PubMed Health⌕ Search

Biomedical subjects

M D Shelby

Publications and source records attributed to M D Shelby.

At least 19 recordsLinked to original sources
PubMed1995

Study design and sample sizes for a lacI transgenic mouse mutation assay.

Design features that adjust and account for excess variation in a transgenic mouse mutation assay based on a lacI target transgene from E. coli are considered. These features include proper identification of plate, packaging reaction, and animal identifier codes throughout the experimental and analysis phases of the study, "blocking" of exposed and unexposed animals when preparing and plating multiple packaging reactions from the same genomic DNA sample, separating sectored mutant plaques and complete mutant plaques before performing any quantitative analyses, and testing for sources of excess variation attributable to features of the experimental protocol--such as plate-to-plate (within packaging reactions), packaging reaction-to-packaging reaction (within animals), and animal-to-animal (within study). Control and ethylnitrosourea-treated animal data are presented from a fully designed study in the lacI assay. The study design incorporates many of these experimental principles. Statistical methods to identify excess variability are noted, and the designed study data are used to illustrate the types of variability encountered in practice. A standard statistical test for two-sample testing is highlighted, from which recommendations are made for sample size selection in future studies.

Analysis of Variance↗
PubMed1995

Comparison of results from mouse bone marrow chromosome aberration and micronucleus tests.

Tests for the induction of chromosomal aberrations (ABS) and micronuclei (MN) in bone marrow cells of mice have been conducted on 65 chemicals. Although these tests were not conducted with the purpose of comparing the outcomes of these two in vivo genetic toxicity endpoints, the availability of these test results permits such a comparison. Based on studies to date, results from the 2 tests agree for more than 80% of the chemicals; 17 gave positive results in both tests, and 36 gave negative results in both. Seven chemicals were positive only for ABS and 5 were positive only for MN. Three chemicals that were originally concluded to be positive for ABS but not for MN were found to induce MN when the MN protocol was modified to more closely reflect the ABS protocol. Among the 12 chemicals for which there are discrepant results, there are only 2 for which the difference is convincing. One of these, selenium sulfide (MN negative, ABS positive) remains an enigma; further studies are being conducted. The second, isoprene (MN positive, ABS negative) will be difficult to pursue because the studies reported here were done by inhalation exposure. Based on the outcomes of these comparisons, protocol factors, rather than endpoint specificity, appear to be the major source of discrepant test results. Thus, these results do not support a recommendation that both tests be conducted in a primary testing scheme for genetic toxicity.

Animals↗
PubMed1995

Mutagenesis and human genetic disease: an introduction.

This special issue attempts to provide a fresh perspective on the importance of germ-cell mutagenesis studies and restate the questions and challenges inherent in efforts to minimize the incidence of human genetic diseases. We are working in a time when rapidly advancing molecular technologies provide the tools that permit a more detailed understanding of germ-cell mutagenesis and genetic disease. Meanwhile, discoveries of new genetic disease phenomena challenge our abilities to conceive and develop research models for their study. It is hoped that the collection of articles in this issue will serve to stimulate interest in scientists of varied disciplines and help focus those interests on the issues surrounding the relationship between environmental mutagens and human genetic disease.

Aneuploidy↗
PubMed1995

Phenolphthalein: induction of micronucleated erythrocytes in mice.

Phenolphthalein was tested for the induction of micronucleated erythrocytes in mice. Results of an initial investigation revealed significant, dose-related increases in micronucleated polychromatic erythrocytes (MN-PCE) and normochromatic erythrocytes (MN-NCE) in peripheral blood samples of male and female mice exposed to 0.6% to 5% phenolphthalein (approximately 1100 to 10,000 mg/kg/day) in feed for 90 days (Dietz et al., 1992). Results from a second long-term feed study with Swiss CD-1 mice confirmed this effect. However, administration of comparable doses of phenolphthalein by corn oil gavage on two consecutive days gave negative results in a mouse bone marrow micronucleus test. Subsequent tests were performed to clarify the conflicting results seen in the chronic exposure, dosed-feed, peripheral blood studies and the acute, corn oil gavage, bone marrow studies. Phenolphthalein was administered to male B6C3F1 mice in feed (3%) for 14 days. Peripheral blood samples taken at 4, 7, and 14 days all showed significant increases in micronucleated PCE; bone marrow samples taken on days 7 and 14 also were clearly positive for micronucleus induction. Therefore, comparable results were obtainable from both bone marrow and peripheral blood analyses. Because of the negative results in the two-exposure gavage test, additional tests were then designed to investigate the effects of bolus vs continuous dosing, feeding vs gavage administration, and corn oil vs feed as a carrier for phenolphthalein. Results of these tests indicated that the rate of exposure to phenolphthalein affects the frequency of induced MN-PCE and that micronucleated erythrocytes can be induced by phenolphthalein either by feeding or by corn oil gavage administration. In all the acute exposure studies, relatively high doses of phenolphthalein (2000-6000 mg/kg/day for at least 2 days) were required to induce micronuclei. The positive results obtained with phenolphthalein in vivo were consistent with the results of an in vitro chromosomal aberration test in Chinese hamster ovary cells, where dose-related increases in aberrations were noted only in cells treated in the presence of induced rat liver S9.

