title: Influence of CLOCK on cytotoxicity induced by diethylnitrosamine in mouse primary hepatocytes.
abstract: The Clock gene is a core clock factor that plays an essential role in generating circadian rhythms. In the present study, it was investigated whether the Clock gene affects the response to diethylnitrosamine (DEN)-induced cytotoxicity using mouse primary hepatocytes. DEN-induced cytotoxicity, after 24h exposure, was caused by apoptosis in hepatocytes isolated from wild-type mouse. On the other hand, Clock mutant mouse (Clk/Clk) hepatocytes showed resistance to apoptosis. Because apoptosis is an important pathway for suppressing carcinogenesis after genomic DNA damage, the mechanisms that underlie resistance to DEN-induced apoptosis were examined in Clk/Clk mouse hepatocytes. The mRNA levels of metabolic enzymes bioactivating DEN and apoptosis-inducing factors before DEN exposure were lower in Clk/Clk cells than in wild-type cells. The accumulation of p53 and Ser15 phosphorylated p53 after 8h DEN exposure was seen in wild-type cells but not in Clk/Clk cells. Caspase-3/7 activity was elevated during 24h DEN exposure in wild-type cells but not in Clk/Clk cells. In addition, resistance to DEN-induced apoptosis in Clk/Clk cells affected the cell viability. These studies suggested that the lower expression levels of metabolic enzymes bioactivating DEN and apoptosis inducing factors affected the resistance to DEN-induced apoptosis in Clk/Clk cells, and the Clock gene plays an important role in cytotoxicity induced by DEN.
authors.affiliation: Division of Clinical Pharmacy, Department of Medico-Pharmaceutical Sciences, Faculty of Pharmaceutical Sciences, Kyushu University, Fukuoka, Japan. matunaga@phar.kyushu-u.ac.jp
authors.forename: Naoya
authors.initials: N
authors.lastname: Matsunaga
authors.forename: Yumiko
authors.initials: Y
authors.lastname: Kohno
authors.forename: Keisuke
authors.initials: K
authors.lastname: Kakimoto
authors.forename: Akane
authors.initials: A
authors.lastname: Hayashi
authors.forename: Satoru
authors.initials: S
authors.lastname: Koyanagi
authors.forename: Shigehiro
authors.initials: S
authors.lastname: Ohdo
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000187:drug effects / Q000235:genetics; D056926:CLOCK Proteins / Q000172:deficiency / Q000235:genetics* / Q000502:physiology; D002470:Cell Survival / Q000187:drug effects / Q000235:genetics; D002478:Cells, Cultured; D003603:Cytotoxins / Q000096:biosynthesis / Q000633:toxicity*; D018736:DNA Adducts / Q000096:biosynthesis; D004052:Diethylnitrosamine / Q000633:toxicity*; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D022781:Hepatocytes / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism*; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D008813:Mice, Inbred ICR; D008817:Mice, Mutant Strains; D009154:Mutation / Q000187:drug effects; D034741:RNA, Small Interfering / Q000633:toxicity
chemical_list: D003603:Cytotoxins; D018736:DNA Adducts; D034741:RNA, Small Interfering; D004052:Diethylnitrosamine; D056926:CLOCK Proteins; C538944:Clock protein, mouse