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Biomedical subjects

O Nikaido

Publications and source records attributed to O Nikaido.

At least 73 records · Page 4Linked to original sources
PubMed1995

Correlation of UVC and UVB cytotoxicity with the induction of specific photoproducts in T-lymphocytes and fibroblasts from normal human donors.

By using specific monoclonal antibodies in situ and a computer-assisted image analysis system we have determined the relative induction of cyclobutane dimers, (6-4) photoproducts and Dewar isomers in human mononuclear cells and fibroblasts following irradiation with UVC, broad-spectrum UVB and narrow-spectrum UVB. The lamps produced these lesions in different proportions, with broad-spectrum UVB inducing a greater combined yield of (6-4) photoproducts and Dewar isomers per cyclobutane dimer than UVC or narrow-spectrum UVB. The relative induction ratios of (6-4) photoproducts compared to cyclobutane dimers were 0.15, 0.21 and 0.10 following irradiation with UVC, broad- or narrow-spectrum UVB, respectively. Although Dewar isomers were induced by UVC, their relative rate of formation compared to cyclobutane dimers was significantly greater after irradiation with either broad-spectrum or narrow-spectrum UVB. These values were 0.001, 0.07 and 0.07, respectively. With each lamp source, we have determined the survival of normal human T-lymphocytes and fibroblasts at fluences, which induce equivalent yields of cyclobutane dimers, (6-4) photoproducts or (6-4) photoproducts plus Dewar isomers. Killing of fibroblasts appears to be associated with (6-4) photoproduct formation, whereas killing of T-lymphocytes seems to be mediated by combined (6-4) plus Dewar yields. These results emphasize the need to study the biological effects of UVB because cellular responses may be different from those following UVC irradiation.

Antibodies, Monoclonal↗
PubMed1994

Detection of active UV-photoproduct repair in monkey skin in vivo by quantitative immunohistochemistry.

Ultraviolet-induced cyclobutane pyrimidine dimers (CPDs) and pyrimidine-pyrimidone (6-4)photoproducts in DNA were quantitatively measured in monkey skin using an immunohistochemical method with two specific monoclonal antibodies. The skins of Cynomolgus monkeys (Macaca fascicularis) were irradiated with UV light and processed for preparation of conventional formalin-fixed, paraffin-embedded histological sections. Both of the photoproducts were detectable in the nuclei of epidermal cells at doses of 500 J/m2 for UVB and 50 J/m2 for UVC, respectively, nuclear staining being clearly dose-dependent. Time course studies also showed a statistically significant decrease in nuclear staining with time after exposure to either UVB or UVC irradiation. Although only 30% of CPDs were removed from DNA in the first 24 h, about half of the (6-4) photoproducts were repaired within 3 h post-UV irradiation. Staining completely disappeared by 48 h in the (6-4) photoproduct case and by 72 h in the case of CPDs. The results suggest that epidermal cells of monkey skin can efficiently repair UV-photoproducts in DNA, but that the capacity is slightly less than in man.

Animals↗
PubMed1994

Removal of UV-induced DNA lesions in mouse epidermis soon after irradiation.

Induction and removal of cyclobutane thymine dimers and (6-4)photoproducts were studied in epidermal DNA isolated from UV-exposed hairless mice. For the detection of DNA damage, lesion-specific monoclonal antibodies were used in an immunoslotblot assay. Following the exposure of mice to 3.0 kJ m-2 UV-B, substantial removal of both thymine dimers (66%) and (6-4)photoproducts (77%) was observed at 24 h after irradiation. No removal, however, was detected at 4 h after irradiation. In contrast, immunofluorescence data obtained previously showed a rapid initial dimer removal after irradiation with 1.0 kJ m-2 UV-B (A.A. Vink, R.J.W. Berg, F.R. De Gruijl, L. Roza and R.A. Baan, Carcinogenesis, 12 (1991) 861-864). Reinvestigation of the removal of dimers and (6-4)-photoproducts shortly after three different UV doses showed a rapid decreases of both lesions at 2 h after irradiation with 1.0 kJ m-2. The results obtained after irradiations with 2.0 and 3.0 kJ m-2 UV-B suggest a saturation of repair already at 2.0 kJ m-2. Cyclobutane dimers were found to be removed at a lower rate than (6-4)photoproducts.

