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R W Tennant

Publications and source records attributed to R W Tennant.

At least 37 records · Page 2Linked to original sources
PubMed1998

Effect of the viable-yellow (A(vy)) agouti allele on skin tumorigenesis and humoral hypercalcemia in v-Ha-ras transgenic TGxAC mice.

We previously reported that papillomas can arise from the follicular epithelium of v-Ha-ras transgenic TGxAC mice. Since the viable-yellow mutation (A(vy)) of the mouse agouti gene which regulates coat color pigmentation by acting within the micro-environment of the hair follicle has been shown to function as a tumor promoter in the liver, we hypothesized that it may also play a role in TGxAC skin tumorigenesis. Endogenous agouti protein product was detected in the outer root sheath of anagen hair follicles following plucking of the hair shaft, but not in the interfollicular epithelium, in TGxAC mice on an FVB/N genetic background. It was also detected in papillomas from these mice produced by 12-O-tetradecanoylphorbol-13-acetate (TPA) treatment or plucking. Expression of the A(vy) allele in the v-Ha-ras transgenic TGxAC mouse line results in an approximately 2-fold increase in papilloma development compared with controls which did not carry the A(vy) allele following twice-weekly treatment with 1.25, 2.5 or 5.0 microg TPA. In addition, TPA-treated, papilloma-bearing F1 mice which carried the A(vy) allele, but not F1 mice which did not carry the A(vy) allele, exhibited a syndrome of humoral hypercalcemia mediated by parathyroid hormone-related protein (PTHrP) that led to weight loss, hypercalcemia and hypophosphatemia. Thus, we conclude that the A(vy) allele can influence the development of skin tumors and PTHrP-mediated humoral hypercalcemia in v-Ha-ras transgenic TGxAC mice.

Agouti Signaling Protein↗
PubMed1998

Evaluation and validation issues in the development of transgenic mouse carcinogenicity bioassays.

Transgenic mouse models have emerged as plausible alternatives to long-term bioassays for carcinogenicity. Three transgenic lines evaluated to date have shown a clear capability to discriminate between carcinogens and noncarcinogens, using long-term bioassay results as the standard. The data also suggest that the transgenic lines will not fully duplicate long-term bioassay results. It is proposed that these models do not respond to chemicals that have induced highly restricted species or strain-specific tumor responses in mice or rats. Rather, the value of the transgenic models is predicated on a preferential response to transspecies carcinogens (i.e., those positive in both rats and mice, often including tumors in the same tissues). Thus, although results in transgenic models may not be completely concordant with long-term bioassays, the data can be used effectively in chemical and drug safety assessments. Further, it is proposed that validation of the models is readily achievable via ongoing studies. Validation of any alternative model is best achieved by sufficient mechanistic understanding of the model to reasonably predict the outcome of bioassays conducted in the models and use all available information on the drug or chemical. This goal can now be met with the transgenic mouse lines.

Animal Testing Alternatives↗
PubMed1997

Kinetics of wound-induced v-Ha-ras transgene expression and papilloma development in transgenic Tg.AC mice.

The Tg.AC transgenic mouse, which harbors an activated v-Ha-ras coding region that is fused to an embryonic zeta globin transcriptional control region and a 3' simian virus 40 polyadenylation sequence, rapidly develops epidermal papillomas in response to topical application of chemical carcinogens or tumor promoters or to full-thickness wounding of the dorsal skin. In this report, we investigated the localization and temporal induction of v-Ha-ras transgene expression after full-thickness wounding of Tg.AC mouse skin. Surgically inflicted full-thickness incisions 3 cm long yielded four to six papillomas per Tg.AC mouse by 5 wk after wounding. Similar wounding of the FVB/N isogenic host strain did not produce tumors, which implicates a causal role for the v-Ha-ras transgene. Reverse transcription-polymerase chain reaction assays detected the v-Ha-ras transgene transcript in total RNA samples isolated from wound-associated tissue 3 and 4 wk after wounding. Tissues 1-2 wk after wounding and all non-wound-associated tissues were negative for transgene expression. In situ hybridization experiments using transgene-specific 35S-labeled antisense RNA probes localized transgene expression to the basal epidermal cells in wound-induced papillomas. Adjacent normal and hyperplastic skin tissues were negative for transgene expression by this assay. This work supports the hypothesis that the wound repair response leads to the transcriptional activation and continued expression of the v-Ha-ras transgene in specific cells in the skin, which alters normal epithelial differentiation and ultimately results in neoplastic growth.

