Development of a novel mouse model of hepatocellular carcinoma with nonalcoholic steatohepatitis using a high-fat, choline-deficient diet and intraperitoneal injection of diethylnitrosamine.
BACKGROUND
The incidence of hepatocellular carcinoma with nonalcoholic steatohepatitis is increasing, and its clinicopathological features are well established. Several animal models of nonalcoholic steatohepatitis have been developed to facilitate its study; however, few fully recapitulate all its clinical features, which include insulin resistance, inflammation, fibrosis, and carcinogenesis. Moreover, these models require a relatively long time to produce hepatocellular carcinoma reliably. The aim of this study was to develop a mouse model of hepatocellular carcinoma with nonalcoholic steatohepatitis that develops quickly and reflects all clinically relevant features.


METHODS
Three-week-old C57BL/6J male mice were fed either a standard diet (MF) or a choline-deficient, high-fat diet (HFCD). The mice in the MF + diethylnitrosamine (DEN) and HFCD + DEN groups received a one-time intraperitoneal injection of DEN at the start of the respective feeding protocols.


RESULTS
The mice in the HFCD and HFCD + DEN groups developed obesity early in the experiment and insulin resistance after 12 weeks. Triglyceride levels peaked at 8 weeks for all four groups and decreased thereafter. Alanine aminotransferase levels increased every 4 weeks, with the HFCD and HFCD + DEN groups showing remarkably high levels; the HFCD + DEN group presented the highest incidence of nonalcoholic steatohepatitis. The levels of fibrosis and steatosis varied, but they tended to increase every 4 weeks in the HFCD and HFCD + DEN groups. Computed tomography scans indicated that all the HFCD + DEN mice developed hepatic tumors from 20 weeks, some of which were glutamine synthetase-positive.


CONCLUSIONS
The nonalcoholic steatohepatitis-hepatocellular carcinoma model we describe here is simple to establish, results in rapid tumor formation, and recapitulates most of the key features of nonalcoholic steatohepatitis. It could therefore facilitate further studies of the development, oncogenic potential, diagnosis, and treatment of this condition.
Department of Surgery, School of Medicine, Keio University, 35 Shinanomachi, Shinjuku-ku, Tokyo, 160-8582, Japan.
Norihiro
N
Kishida
Department of Surgery, School of Medicine, Keio University, 35 Shinanomachi, Shinjuku-ku, Tokyo, 160-8582, Japan.|Chugai Pharmaceutical Endowed Research Chair in Molecular Targeted Therapy of Gastrointestinal Cancer, School of Medicine, Keio University, Tokyo, Japan.
Sachiko
S
Matsuda
Department of Surgery, School of Medicine, Keio University, 35 Shinanomachi, Shinjuku-ku, Tokyo, 160-8582, Japan. laplivertiger@gmail.com.
Osamu
O
Itano
Department of Surgery, School of Medicine, Keio University, 35 Shinanomachi, Shinjuku-ku, Tokyo, 160-8582, Japan.
Masahiro
M
Shinoda
Department of Surgery, School of Medicine, Keio University, 35 Shinanomachi, Shinjuku-ku, Tokyo, 160-8582, Japan.
Minoru
M
Kitago
Department of Surgery, School of Medicine, Keio University, 35 Shinanomachi, Shinjuku-ku, Tokyo, 160-8582, Japan.
Hiroshi
H
Yagi
Department of Surgery, School of Medicine, Keio University, 35 Shinanomachi, Shinjuku-ku, Tokyo, 160-8582, Japan.
Yuta
Y
Abe
Department of Surgery, School of Medicine, Keio University, 35 Shinanomachi, Shinjuku-ku, Tokyo, 160-8582, Japan.
Taizo
T
Hibi
Department of Pathology, School of Medicine, Keio University, Tokyo, Japan.
Yohei
Y
Masugi
Department of Surgery, Kawasaki Municipal Hospital, Kawasaki-ku, Japan.
Koichi
K
Aiura
Department of Pathology, School of Medicine, Keio University, Tokyo, Japan.
Michiie
M
Sakamoto
Department of Surgery, School of Medicine, Keio University, 35 Shinanomachi, Shinjuku-ku, Tokyo, 160-8582, Japan.
Yuko
Y
Kitagawa
D000818:Animals; D015415:Biomarkers / Q000097:blood; D001835:Body Weight; D006528:Carcinoma, Hepatocellular / Q000097:blood / Q000150:complications* / Q000473:pathology; D002796:Choline Deficiency; D059305:Diet, High-Fat; D004052:Diethylnitrosamine; D004195:Disease Models, Animal; D007150:Immunohistochemistry; D007274:Injections, Intraperitoneal; D007333:Insulin Resistance; D008113:Liver Neoplasms / Q000097:blood / Q000150:complications* / Q000473:pathology; D008297:Male; D008810:Mice, Inbred C57BL; D065626:Non-alcoholic Fatty Liver Disease / Q000097:blood / Q000150:complications* / Q000473:pathology; D009929:Organ Size
D015415:Biomarkers; D004052:Diethylnitrosamine
Diethylnitrosamine; Hepatocellular carcinoma; High-fat choline-deficient diet; Mouse model; Nonalcoholic steatohepatitis