Characterization of the enhancing effect of protamine on the proliferative activity of hepatocyte growth factor in rat hepatocytes.
OBJECTIVE
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 alpha-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 (125)I-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.


CONCLUSIONS
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.
Liu
Ke-Xin
KX
Graduate School of Pharmaceutical Sciences, University of Tokyo, Hongo, Bunkyo-ku, Tokyo, 113-0033, Japan.
Kato
Yukio
Y
Matsumoto
Kunio
K
Nakamura
Toshikazu
T
Kaku
Taiichi
T
Sugiyama
Yuichi
Y
D015058:1-Naphthylisothiocyanate / Q000633:toxicity; D000818:Animals; D049109:Cell Proliferation / Q000187:drug effects*; D002478:Cells, Cultured; D004261:DNA Replication / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D006493:Heparin / Q000031:analogs & derivatives / Q000378:metabolism; D017228:Hepatocyte Growth Factor / Q000378:metabolism*; D022781:Hepatocytes / Q000187:drug effects* / Q000378:metabolism; D006801:Humans; D008099:Liver / Q000187:drug effects* / Q000378:metabolism; D008297:Male; D010766:Phosphorylation; D011479:Protamines / Q000493:pharmacokinetics / Q000494:pharmacology*; D011509:Proteoglycans / Q000378:metabolism; D019859:Proto-Oncogene Proteins c-met / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D011994:Recombinant Proteins / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D013997:Time Factors
C066855:HGF protein, human; D011479:Protamines; D011509:Proteoglycans; D011994:Recombinant Proteins; C036090:heparin proteoglycan; D015058:1-Naphthylisothiocyanate; D017228:Hepatocyte Growth Factor; D006493:Heparin; D019859:Proto-Oncogene Proteins c-met