Animals↗
PubMed1997

Decreases in phorbol ester-induced papilloma development in v-Ha-ras transgenic TG.AC mice during reduced gene dosage of bcl-2.

We have demonstrated that induction of transgene expression in the v-Ha-ras-transgenic TG.AC mouse is a critical event in skin tumorigenesis and that cutaneous papillomas arise from follicular epidermis after treatment with chemical carcinogens. The sensitivity of TG.AC mice to skin tumorigenesis, coupled with their low incidence of spontaneous skin tumors, makes this strain a good model for identifying carcinogens and for investigating the roles that other genes may play in the development of skin neoplasia. To investigate the possible involvement of the bcl-2 gene in skin tumorigenesis in the TG.AC mouse, we crossed heterozygous bcl-2-knockout mice (C57BI/6, 129 background) with TG.AC mice (FVB/N background). Female mice were genotyped by using a neo cassette to identify bcl-2-deficient mice. In addition, homozygous TG.AC mice were bred with FVB/N mice to generate hemizygous TG.AC mice on an FVB/N background to serve as a gene-dosage control. The F1 progeny consisted of FVB/N(v-Ha-ras+/-):C57BI/6,129(bcl-2+/+),FVB/N(v-Ha-ra s+/-):C57BI/6,129(bcl-2+/-), and FVB/N(v-Ha-ras+/-,bcl-2+/+). Ten-week-old mice were dosed twice weekly for 10 wk with acetone, 1.25 microg of 7,12-tetradecanoylphorbol-13-acetate (TPA), or 2.5 microg of TPA, and papillomas were counted weekly. Papillomas were analyzed for ras transgene and bcl-2 expression by reverse transcription-polymerase chain reaction, v-Ha-ras expression by in situ hybridization, and proliferating cell nuclear antigen expression by immunohistochemical analysis. Fewer papillomas (P < 0.05) were observed at the low dose of TPA (1.25 microg) in mice carrying the bcl-2 knockout allele than in the wild-type mice, suggesting that reduction of the bcl-2 gene product affects the susceptibility of TG.AC mice to TPA-induced papillomas. However, at the high dose of TPA (2.5 microg), there was no difference in papilloma response between knockout and wild-type mice, regardless of strain background. This suggests that at the higher dose of TPA, the effect of reduction in bcl-2 gene product was obscured. These results support the hypothesis that bcl-2 plays a limited role in skin tumorigenesis in the TG.AC mouse.

Animals↗
PubMed1997

T25: a simplified carcinogenic potency index: description of the system and study of correlations between carcinogenic potency and species/site specificity and mutagenicity.

A simplified carcinogenic potency index, the T25, is proposed as a practical method for the inclusion of potency considerations in carcinogen classification systems. The T25 is the chronic daily dose in mg per kg bodyweight which will give 25% of the animals tumours at a specific tissue site, after correction for spontaneous incidence, within the standard life span of that species. Calculated T25 values of a set of 113 US National Cancer Institute/National Toxicology Program (NC/NTP) carcinogens showed excellent correlation (correlation coefficient 0.96, P < 0.0001) with the carcinogenic potency index TD50 of Peto et al. (1984). The mean of T25 values for 51 transspecies, multiple common site NCI/NTP carcinogens were 10-fold lower than those for 62 NCI/NTP single species, single site carcinogens. For these 113 carcinogens, the mean T25 values were approximately 3-fold lower for agents that were also mutagenic in Salmonella compared to the non-mutagenic agents.

Abdominal Neoplasms↗
PubMed1996

Evaluation of transgenic mouse bioassays for identifying carcinogens and noncarcinogens.

