Activation of glomerular mesangial cells by hepatocyte growth factor through tyrosine kinase and protein kinase C.
Hepatocyte growth factor (HGF) induces mitogenesis, chemotaxis, and tubule formation in renal epithelial cells. This study examined the effects of wortmannin and protein kinase C (PKC) inhibitors on HGF-mediated changes in metabolic activity in glomerular mesangial cells and renal epithelial carcinoma A498 cells. The extracellular acidification rate of transformed mouse glomerular mesangial cells and A498 cells was measured as an index of metabolic activity with a microphysiometer. HGF increased the acidification rate of mesangial cells and A498 cells in a concentration-dependent fashion that was inhibited completely by the tyrosine kinase inhibitor tyrophostin-23 (100 microM). The PKC inhibitors RO-32-0432 and SKF-57048 also inhibited HGF-induced acidification. The IC50 values for SKF-57048 were 59 +/- 2 and 20 +/- 10 nM in mesangial cells and A498 cells, respectively (P < 0.05). 12-O-Tetradecanoylphorbol 13-acetate (TPA), a phorbol ester that activates PKC, increased acidification in mesangial and epithelial cells similar to HGF. Wortmannin, an inhibitor of phosphatidylinositol (PI) 3-kinase (IC50 value 1-10 nM), inhibited HGF-induced acidification with an IC50 of 93 +/- 31 and 9 +/- 1 nM in mesangial and A498 cells, respectively (P < 0.05). In contrast, there was no significant difference in the IC50 value of wortmannin for epidermal growth factor (EGF)-induced acidification between mesangial and A498 cells (23 +/- 9 vs 14 +/- 1 nM, respectively). Because the IC50 value for wortmannin in inhibiting HGF but not EGF-induced acidification was an order of magnitude higher in mesangial cells than in epithelial A498 cells, a wortmannin-sensitive PI 3-kinase pathway may not be involved in HGF-mediated acidification in mesangial cells.
Laping
N J
NJ
Department of Renal Pharmacology, SmithKline Beecham Pharmaceuticals, King of Prussia, PA 19406, USA.
Olson
B A
BA
DeWolf
R E
RE
Albrightson
C R
CR
Fredrickson
T
T
King
C
C
Chirivella
M
M
Ziyadeh
F N
FN
Nambi
P
P
D000730:Androstadienes / Q000494:pharmacology; D000818:Animals; D002396:Catechols / Q000494:pharmacology; D002478:Cells, Cultured; D015925:Cryopreservation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004815:Epidermal Growth Factor / Q000494:pharmacology; D005920:Glomerular Mesangium / Q000166:cytology / Q000187:drug effects / Q000502:physiology*; D017228:Hepatocyte Growth Factor / Q000494:pharmacology*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007680:Kidney Neoplasms; D007700:Kinetics; D051379:Mice; D008815:Mice, Inbred Strains; D009570:Nitriles / Q000494:pharmacology; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D016133:Polymerase Chain Reaction; D011493:Protein Kinase C / Q000378:metabolism*; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D019859:Proto-Oncogene Proteins c-met / Q000096:biosynthesis; D012333:RNA, Messenger / Q000096:biosynthesis; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014158:Transcription, Genetic / Q000187:drug effects; D014407:Tumor Cells, Cultured; D020032:Tyrphostins; D000077191:Wortmannin
D000730:Androstadienes; D002396:Catechols; D004791:Enzyme Inhibitors; D009570:Nitriles; D012333:RNA, Messenger; D020032:Tyrphostins; D004815:Epidermal Growth Factor; D017228:Hepatocyte Growth Factor; D019869:Phosphatidylinositol 3-Kinases; D011505:Protein-Tyrosine Kinases; D019859:Proto-Oncogene Proteins c-met; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate; C060641:tyrphostin A23; D000077191:Wortmannin