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Quantitative Studies of Transformation by Chemical Carcinogens and Ultraviolet Radiation Using a Subclone of BHK₂₁ Clone 13 Syrian Hamster Cells

Environmental Mutagenesis Branch, National Institute of Environmental Health Sciences, NIH, Research Triangle Park, North Carolina 27709 [Y. I., J. A. E., M. W. L.], and Microbiological Associates, Bethesda, Maryland 20014 [N. K. M.]

Transformation (growth in soft agar) and survival of a subclone of BHK₂₁ C13 were evaluated after treatment with N-acetoxy-2-acetylaminofluorene, N-methyl-N'-nitro-N-nitrosoguanidine, 4-nitroquinoline 1-oxide, and ultraviolet radiation. Because cells of this subclone are capable of dividing several times in soft agar before dying, transformation is expressable in these cells without the intermediate step of plating in liquid media with a solid plastic or glass substrate. Thus a quantitative assessment of transformation may be obtained by treating these cells, trypsinizing, and plating directly in soft agar. The spontaneous transformation frequency of this subclone is 5 to 10 x 10⁶. As a function of molar dose, 4-nitroquinoline 1-oxide is the most effective at inducing transformation while N-acetoxy-2-acetylaminofluorene and N-methyl-N'-nitro-N-nitrosoguanidine are about equally effective. Acetylaminofluorene, a precarcinogen, at concentrations up to 20 µM did not increase the frequency of transformants. If transformation is scored on the basis of number of cells inoculated and under conditions of high relative survival, the number of transformants is seen to increase with the concentration of chemical or fluence of UV. These data indicate that these agents induce transformation rather than select for existing transformants. If transformation is scored as a function of survival, N-methyl-N'-nitro-N-nitrosoguanidine and 4-nitroquinoline 1-oxide are the most effective transforming agents, N-acetoxy-2-acetylaminofluorene is intermediate, and ultraviolet radiation gives the lowest transformation frequency. Isolation of transformed colonies (both induced and spontaneous) and s.c. injection into Syrian hamsters resulted in tumors from about one-half of the isolated clones. Untreated cells produced no tumors under the conditions used. Both untransformed and transformed cells were free of a variety of viruses.

¹ Present address: Brookhaven National Laboratory, Upton, N. Y. 11973.

² Present address: NIOSH, Morgantown, W. V. 26505.

³ To whom requests for reprints should be addressed, at Department of Pathology, Washington University School of Medicine, St. Louis, Mo. 63110.

Received 10/21/76. Accepted 3/30/77.