Transgenic rats carrying copies of the human c-Ha- ras proto-oncogene exhibit enhanced susceptibility to N -butyl- N -(4-hydroxybutyl)nitrosamine bladder carcinogenesis

Abstract

We have established a transgenic rat line carrying three copies of the human c-Ha-ras proto-oncogene with its own original promoter region, Jcl/SD-TgN(HrasGen)128Ncc (Hras128) rat. c-Ha-ras protein from expression of transduced and endogenous c-Ha-ras genes could be detected in the bladder epithelium of untreated transgenic rats. To examine their susceptibility to N-butyl-N-(4-hydroxybutyl)nitrosamine (BBN)-induced urinary bladder carcinogenesis, male transgenic and wild-type littermates were treated with 0.05% BBN in their drinking water for 10 weeks and then killed at week 20. The numbers and volumes of total macroscopic bladder tumors including both transitional cell papillomas and carcinomas (TCC) per rat were much greater in Hras128 rats than in their wild-type counterparts. The numbers of carcinomas per rat were also significantly greater in Hras128 rats. Two cases of TCC exhibiting invasion of the bladder muscle layer, which is extremely rare in the wild-type animals under the experimental conditions used, were also observed in Hras128 rats. The GGC-->GAC mutations at codon 12 of the transgene were observed in only two TCC out of 21 bladder tumors (9.5%), assessed by RFLP analysis and direct sequencing. SSCP analysis did not show any endogenous c-Ha-ras gene mutations. One of 25 tumors (4.0%) in wild-type rats had an endogenous c-Ha-ras gene mutation at codon 12 that was detected (GGA-->GAA) by single-strand conformation polymorphism and direct sequencing. These results indicate that the Hras128 rat is highly susceptible to BBN carcinogenesis and may be utilized as a rat model for analysis of bladder tumor development. The mutation findings indicate that the enhanced tumor development is not primarily due to mutations occurring in the transgene.

Transgenic Rats Carrying Human c‐Ha‐ras Proto‐oncogene Are Highly Susceptible to N‐Nitrosomethylbenzylamine Induction of Esophageal Tumorigenesis

“…1) Similarly, the transgene mutation rate in urinary bladder lesions induced by another nitrosamine, N-butyl-N-(4-hydroxybutyl)nitrosamine, was not predominant. 37) In contrast, the mutated band densities observed in esophageal tumors in the current study indicated the tumors to contain cells with mutated transgene as the major population (Fig. 1).…”

“…[26][27][28] In contrast, mutations in the c-Ha-ras gene are rare in bladder or liver tumors induced by carcinogens in rats. [34][35][36] We have found that our Hras128 strain is highly susceptible to MNU-induced mammary, 9) N-butyl-N-(4-hydroxybutyl)nitrosamine-induced urinary bladder 37) and, in the present work, NMBA-induced esophageal carcinogenesis. The results indicate that mutations of c-Ha-ras can play important roles in neoplasia in the esophagus, in contrast to the breast and urinary bladder.…”

Possible enhancing effects of atrazine and nonylphenol on 7,12‐dimethylbenz[a]anthracene‐induced mammary tumor development in human c‐Ha‐ras proto‐oncogene transgenic rats

“…Human c-Ha-ras proto-oncogene Tg and non-Tg rats I 5 To whom correspondence should be addressed. E-mail: htsuda@med.nagoya-cu.ac.jp bred by CLEA Japan Inc., Tokyo 32,34) at 7 weeks of age were maintained in plastic cages in an air conditioned room at 22±2°C and 55±10% humidity. The animals were all allowed free access to pellet basal diet (Oriental Yeast Co., Ltd., Tokyo), from which soy constituents, isoflavones which were known to act as estrogens, had been eliminated (Table 1), and to tap water.…”

“…Most of the tumor cells contained mutated transgene, but not endogenous Hras gene, indicating that activation of the transgene plays an important role in such rapid development of tumors. The animals have also been found to be susceptible to N-butyl-N-(4-hydroxybutyl)nitrosamine urinary bladder, 34) N-nitrosomethylbenzylamine esophageal, 35) and DMBA skin carcinogenesis. 36) We recently showed that the transgenic (Tg) rats have a potential for use in medium-term bioassay models to test for the modifying effects of estrogenic environmental compounds on mammary tumor development.…”

Preferential mammary carcinogenic effects of 2‐amino‐1‐methyl‐6‐phenylimidazo[4,5‐b]pyridine (PhIP) in human c‐Ha‐ras proto‐oncogene transgenic rats

“…7) In humans, Ki-ras mutations are preferentially found in colon, 8,9) pancreatic duct 10) and bile duct carcinomas, 11) and N-ras mutations are typically seen in myeloid leukemia. 12) We have generated a transgenic rat line, harboring copies of a human c-Ha-ras proto-oncogene (Hras128 rat), and have shown that these animals are highly susceptible to mammary, 13,14) bladder 15) and esophageal 16) carcinogens.…”