Elevated levels of circulating ITIH4 are associated with hepatocellular carcinoma with nonalcoholic fatty liver disease: from pig model to human study | BMC Cancer | Full Text

Elevated levels of circulating ITIH4 are associated with hepatocellular carcinoma with nonalcoholic fatty liver disease: from pig model to human study | BMC Cancer | Full Text

BMC Cancer

Elevated levels of circulating ITIH4 are associated with hepatocellular carcinoma with nonalcoholic fatty liver disease: from pig model to human study

• Naohiko Nakamura,
• Etsuro HatanoEmail author,
• Kohta Iguchi,
• Motohiko Sato,
• Hiroaki Kawaguchi,
• Iwao Ohtsu,
• Takaki Sakurai,
• Nobuhiro Aizawa,
• Hiroko Iijima,
• Shuhei Nishiguchi,
• Takuya Tomono,
• Yukihiro Okuda,
• Seidai Wada,
• Satoru Seo,
• Kojiro Taura,
• Shinji Uemoto and
• Masaya IkegawaEmail author

BMC Cancer201919:621

https://doi.org/10.1186/s12885-019-5825-8

©  The Author(s). 2019

• Received: 1 April 2019
• Accepted: 12 June 2019
• Published: 25 June 2019

Open Peer Review reports

Abstract

Background

Noninvasive biomarkers are urgently needed for optimal management of nonalcoholic fatty liver disease (NAFLD) for the prevention of disease progression into nonalcoholic steatohepatitis (NASH) and hepatocellular carcinoma (HCC). In order to identify the biomarkers, we generated the swine hepatocellular carcinoma (HCC) model associated with NAFLD and performed serum proteomics on the model.

Methods

Microminipigs were fed a high-fat diet to induce NAFLD and a normal diet as the control. To induce HCC, diethylnitrosamine was intraperitoneally administered. Biopsied liver samples were histopathologically analyzed every 12 weeks. Serum proteins were separated by blue native two-dimensional gel electrophoresis and proteins of interest were subsequently identified by MALDI-TOF MS/MS. Human serum samples were analyzed to validate the candidate protein using antibody-mediated characterization.

Results

In the NAFLD pigs, hepatic histology of nonalcoholic steatohepatitis (NASH) was observed at 36 weeks, and HCC developed at 60 weeks. Among serum proteins identified with MALDI-TOF MS/MS, serum inter-alpha-trypsin inhibitor heavy chain 4 (ITIH4), an acute response protein which is secreted primarily by liver, was identified as the most characteristic protein corresponding with NAFLD progression and HCC development in the NAFLD pigs. With immunoassay, serum ITIH4 levels in the NAFLD pigs were chronologically increased in comparison with those in control animal. Furthermore, immunohistochemistry showed ITIH4 expression in hepatocytes also increased in both the cancer lesions and parenchyma as NAFLD progressed. Human study is also consistent with this observation because serum ITIH4 levels were significantly higher in HCC-NAFLD patients than in the simple steatosis, NASH, and virus-related HCC patients. Of note, HCC-NAFLD patients who had higher serum ITIH4 levels exhibited poorer prognosis after hepatectomy.

Conclusions

We established an HCC pig model associated with NAFLD. Serum proteomics on the swine HCC with NAFLD model implicated ITIH4 as a non-invasive biomarker reflecting NAFLD progression as well as subsequent HCC development. Most importantly, the results in the swine study have been validated in human cohort studies. Dissecting speciation of serum ITIH4 promises to have clinical utility in monitoring the disease.

Keywords

• Nonalcoholic fatty liver disease
• Hepatocellular carcinoma
• Pig model
• Inter-alpha-trypsin inhibitor heavy chain 4
• Blue native/SDS gel electrophoresis
• MALDI-TOF MS/MS