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Biomedical subjects

F Mendez

Publications and source records attributed to F Mendez.

At least 37 records · Page 2Linked to original sources
PubMed1986

X-ray-induced base sequence damage in primate alphoid DNA.

Radiation-induced single-strand breaks were found throughout the 172 bp repeat units of African green monkey component alpha DNA. Two kinds of 3'-ends of 5'-32P-labeled restriction fragments were found, as previously described by others. After irradiation in vitro, the yield of single-strand breaks was 4 X 10(-5) breaks/nucleotide/Gy, as determined by analyses in DNA sequencing type gels. Protection from X-ray damage was found when the DNA received 150 Gy in the presence of 2-mercaptoethanol. The results demonstrate a very sensitive quantitative means to study the role of indirect effects of ionizing radiation on strand-break induction and protection at the base sequence level. Component alpha DNA was isolated from irradiated CV-1 cells and was analyzed for single-strand breaks. Under these conditions the frequency of breaks was less than the frequency obtained when purified DNA was irradiated. The methodology is presented because of its relevance to the study of DNA strand breakage in living cells.

Animals↗
PubMed1985

IgG binding enhances DNAase I sensitivity of N-acetoxy-N-2-acetylaminofluorene-modified phi X-174 RF DNA.

DNA restriction fragments of phi X-174 RF were modified with the carcinogen, N-acetoxy-N-2-acetylaminofluorene (N-Aco-AAF). Immune complexes of 5'-32P-labeled AAF-modified DNA and rabbit immunoglobulin (IgG) against AAF-guanosine were specifically bound by surface membranes of Cowan I strain micrococci whose protein A binds the Fc portion of IgG. DNAase I sensitivity of the bound DNA was 20-fold greater than in solution, but the normal pattern of hydrolysis was not altered, as determined in sequencing gels. Nonadducted DNA ligated to AAF-modified DNA acquired the enhanced sensitivity to DNAase I hydrolysis when the ligation hybrid was immunobound.

2-Acetylaminofluorene↗
PubMed1983

Site specific cleavage of phi X-174 replicative form DNA after modification by N-acetoxy-N-2-acetylaminofluorene.

Three kinds of structural disturbances were found in an 88 base pair (bp) fragment of phi X-174 DNA after exposure to N-acetoxy-N-2-acetylaminofluorene (N-Aco-AAF). (i) Frequent strand scissions at two specific guanine sites on the 5' 32P-end-labeled fragment were identified by base sequence analysis. Scissions at these two sites were induced at neutral pH and they were not increased by treatment with apurinic endonuclease. They are an immediate consequence of N-Aco-AAF action and are not primarily apurinic sites. (ii) Alkali treatment with 1 M piperidine at 90 degrees C induced strand scissions at every guanine, demonstrating adduct slices, depurination and strand scissions. (iii) Adducted DNA was sensitive to single-strand specific nuclease digestion, suggesting unwound DNA. These studies indicate the prediliction of N-Aco-AAF for certain DNA sites and they suggest three kinds of DNA modifications which can be expected after adduction by this carcinogen. Some of the sites may be premutational carcinogen-induced DNA structural modifications.

2-Acetylaminofluorene↗
PubMed1982

Morphological transformation, DNA strand separation, and antinucleoside immunoreactivity following exposure of cells to intercalating drugs.

The intercalating drugs quinacrine and proflavine induced increases in single-stranded DNA detected in the nuclei of mouse BALB/c 3T3 1--13 cells. The denatured DNA was detected by fluorescein-labeled antinucleoside antibodies, which bind to single-stranded but not double-stranded DNA. Exposure of cells to the potent mutagen proflavine increased the fraction of immunoreactive nuclei from 0.65 to 0.8. With the weaker mutagen quinacrine, higher concentrations were needed to induce increases in immunoreactivity. Both intercalating drugs rapidly induced morphological transformation in mouse 3T3 cells. Treatment with proflavine resulted in higher transformation frequencies than were found with quinacrine. Significant increases in cell transformation frequency were observed at the concentrations which induced high levels of immunoreactivity. These results suggest that DNA strand separation is itself, or at least accompanies, an early step in cell transformation by intercalating drugs.

Animals↗
PubMed1981

Immunocytological detection of AAF-DNA adducts in HeLa cell nuclei.

Acetylaminofluorene-DNA adducts (AAF-DNA) were detected in the nuclei of HeLa cells exposed to N-acetoxy-2-acetylaminofluorene (N-Ac-AAF), using an immunocytological technique and specific antibodies directed against AAF modified DNA. The proportion of cells exhibiting specific nuclear immunoreactivity was dose-dependent. The time course of disappearance of adduct specific nuclear immunoreactivity was compared with removal of N-(deoxyguanosin-8-yl)-2-acetylaminofluorene (dG-C8-AAF) and other adducts.