Animals↗
PubMed1995

Dominant lethal and heritable translocation tests with chlorambucil and melphalan in male mice.

Chemicals used in the treatment of cancer include several that are potent mutagens in a range of in vitro and in vivo assays. For some, genetic effects have also been demonstrated in humans, detected as chromosomal aberrations in peripheral lymphocytes. Because (1) many of these agents are confirmed mutagens, (2) humans are exposed to them in relatively high doses, and (3) an increasing number of early cancer victims are surviving to reproductive age, it is important that information be available on the genetic and reproductive hazards associated with exposure to these agents. Chlorambucil and melphalan are structurally related chemicals that are included in our efforts to identify and assess such hazards among cancer chemotherapy agents. To date, both have been reported to induce specific locus mutations in germ cells of male mice (Russell et al., 1989; Russel et al., 1992b) and melphalan is one of very few chemicals shown to induce such mutations in spermatogonial stem cells. More recently, both chemicals were found to have strong reproductive effects in female mice (Bishop and Generoso, 1995, in preparation). In the present studies, these chemicals were tested for the induction of dominant lethal mutations and heritable translocations in male mice. Both chemicals were found to have reproductive effects attributable to cytotoxicity in specific male germ cell stages and to induce dominant lethal mutations and heritable translocations in postmeiotic germ cells, particularly in mid to early stage spermatids. Thus, relatively extensive data are now available for assessing the genetic and reproductive hazards that may result from therapeutic exposures to these chemicals.

Animals↗
PubMed1994

Sources of variability in data from a lacI transgenic mouse mutation assay.

Experimental features of a transgenic mouse mutation assay based on a lacI target transgene from Escherichia coli are considered in detail. Sources of variability in the experimental protocol that can affect the statistical nature of the observations are examined with the goal of identifying sources of excess variation in the observed mutant fractions. The sources include plate-to-plate (within packages), package-to-package (within animals), and animal-to-animal (within study) variability. Data from two laboratories are evaluated, using various statistical methods to identify excess variability. Results suggest only scattered patterns of excess variability, except possibly in those cases where genomic DNA from test animals is stored for extended periods (e.g., > 90 days) after isolation from tissues. Further study is encouraged to examine the validity and implications of this time/storage-related effect.

9,10-Dimethyl-1,2-benzanthracene↗
PubMed1994

Human germ cell mutagens.

Human genetic disorders constitute a major public health burden in this country and around the world. The possibility that exposures to mutagenic environmental agents may result in induced genetic damage in human germ cells and thereby increase the incidence of genetic disease has been investigated in research laboratories and in epidemiology studies for decades. The capacity of ionizing radiation and some chemicals to induce transmissible genetic damage in the germ cells of laboratory mammals has been clearly demonstrated and extensively investigated. To date, no clear evidence of such effects in humans has been reported although increased frequencies of chromosomal aberrations have been detected in human sperm following exposures to radiation or chemotherapeutic agents. Evolving methods to detect molecular changes in DNA offer to improve our abilities to detect induced genetic changes. The integration of these methods into mutation epidemiology studies promises to help resolve some of the questions regarding human genetic risk.

Animals↗
PubMed1994

Summary report of the Working Group on Mammalian Germ Cell Tests.

