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A Cholecystokinin Receptor Antagonist Halts Nonalcoholic Steatohepatitis and Prevents Hepatocellular Carcinoma

  • Robin D. Tucker
  • Victor Ciofoaia
  • Sandeep Nadella
  • Martha D. Gay
  • Hong Cao
  • Matthew Huber
  • Anita Safronenka
  • Narayan Shivapurkar
  • Bhaskar Kallakury
  • Annie J. Kruger
  • Alexander H. K. Kroemer
  • Jill P. SmithEmail author
Original Article

Abstract

Background and Aims

Nonalcoholic steatohepatitis (NASH) is a common inflammatory liver condition that may lead to cirrhosis and hepatocellular carcinoma (HCC). Risk factors for NASH include a saturated fat diet, altered lipid metabolism, and genetic and epigenetic factors, including microRNAs. Serum levels of cholecystokinin (CCK) are elevated in mice and humans that consume a high-saturated fat diet. CCK receptors (CCK-Rs) have been reported on fibroblasts which when activated can induce fibrosis; however, their role in hepatic fibrosis remains unknown. We hypothesized that elevated levels of CCK acting on the CCK-Rs play a role in the development of NASH and in NASH-associated HCC.

Methods

We performed a NASH Prevention study and Reversal study in mice fed a saturated fat 75% choline-deficient–ethionine-supplemented (CDE) diet for 12 or 18 weeks. In each study, half of the mice received untreated drinking water, while the other half received water supplemented with the CCK-R antagonist proglumide. CCK-R expression was evaluated in mouse liver and murine HCC cells.

Results

CCK receptor antagonist treatment not only prevented NASH but also reversed hepatic inflammation, fibrosis, and steatosis and normalized hepatic transaminases after NASH was established. Thirty-five percent of the mice on the CDE diet developed HCC compared with none in the proglumide-treated group. We found that CCK-BR expression was markedly upregulated in mouse CDE liver and HCC cells compared with normal hepatic parenchymal cells, and this expression was epigenetically regulated by microRNA-148a.

Conclusion

These results support the novel role of CCK receptors in the pathogenesis of NASH and HCC.

Keywords

Hepatocellular carcinoma Nonalcoholic steatohepatitis Cholecystokinin Fibrosis Receptors 

Abbreviations

ALT

Alanine aminotransferase

AST

Serum aspartate aminotransferase

CCK

Cholecystokinin

CCK-AR

CCK-A receptor

CCK-BR

CCK-B receptor

CDE

Choline-deficient–ethionine-supplemented diet

CDE/Reg

Choline-deficient–ethionine-supplemented diet and regular untreated water

CDE/Prog

Choline-deficient–ethionine-supplemented diet and proglumide-treated water

CON/Reg

Control diet and regular untreated water

CON/Prog

Control diet and proglumide-treated water

DILI

Drug-induced liver injury

FAP

Fibroblast-activated protein

FAs

Fatty acids

GI

Gastrointestinal

HCC

Hepatocellular carcinoma

KO

Knockout

miR

MicroRNA

NAFLD

Nonalcoholic fatty liver disease

NASH

Nonalcoholic steatohepatitis

PCR

Polymerase chain reaction

PIVENS

Pioglitazone, vitamin E, or placebo for nonalcoholic steatohepatitis

qRT-PCR

Quantitative real-time polymerase chain reaction

Notes

Acknowledgments

We appreciate the staff in the Lombardi Georgetown Histology Core facility and technical assistance of the Department of Comparative Medicine.

Author’s contribution

JPS conceived and designed the study. RT, VC, SN, MG, HC, MH, AS, NS, BK, and JPS carried out acquisition of data. RT, JPS, HC, AHKK, MG, and AJK analyzed and interpreted the data. JPS and RT drafted the manuscript. All authors critically revised and approved the manuscript. HC, MG, MH, NS, and JPS carried out statistical analysis. JPS obtained funding.

Funding

The authors were supported by the AGA Elsevier Pilot Research Award and also supported by the Ruesch Center for the Cure of Gastrointestinal Cancers, Georgetown University Medical Center. Postdoctoral support was provided by a NIH training grant to Martha Gay TL1TR001431. These studies were conducted in part at the Lombardi Comprehensive Cancer Center Histopathology & Tissue Shared resource and in the Preclinical Imaging Research Laboratory which is supported in part by NIH/NCI Grant P30-CA051008.

Compliance with Ethical Standards

Conflict of interest

Jill P. Smith has intellectual property on the use of CCK receptor antagonists in NASH. Sandeep Nadella and Jill P. Smith are co-inventors on a pending patent for proglumide and fibrosis. None of the other authors have conflicts to report.

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Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  • Robin D. Tucker
    • 1
  • Victor Ciofoaia
    • 2
  • Sandeep Nadella
    • 2
  • Martha D. Gay
    • 2
  • Hong Cao
    • 2
  • Matthew Huber
    • 2
  • Anita Safronenka
    • 3
  • Narayan Shivapurkar
    • 2
  • Bhaskar Kallakury
    • 1
  • Annie J. Kruger
    • 2
  • Alexander H. K. Kroemer
    • 4
  • Jill P. Smith
    • 2
    Email author
  1. 1.Department of PathologyGeorgetown UniversityWashingtonUSA
  2. 2.Department of MedicineGeorgetown UniversityWashingtonUSA
  3. 3.Department of BiochemistryGeorgetown UniversityWashingtonUSA
  4. 4.Department of Transplant SurgeryGeorgetown UniversityWashingtonUSA

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