Lactic acid bacteria (LAB) are proposed to have several beneficial effects, including the inactivation of carcinogens. We have studied the potential of Lactobacillus acidophilus (from a commercially available yogurt), Lactobacillus gasseri (P79), Lactobacillus confusus (DSM20196), Streptococcus thermophilus (NCIM 50083), Bifidobacterium breve and Bifidobacterium longum (from human infant stool) to prevent the induction of DNA damage by N-methyl-N'-nitro-N-nitrosoguanidine (MNNG, 7.5 mg/kg body wt) in colon cells of the rat. Using the new technique of single cell microgel electrophoresis, all investigated strains were antigenotoxic toward MNNG after a single dose of 10(10) viable cells/kg body wt p.o. eight hours before the carcinogen. One-half and one-tenth of this initial dose resulted in a loss of protective activity. High doses of heat-treated L. acidophilus strains were also not antigenotoxic. One mechanism of the preventive effect could be that bacterial metabolites or components are responsible. Accordingly, selected examples were investigated in vitro in colon cells of the rat. Metabolically active L. acidophilus cells, as well as an acetone extract of the culture, prevented MNNG-induced DNA damage. Different cell fractions from L. acidophilus (cytoplasm, cell wall skeleton, cell wall) were devoid of antigenotoxic activity, whereas the peptidoglycan fraction and whole freeze-dried cells were antigenotoxic. As a second carcinogen, 1,2-dimethylhydrazine (DMH) was used. A dose- and time-response study was first performed to assess the effects of DMH in several segments of the gastrointestinal (GI) tract. Exposure for 16 hours to 15 or 25 mg DMH/kg body wt p.o. induced DNA damage in cells of the distal colon of rats, whereas no cytotoxicity was seen. Pretreatment orally with LAB on four consecutive mornings before DMH gavage (8 hours after the last LAB application) revealed that L. acidophilus, L. confusus, L. gasseri, B. longum, and B. breve inhibited the genotoxic effect of DMH. One of four S. thermophilus and one of three Lactobacillus delbrueckeii ssp. bulgaricus strains were also protective. Heat-treated L. acidophilus did not inhibit DMH-induced genotoxicity. A few aliquots of the colon cells were processed immunohistochemically for the presence of the "proliferation cell nuclear antigen" (PCNA). DMH treatment did not increase PCNA, nor was there any modulation by LAB. The effect of L. acidophilus on foreign compound-metabolizing enzymes (Phase I and Phase II) in liver and colon cells of rats revealed only one parameter to be modulated, namely, a two- to three-fold increase in the levels of NADPH-cytochrome P-450 reductase. The meaning of this finding, in terms of possible chemoprevention by LAB, remains unclear. In conclusion, our studies show that most, but not all, LAB tested could strongly inhibit genotoxicity in the GI tract of the rat and that viable LAB organisms are required for the protective effect in vivo. The comet assay technique is a powerful tool to elucidate such in vivo antigenotoxic act...

Lactic acid-producing bacteria prevent carcinogen-induced preneoplastic lesions and tumors in rat colon. Because the mechanisms responsible for these protective effects are unknown, two strains of lactic acid bacteria, Lactobacillus delbrueckii ssp. bulgaricus 191R and Streptococcus salivarius ssp. thermophilus CH3, that are used to produce yogurt, were investigated in vitro and in vivo to elucidate their potential to deactivate carcinogens. Using the "Comet assay" to detect genetic damage, we found that L. bulgaricus 191R applied orally to rats could prevent 1, 2-dimethylhydrazine-induced DNA breaks in the colon in vivo, whereas St. thermophilus CH3 were not effective. However, in vitro, both strains prevented DNA damage induced by N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) in isolated primary rat colon cells. Extracts prepared from milk fermented with St. thermophilus CH3 were as efficient in deactivating MNNG as was L-cysteine. Isolated metabolites arising from bacteria during fermentation in the colon or in milk [L(+) lactate, D(-) lactate, palmitic acid and isopalmitic acid] were not effective. We postulate that thiol-containing breakdown products of proteins, via catalysis by bacterial proteases, could be one mechanism by which MNNG or other carcinogens are deactivated in the gut lumen resulting in reduced damage to colonic mucosal cells.

The influence of refrigeration, freezing, repetitive freezing-thawing, and irradiation on meat quality were evaluated by detecting DNA damage to beef muscle tissues using the single cell gel electrophoresis assay (Comet assay). Comet tail lengths indicating DNA damage levels increased with refrigeration time up to 10 days. In the case of frozen storage, the damage was increased only for the first 15 days of storage, but after that, decreased slightly and the tail lengths at 15 days were similar to those of 3,5 days refrigerated storage. Tail lengths resulting from repetitive freezing-thawing were increased after two cycles of treatment and three cycles produced longer tails than those that had been refrigerated for10 days or kept in frozen storage for 60 days. Irradiation from 1 to 10 kGy caused the most serious DNA damage among the treatments compared in this study. The two kinds of muscle tissue were similar in the pattern of results obtained from the Comet assay.The mean tail length of the comets reflected approximately the increasing patterns of DNA damage, but the differences of values between the extents of treatment were somewhat insignificant and anomalous. Therefore, we used the concept of relative damage index (RDI) calculated from per cent cells data for each tail length range and representing a state of DNA damage. Significantly different values in RDI's made it possible to discriminate samples according to the treatment levels within each treatment.

The influence of refrigeration, freezing, repetitive freezing-thawing, and irradiation on meat quality were evaluated by detecting DNA damage to beef muscle tissues using the single cell gel electrophoresis assay (Comet assay). Comet tail lengths indicating DNA damage levels increased with refrigeration time up to 10 days. In the case of frozen storage, the damage was increased only for the first 15 days of storage, but after that, decreased slightly and the tail lengths at 15 days were similar to those of 3,5 days refrigerated storage. Tail lengths resulting from repetitive freezing-thawing were increased after two cycles of treatment and three cycles produced longer tails than those that had been refrigerated for10 days or kept in frozen storage for 60 days. Irradiation from 1 to 10 kGy caused the most serious DNA damage among the treatments compared in this study. The two kinds of muscle tissue were similar in the pattern of results obtained from the Comet assay.The mean tail length of the comets reflected approximately the increasing patterns of DNA damage, but the differences of values between the extents of treatment were somewhat insignificant and anomalous. Therefore, we used the concept of relative damage index (RDI) calculated from per cent cells data for each tail length range and representing a state of DNA damage. Significantly different values in RDI's made it possible to discriminate samples according to the treatment levels within each treatment.

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