of chick embryos infected at dilutions 10(-1) to 10(-9) yielded detectable virus. Hemagglutination (HA) was weak but could be improved by trypsinization of allantoic fluids. Neuraminidase (NA) activity was barely detectable. Hemolysis (HE) was absent. Propagation of Sendaias virus at 33 degrees C showed no change from weak HA and NA activities but HE activity was now apparent which was temperature sensitive. Mortality of infected chick embryos increased to 100 per cent. HE activity and lethality for chick embryos was thermolabile at 45 degrees C.

Cell Line↗
PubMed1977

DNA damage in ultraviolet-irradiated HeLa and CHO-Kl cells examined by alkaline lysis and hydroxyapatite chromatography.

DNA damage and repair activity have been examined in cultured mammalian cells by the technique of alkaline lysis followed by analysis of the single-strand content of DNA by hydroxyapatite chromatography; the rate of unwinding of DNA in alkali is related to the number of single-strand breaks present. When HeLa cells are ultraviolet-irradiated and incubated, on subsequent alkaline lysis the DNA unwinds at a faster rate than the DNA of unirradiated cells. The rate is increased if inhibitors of DNA synthesis (hydroxyurea, deoxyadenosine or 1-beta-D-arabinofuranosyl cytosine) are present during the incubation. Similar effects are seen in CHO-Kl cells synchronised in G1 phase, though mitotic CHO-Kl cells show little effect of post-irradiation incubation or of the presence of inhibitors. It appears that agents known to block replicative DNA synthesis can also inhibit repair DNA synthesis following ultraviolet irradiation, leading to an accumulation of single-strand breaks in DNA produced by repair nuclease activity. This phenomenon is probably responsible for the retarded sedimentation of such DNA on alkaline sucrose gradients.

Alkalies↗
PubMed1977

Interferon production and response to exogenous interferon in two cell lines of mouse brain origin persistently infected with Sendai virus.

Two persistently infected cell lines established from C3H mouse brain cells infected in vivo with Sendai virus were shown to differ with respect to interferon (IF) production and response to exogenous IF. MB/Sen carrier cells contained 1-5 per cent antigen positive cells when examined by immunofluorescence, and virus was occasionally recovered from the culture medium. MB/SenAS carrier cells were maintained with 0.16 per cent Sendai antiserum in the supernatant medium. All MB/SenAS cells contained viral antigen and infectious virus was present in the culture medium. MB/Sen released IF spontaneously into the culture medium. Further IF production could be stimulated in MB/Sen by superinfection with Newcastle disease virus (NDV) or vesicular stomatitis virus (VSV). Exogenous IF provided good protection against VSV challenge. In contrast, MB/SenAS produced no IF spontaneously but could be stimulated by NDV and VSV to produce IF. Exogenous IF failed to reduce the amount of VSV released into the supernatant fluid. Replication of VSV was restricted in MB/SenAS as shown by a 2.3 log10 lower virus yield compared to MB/Sen.

Animals↗
PubMed1977

The inhibition of repair in UV irradiated human cells.

We have used three different assay procedures to determine the effects of hydroxyurea on excision repair in UV-irradiated HeLa cells. The results were as follows. (a) At the cytological level, incubation of UV-irradiated metaphase cells with hydroxyurea caused chromosome decondensation. (b) Using a modified alkaline sucrose gradient sedimentation technique involving minimal lysis before centrifugation, we found a marked retardation in the sedimentation of DNA from UV-irradiated cells incubated for a short period with hydroxyurea. (c) The effect of hydroxyurea on the incorporation of[3H]thymidine by UV-irradiated G1 cells was found to depend on the concentration of thymidine present in the medium. Normal primary human cells resemble HeLa cells in the response of chromosomes and DNA to UV plus hydroxyurea. Xeroderma pigmentosum cells, deficient in excision repair, are not sensitive to hydroxyurea in our assays. Chromosome decondensation and retarded DNA sedimentation occur also after incubation of irradiated HeLa cells with deoxyadenosine, but not thymidine, at concentrations which inhibit semiconservative DNA synthesis. The effects of hydroxyurea or deoxyadenosine on chromosomes and DNA are not seen if all four deoxyribonucleoside precursors of DNA are supplied exogenously. These results point to an inhibition of repair DNA synthesis by hydroxyurea (or deoxyadenosine), at the level of the supply of DNA precursors, i.e. in the same way that these agents inhibit semiconservative DNA synthesis. In the presence of these inhibitors, single-strand gaps accumulate in the DNA.

Cell Division↗
PubMed1977

Effects of ultraviolet light on synchronized Chinese hamster ovary cells; potentiation by hydroxyurea.

We have examined the effects of hydroxyurea on u.v.-irradiated Chinese hamster CHO-KI cells. Ultraviolet irradiation followed by incubation with hydroxyurea causes only slight disruption of DNA and chromosome structure in CHO-KI cells compared with HeLa cells. There is, however, a clear potentiation by hydroxyurea of the u.v. killing of CHO-KI cells, which is most pronounced at those points in the cycle which are reported to have small DNA precursor pools. This potentiation is reduced when DNA precursors are provided together with hydroxyurea. These data are discussed in terms of an uncoupling of excision and repair DNA synthesis.