The two tests considered by the Working Group were the mammalian germ cell cytogenetic assay and the rodent dominant lethal test. It was agreed that both tests were mainly used for identification of germ cell hazards, however, that the commonly applied protocol of the dominant lethal assay often supplied information for hazard characterization such as sensitivity of particular developmental stages of male germ cells. No particular species or strains were indicated. Concurrent solvent controls were regarded as indispensable for both tests. In the discussion of the mammalian germ cell cytogenetic assay, harmonization was obtained to a large extent with the cytogenetic bone marrow assay regarding the number of animals (5), the number of cells analyzed per animal (200), the highest exposure dose (MTD) and sampling times (twice within 24 and 48 h after dosing). However, it was pointed out that only the single acute exposure was adequate for the mammalian germ cell cytogenetic assay. Furthermore, it was stated that only structural chromosome aberrations could be analyzed and that it was not informative to score polyploidies or aneuploidies. In the discussion of the rodent dominant lethal test, it was stated that the assay was generally performed with treated males, however, increasing concern about female specific effects required that a protocol for female dominant lethal testing should be developed and validated. Acute and subacute treatment schedules were considered equally acceptable. It was regarded as highly important that the entire male germ cell development from meiosis to mature sperm was covered in the test protocol either by the appropriate mating schedules after single dosing or by subchronic dosing during the respective period. Postimplantation loss, preimplantation loss and fertility rate were the main parameters to be assessed in the rodent dominant lethal tests. It was agreed that the size of the experiment depended on the spontaneous frequency of dead implants, the mating scheme and the statistical design of the experiment.

Aneuploidy↗
PubMed1993

Evaluation of a three-exposure mouse bone marrow micronucleus protocol: results with 49 chemicals.

Forty-nine chemicals were tested in a mouse bone marrow micronucleus test that employed three daily exposures by intraperitoneal injection. Bone marrow samples were obtained 24 hr following the final exposure. Twenty-five rodent carcinogens and 24 noncarcinogens were selected randomly from the 44 carcinogens and 29 noncarcinogens used by Tennant et al. (Science 236:933-941, 1987) to evaluate the performance of four in vitro genetic toxicity tests. As in that study of in vitro tests, the micronucleus tests were conducted with coded chemicals and test results (positive or negative) were determined prior to decoding. This study was conducted as part of an effort to assess the ability of the micronucleus test to discriminate between rodent carcinogens and noncarcinogens and to determine its potential role, in combination with other short-term tests, in identifying genotoxic chemicals that present a carcinogenic hazard. Nine chemicals were judged to be positive in the micronucleus test. This relatively low number of positive results, along with published and unpublished results from rodent micronucleus and chromosome aberration assays on several of these 49 chemicals, contributed to the conclusion that a single micronucleus test protocol is not adequate to detect all chemicals capable of inducing chromosomal damage in the bone marrow. However, a combination of two relatively simple assays such as the Salmonella and micronucleus tests can provide important information on the genetic toxicity of test chemicals and may provide guidance on the need for and the nature and extent of future toxicity studies.

Animals↗
PubMed1993

Genetic toxicity of fluoride.

F- is not mutagenic in standard bacterial systems, but produces chromosome aberrations and gene mutations in cultured mammalian cells. Although there is disagreement in the literature concerning the ability of F- to induce chromosome aberrations in cultured human and rodent cells, the weight of the evidence leads to the conclusion that F- exposure results in increased chromosome aberrations in these test systems. NaF induced primarily chromatid gaps and chromatid breaks, indicating that the rodent cells are responsive in the G2 stage of the cell cycle. In contrast, studies with synchronized human cells indicated that the S phase was the most sensitive. If F- does have a cell cycle-specific effect, it could be expected that differences in the cell treatment and harvest protocols could lead to conflicting results for the induction of chromosome aberrations. Gene mutations were produced in cultured rodent and human cells in the majority of the studies. Unfortunately, a number of the in vitro and in vivo cytogenetic studies are of questionable utility because of the protocols used, the quality of the responses reported, or the interpretations of the data. The conflicting results in the in vivo cytogenetic studies are difficult to reconcile. There are reports of increased chromosome aberrations in rat bone marrow and testes, but other studies, using similar protocols and dose ranges, have reported no induced chromosome damage. Although some of the studies were performed at toxic levels of F-, other studies, including those that showed positive results, were at F- concentrations (1-5 ppm) equivalent to human exposure levels. In the majority of studies that were reported to be positive, there were high background frequencies, or the investigators reported categories of nuclear or chromosome damage that are difficult to interpret. Interestingly, many of the positive results were obtained when anaphase cells were scored, whereas similar treatment protocols in other laboratories yielded negative results when metaphase cells were the only cell type examined. It is difficult, without additional data, to determine the reasons for finding chromosome breaks in anaphase, but not metaphase, cells. Other reports have presented insufficient information to allow adequate evaluations. Therefore, at this time, the question of whether F- produces chromosome damage in vivo should be considered unresolved.

Animals↗
PubMed1993

Fertility, reproduction, and genetic disease: studies on the mutagenic effects of environmental agents on mammalian germ cells.