2-Acetylaminofluorene↗
PubMed1980

DNA of HeLa cells during caffeine-promoted recovery from X-ray induced G2 arrest.

Progression of X-irradiated HeLa cells from G2 arrest through mitosis was promoted by 1mM caffeine. Caffeine promoted the return from abnormally high levels of radiation-induced immunoreactivity to antinucleoside antibodies, which indicates persistent DNA strand separation, to the low levels normally found in G2. With caffeine, the irradiated cells progressed through mitosis, producing daughter cells with the normal G1 content of DNA. Without caffeine, the DNA content of individual radiation-arrested cells retained G2 values and the abnormally high levels of immunoreactivity to antinucleoside antibodies.

Antibodies, Neoplasm↗
PubMed1979

Mutagen-induced disturbances in the DNA of human lymphocytes detected by antinucleoside antibodies.

The alkylating mutagens N-methyl-N'-nitro-N-nitrosoguanidine, methyl methanesulfonate, and N-nitroso-methylurea induced immunoreactivity to antinucleoside antibodies in human peripheral blood lymphocytes in vitro. This could also be detected in lymphocytes taken from a patient soon after i.v. administration of cyclophosphamide. The immunoreactivity response, which indicates denatured DNA or DNA single-strand breaks, was scored by immunofluorescent or immunoperoxidase techniques. Examination of blood from 10 normal subjects showed that 32 +/- 4% (S.E.) of resting peripheral blood lymphocytes were immunoreactive to antinucleoside antibodies. We have shown that these naturally occurring immunoreactive lymphocytes are largely accounted for by a subpopulation of thymus-derived lymphocytes bearing the Fc receptor for immunoglobulin M. The presence of these cells did not interfere with the use of peripheral blood lymphocytes for in vitro measurement of additional immunoreactivity caused by alkylating mutagens. The response proved to be dose dependent; up to 90% of lymphocytes could be rendered immunoreactive. Parallel studies with HeLa cells showed a similar dose-response relationship between mutagen action and immunoreactivity. With some agents, the immunoreactivity technique detected effects at lower concentrations than could be detected by HeLa cell survival studies. With N-nitrosomethylurea, measurement of DNA repair synthesis by [3H]thymidine autoradiography showed that in HeLa cells these two parameters of response to DNA damage increased in parallel. Our results provide a new basis for detecting the action of alkylating mutagens on human lymphocytes in vitro or in vivo.

Alkylating Agents↗
PubMed1978

Early steps in mutagenesis by hycanthone.

Hycanthone, the most potent mutagen in a series of nine thiaxanthenones, is a potent inducer of nuclear immunoreactivity to antinucleoside antibodies in HeLa cells. This response indicates exposure of single-stranded DNA regions. All classes of mutagens thus far tested share this property with hycanthone. Immunoreactivity to antinucleoside antibodies was also induced by brief exposure to hycanthone, 3 microgram/ml, in human fibroblasts from three normal subjects and in fibroblasts from seven patients with DNA repair deficiencies. Unlike those of many other mutagens, the metabolic effects and immunoreactivity induction of hycanthone were readily reversible. No evidence for covalent attachment of [3H]hycanthone to HeLa macromolecules could be found. Induction of DNA repair synthesis could not be detected by autoradiography after exposure of cells to hycanthone. Exposure of single-stranded DNA regions appears to be an important feature of the mechanism of action of hycanthone as a mutagen. Both hycanthone and lucanthone intercalate with DNA, but hycanthone was much less active than was lucanthone in reducing the rapid sedimentation of cell lysate DNA in alkaline sucrose gradients. Similarities and differences, therefore, have been found in the way the potent and the weak mutagen affect DNA of HeLa cells. This may provide clues to understanding the mechanism of mutagenesis by thiaxanthenones and other mutagens.

Antibodies↗
PubMed1976

Deficit in DNA content relative to histones in X-irradiated HeLa cells.

The DNA and histone content of HeLa S-3 cell cultures was measured by direct mass assays 21 hours after 1000 rad of X-irradiation, when the cells were arrested in G2 phase. The nuclear DNA content of such cultures was found to be deficient (73% of control values). In contrast, the synthesis of nuclear histones persisted, and the total histone content was close to 100% of control values. When synchronously-growing cultures were irradiated in mid-S phase and examined 3-5 hours later in G2 phase, both DNA and histone content were equal to control values.

DNA↗