title: Detection of Pig-a gene mutants in rat peripheral blood following a single urethane treatment: A comparison of the RBC Pig-a and PIGRET assays.
abstract: The rat red blood cell (RBC) Pig-a assay has been recommended by an expert working group of the International Workshop on Genotoxicity Testing as a potential new method to evaluate in vivo gene mutations in regulatory genotoxicity risk assessments. In a collaborative study in Japan, an improved Pig-a assay using reticulocytes (PIGRET assay) with magnetic enrichment of CD71-positive cells was evaluated, and it was revealed that this assay could detect the mutagenicity of chemicals earlier than the RBC Pig-a assay could. To verify further the suitability of the PIGRET assay for an in vivo short-term genotoxicity screening test, a joint research study was conducted by the Japanese Environmental Mutagen Society, and 24 compounds were evaluated. One of the compounds evaluated in this study was urethane, a multi-organ rodent carcinogen. Urethane (250, 500, and 1000mg/kg body weight) was orally administered once to 8-week-old male Crl:CD (SD) rats. Blood samples were collected at 1, 2, and 4 weeks after the administration and processed for the RBC Pig-a and PIGRET assays. In the PIGRET assay, the Pig-a mutant frequency (MF) significantly increased at both 2 and 4 weeks after the treatment of 1000mg/kg of urethane. However, in the RBC Pig-a assay, a significant increase in the Pig-a MF was observed only at 1 week after the treatment with 500mg/kg, but the MF value was within our historical control range; therefore, it was judged to be negative. These results suggest that the PIGRET assay might be useful for evaluating the in vivo mutagenicity more clearly than the RBC Pig-a assay after a single treatment of test compounds.
authors.affiliation: Yakult Central Institute, Yakult Honsha Co., Ltd., 5-11 Izumi, Kunitachi-shi, Tokyo 186-8650, Japan. Electronic address: kazunori-narumi@yakult.co.jp.
authors.forename: Kazunori
authors.initials: K
authors.lastname: Narumi
authors.affiliation: Yakult Central Institute, Yakult Honsha Co., Ltd., 5-11 Izumi, Kunitachi-shi, Tokyo 186-8650, Japan.
authors.forename: Yohei
authors.initials: Y
authors.lastname: Fujiishi
authors.affiliation: Yakult Central Institute, Yakult Honsha Co., Ltd., 5-11 Izumi, Kunitachi-shi, Tokyo 186-8650, Japan.
authors.forename: Emiko
authors.initials: E
authors.lastname: Okada
authors.affiliation: Yakult Central Institute, Yakult Honsha Co., Ltd., 5-11 Izumi, Kunitachi-shi, Tokyo 186-8650, Japan.
authors.forename: Wakako
authors.initials: W
authors.lastname: Ohyama
mesh_terms: D000818:Animals; D001835:Body Weight / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D004912:Erythrocytes / Q000187:drug effects*; D008297:Male; D008565:Membrane Proteins / Q000097:blood / Q000235:genetics*; D009152:Mutagenicity Tests / Q000379:methods*; D009153:Mutagens / Q000633:toxicity*; D009154:Mutation; D051381:Rats; D017207:Rats, Sprague-Dawley; D012156:Reticulocytes / Q000187:drug effects*; D014520:Urethane / Q000633:toxicity*
chemical_list: D008565:Membrane Proteins; D009153:Mutagens; C080086:phosphatidylinositol glycan-class A protein; D014520:Urethane
keywords: In vivo mutation assay; Peripheral blood; Pig-a gene; Reticulocyte; Urethane