Animals↗
PubMed1994

Photorepair and excision repair removal of UV-induced pyrimidine dimers and (6-4) photoproducts in the tail fin of the medaka, Oryzias latipes.

Induction and repair of UV-B induced DNa damage in the tail fin of the Medaka, were examined immunohistochemicaly and by the enzyme-linked immunosorbent assay (ELISA). UV-induced DNA damage was detected only in the outermost layer of epithelial cells and did not differ in fishes having different degree of melanization. Both pyrimidine dimers and (6-4) photoproducts in the fin cells were removed by excision repair in the dark, the excision of (6-4) photoproducts being about twice as efficient as that of pyrimidine dimers. The rate of excision repair of UV-induced lesions in fin tissue was three to four times that in cultured Medaka cells, OL32. In the fin cells, reductions in the numbers of pyrimidine dimers and (6-4) photoproducts were seen after treatment with fluorescent light, whereas less reductions of pyrimidine dimers and no reductions of (6-4) photoproducts were observed in OL32 cells.

Animals↗
PubMed1993

Identification of cellular defect in UVS1, a UV-sensitive Chinese hamster ovary mutant cell line.

UVS1 is an intermediately UV-sensitive Chinese hamster ovary mutant originally isolated by its hypersensitivity to an anticancer drug, 1-[(4-amino-2-methyl-5-pyrimidinyl)methyl]-3-(2-chloroethyl)-3-nitrosour ea hydrochloride. By cell fusion analysis, UVS1 complemented the UV sensitivity of the mouse lymphoma cell line US31 from the eighth complementation group of UV-sensitive rodent cell lines. By enzyme-linked immunosorbent assay we found that within 3 h after UV irradiation both pyrimidine dimers and (6-4)photoproducts in UVS1 were not removed from chromosomal DNA in UVS1 at all. Twenty-four h after UV irradiation the removal rate of (6-4)photoproducts was intermediate between CHO9, the parental cell line, and 43-3B, a UV-hypersensitive Chinese hamster ovary mutant of the complementation group 1, whereas the pyrimidine dimers in UVS1 were removed less efficiently as 43-3B. Alkaline elution assay showed that the incising activity to damaged DNA after UV irradiation of UVS1 was as low as that of 43-3B. The number of 1-[(4-amino-2-methyl-5-pyrimidinyl)methyl]-3-(2-chloroethyl)-3-nitrosour ea hydrochloride-induced DNA interstrand cross-links of UVS1 was almost equal to that of 43-3B and about 1.5 times more than that of CHO9, suggesting that the gene products defective in UVS1 and 43-3B are essential for the excision repair of DNA damages produced by 1-[(4-amino-2-methyl-5-pyrimidinyl)methyl]-3-(2-chloroethyl)-3-nitrosour ea hydrochloride.

Animals↗
PubMed1993

Hypersensitivity of human lymphocytes to UV-B and solar irradiation.

T-lymphocytes from three normal human donors, irradiated with broad-spectrum UV-B (peak emission, 312 nm), are 20-fold more sensitive than fibroblasts from four normal donors in a clonogenic assay. We have compared the formation of thymine cyclobutane dimers and pyrimidine-(6-4)-pyrimidone photoproducts following irradiation by UV-C (254 nm) and UV-B and studied killing at doses giving equal dimer formation. UV-B killing of fibroblasts appears to be associated with dipyrimidine photoproduct formation, whereas UV-B killing of lymphocytes is mediated by nondimer damage. Strand breakage following UV-B irradiation measured using the "Comet" assay (single cell gel electrophoresis) reflects this nondimer damage and has kinetics consistent with excisable damage. Lymphocytes from three excision-deficient xeroderma pigmentosum donors show reduced strand breakage and increased killing following UV-B irradiation, compared with lymphocytes from normal donors. We therefore suggest that UV-B kills human lymphocytes by excisable nondimer damage and that xeroderma pigmentosum lymphocytes are defective in its repair. The putative nondimer damage does not appear to be associated with radical attack, and the strand breakage is not a manifestation of apoptosis. A 1-min exposure of human lymphocytes in vitro to natural sunlight is sufficient to produce damage measurable by the Comet assay.

