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Characterization of the Enhancing Effect of Protamine on the Proliferative Activity of Hepatocyte Growth Factor in Rat Hepatocytes

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Abstract

Purpose

The aim of the present study was to characterize the mechanism of the stimulatory effect of protamine on HGF activity.

Methods

The enhancing effects of protamine on the proliferative activity of HGF were investigated in vivo, in primary cultured rat hepatocytes, and in perfused rat liver.

Results

In α-naphthylisothiocyanate-intoxicated rats, pretreatment with protamine increased HGF-induced autophosphorylation of the HGF receptor in liver. The maximum enhancing effect of protamine on HGF-induced DNA synthesis of hepatocytes required a 10 min-pretreatment period both in vivo and in vitro, and the stimulatory effect of protamine was not observed when it was administered simultaneously with HGF. Preperfusion of the liver with protamine for 10 min decreased the non-saturable portion of hepatic clearance for 125I-HGF, which is mainly mediated by cell-surface heparan-sulfate proteoglycan (HSPG). Inhibition of HGF binding to heparin by protamine was confirmed using heparin-coated sepharose. This inhibition also required 10 min of pretreatment, for protamine to bind heparin.

Conclusion

The enhancing effect of protamine on the mitogenic activity of HGF on hepatocytes requires pretreatment with protamine for a short period presumably required for its binding to cell-surface heparin, implying possible regulation of c-met autophosphorylation by HSPG.

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References

  1. T. Nakamura, T. Nishizawa, M. Hagiya, T. Seki, M. Shimonishi, A. Sugimura, K. Tashiro, and S. Shimizu. Molecular cloning and expression of human hepatocyte growth factor. Nature. 342:440–443 (1989). doi:10.1038/342440a0.

    Article  PubMed  CAS  Google Scholar 

  2. T. Nakamura, H. Teramoto, A. Icto, and A. Ichihara. Purification and characterization of a growth factor from rat platelets for mature parenchymal hepatocytes in primary cultures. Proc. Natl. Acad. Sci. U. S. A. 83:6489–6493 (1986). doi:10.1073/pnas.83.17.6489.

    Article  PubMed  CAS  Google Scholar 

  3. Y. Ishiki, H. Ohnishi, Y. Muto, K. Matsumoto, and T. Nakamura. Direct evidence that hepatocyte growth factor is a hepatotrophic factor for liver regeneration and for potent anti-hepatitis action in vivo. Hepatology. 16:1227–1235 (1992).

    PubMed  CAS  Google Scholar 

  4. Y. Kato, K. X. Liu, T. Nakamura, and Y. Sugiyama. Heparin–hepatocyte growth factor complex with low plasma clearance and retained hepatocyte proliferating activity. Hepatology. 20:417–424 (1994).

    Article  PubMed  CAS  Google Scholar 

  5. J. K. Spix, E. Y. Chay, E. R. Block, and J. K. Klarlund. Hepatocyte growth factor induces epithelial cell motility through transactivation of the epidermal growth factor receptor. Exp. Cell Res. 313:3319–3325 (2007). doi:10.1016/j.yexcr.2007.06.006.

    Article  PubMed  CAS  Google Scholar 

  6. Y. Matsumoto, T. Motoki, S. Kubota, M. Takigawa, H. Tsubouchi, and E. Gohda. Inhibition of tumor–stromal interaction through HGF/Met signaling by valproic acid. Biochem. Biophys. Res. Commun. 366:110–116 (2008). doi:10.1016/j.bbrc.2007.11.089.

    Article  PubMed  CAS  Google Scholar 

  7. A. Catizone, G. Ricci, J. D. Bravo, and M. Galdieri. Hepatocyte growth factor modulates in vitro survival and proliferation of germ cells during postnatal testis development. J. Endocrinol. 189:137–146 (2006). doi:10.1677/joe.1.06528.

    Article  PubMed  CAS  Google Scholar 

  8. E. Cacci, M. Salani, S. Anastasi, I. Perroteau, G. Poiana, S. Biagioni, and G. Augusti-Tocco. Hepatocyte growth factor stimulates cell motility in cultures of the striatal progenitor cells ST14A. J. Neurosci. Res. 74:760–768 (2003). doi:10.1002/jnr.10799.

    Article  PubMed  CAS  Google Scholar 

  9. M. Johansson, G. Mattsson, A. Andersson, L. Jansson, and P. O. Carlsson. Islet endothelial cells and pancreatic beta-cell proliferation: studies in vitro and during pregnancy in adult rats. Endocrinology. 147:2315–2324 (2006). doi:10.1210/en.2005-0997.