Because genetically based diseases have a major impact on human health, the National Institute of Environmental Health Sciences (NIEHS) has conducted a research and testing program for more than a decade to address chemical induction of heritable genetic damage in the germ cells of mammals. Although most genetic disease results from preexisting mutations, a portion is due to the occurrence of new mutations. The supposition that exposure to mutagenic chemicals contributes to the occurrence of new mutations in the human population is strongly supported by the results from animal models. Such studies clearly demonstrate the potential of environmental chemicals to induce mutations in both somatic and reproductive cells of mammals. This NIEHS program has become a leader in the identification of genetic hazards in the environment and in the acquisition of animal model data used by regulatory agencies in assessing genetic risks to human health.

Animals↗
PubMed1992

ENU mutagenesis in the mouse electrophoretic specific-locus test. 2. Mutational studies of mature oocytes.

Experiments were conducted using the biochemical specific-locus test to assess the mutagenicity of N-ethyl-N-nitrosourea (ENU) in mature oocytes of mice. C57Bl/6J females were treated with 100 mg/kg ENU by intraperitoneal injection and mated to untreated DBA males for 1 week following treatment. 1447 progeny were screened for evidence of mutations affecting the electrophoretic mobility of 32 different proteins; two mutants were detected by electrophoretic analyses. These results provide evidence that ENU is a germ-cell mutagen in mouse mature oocytes, although the frequency of mutants is somewhat lower than that obtained from spermatogonia treated with the same dose.

Animals↗
PubMed1992

In vivo cytogenetic activity of diphenylhydantoin in mice.

Diphenylhydantoin was tested in vivo in mice using a variety of cytogenetic endpoints to evaluate its genotoxicity. Injected doses of 125, 250 and 500 mg/kg failed to increase the number of chromosome aberrations in marrow cells at 17 h post-treatment, and 37.5, 75 and 150 mg/kg doses were likewise ineffective at 36 h. SCEs were significantly increased by doses of 125 mg/kg (but not 250 mg) after 23 h and modestly, in relation to dose, at 42 h. No increase in the number of micronuclei among marrow PCEs was seen following single i.v. injections ranging from 0.1 to 20 mg/kg. Three daily i.p. injections of doses up to 70 mg/kg also failed to increase the number of micronuclei in either marrow or peripheral blood PCEs. Some cytotoxic effect was evident following relatively high doses.

Animals↗
PubMed1992

Ethylene dibromide: negative results with the mouse dominant lethal assay and the electrophoretic specific-locus test.

Ethylene dibromide (1,2-dibromoethane; EDB) was tested for the induction of dominant lethal and electrophoretically-detectable specific-locus mutations in the germ cells of DBA/2J male mice. Males were treated with a single intraperitoneal injection of 100 mg/kg EDB and mated to two C57BL/6J females. In the dominant lethal assay, matings were carried out to measure the effect of EDB on meiotic and postmeiotic stages; germ cells representing spermatogonial stem cells were analyzed in the electrophoretic specific-locus test. Neither of these germ cell tests produced any evidence that EDB is a germ cell mutagen. It appears from these data and those reported in the literature that EDB, a genotoxic carcinogen that affects male fertility in some mammalian species, is not mutagenic in the germ cells of the male mouse.

Animals↗
PubMed1992

Melphalan, a second chemical for which specific-locus mutation induction in the mouse is maximum in early spermatids.

Melphalan (MLP), a bifunctional alkylating agent structurally related to the highly mutagenic chemical chlorambucil (CHL), was found to induce high frequencies of specific-locus mutations in postspermatogonial germ cells of the mouse, and to be one of only a few chemicals that is also mutagenic in spermatogonial stem cells. Productivity patterns following MLP exposures resembled those that had been found for CHL. Mutation rates in successive male germ-cell stages were measured at three MLP-exposure levels in a total of 95,375 offspring. While the induced (experimental minus historical-control) mutation rate is relatively low in stem-cell spermatogonia (1.2 x 10(-5) per locus at a weighted-mean exposure of 7.3 mg/kg), it is about 5 times higher in poststem-cell stages overall, and peaks at 26.7 x 10(-5) per locus in early spermatids at a weighted-mean exposure of only 5.7 mg/kg. This "type-2 pattern" of mutation yield (Russell et al., 1990), i.e., peak sensitivity in early spermatids, has heretofore been found for only one other chemical, CHL. Mutation-rate data earlier reported for CHL (Russell et al., 1989) were augmented in the present study for comparison with MLP-induced rates. Because of the greater toxicity of MLP, average exposures used for this chemical were only about one-half of those for CHL. When MLP and CHL mutation rates are extrapolated to equimolar doses, they appear very similar for poststem-cell stages overall. However, in the case of CHL, a somewhat higher proportion of the mutations is induced in early spermatids than in the case of MLP.

Animals↗