Apoptosis↗
PubMed1993

A new photoreactivating enzyme that specifically repairs ultraviolet light-induced (6-4)photoproducts.

Cyclobutane pyrimidine dimers (CPDs) and pyrimidine (6-4) pyrimidone photoproducts ((6-4)photoproducts) are the two major classes of cytotoxic, mutagenic and carcinogenic DNA photoproducts produced by ultraviolet light irradiation of cells. The phenomenon of photoreactivation, the reduction of the lethal and mutagenic effects of ultraviolet radiation by simultaneous or subsequent irradiation with near ultraviolet or visible light, has been identified in several organisms and in some cases the enzymes that catalyse this process have been characterized in sufficient detail. CPDs are the only known substrate for the photoreactivating enzymes so far analysed and enzymatic photoreactivation of (6-4)photoproducts has not yet been reported. We report here that an enzyme that catalyses the light-dependent repair of (6-4)photoproduct exists in Drosophila melanogaster. This is, to our knowledge, the first report of such photoreactivating activity specific for (6-4)photoproducts in any organism.

Animals↗
PubMed1993

A xeroderma pigmentosum complementation group A related gene: confirmation using monoclonal antibodies against the cyclobutane dimer and (6-4) photoproduct.

Xeroderma pigmentosum complementation group A was partially complemented by a cosmid genomic clone containing a 42-kb human DNA insert selected with a cDNA clone that we obtained through cDNA competition between the repair-proficient and repair-deficient cell line. The relationship between these two clones was confirmed using PCR amplifications. The enhancement in DNA-repair capacity of the transformants was assessed with the monoclonal antibodies specific for cyclobutane dimers and (6-4) photoproducts and partially correct the xeroderma pigmentosum complementation group A defect. Furthermore, the level of the photoproduct-repair capacity is in agreement with the survival enhancement calculated from the D37 values. This gene was mapped to chromosome 8, suggesting that this may represent one of the defective gene(s) in xeroderma pigmentosum complementation group A.

Antibodies, Monoclonal↗
PubMed1993

DNA damage in UV-irradiated human skin in vivo: automated direct measurement by image analysis (thymine dimers) compared with indirect measurement (unscheduled DNA synthesis) and protection by 5-methoxypsoralen.

The incidence of the various types of skin cancer in the general population has been increasing at an annual rate of 2-8% over the past 2 decades. In spite of considerable media coverage on the risk of skin cancer the acquisition of a suntan is still very popular. Thus the UV exposures required for tanning pose a serious carcinogenic risk, particularly to individuals who tan poorly. On the other hand, the presence of natural skin pigment, or the ability to tan easily can protect the skin against some of the harmful effects of subsequent UV exposures (Kollias et al. 1991). We have recently shown (Young et al. 1991) in human volunteers, using an indirect measurement of DNA damage (unscheduled DNA synthesis (UDS) detected by autoradiography), that a tan induced by UV in the presence of a UVB sunscreen (Parsol MCX) preparation containing 5-methoxypsoralen (5-MOP) is more effective at protecting the skin against a subsequent DNA-damaging challenge dose of UV than a tan induced by UV alone, particularly in individuals who tan poorly. No such protection was seen with the same sunscreen lacking 5-MOP. 5-MOP is an ingredient in natural citrus oils and in many other plants. Here we show the same pattern of protective action when measuring, for the first time, DNA damage directly using a monoclonal antibody to thymine dimers (a major category of DNA lesion induced by UV radiation) on fixed human skin sections and automated image analysis. There is a good correlation between UV exposure dose and the levels of thymine dimers in epidermal nuclei. The levels of thymine dimers (measured as absorption by the mean integrated optical density (IOD)) also correlated well with the levels of UDS (grains per nucleus). These findings are of importance in the comparative risk-benefit assessment of sunscreens with and without 5-MOP. The techniques described have applications for measuring other DNA lesions following UV and other exposures.

5-Methoxypsoralen↗
PubMed1993

Establishment and characterization of a monoclonal antibody recognizing the Dewar isomers of (6-4)photoproducts.

