Tgf-beta auto-induction and connective tissue growth factor expression in human renal tubule epithelial cells requires N-ras.
OBJECTIVE
Transforming growth factor (TGF) beta is strongly implicated in the progression of renal fibrosis. TGFbeta1 is reported to cause epithelial-mesenchymal transition, inhibition of epithelial cell proliferation, increased apoptosis, auto-induction of TGFbeta production and induction of secondary mediators of tissue fibrosis such as connective tissue growth factor (CTGF, CCN2). The aims of this study were to investigate the role of the Ras/MAP kinase pathway in TGFbeta1 inhibition of proliferation, TGFbeta auto-induction and TGFbeta1-induced CTGF expression in HKC human renal tubule epithelial cells.


RESULTS
TGFbeta1 (0-25 ng/ml) inhibited proliferation of HKC cells and at 25 ng/ml also induced apoptosis. After 5-10 min of incubation, TGFbeta1 increased cellular levels of phospho-ERK1/2 and phospho-AKT with a bell-shaped dose-response curve with a maximally effective concentration of 2.5 ng/ml. TGFbeta3 caused an increase in extracellular TGFbeta1, which was significantly reduced in the presence of PD 98059. TGFbeta1 increased cellular and secreted CTGF protein in HKC cells in a MEK-dependent manner. To identify the Ras isoform involved, specific antisense oligonucleotides targeted to Ha-Ras, Ki-Ras and N-Ras were employed. Only inhibition of N-Ras resulted in a significant reduction of auto-induced TGFbeta1 secretion and TGFbeta1-induced cellular and secreted CTGF.


CONCLUSIONS
These results establish that the Ras/MAP kinase pathway, specifically through N-Ras, mediates TGFbeta1 auto-induction and TGFbeta1-induced CTGF expression in human renal tubule epithelial cells.
Dockrell
Mark E C
ME
Department of Renal Medicine, King's College London School of Medicine, London, UK.
Phanish
Mysore K
MK
Hendry
Bruce M
BM
D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D055513:Connective Tissue Growth Factor / Q000096:biosynthesis*; D004847:Epithelial Cells / Q000378:metabolism; D005419:Flavonoids / Q000494:pharmacology; D006801:Humans; D007687:Kidney Tubules, Proximal; D020935:MAP Kinase Signaling System / Q000502:physiology; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D020928:Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D053773:Transforming Growth Factor beta1 / Q000378:metabolism / Q000502:physiology*; D053782:Transforming Growth Factor beta3 / Q000502:physiology*; D018631:ras Proteins / Q000502:physiology*
D005419:Flavonoids; D053773:Transforming Growth Factor beta1; D053782:Transforming Growth Factor beta3; D055513:Connective Tissue Growth Factor; D019950:Mitogen-Activated Protein Kinase 1; D020928:Mitogen-Activated Protein Kinases; D018631:ras Proteins; C093973:2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one