    Article  PubMed  CAS  Google Scholar 

  10. K. X. Liu, Y. Kato, T. Terasaki, T. Nakamura, and Y. Sugiyama. Change in hepatic handling of hepatocyte growth factor during liver regeneration in rats. Am. J. Physiol. 269:G745–G753 (1995).

    PubMed  CAS  Google Scholar 

  11. K. X. Liu, Y. Kato, M. Yamazaki, O. Higuchi, T. Nakamura, and Y. Sugiyama. Decrease in the hepatic uptake clearance of hepatocyte growth factor (HGF) in CCl4-intoxicated rats. Hepatology. 17:651–660 (1993). doi:10.1002/hep.1840170420.

    Article  PubMed  CAS  Google Scholar 

  12. S. Hagiwara, T. Otsuka, Y. Yamazaki, T. Kosone, N. Sohara, T. Ichikawa, K. Sato, S. Kakizaki, H. Takagi, and M. Mori. Overexpression of NK2 promotes liver fibrosis in carbon tetrachloride-induced chronic liver injury. Liver Int. 28:126–131 (2007).

    PubMed  Google Scholar 

  13. Y. H. Lee, Y. J. Suzuki, A. J. Griffin, and R. M. Day. Hepatocyte growth factor regulates cyclooxygenase-2 expression via beta-catenin, Akt, and p42/p44 MAPK in human bronchial epithelial cells. Am. J. Physiol. 294:L778–L786 (2008).

    CAS  Google Scholar 

  14. E. Vigna, L. Naldini, L. Tamagnone, P. Longati, A. Bardelli, F. Maina, C. Ponzetto, and P. M. Comoglio. Hepatocyte growth factor and its receptor, the tyrosine kinase encoded by the c-MET proto-oncogene. Cell Mol. Biol. 40:597–604 (1994).

    PubMed  CAS  Google Scholar 

  15. C. C. Lee, and K. M. Yamada. Alternatively spliced juxtamembrane domain of a tyrosine kinase receptor is a multifunctional regulatory site. Deletion alters cellular tyrosine phosphorylation pattern and facilitates binding of phosphatidylinositol-3-OH kinase to the hepatocyte growth factor receptor. J. Biol. Chem. 270:507–510 (1995). doi:10.1074/jbc.270.2.507.

    Article  PubMed  CAS  Google Scholar 

  16. A. Bardelli, C. Ponzetto, and P. M. Comoglio. Identification of functional domains in the hepatocyte growth factor and its receptor by molecular engineering. J. Biotechnol. 37:109–122 (1994). doi:10.1016/0168-1656(94)90002-7.

    Article  PubMed  CAS  Google Scholar 

  17. K. X. Liu, Y. Kato, M. Narukawa, D. C. Kim, M. Hanano, O. Higuchi, T. Nakamura, and Y. Sugiyama. The importance of the liver in the plasma clearance of hepatocyte growth factor in rats. Am. J. Physiol. 263:G642–G649 (1992).

    PubMed  CAS  Google Scholar 

  18. K. X. Liu, Y. Kato, T. Terasaki, S. Aoki, K. Okumura, T. Nakamura, and Y. Sugiyama. Contribution of parenchymal and non-parenchymal liver cell to the clearance of hepatocyte growth factor from the circulation in rats. Pharm. Res. 12:737–1740 (1995).

    Google Scholar 

  19. K. X. Liu, Y. Kato, T. I. Kaku, K. Okumura, T. Nakamura, and Y. Sugiyama. Protamine enhances the proliferative activity of hepatocyte growth factor. Am. J. Physiol. 274:G21–G28 (1998).

    PubMed  CAS  Google Scholar 

  20. K. X. Liu, Y. Kato, T. I. Kaku, T. Nakamura, and Y. Sugiyama. Existence of two nonlinear elimination mechanisms for hepatocyte growth factor in rats. Am. J. Physiol. 273(5 Pt 1):E891–E897 (1997).

    PubMed  CAS  Google Scholar 

  21. L. E. Kemp, B. Mulloy, and E. Gherardi. Signalling by HGF/SF and Met: the role of heparin sulphate co-receptors. Biochem. Soc. Trans. 34(Pt 3):414–417 (2006). doi:10.1042/BST0340414.