Data supporting the use of transgenic lines to identify carcinogens and noncarcinogens are thus far based on a limited number of chemicals for which there are also long-term bioassay results in rats and/or mice. Six chemicals have been tested in the heterozygous p53-deficient mice and 13 in the Tg.AC line. The results show that the p53def responds rapidly to mutagenic carcinogens and the Tg.AC responds rapidly to both mutagenic and nonmutagenic carcinogens. Neither transgenic line responded to the noncarcinogens that were tested. The p53def line failed to respond to two nonmutagenic carcinogens (N-methyloacrylamide and reserpine), the Tg.AC line failed to respond to ethyl acrylate, a nonmutagenic chemical that induced tumors of the forestomach when administered by gavage, and to triethanolamine that caused an increase in hepatocellular tumors in B6C3F1 mice via skin painting. Both of the latter chemicals are examples of highly specific responses related to either route of administration or to strain susceptibility. Further efforts to evaluate the range of chemicals to which these transgenic lines respond are currently in progress.

Animals↗
PubMed1996

Association of tumor development with increased cellular proliferation and transgene overexpression, but not c-Ha-ras mutations, in v-Ha-ras transgenic Tg.AC mice.

The transgenic mouse line Tg.AC carries a v-Ha-ras gene fused to a fetal (zeta) globin promoter and uniquely responds to chemical carcinogens and tumor promoters by the induction of epidermal papillomas. Although the transgene was not constitutively expressed in non-tumor-bearing tissues, expression was induced by exposure to selected chemicals. Tg.AC transgenic mice on the FVB/N background developed occasional spontaneous tumors, including odontomas, squamous cell carcinoma of the salivary gland, leukemias and a rare ovarian yolk sac carcinoma. Both spontaneous and induced tumors are associated with expression of the transgene and, as determined by in situ hybridization, transgene expression is localized to proliferative areas of the tumors. Sequence analysis of the endogenous c-Ha-ras gene in both induced and spontaneous tumors revealed no mutations in codons 12, 59 or 61. These results suggest that expression of the v-Ha-ras transgene induces proliferation of specific cells in diverse tissues which then acquire neoplastic properties.

Animals↗
PubMed1996

Predictions for the outcome of rodent carcinogenicity bioassays: identification of trans-species carcinogens and noncarcinogens.

Thirty chemicals or substances currently undergoing long-term carcinogenicity bioassays in rodents have been used in a project to further evaluate methods and information that may have the capability of predicting potential carcinogens. In our predictions the principal information used includes structural alerts and in vitro test results for Salmonella mutagenicity, relative subchronic toxicity, and the sites and types of pathology found in subchronic (90-day) studies. This group of chemicals differs significantly from those used previously to evaluate predictive methods in that 23 of 30 are defined as nonmutagenic by conventional criteria. The goal of this predictive effort is to identify categorically the chemicals that have the capacity to induce cancers in both rats and mice (trans-species carcinogens) and those that are not carcinogenic in either rats or mice. Chemicals that show properties that may be associated with tumor induction in either species, i.e., species-specific cancers, are categorized as being of "uncertain predictability." This category includes chemicals believed to have limited carcinogenic potential that is manifested principally as a consequence of the genetic background of the test strain of inbred rodent.

Animals↗
PubMed1995

Genetic alterations cooperate with v-Ha-ras to accelerate multistage carcinogenesis in TG.AC transgenic mouse skin.