We established a monoclonal antibody (DEM-1) that recognizes UV-induced DNA damage other than cyclobutane pyrimidine dimers or (6-4)photoproducts. The binding of DEM-1 antibody to 254 nm UV-irradiated DNA increased with subsequent exposure to UV wavelengths longer than 310 nm, whereas that of the 64M-2 antibody specific for the (6-4)photoproduct decreased with this treatment. Furthermore, the increase in DEM-1 binding was inhibited by the presence of the 64M-2 antibody during the exposure. We concluded that the DEM-1 antibody specifically recognized the Dewar photoproduct, which is the isomeric form of the (6-4)photoproduct. The DEM-1 antibody, however, also bound to DNA irradiated with high fluences of 254 nm UV, suggesting that 254 nm UV could induce Dewar photoproducts without subsequent exposure to longer wavelengths of UV. Furthermore, an action spectral study demonstrated that 254 nm was the most efficient wavelength for Dewar photoproduct induction in the region from 254 to 365 nm, as well as cyclobutane dimers and (6-4)photoproducts, although the action spectrum values in the UV-B region were significantly higher compared with those for cyclobutane dimer and (6-4)photoproduct induction.

Animals↗
PubMed1993

Cloning of VH and VL genes for antibodies specific for pyrimidine photo dimers.

We have prepared two types of cDNAs encoding mouse immunoglobulin heavy (VH) and light (VL) chain variable (V) domains against UV-induced DNA damage by the polymerase chain reaction (PCR). The amplified cDNA was cloned for sequencing and expression, and the nucleotide sequences of the V domains were determined. The deduced amino acid sequences of the complementarity-determining regions (CDRs) indicated that CDR3 of the VH domain and CDR2 of the VL domain might be concerned with the recognition of photolesions.

Antibodies↗
PubMed1992

Repair of thymine dimers and (6-4) photoproducts in group A xeroderma pigmentosum cell lines harboring a transferred normal chromosome 9.

Transfer of a normal chromosome 9 into a xeroderma pigmentosum (XP)-A cell line partially restored its DNA repair activity. XP-A cell lines harboring a transferred chromosome were much more UV-resistant than parental XP-A cells but still more UV-sensitive than normal cells. The amount of UV-induced unscheduled DNA synthesis was only one-third of that in normal cells. The repair of thymine dimers and (6-4) photoproducts in these cell lines was analyzed by using monoclonal antibodies raised against them. Although these XP-A cell lines carrying a normal chromosome 9 could repair (6-4) photoproduct with a little lower efficiency than normal cells, the repair of thymine dimers was completely absent in these cells. The present results suggest a gene-dosage effect in DNA excision repair mechanisms in human cells or a rather complicated mechanism which involves two or more pathways.

Cell Line↗
PubMed1992

High resistance of cultured Mongolian gerbil cells to X-ray-induced killing and chromosome aberrations.

The Mongolian gerbil (Meriones unguiculatus) is known to be one of the most radioresistant animals. We have examined the X-ray sensitivity of normal diploid fibroblasts from Mongolian gerbil embryos compared with those of cultured embryo cells obtained from various laboratory animals and a normal human. There was a wide difference in X-ray sensitivity for cell killing among different mammalian species. The D0 values for Mongolian gerbil cells ranged from 2.08 to 2.28 Gy, values which are twice as high as those for human cells. The mean D0 value for human cells was 1.06 Gy. Mouse, rat, Chinese hamster, and Syrian/golden hamster cells showed similar D0 values ranging from 1.30 to 1.56 Gy. When cells were irradiated with X rays, ten times more chromosome aberrations were detected in human cells than in Mongolian gerbil cells. The frequencies of chromosome aberrations in other rodent cells were between the values for cells from humans and those from gerbils. These data indicate that the Mongolian gerbil cells are resistant to X-ray-induced cell killing and chromosome aberrations, and that the radiation sensitivity of mammalian cells in primary culture may be reflected by their radioresistance in vivo.

Animals↗
PubMed1992

Further characterization of monoclonal antibody indicates specificity for (6-4)-dipyrimidine photoproducts.

