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In vitro and in vivo characteristics of hepatic oval cells modified with human hepatocyte growth factor

Abstract

Hepatocyte growth factor (HGF) is a multifunctional growth factor that controls cell scattering. It has been suggested that it regulates the proliferation of hepatic oval cells (HOCs). Using a HOC line that stably expresses the human HGF gene (hHGF), we investigated the in vitro proliferation and differentiation characteristics of hHGF-modified HOCs and explored their potential capacity for intrahepatic transplantation. A modified 2-acetylaminofluorene and partial hepatectomy (2-AAF/PH) model was established to activate the proliferation of oval cells in the rat liver. HOCs were transfected with the pBLAST2-hHGF plasmid and hHGF-carrying HOCs were selected based on blasticidin resistance. The level of hHGF secretion was determined via ELISA. Cell proliferation was determined using the MTT assay. Differentiation was induced by growth factor withdrawal. A two-cuff technique was used for orthotopic liver transplantation, and HOCs or hHGF-modified HOCs were transplanted into the recipients. The levels of biochemical indicators of liver function were measured after transplantation. An HOC line stably expressing hHGF was established. The transfected line showed greater hHGF secretion than normal HOCs. The hHGF gene promoted the proliferation capability of HOCs by reducing the peak time in vitro. The hHGF-modified HOCs differentiated into hepatocytes and bile duct epithelial cells upon growth factor withdrawal in vitro. In addition, hHGF-modified HOC transplantation significantly prolonged the median survival time (MST) and improved the liver function of recipients compared to HOC transplant recipients and nontransplanted controls. Our results indicate that hHGF-modified HOCs may have valuable properties for therapeutic liver regeneration after orthotopic liver transplantation.

Abbreviations

2-AAF:

2-acetylaminofluorene

ALB:

albumin

ALP:

alkaline phosphate

ALT:

alanine aminotransferase

ChE:

cholinesterase

DBil:

direct bilirubin

EGF:

epidermal growth factor

ELISA:

enzyme-linked immunoassay

GGT:

γ-glutamyltransferase

HGF:

hepatocyte growth factor

hHGF:

human hepatocyte growth factor

HOCs:

hepatic oval cells

I/R:

ischemia/reperfusion

LIF:

leukemia inhibitory factor

MST:

median survival time

PH:

partial hepatectomy

References

  1. 1.

    Faris, R.A., Konkin, T. and Halpert, G. Liver stem cells: a potential source of hepatocytes for the treatment of human liver disease. Artif. Organs 25 (2001) 513–521.

    CAS  PubMed  Article  Google Scholar 

  2. 2.

    Fausto, N. and Campbell, J.S. The role of hepatocytes and oval cells in liver regeneration and repopulation. Mech. Dev. 120 (2003) 117–130.

    CAS  PubMed  Article  Google Scholar 

  3. 3.

    Zimmermann, A. Regulation of liver regeneration. Nephrol. Dial. Transplant. 19Suppl 4 (2004) iv6–10.

    CAS  PubMed  Google Scholar 

  4. 4.

    Oertel, M. and Shafritz, D.A. Stem cells, cell transplantation and liver repopulation. Biochim. Biophys. Acta 1782 (2008) 61–74.

    CAS  PubMed Central  PubMed  Article  Google Scholar 

  5. 5.

    Williams, J.M., Oh, S.H., Jorgensen, M., Steiger, N., Darwiche, H., Shupe, T. and Petersen, B.E. The role of the Wnt family of secreted proteins in rat oval “stem” cell-based liver regeneration: Wnt1 drives differentiation. Am. J. Pathol. 176 (2010) 2732–2742.

    CAS  PubMed  Article  Google Scholar 

  6. 6.

    Matsumoto, K. and Nakamura, T. Emerging multipotent aspects of hepatocyte growth factor. J. Biochem. 119 (1996) 591–600.

    CAS  PubMed  Article  Google Scholar 

  7. 7.

    Huh, C.G., Factor, V.M., Sanchez, A., Uchida, K., Conner, E.A. and Thorgeirsson, S.S. Hepatocyte growth factor/c-met signaling pathway is required for efficient liver regeneration and repair. Proc. Natl. Acad. Sci. USA 101 (2004) 4477–4482.

    CAS  PubMed  Article  Google Scholar 

  8. 8.

    Matsumoto, K. and Nakamura, T. Hepatocyte growth factor: molecular structure, roles in liver regeneration, and other biological functions. Crit. Rev. Oncog. 3 (1992) 27–54.

    CAS  PubMed  Google Scholar 

  9. 9.

    Hasuike, S., Ido, A., Uto, H., Moriuchi, A., Tahara, Y., Numata, M., Nagata, K., Hori, T., Hayashi, K. and Tsubouchi, H. Hepatocyte growth factor accelerates the proliferation of hepatic oval cells and possibly promotes the differentiation in a 2-acetylaminofluorene/partial hepatectomy model in rats. J. Gastroenterol. Hepatol. 20 (2005) 1753–1761.

    CAS  PubMed  Article  Google Scholar 

  10. 10.

