Use of fluorescence in situ hybridization (FISH) to study chromosomal damage induced by radiation and bromodeoxyuridine in human colon cancer cells.

PURPOSE: Although the thymidine analog radiation sensitizer bromodeoxyuridine (BrdUrd) increases radiation-induced chromosomal aberrations, it is not known whether these aberrations are uniformly distributed among chromosomes. Using fluorescence in situ hybridization, we carried out a study to test the hypothesis that BrdUrd-induced radiosensitization may be mediated by nonuniform chromosomal damage. METHODS AND MATERIALS: Log phase HT29 human colon cancer cells were exposed to 10 microM BrdUrd (or media alone) for one cell cycle, and the G1 cells were separated by centrifugal elutriation. Half of the control and BrdUrd samples were irradiated with 8 Gy. Cells were then incubated for 24-28 h, and metaphase spreads were prepared. Fluorescence in situ hybridization was performed using paint probes for chromosomes 1 and 4. RESULTS: We found that radiation induced 0.20 aberrations per chromosome in chromosome 4. Based on the ratio of the relative lengths of chromosome 1-4 (1.34), it was predicted that chromosome 1 would have approximately 0.26 aberrations per chromosome. However, we observed 0.39 aberrations per chromosome 1, which was significantly greater than the predicted (p < 0.001 by chi-square). Incubation with BrdUrd prior to irradiation significantly increased the aberrations found in chromosome 1 (by a factor of 1.4) and chromosome 4 (by a factor of 1.9) compared to radiation alone (p < 0.001) for both chromosome 1 and 4). CONCLUSION: This study demonstrates that individual chromosomes in human colon cancer cells show significantly different rates of aberration after irradiation. Furthermore, the BrdUrd-mediated increase in radiation-induced chromosomal aberrations may not be uniform among chromosomes.

The effects of selenium on the emergence of aflatoxin B1-induced enzyme-altered foci in rat liver.

The effects of selenium on the emergence of aflatoxin B1 (AFB1)-induced enzyme-altered foci were studied in male Sprague-Dawley rats. Animals were fed a selenium-deficient diet and supplemented with 5.0, 2.0, and 0.2, or 0 ppm selenium in drinking water for 3 weeks prior to initiation with 2.0 mumol/kg AFB1. After a 1-week period of selenium normalization, the animals were placed on a diet of ordinary rat chow, and were administered a promoting regimen of 500 ppm phenobarbital in drinking water for 1 week, after which time each rat received a two-thirds partial hepatectomy. The promoting regimen of phenobarbital in tap water was then reduced to 100 ppm and continued for 7 weeks. Subsequently, the rats were sacrificed, their livers excised, and fresh frozen sections prepared and stained histochemically to demonstrate areas of gamma-glutamyl transpeptidase (GGT) activity. Selenium supplementation was observed to diminish the induction of GGT-positive foci, especially at the 5.0-ppm level. These data suggest that selenium is able to protect against the hepatocarcinogenic effects of AFB1 in the rat, and that the enzyme-altered foci bioassay may be a useful technique in assessing the interaction of selenium on the process of hepatocarcinogenesis.

Methapyrilene effects on initiation and promotion of gamma-glutamyl-transpeptidase positive foci in rat liver.

The initiating and promoting effects of methapyrilene were evaluated using the hepatic enzyme-altered foci bioassay. Male Sprague-Dawley rats were partially hepatectomized and 24 hours later were administered either methapyrilene or nitrosodiethylamine by oral gavage. Subsequently, the animals were promoted with 500 ppm2 phenobarbital or 200 ppm methapyrilene in drinking water for eight weeks. Fresh frozen sections of liver were then stained and scored for gamma-glutamyl-transpeptidase (GGT)-positive foci. Methapyrilene, when tested as the nominal initiator at a single dose of 50 mg/kg or at two doses of 130 mg/kg and promoted by phenobarbital, did not induce foci above background levels. However, when substituted for phenobarbital as the promoting agent following nitrosodiethylamine initiation, methapyrilene enhanced enzyme-altered foci formation to an equal or greater extent than did the promoter phenobarbital. These results suggest that the carcinogenic effects of methapyrilene may be related to its ability to enhance hepatic tumorigenesis.