Monoclonal antibody aUVssDNA-1 is produced by hybridoma cell line 25JF.C3B6 originally selected from cell fusions using spleen cells from mice immunized with UV-irradiated polydeoxynucleotides (Strickland and Boyle, Photochem. Photobiol. 34, 595-601, 1981). Original and subsequent studies of the binding characteristics of aUVssDNA-1 indicated that it was specific for cyclobuta-dithymidine photoproducts. Those investigations examined action spectrum, short-wavelength photo-reversal, nucleotide sequence effects, and photoreactivation using E. coli photolyase and incandescent light. However, the more recent studies reported here examined acetophenone-UV-B photosensitization, UV-B photoisomerization, and photoreactivation using cloned E. coli photolyase and filtered incandescent light. The results indicate that aUVssDNA-1 recognizes photoproducts with characteristics of (6-4)-dipyrimidines. Thus, previous studies in which relatively rapid repair of cyclobuta-dithymidine photoproducts was inferred using this antibody, require re-interpretation in light of these new findings.

Animals↗
PubMed1991

Specificity of the sex-influenced esterase (ES-SI) isozyme in rat liver.

A carboxylesterase which shares common antigenicity with sex-influenced esterase (ES-SI) was found in both the male and the female liver of the rat but not in the following tissues: erythrocyte, heart, kidney, lung, spleen, small intestine, testis, thymus, and lymph node. Subcellular fractionation showed the esterase localizes in the microsome-rich fraction. The strain distribution of the presence or absence of the esterase in inbred rats was identical to that of ES-SI, although in adult males a considerable amount of the esterase exists, unlike ES-SI. The esterase had a higher isoelectric point than ES-SI but after neuraminidase treatment the difference disappeared, suggesting that the esterase has a sialic acid moiety. Because this esterase has different properties from those previously reported, it is proposed that it is designated liver-ES-SI. The common antigenicity and similar strain distribution between ES-SI and liver-ES-SI suggest that liver-ES-SI is a precursor molecule of ES-SI and therefore the two esterases are products of a single gene.

Animals↗
PubMed1991

Suppression of differentiation phenotypes in myogenic cells: association of aneuploidy and altered regulation of c-myc gene expression.

We isolated 10 myoblast clones from Syrian/golden hamster embryo (SHE) cells irradiated with ultraviolet light. They were originally isolated as anchorage-independent clones. All clones showed characteristic morphology of myoblast in culture and formed swirled myofiber colonies specific to myoblast macrocolonies. However, in immunostaining experiments using anti-fast myosin antibody, we found that 4 of 10 myoblast clones were myosin negative (Myo-), while six were myosin positive (Myo+). Western blot analysis confirmed the disappearance of the 200-kDa myosin-specific band in the 4 Myo- clones. Furthermore, these 4 Myo- clones lost the ability to form multinucleated myotubes. Karyotype analysis revealed that all Myo- clones had trisomy of chromosome 7, while Myo+ clones showed no apparent karyotypic change from normal SHE cells. Compared with that of SHE cells, the transcriptional level of the myc gene in Myo+ clones was augmented, but there was no increase of myc gene expression in Myo- clones. Furthermore, the introduction of activated myc gene partially converted the Myo- phenotype to Myo+. These results suggest that trisomy of chromosome 7 and a deficiency in enhanced expression of the myc oncogene are associated with the suppression of both the production of myosin and the formation of multinucleated cells.

Aneuploidy↗
PubMed1991

Establishment of a monoclonal antibody recognizing cyclobutane-type thymine dimers in DNA: a comparative study with 64M-1 antibody specific for (6-4)photoproducts.

We obtained a monoclonal antibody (TDM-1) binding to 313-nm UV-irradiated DNA in the presence of acetophenone. The binding of TDM-1 to 254-nm UV-irradiated DNA was not reduced with the subsequent irradiation of 313-nm UV. Furthermore, the treatment of UV-irradiated DNA with photolyase from E. coli and visible light exposure reduced both the antibody binding and the amount of thymine dimers in the DNA. A competitive inhibition assay revealed that the binding of TDM-1 to UV-irradiated DNA was inhibited with photolyase, but not with 64M-1 antibody specific for (6-4)photoproducts. These results suggest that TDM-1 antibody recognizes cyclobutane-type thymine dimers in DNA. Using TDM-1 and 64M-1 antibodies, we differentially measured each type of damage in DNA extracted from UV-irradiated mammalian cells. Repair experiments confirm that thymine dimers are excised from UV-irradiated cellular DNA more slowly than (6-4)photoproducts, and that the excision rates of thymine dimers and (6-4)photoproducts are lower in mouse NIH3T3 cells than in human cells.

Acetophenones↗