    Shiota, G., Kunisada, T., Oyama, K., Udagawa, A., Nomi, T., Tanaka, K., Tsutsumi, A., Isono, M., Nakamura, T., Hamada, H., Sakatani, T., Sell, S., Sato, K., Ito, H. and Kawasaki, H. In vivo transfer of hepatocyte growth factor gene accelerates proliferation of hepatic oval cells in a 2-acetylaminofluorene/partial hepatectomy model in rats. FEBS. Lett. 470 (2000) 325–330.

    CAS  PubMed  Article  Google Scholar 

  11. 11.

    Kato, H., Shimomura, T., Murai, R., Gonda, K., Ishii, K., Yoshida, Y., Kanbe, T., Hashiguchi, K., Sakabe, T., Takubo, K., Okano, J., Tsuchiya, H., Hoshikawa, Y., Kurimasa, A. and Shiota, G. Regulation of hepatic oval cell proliferation by adenoviral mediated hepatocyte growth factor gene transfer and signal transduction inhibitors. Hepatogastroenterology 54 (2007) 821–825.

    CAS  PubMed  Google Scholar 

  12. 12.

    Li, Z., Li, L., Ran, J.H., Zhang, S.N., Liu, J., Li, L.B. and Chen, J. Experimental study on hepatic oval cells to establish proliferation model and isolation, culture and differentiation in vitro in adult rat. China Journal of Modern Medicine 5 (2010) 31–34.

    Google Scholar 

  13. 13.

    Savatier, P., Lapillonne, H., van Grunsven, L.A., Rudkin, B.B. and Samarut, J. Withdrawal of differentiation inhibitory activity/leukemia inhibitory factor up-regulates D-type cyclins and cyclin-dependent kinase inhibitors in mouse embryonic stem cells. Oncogene 12 (1996) 309–322.

    CAS  PubMed  Google Scholar 

  14. 14.

    Leary, A.G., Wong, G.G., Clark, S.C., Smith, A.G. and Ogawa, M. Leukemia inhibitory factor differentiation-inhibiting activity/human interleukin for DA cells augments proliferation of human hematopoietic stem cells. Blood 75 (1990) 1960–1964.

    CAS  PubMed  Google Scholar 

  15. 15.

    Xie, R. and Xu, S.R. Two-cuff technique was applied in the establishment of orthotopic liver transplantation in rats Journal of Jiangsu University (Medicine Edition) 16 (2006) 392–395.

    Google Scholar 

  16. 16.

    Li, Z., Chen, J., Li, L., Ran, J.H., Li, X.H., Liu, Z.H., Liu, G.J., Gao, Y.C., Zhang, X.L. and Sun, H.D. Human hepatocyte growth factor (hHGF)-modified hepatic oval cells improve liver transplant survival. PLoS One 7 (2012) e44805.

    CAS  PubMed Central  PubMed  Article  Google Scholar 

  17. 17.

    Ma, P.C., Maulik, G., Christensen, J. and Salgia, R. c-Met: structure, functions and potential for therapeutic inhibition. Cancer Metastasis Rev. 22 (2003) 309–325.

    CAS  PubMed  Article  Google Scholar 

  18. 18.

    Schmidt, C., Bladt, F., Goedecke, S., Brinkmann, V., Zschiesche, W., Sharpe, M., Gherardi, E. and Birchmeier, C. Scatter factor/hepatocyte growth factor is essential for liver development. Nature 373 (1995) 699–702.

    CAS  PubMed  Article  Google Scholar 

  19. 19.

    Trusolino, L., Bertotti, A. and Comoglio, P.M. MET signalling: principles and functions in development, organ regeneration and cancer. Nat. Rev. Mol. Cell Biol. 11 (2010) 834–848.

    CAS  PubMed  Article  Google Scholar 

  20. 20.

    Montalvo-Jave, E.E., Escalante-Tattersfield, T., Ortega-Salgado, J.A., Pina, E. and Geller, D.A. Factors in the pathophysiology of the liver ischemiareperfusion injury. J. Surg. Res. 147 (2008) 153–159.

    CAS  PubMed Central  PubMed  Article  Google Scholar 

  21. 21.

    Kitta, K., Day, R.M., Ikeda, T. and Suzuki, Y.J. Hepatocyte growth factor protects cardiac myocytes against oxidative stress-induced apoptosis. Free Radic. Biol. Med. 31 (2001) 902–910.

    CAS  PubMed  Article  Google Scholar 

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Correspondence to Zhu Li.

Additional information

Zhu Li and Juan Chen contributed equally to this study

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Li, Z., Chen, J., Li, L. et al. In vitro and in vivo characteristics of hepatic oval cells modified with human hepatocyte growth factor. Cell Mol Biol Lett 18, 507–521 (2013). https://doi.org/10.2478/s11658-013-0104-1

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Key words

  • Hepatic oval cells
  • Hepatocyte growth factor
  • pBLAST2-hHGF
  • Plasmid transfection
  • Proliferation
  • Differentiation
  • Liver transplantation
  • Rats
  • In vitro
  • Liver regeneration