Hepatocarcinogenic Susceptibility of Fenofibrate and Its Possible Mechanism of Carcinogenicity in a Two-Stage Hepatocarcinogenesis Model of rasH2 Mice

Abstract

Fenofibrate (FF) has previously been shown to induce hepatocellular neoplasia in a conventional mouse bioassay (NDA 1993), but there has been no report to examine the carcinogenic susceptibility of rasH2 mice to this chemical. In the present study, male rasH2 mice were subjected to a twothirds partial hepatectomy (PH), followed by an N-diethylnitrosamine (DEN) initiation twenty-four hours after PH, and given a diet containing 0, 1200, or 2400 ppm FF for seven weeks. The incidences of preneoplastic foci were significantly increased in mice from the FF-treated groups. Immunohistochemistry revealed that significant increases in proliferating cell nuclear antigen (PCNA)-positive cells and cytokeratin 8/18 positive foci were observed in FF-treated groups. In addition, the transgene and several downstream molecules such as c-myc, c-jun, activating transcription factor 3 (ATF3), and cyclin D1 were overexpressed in these groups. These results suggest that the hepatocarcinogenic activity of rasH2 mice to FF can be detected in this hepatocarcinogenesis model and that up-regulation of genes for the ras/MAPK pathway and cell cycle was probably involved in the hepatocarcinogenic mechanism of rasH2 mice.

Cytokeratin 8/18 is a Useful Immunohistochemical Marker for Hepatocellular Proliferative Lesions in Mice

“…In our previous study, we performed a two-stage liver carcinogenesis experiment in transgenic mice carrying human prototype of c-Ha-ras gene (rasH2 mice) administered fenofibrate for 8 weeks after DEN initiation and reported that the numbers of CK8/18-positive foci were significantly increased in the livers of rasH2 mice [18]. In the present study, more hepatocellular foci were demonstrated in paraffin-embedded sections by CK8/18 immunohistochemistry than HE staining, and the total multiplicity of hepatocellular foci in HE-stained sections and positive for CK8/18 was 10.17 and 18.50, respectively, in Experiment I and 4.47 and 23.17, respectively, in Experiment II, although there were some foci being negative for CK8/18 in 5 of 6 mice in Experiment II.…”

“…It has been shown that cytokeratin (CK) 8/18 overexpression may drive neoplastic transformation of preneoplastic cells in GST-P-positive foci during rat hepatocarcinogenesis [16]. Moreover, we previously reported that hepatocellular altered foci induced in rasH2 mice given fenofibrate for 8 weeks after N-diethylnitrosamine (DEN) initiation were immunohistochemically stained with CK8/18 [18].Piperonyl butoxide (PBO) is a pesticide synergist that is widely used with pyrethroids for grain protection and as a domestic insecticide. PBO is a hepatocarcinogen in F344 rats fed a diet containing 1.2% PBO for 2 years [29] and CD-1 mice fed a diet containing 300 mg/kg/day PBO for 79 weeks [4].…”

“…It has been shown that cytokeratin (CK) 8/18 overexpression may drive neoplastic transformation of preneoplastic cells in GST-P-positive foci during rat hepatocarcinogenesis [16]. Moreover, we previously reported that hepatocellular altered foci induced in rasH2 mice given fenofibrate for 8 weeks after N-diethylnitrosamine (DEN) initiation were immunohistochemically stained with CK8/18 [18].…”

Gene expression analysis in the lung of the rasH2 transgenic mouse at week 4 prior to induction of malignant tumor formation by urethane and N-methylolacrylamide

“…indicated that the expressions of genes related to blood vessel formation and MAPK cascade activation were significantly upregulated in splenic hemangiosarcomas in rasH2 Tg mice treated with troglitazone and UR. Kawai et al (2008) showed enhanced mRNA expressions of the ras/MAPK pathway and cell cycle genes in the livers of the rasH2 Tg mice treated with FF for 7 weeks after DEN initiation. Most of the toxicogenomic studies in the rasH2 Tg mice were conducted with tissues showing obvious cancerous changes as described above.…”

“…They also demonstrated that 5 injections of urethane (UR) at 2-day intervals induced the formation of lung tumors at 5 weeks after the UR treatment (Okamura et al, 2006). In twostage carcinogenesis models established using rasH2 Tg mice, 16 weeks of troglitazone administration after UR initiation induced the formation of splenic hemangiosarcomas and 7 weeks of fenofibrate (FF) administration after N-diethylnitrosamine (DEN) initiation induced hepatocarcinogenesis (Kawai et al, 2008).…”