Animals↗
PubMed1976

Comparative effects of glycerol and dextrose on porphyrin precursor excretion in acute intermittent porphyria.

The effects of dietary manipulations on excretion of the porphyrin precursors, delta-aminolevulinic acid (ALA), and porphobilinogen (PBG) were studied in eight patients with acute intermittent porphyria. Three diet periods of 9-17 days comprised each study. In each patient, a "baseline" protein, fat, and carbohydrate intake was kept constant throughout. In addition, during the first diet period each patient received 150 g dextrose; during the second, this was replaced by an isocaloric amount of neutral fat; and during the third, the fat was replaced by 150 g glycerol. In each of the patients, three comparisons of the effect of diet on both ALA and PBG excretion were made: (1) 300 g carbohydrate versus 150 g carbohydrate (dextrose versus fat), (2) 150 g carbohydrate + 150 g glycerol versus 150 g carbohydrate (glycerol versus fat), and (3) 300 g carbohydrate versus 150 g carbohydrate + 150 g glycerol (dextrose versus glycerol). For each of these three diet comparisons, there are sixteen individual comparisons possible for the effect of diet on porphyrin precursor excretion, eight for ALA and eight for PBG. Thus, the mean values for ALA and PBG excretions during each of the diet periods are statistically compared internally within each individual patient. Increasing carbohydrate intake from 150 g to 300 g by isocaloric substitution of dextrose for fat produced a significant (p less than 0.05) decline in eight of the sixteen comparisons of ALA and PBG excretion. Addition of 150 g glycerol by isocaloric substitution for fat caused a significant (p less than 0.05) decline in nine of the sixteen possible comparisons. In the sixteen comparisons of isocaloric dextrose and isocaloric glycerol-substituted diets, dextrose produced significantly (p less than 0.05) lower porphyrin precursor excretion in four cases and glycerol produced significantly (p less than 0.05) lower values in five. One patient showed no significant change on any of the diets. Of the four patients having symptoms believed referrable to porphyria during the study, three reported an improvement in symptoms during the high glycerol intake. The effects of dietary perturbations on porphyrin precursor excretion in acute intermittent porphyria are variable, but glycerol appears to be capable of decreasing the excretions and may prove useful in treating some of these patients.

Acute Disease↗
PubMed1975

Characterization of an in vitro persistent-state measles virus infection: establishment and virological characterization of the BGM/MV cell line.

The parameters of a persistent-state measles virus infection in BGM/MV cells were examined. The BGM/MV cell line was established by cocultivation of measles virus-infected primary C3H mouse brain cells with a stable line of African green monkey kidney cells (BGM). Initially, a morphologically mixed population of cells existed:BGM-like (epithelioid) and fibroblasts. Gradually the fibroblasts were replaced by BGM-like cells, resulting in a morphologically homogeneous population. Measles cytopathic effect was noted 2 days after initiation of this culture and persisted for approximately 290 days. The time of disappearance of viral cytopathic effect corresponded to the time at which morphological homogeneity was reached. Low titers of infectious measles virus were detected in the BGM/MV culture up to 20 days postseeding; thereafter none was observed. After 440 days in culture, 100% of BGM/MV cells demonstrated intractyoplasmic measles antigen by immunofluorescence. Nuclear fluorescence was never observed. Electron microscopy revealed the presence of measles virus mucleocapsid within the almost completely filling the cytoplasm of BGM/MV cells. The plasma membrane of these cells appeared normal; no maturing or budding particles were observed. Measles virus hemagglutinin was not detected in either clarified cell lysates or in supernatant culture fluids. Cell membrane alteration by measles virus was detected in less than 1% of these cells by hemadsorption and by membrane immunofluorescence. The hemadsorption activity of the cells could be enhanced (30 to 70%) by treatment with actinomycin D or enucleation with cytochalasin B; these treatments, however, were unsuccessful in inducing detectable levels of measles hemagglutinin. Treatment of BGM/MV cells with 5-bromo-2'-deoxyuridine (BUdR) at 5 to 50 mug/ml and cytosine arabinoside at 1 to 50 mug/ml failed to enhance hemadsorption activity. Doses of 5-bromo-2'-deoxyuridine ranging from 5 to 200 mug/ml and of actinomycin D ranging from 0.1 to 10 mug/ml were ineffective in inducing the synthesis of infectious virus. Various physical methods of induction of infectious virus was also unsuccessful.

Animals↗
PubMed1975

Characterization of an in vitro persistent-state measles virus infection: species characterization and interference in the BGM/MV cell line.

Serological methods of mixed agglutination and indirect immunofluorescence showed the BGM/MV cell line to possess monkey antigens. As a means of further characterizing the species constitution of the BGM/MV cell line, the species specificity of viral-induced interferon from these cells, as well as the response of these cells to exogenous interferons, was determined. Low titers of spontaneously elaborated interferon capable of protecting monkey but not mouse cells were detected in BGM/MV culture fluids. Interferon induced by Newcastle disease virus infection of BGM/MV cells was capable of conferring an antiviral state on monkey and, to a lesser extent, on mouse cells. Exogenous interferons of both homologous (BGM/MV) and heterologous sources failed to confer an antiviral state on BGM/MV cells. BGM/MV cells were found to be partially refractive to superinfection with measles virus but freely replicated mumps and vesicular stomatitis virus.

Animals↗