Molecular changes in the early stage of colon carcinogenesis in rats treated with azoxymethane.
To elucidate early molecular events related to colon carcinogenesis, we examined alterations in the expression of colon cancer-related genes such as cyclooxygenase (COX)-2, APC and c-Myc, cell proliferation and apoptosis in the background colon mucosa, and K-ras mutation at aberrant crypt foci (ACF) in the colons of azoxymethane (AOM)-treated rats 4 weeks after the first exposure to AOM. About 40 ACF/colon were induced in the colons of rats treated with AOM (Group 1); however, rats not treated with AOM (Group 2) showed no ACF formation in the colon. The level of AgNORs in the colonic mucosa was significantly higher in Group 1 than in Group 2 (P<0.01). The colonic mucosa in Group 1 looked macroscopically and histologically normal, but the proliferative activity of the mucosa of rats treated with AOM was clearly elevated. COX-2 mRNA expression was not detected in normal colonic mucosa in Group 2, but 3 out of 10 rats in Group 1 showed COX-2 mRNA expression in their colons by reverse transcription (RT)-polymerase chain reaction (PCR). There was a tendency toward an increased expression level of COX-2 in the AOM-treated group. The level of APC mRNA expression in Group 1 was significantly lower than that in Group 2 (P<0.01). Moreover, the level of c-Myc mRNA expression in Group 1 was significantly higher than that in Group 2 (P<0.01). An average of 0.034+/-0.006% apoptosis in colonic mucosa was detected in Group 1; the incidence of apoptosis in Group 2 was 0.021+/-0.005%. The difference between Groups 1 and 2 was significant (P<0.01). These results indicate that apoptosis was possibly induced to eliminate cells damaged by AOM administration. Six out of 22 (27%) ACF with 4 or more crypts showed K-ras mutations at codon 12; all mutations were G to A transitions (GGT to GAT). ACF with 1-3 crypts showed no mutations in the K-ras gene. In conclusion, AOM caused an increase in COX-2 and c-Myc mRNA expression, a decrease in APC mRNA expression, induction of apoptosis in normal-appearing colonic mucosa, and a K-ras mutation in ACF with 4 or more crypts. These findings may help to identify key targets in the early steps of colon carcinogenesis, against which drugs that would be broadly effective for chemoprevention of colon cancer could be developed.
Kishimoto
Y
Y
Dept of Clinical Pharmacology, Faculty of Medicine, Tottori University, Yonago, Japan. ykishimo@grape.med.tottori-.ac.jp
Morisawa
T
T
Hosoda
A
A
Shiota
G
G
Kawasaki
H
H
Hasegawa
J
J
D025601:Adenomatous Polyposis Coli Protein / Q000235:genetics / Q000378:metabolism; D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D001397:Azoxymethane / Q000633:toxicity*; D002273:Carcinogens / Q000633:toxicity*; D003106:Colon / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D003110:Colonic Neoplasms / Q000139:chemically induced / Q000378:metabolism* / Q000473:pathology; D051546:Cyclooxygenase 2; D011905:Genes, ras / Q000235:genetics; D020287:In Situ Nick-End Labeling; D007413:Intestinal Mucosa / Q000187:drug effects / Q000378:metabolism; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism; D008297:Male; D009697:Nucleolus Organizer Region / Q000378:metabolism; D016133:Polymerase Chain Reaction; D018807:Polymorphism, Single-Stranded Conformational; D011230:Precancerous Conditions / Q000139:chemically induced / Q000378:metabolism* / Q000473:pathology; D011451:Prostaglandin-Endoperoxide Synthases / Q000235:genetics / Q000378:metabolism; D016271:Proto-Oncogene Proteins c-myc / Q000235:genetics / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D011916:Rats, Inbred F344
D025601:Adenomatous Polyposis Coli Protein; D002273:Carcinogens; D007527:Isoenzymes; D016271:Proto-Oncogene Proteins c-myc; D012333:RNA, Messenger; D051546:Cyclooxygenase 2; D011451:Prostaglandin-Endoperoxide Synthases; D001397:Azoxymethane