TG.AC transgenic mice harbor a v-Ha-ras transgene and retain two normal c-Ha-ras alleles and are susceptible to skin tumor formation by 12-O-tetradecanoylphorbol-13-acetate (TPA). To determine whether normal c-Ha-ras antagonizes the oncogenic potential of the v-Ha-ras transgene and/or whether additional non-Ha-ras 7,12-dimethylbenz(a)anthracene (DMBA) initiation target genes exist in mouse skin, which could cooperate with v-Ha-ras to increase the frequency of initiation, rate of promotion, or risk of malignant conversion, we treated TG.AC mouse skin with a single subthreshold dose of DMBA. This was followed by limited TPA or diacylglycerol promotion to select for cells with additional genetic alterations over those cells containing the v-Ha-ras transgene only. DMBA-treated/TPA-promoted TG.AC mice demonstrated a 10-fold increase in the average number of papillomas per mouse, a greater incidence of papilloma bearing-mice, and an increased papilloma growth rate when compared to acetone-treated/TPA-promoted TG.AC mice. These profound changes in papilloma frequency and growth occurred in the absence of the characteristic DMBA-induced A182-->T mutation in c-Ha-ras and immunohistochemical nuclear staining for p53 protein. DMBA-treated/acetone-promoted TG.AC mice did not develop any tumors. Limited promotion with the model diacylglycerol, sn-1,2-didecanoylglycerol, similarly produced an average of 10-fold more papillomas in DMBA-treated mice than in acetone-treated/sn-1,2-didecanoylglycerol-promoted TG.AC mice. DMBA-treated/TPA-promoted TG.AC mice developed their first malignancy by 16 weeks, and by 30 weeks, 50% of the mice developed malignancies, whereas no malignancies were observed in acetone-treated/TPA-promoted TG.AC mice. These results indicate that there exist unidentified DMBA initiation target genes in TG.AC mouse skin that cooperate with mutant Ha-ras to increase papilloma frequency, growth, and malignant conversion, and that promoter treatment can influence malignant conversion by selecting for cells with multiple genetic alterations.

9,10-Dimethyl-1,2-benzanthracene↗
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

Identifying chemical carcinogens and assessing potential risk in short-term bioassays using transgenic mouse models.

Cancer is a worldwide public health concern. Identifying carcinogens and limiting their exposure is one approach to the problem of reducing risk. Currently, epidemiology and rodent bioassays are the means by which putative human carcinogens are identified. Both methods have intrinsic limitations: they are slow and expensive processes with many uncertainties. The development of methods to modify specific genes in the mammalian genome has provided promising new tools for identifying carcinogens and characterizing risk. Transgenic mice may provide advantages in shortening the time required for bioassays and improving the accuracy of carcinogen identification; transgenic mice might now be included in the testing armamentarium without abandoning the two-year bioassay, the current standard. We show that mutagenic carcinogens can be identified with increased sensitivity and specificity using hemizygous p53 mice in which one allele of the p53 gene has been inactivated. Furthermore, the TG.AC transgenic model, carrying a v-Ha-ras construct, has developed papillomas and malignant tumors in response to a number of mutagenic and nonmutagenic carcinogens and tumor promoters, but not to noncarcinogens. We present a decision-tree approach that permits, at modest extra cost, the testing of more chemicals with improved ability to extrapolate from rodents to humans.

Animals↗
PubMed1995

A transgenic mouse model (TG.AC) for skin carcinogenesis: inducible transgene expression as a second critical event.

The v-Ha-ras transgenic TG.AC mouse line behaves as a genetically initiated model for mouse skin tumorigenesis with enhanced susceptibility to skin carcinogens. TG.AC mice develop epidermal papillomas in fewer than 20 weeks in response to the topical application of a variety of chemicals such as complete carcinogens, phorbol ester-type tumor promoters, and nonphorbol ester-type tumor promoters as well as to full-thickness skin wounds or plucking of the dorsal hair. We have found that the pedunculated epidermal papillomas can arise as focal hyperplasias from the permanent portion of the follicular epithelium. Expression of the v-Ha-ras transgene serves as a marker for tumor development since it is expressed at significant levels in the papilloma precursors, focal follicular hyperplasias, and the papillomas but not in the surrounding skin. Transgene expression colocalizes with increased cell proliferation in the papillomas as compared to non-tumor bearing surrounding skin. Malignant skin tumors, primarily squamous cell carcinomas and sarcomas, develop from sites of papilloma development in approximately 40% of papilloma bearing mice. As well as significant levels of transgenic v-Ha-ras expression, some of the malignancies also exhibit karyotypic changes which include trisomy of chromosome six or fifteen, but not chromosome seven. We believe that the TG.AC mouse line serves not only as a model for studying the mechanisms of skin tumorigenesis, but will also be a useful adjunct to the two year NTP toxicity/carcinogenicity studies by identifying carcinogens in fewer than 20 weeks.

