Relationship between exposure to TPA and appearance of transformed cells in MNNG-initiated transformation of BALB/c 3T3 cells.
In the BALB/c-3T3-cell transformation system, the effect of 12-O-tetradecanoylphorbol-13-acetate (TPA) exposure on the appearance of transformed cells was examined in order to investigate the mechanisms of in vitro tumor promotion. Optimal duration of TPA exposure on N-methyl-N'-nitro-N-nitrosoguanidine(MNNG)-initiated cells was at least 11 days. To investigate the effect of transformation frequencies of altering inoculating cell density at the replating of MNNG-exposed cells and of altering the time of starting TPA exposure, MNNG-exposed cells were replated at various inoculum sizes. With lower inoculum sizes (1 x 10(3) to 3 x 10(4) cells/dish), maximum TPA-induced transformation occurred for TPA commencement at confluence, while with higher inoculum size (1 x 10(5) cells/dish), maximum transformation frequency was observed when TPA exposure was started on day 7 after replating, being some 2 days after confluence. This may suggest that there are different mechanisms involved, depending on inoculum size, and that these may involve cell-cell interactions (at lower inoculum) and mutation expression periods (at higher inoculum). By means of redispersion experiments, it was demonstrated that the appearance of transformed cells begins on about day 7 after replating at a cell density of 1 x 10(4) cells/dish. These results suggest the usefulness of the replating method for optimizing transformation in the BALB/c-3T3-cell transformation assay, and provide insight into the time frame of expression of MNNG-initiated transformants and TPA-induced expansion of these transformants.
Tsuchiya
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Safety Evaluation Center, Central Research Laboratory, Showa Denko K.K., Chiba, Japan. tsuchiya@ctl.sdk.co.jp
Umeda
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D016475:3T3 Cells / Q000187:drug effects* / Q000473:pathology; D000818:Animals; D002273:Carcinogens / Q000494:pharmacology*; D002452:Cell Count; D002461:Cell Line, Transformed / Q000187:drug effects; D002471:Cell Transformation, Neoplastic / Q000139:chemically induced*; D008769:Methylnitronitrosoguanidine / Q000494:pharmacology*; D051379:Mice; D008807:Mice, Inbred BALB C; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*; D013997:Time Factors
D002273:Carcinogens; D008769:Methylnitronitrosoguanidine; D013755:Tetradecanoylphorbol Acetate