Inhibition of morphological transformation induced withN-methyl-N′-nitro-N-nitrosoguanidine in cultures of hamster embryo cells by 5′-bromo-2′-deoxyuridine-photolysis

Abstract

The present study was performed in order to determine whether type III transformed foci induced by N‐methyl‐N′‐nitro‐N‐nitrosoguanidine originate from the small subpopulation of cells stimulated by the carcinogen to enter DNA synthesis. During the last 30 min of variable treatment periods using different doses of N‐methyl‐N′‐nitro‐N‐nitrosoguanidine, administered alone or in association with the thymidine analogue, 5′‐bromo‐2′‐deoxyuridine (0.98 × 10⁻⁵ M), the density‐inhibited monolayers of hamster embryo cells were exposed to fluorescent light and then assayed for abnormal growth patterns by the focus formation method. Mock‐irradiated cultures as well as monolayers whose medium lacked N‐methyl‐N′‐nitro‐N‐nitro‐soguanidine, 5′‐bromo‐2′‐deoxyuridine, or both, served as controls. The cytotoxicity of 5′‐bromo‐2′‐deoxyuridine + N‐methyl‐N′‐nitro‐N‐nitrosoguanidine + photolysis (BMP) protocol on confluent as well as logarithmically growing hamster embryo cells was estimated in single‐cell survival experiments. Plating efficiency determinations have demonstrated that, unlike their actively growing counterparts, confluent hamster embryo cell mono‐layers are extremely resistant to the cytotoxic effects of the BMP protocol. The quantitative transformation assays indicated that: (1) in non‐illuminated cultures addition of 5′‐bromo‐2′‐deoxyuridine to carcinogen‐containing medium does affect transformation frequency of hamster embryo cells in the sense that the incidence of type III foci did not subside at later intervals during the post‐carcinogen administration period as it did in the absence of the analogue; (2) irradiation of N‐methyl‐N′‐nitro‐N‐nitrosoguanidine and halogenated pyrimidine analogue‐treated cultures with fluorescent light practically suppressed transformation; (3) analogueadded and analogue‐removed experiments pointed out that the event(s) on which 5′‐bromo‐2′‐deoxy‐uridine fluorescent light sensitization of morphological transformation largely depends, takes place between 5 and 15 h after N‐methyl‐N′‐nitro‐N‐nitrosoguanidine administration, i.e., during the period of maximal carcinogen‐stimulated DNA synthesis; and (4) neither fluorescent light nor 5′‐bromo‐2′‐deoxyuridine, singly or in combination, were able to transform cultures of hamster embryo cells. These findings are strong indirect arguments for the concept that carcinogen‐induced DNA synthesis and the initiation of transformed clones are causally related.