    PubMed  CAS  Google Scholar 

  22. K. Matsumoto, and T. Nakamura. NK4 gene therapy targeting HGF-Met and angiogenesis. Front. Biosci. 13:1943–1951 (2008). doi:10.2741/2813.

    Article  PubMed  CAS  Google Scholar 

  23. Y. L. Yin, H. L. Chen, H. M. Kuo, and S. P. He. NK3 and NK4 of HGF enhance filamin production via STAT pathway, but not NK1 and NK2 in human breast cancer cells. Acta Pharmacol. Sin. 29:728–735 (2008). doi:10.1111/j.1745-7254.2008.00799.x.

    Article  PubMed  CAS  Google Scholar 

  24. A. Krishnan, K. Viker, H. Rietema, M. Telgenkamp, B. Knudsen, and M. Charlton. Prolonged engraftment of human hepatocytes in mice transgenic for the deleted form of human hepatocyte growth factor. Hepatol. Res. 37:854–862 (2007). doi:10.1111/j.1872-034X.2007.00139.x.

    Article  PubMed  CAS  Google Scholar 

  25. V. C. Yang, Y. Y. Fu, C. C. Teng, S. C. Ma, and J. N. Shanberge. A method for the quantitation of protamine in plasma. Thromb. Res. 74:427–434 (1994). doi:10.1016/0049-3848(94)90158-9.

    Article  PubMed  CAS  Google Scholar 

  26. S. Miyauchi, Y. Sawada, T. Iga, M. Hanano, and Y. Sugiyama. Comparison of the hepatic uptake clearances of fifteen drugs with a wide range of membrane permeabilities in isolated rat hepatocytes and perfused rat livers. Pharm. Res. 10:434–440 (1993). doi:10.1023/A:1018952709120.

    Article  PubMed  CAS  Google Scholar 

  27. M. Kato, Y. Kato, T. Nakamura, and Y. Sugiyama. Efficient extraction by the liver governs overall elimination of hepatocyte growth factor in rats. J. Pharmacol. Exp. Ther. 290:373–379 (1999).

    PubMed  CAS  Google Scholar 

  28. K. Komamura, J. Miyazaki, E. Imai, K. Matsumoto, T. Nakamura, and M. Hori. Hepatocyte growth factor gene therapy for hypertension. Methods Mol. Biol. 423:393–404 (2008). doi:10.1007/978-1-59745-194-9_31.

    Article  PubMed  CAS  Google Scholar 

  29. R. Gong, A. Rifai, Y. Ge, S. Chen, and L. D. Dworkin. Hepatocyte growth factor suppresses proinflammatory NFkappaB activation through GSK3beta inactivation in renal tubular epithelial cells. J. Biol. Chem. 283:7401–7410 (2008). doi:10.1074/jbc.M710396200.

    Article  PubMed  CAS  Google Scholar 

  30. T. H. Cheng, W. T. Lee, J. S. Jeng, C. M. Wu, G. C. Liu, M. Y. Chiang, and Y. M. Wang. Synthesis and characterization of a novel paramagnetic macromolecular complex [Gd(TTDASQ-protamine)]. Dalton Trans. 21:5149–5155 (2006). doi:10.1039/b604783a.

    Article  CAS  Google Scholar 

  31. E. Moreno, J. C. Meneu, J. Calvo, B. Perez, A. G. Sesma, A. Manrique, I. Vegh, A. M. Aragon, M. Grau, A. Gimeno, C. Jimenez, R. Gomez, A. Moreno, M. Abradelo, I. Garcia, and A. de la Calle. Modulation of hepatocyte growth factor plasma levels in relation to the dose of exogenous heparin administered: an experimental study in rats. Transplant. Proc. 37:3943–3947 (2005). doi:10.1016/j.transproceed.2005.10.089.

    Article  PubMed  CAS  Google Scholar 

  32. D. C. West, C. G. Rees, L. Duchesne, S. J. Patey, C. J. Terry, J. E. Turnbull, M. Delehedde, C. W. Heegaard, F. Allain, C. Vanpouille, D. Ron, and D. G. Fernig. Interactions of multiple heparin binding growth factors with neuropilin-1 and potentiation of the activity of fibroblast growth factor-2. J. Biol. Chem. 280:13457–13464 (2005). doi:10.1074/jbc.M410924200.

