Quantitative evaluation of the effects of human carcinogens and related chemicals on human foreskin fibroblasts

Ten compounds representative of diverse classes of chemicals were evaluated for their cytotoxicity and transforming ability to human skin fibroblasts in vitro. Only five of the ten compounds were highly cytotoxic in the 0-100 µg/ml range and their order of cytotoxicity was: 2,5-bis(1-aziridinyl)-3,6-bis(carboethoxyamino)-1,4-benzoquinone (AZQ) > cis platin > bis(chloromethyl)ether (BCME) > acrylonitrile > afatoxin BI (AFBI). The other five compounds, afatoxin B2 (AFB2), methylmethacrylate, 1-naphthylamine (1-NA), 2-naphthylamine (2-NA), and cyclophosphamide, exhibited less than 40% inhibition of colony formation even at 100 µg/ml of the compound (the maximum concentration of AFB2 used was 50 µg/ml due to its low solubility). Anchorage-independent growth of exposed cells in soft agar was used as a biological endpoint for the expression of chemical transformation. AFB₁ had strong transforming ability, whereas AFB₂ was a weak transforming agent. The transforming abilities of acrylonitrile, AZQ, BCME, cis-platin, methylmethacrylate and 2-NA ranged between those of AFBI and AFB2. 1-NA also induced the soft agar growth property in the treated cells even though this compound has not been shown to be carcinogenic. AFB₁, AZQ, cisplatin, cyclophosphamide and 1-NA exhibited a dose dependent increase in soft agar growth frequency for at least three consecutive concentrations. The data suggest that anchorage-independent colony forming ability of exposed cells is a reliable marker to measure the carcinogenic potential of various hazardous chemicals.Abbreviations AZQ 2,5-bis(1-aziridinyl)-3,6-bis(carboethoxyamino)-1,4-benzoquinone - AFB aflatoxin B₁ - AFB2 aflatoxin 132 - AI anchorage independent - B[a]P benzo[a]pyrene - BCME bis(chloromethyl)ether; cis-platin, cis-diammine-dichloroplatinum - CM complete medium - E.D.50 effective dose which produced 50% cytotoxicity - CP cyclophosphamide - HNF human neonatal foreskin - HPLC high pressure liquid chromatography - 1-NA 1-naphthylamine - 2-NA 2-naphthylamine - PDL population doubling     

Induction of hyperplasia and anaplasia by carcinogens in organ cultures of mouse prostate

Summary In an effort to establish a test system to examine the carcinogenic potential of chemicals, mouse prostate explants were maintained as organ cultures and the effects of carcinogenic and noncarcinogenic compounds were examined at various intervals after treatment. The degree of hyperplasia produced by a compound was determined by the colcemid metaphase arrest technique. Extensive hyperplasia of the prostatic epithelium occurred at 8 days after treatment with 3-methylcholanthrene, the 11–12 epoxide of methylcholanthrene, benzo(a)pyrene and N-methyl-N-nitro-N-nitrosoguanidine. At 12 days most carcinogentreated explants were anaplastic. The noncarcinogenic compounds, pyrene and phenanthrene, did not produce a mitotic stimulatory effect on the epithelium of the explants. The data suggest that the organ culture system of mouse prostate may be employed as a test system to obtain preliminary information regarding the carcinogenicity of a compound. This investigation was supported by Contract No. NO1-CP-22064 from the Lung Cancer Segment, Division of Cancer Cause and Prevention, National Cancer Institute, NIH, Department of Health, Education and Welfare.     

J-Shaped Dose-Response Relationship for Tumor Induction by Caffeic Acid in the Rat Forestomach,Modeled by Non-Monotonic Dose Response for DNA Damage and Cell Proliferation

Dose-response relationships for chemical tumor induction can be described with stochastic carcinogenesis models by incorporation of dose-dependent parameters. We used a two-stage mutational model with clonal expansion of intermediate cells to mimic the observed U-shaped (more precisely: J-shaped) dose response for cell division in the forestomach of rats treated with caffeic acid. The question was whether biologically acceptable model parameters for the rates of mutation and cell turnover could be found to fit the experimental data, including tumor incidence in controls and at high dose. Using rates of cell turnover in proportion to the experimental labeling indices for DNA replication, model parameters could fit the tumor incidences within the 95% confidence limits of the experimental data, and the J-shape was retained quantitatively. However, when the smaller confidence limits derived from historical data on the spontaneous forestomach tumor incidence were taken into account, the original model could no longer accommodate all data and had to be expanded. For that, mutation rates were considered to be proportional to both the level of DNA damage and the rate of cell division, and both factors were allowed to be dose dependent. For the DNA damage, an increment over background was used at the higher, cytotoxic dose levels. This model improvement resulted in biologically acceptable parameters to fit all available data, demonstrating its usefulness for further analyses of hormetic dose response.     

Tumorlike Formations in the Olfactory Organ of the Striped Mullet Mugil cephalusfrom Sovetskaya Gavan'' Bay, Sea of Japan

This paper describes tumorlike formations in the olfactory organ of the striped mullet Mugil cephalusinhabiting polluted waters of Zapadnaya Bay (Sovetskaya Gavan' Bay, western coast of Tatar Strait). The structure of these tumorlike formations is characteristic of fibroma. The appearance of the tumors is apparently associated with carcinogenic industrial wastes and radioactive contamination of the bay.     

Invited contribution: An objective approach to the development of short-term tests predictive of carcinogenicity

The Carcinogenicity Prediction and Battery Selection procedure was developed to address two problems: (1) the identification of highly predictive, yet cost-effective, batteries of short-term tests and (2) the objective prediction of the potential carcinogenicity of chemicals based upon the results of short-term tests even when a mixture of positive and negative results is obtained. In the present report the usefulness of the Carcinogenicity Prediction and Battery Selection procedure is demonstrated using benzo[a]pyrene, benzoin and diethylstilbestrol as examples. In addition, its applicability in the analysis of all the possible outcomes of a battery is illustrated together with an analysis of the worth of additional testing.Abbreviations B[a]P benzo[a]pyrene - CASE Computer-Automated Structure Evaluation - CPBS Carcinogenicity Prediction and Battery Selection - DEHP diethylhexylphthalate - DES diethylstilbestrol - NTA nitrilotriacetate - TCDD 2,3,7,8-tetrachlorodibenzo-p-dioxin     

Organ culture of chicken cecum: Morphologic and physiologic observations after 24 and 48 h of culture

Summary Cecum from chickens, 4 wk old, can best be maintained for 24 h in a serum-free organ culture system using Trowell T8 medium agar sheet at 25°C. As determined by light microscopy as well as scanning and transmission electron microscopy, mucosal architecture involving classic ultrastructure of chicken cecal mucosa was preserved. Protein content of cecal explants did not change up to 48 h of culture. DNA content of cecal explants did not change up to 24 h of culture but decreased significantly to two-thirds of control in 48 h of culture. Based on the morphologic and physiologic findings, it became evident that this organ culture system using Trowell T8 medium at 25°C can be successfully used as an in vitro experimental model for as long as 24 h. The organ culture system could be a useful tool, from the structural integrity of ceca observed in this study, in investigating mucosal function and mucosal response to drugs, carcinogens, trophic factors, and pathogens.