Angiotensin II stimulates alpha3(IV) collagen production in mouse podocytes via TGF-beta and VEGF signalling: implications for diabetic glomerulopathy.
BACKGROUND
The podocyte is bathed in an angiotensin II (AngII)-rich ultrafiltrate, but the impact of AngII on podocyte pathobiology is not well known. Because podocytes play a direct role in the glomerular basement membrane (GBM) thickening of diabetes, the alpha3(IV) collagen chain was examined. Podocyte expression of alpha3(IV) collagen may involve the transforming growth factor-beta (TGF-beta) and vascular endothelial growth factor (VEGF) systems.


METHODS
Cultured mouse podocytes were treated with various doses of AngII for selected periods of time, with or without inhibitors of TGF-beta and VEGF signalling, SB-431542 and SU5416, respectively. TGF-beta1 and VEGF were assayed by enzyme-linked immunosorbent assay (ELISA); alpha3(IV) collagen, TGF-beta type II receptor and phospho-Smad2 were assayed by immunoblotting.


RESULTS
AngII >or=10(-10) M was found to stimulate the production of alpha3(IV) collagen significantly in as short a time as 3 h. The expression of alpha3(IV) collagen was influenced by the TGF-beta system, but AngII did not increase the podocyte's production of TGF-beta1 ligand; rather, it increased the expression of the TGF-beta type II receptor and activated the TGF-beta signalling system through Smad2. Despite the TGF-beta receptor upregulation, synergy between AngII and TGF-beta1 to boost alpha3(IV) collagen production was not observed. However, blockade of TGF-beta signalling with SB-431542 prevented AngII from stimulating alpha3(IV) collagen production. Podocyte expression of alpha3(IV) collagen was also increased by the autocrine activity of VEGF. Podocytes were stimulated to secrete VEGF by 10(-10) M or higher AngII after 48 h. Blockade of the endogenous VEGF activity by SU5416 prevented AngII-stimulated alpha3(IV) collagen production.


CONCLUSIONS
AngII stimulates the podocyte to produce alpha3(IV) collagen protein via mechanisms involving TGF-beta and VEGF signalling. Alterations in alpha3(IV) collagen production may contribute to GBM thickening and perhaps proteinuria in diabetes.
Chen
Sheldon
S
Renal-Electrolyte and Hypertension Division of the University of Pennsylvania, Philadelphia, PA, USA. chens@mail.med.upenn.edu
Lee
Joseph S
JS
Iglesias-de la Cruz
M C
MC
Wang
Amy
A
Izquierdo-Lahuerta
Adriana
A
Gandhi
Nisha K
NK
Danesh
Farhad R
FR
Wolf
Gunter
G
Ziyadeh
Fuad N
FN
D000804:Angiotensin II / Q000008:administration & dosage / Q000494:pharmacology*; D000818:Animals; D001324:Autoantigens / Q000096:biosynthesis / Q000187:drug effects*; D018929:Cell Culture Techniques; D024141:Collagen Type IV / Q000096:biosynthesis / Q000187:drug effects*; D004305:Dose-Response Relationship, Drug; D004847:Epithelial Cells / Q000187:drug effects* / Q000378:metabolism; D007678:Kidney Glomerulus / Q000166:cytology / Q000187:drug effects*; D051379:Mice; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology; D013997:Time Factors; D016212:Transforming Growth Factor beta / Q000187:drug effects / Q000502:physiology; D042461:Vascular Endothelial Growth Factor A / Q000187:drug effects / Q000502:physiology; D014662:Vasoconstrictor Agents / Q000008:administration & dosage / Q000494:pharmacology*
D001324:Autoantigens; D024141:Collagen Type IV; D016212:Transforming Growth Factor beta; D042461:Vascular Endothelial Growth Factor A; D014662:Vasoconstrictor Agents; C057112:type IV collagen alpha3 chain; C467480:vascular endothelial growth factor A, mouse; D000804:Angiotensin II