    Article  PubMed  CAS  Google Scholar 

  33. D. Naka, T. Ishii, T. Shimomura, T. Hishida, and H. Hara. Heparin modulates the receptor-binding and mitogenic activity of hepatocyte growth factor on hepatocytes. Exp Cell Res. 209:317–324 (1993). doi:10.1006/excr.1993.1316.

    Article  PubMed  CAS  Google Scholar 

  34. R. H. Schwall, L. Y. Chang, P. J. Godowski, D. W. Kahn, K. J. Hillan, K. D. Bauer, and T. F. Zioncheck. Heparin induces dimerization and confers proliferative activity onto the hepatocyte growth factor antagonists NK1 and NK2. J .Cell Biol. 133:709–718 (1996). doi:10.1083/jcb.133.3.709.

    Article  PubMed  CAS  Google Scholar 

  35. M. Machide, K. Kamitori, and S. Kohsaka. Hepatocyte growth factor-induced differential activation of phospholipase Cgamma1 and phosphatidylinositol 3-kinase is regulated by tyrosine phosphatase, SHP-1 in astrocytes. J. Biol. Chem. 275:31392–31398 (2000). doi:10.1074/jbc.M002817200.

    Article  PubMed  CAS  Google Scholar 

  36. M. Hecht, M. Papoutsi, H. D. Tran, J. Wilting, and L. Schweigerer. Hepatocyte growth factor/c-Met signaling promotes the progression of experimental human neuroblastomas. Cancer Res. 64:6109–6118 (2004). doi:10.1158/0008-5472.CAN-04-1014.

    Article  PubMed  CAS  Google Scholar 

  37. V. Crljen, S. Volinia, and H. Banfic. Hepatocyte growth factor activates phosphoinositide 3-kinase C2 beta in renal brush-border plasma membranes. Biochem. J. 365(Pt 3):791–799 (2002).

    PubMed  CAS  Google Scholar 

  38. Y. Okano, K. Mizuno, S. Osada, T. Nakamura, and Y. Nozawa. Tyrosine phosphorylation of phospholipase C gamma in c-met/HGF receptor-stimulated hepatocytes: comparison with HepG2 hepatocarcinoma cells. Biochem. Biophys. Res. Commun. 190:842–842 (1993). doi:10.1006/bbrc.1993.1125.

    Article  PubMed  CAS  Google Scholar 

  39. J. A. Price, J. Caldwell, and N. J. Hewitt. The effect of EGF and the comitogen, norepinephrine, on the proliferative responses of fresh and cryopreserved rat and mouse hepatocytes. Cryobiology. 53:182–193 (2006). doi:10.1016/j.cryobiol.2006.05.008.

    Article  PubMed  CAS  Google Scholar 

  40. V. B. Lokeshwar, S. S. Huang, and J. S. Huang. Protamine enhances epidermal growth factor (EGF)-stimulated mitogenesis by increasing cell surface EGF receptor number. J. Biol. Chem. 264:19318–19326 (1989).

    PubMed  CAS  Google Scholar 

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Author information

Author notes

  1. Kunio Matsumoto

    Present address: Cancer Research Institute, Kanazawa University, Takara-machi, Kanazawa, 920-0934, Japan

  2. Yukio Kato

    Present address: Institute of Medical, Pharmaceutical and Health Sciences, Kanazawa University, Kakuma-machi, Kanazawa, 920-1192, Japan

Authors and Affiliations

  1. Graduate School of Pharmaceutical Sciences, University of Tokyo, Hongo, Bunkyo-ku, Tokyo, 113-0033, Japan

    Ke-Xin Liu, Yukio Kato & Yuichi Sugiyama

  2. Biomedical Research Center, Osaka University School of Medicine, Suita, Osaka, 565-0871, Japan

    Kunio Matsumoto & Toshikazu Nakamura

  3. College of Pharmacy, Dalian Medical University, 9 Western Section, Lvshun South Street, 116044, Dalian, China

    Ke-Xin Liu

  4. Japan Bioproducts Industry Co. Ltd., Tomigaya, Shibuya-ku, Tokyo, 151-0063, Japan

    Ke-Xin Liu & Taiichi Kaku

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  1. Ke-Xin Liu
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  2. Yukio Kato
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Correspondence to Yuichi Sugiyama.

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Liu, KX., Kato, Y., Matsumoto, K. et al. Characterization of the Enhancing Effect of Protamine on the Proliferative Activity of Hepatocyte Growth Factor in Rat Hepatocytes. Pharm Res 26, 1012–1021 (2009). https://doi.org/10.1007/s11095-008-9810-1

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