Animals↗
PubMed1994

Follicular origin of epidermal papillomas in v-Ha-ras transgenic TG.AC mouse skin.

A follicular origin for some skin tumors has been hypothesized in both humans and animal models. Because of its rapid and sensitive response to tumor promoter treatment, a v-Ha-ras transgenic (TG.AC) mouse line was used to determine the origins of epidermal papillomas. Using histological studies and transgene expression as a marker for papilloma development, we determined that pedunculated papillomas arose from focal hyperplasias of the permanent portion of the follicular epithelium in phorbol 12-myristate 13-acetate-treated TG.AC mouse skin. Damage to the hair follicle by depilation was also sufficient to induce papillomas that were histologically indistinguishable from those produced by chemical exposure. Identification of the cellular origins of papillomas in this transgenic mouse model will allow for an analysis of the role of the hair follicle and hair cycle-associated signaling in tumor development.

Animals↗
PubMed1994

Cytogenetic analysis of malignant skin tumors induced in chemically treated TG-AC transgenic mice.

TG.AC mice (which carry a v-Ha-ras transgene) rapidly develop papillomas in response to 12-O-tetradecanoylphorbol-13-acetate (TPA). Approximately 30% of the papillomas are associated with subsequent development of malignancies. Early-passage spindle-shaped tumor cells arising from explant cultures of TPA-induced tumors in TG.AC mice were tumorigenic when transplanted to syngeneic recipients. The v-Ha-ras transgene in the transplanted tumors was expressed at a high level. To identify possible genetic changes associated with the development of malignant tumors, explanted cells were cultured in vitro and assessed for karyotypic changes between the second and third passages by analyzing G-banded metaphase chromosomes. For comparison, skin malignancies were induced in nontransgenic FVB/N mice (parent strain) by 7,12-dimethylbenz[a]anthracene (DMBA) initiation and TPA promotion, and their G-banded metaphase chromosomes were analyzed. Trisomy (in at least 50% of about 30 metaphases) of chromosome 15 (in five of 15 tumors) and chromosome 6 (four of 15 tumors) was observed in TG.AC mice, independent of chemical treatment or tumor type. Of six tumors from DMBA/TPA-treated FVB/N mice, three had trisomy 10 or 15 (or both), and two appeared normal. The absence of trisomy 7 is notable because c-Ha-ras maps to that chromosome. The absence of trisomy 7 in the six FVB/N DMBA/TPA-induced skin malignancies contrasts with DMBA/TPA-induced karyotypic effects in SENCAR mice. Expression of the v-Ha-ras transgene may have precluded the requirement for endogenous mutant ras and allelic imbalance involving chromosome 7 in TG.AC mice, but it could not have in FVB/N mice. These results suggest the possibility that the observed trisomies are consequential, rather than causal, events in the development of TG.AC or FVB/N skin malignancies. Molecular genetic analysis will be required to understand the changes associated with tumorigenesis in this transgenic line as well as in the parent mouse line.

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

Transgenic animals in toxicology.

Recent advances have been made in the characterization of a number of transgenic animal models. These animal models have provided a powerful toxicological tool for studying in vivo chemical effects and have increased our understanding of the role of specific genetic alterations as predisposing factors for chemical carcinogenesis. The goal of this symposium was to introduce the development of transgenic animals and the utilization of transgenics in toxicology research focusing on understanding tissue-specific mutation, chemical effects, and cancer. The production of transgenic animals, including gene insertions and gene knockouts, and the utilization of transgenic technology for studying multistage carcinogenesis and tumor suppressor genes are described. Data on the application and implications of transgenics as a genetic endpoint are also discussed. The use of transgenic animals in toxicology should improve our understanding of the role of specific genetic alterations in the carcinogenic process and lead to improved estimations of human health risks.

Animals↗