Dickkopf-3 in aberrant endothelial secretome triggers renal fibroblast activation and endothelial-mesenchymal transition.
Adaptor Proteins, Signal Transducing
/ physiology
Animals
Endothelium, Vascular
/ metabolism
Epithelial-Mesenchymal Transition
Fibrosis
/ metabolism
Kidney Diseases
/ metabolism
Mice
Mice, Knockout
Proteome
/ analysis
Proteomics
Receptor, Transforming Growth Factor-beta Type II
/ physiology
Sirtuin 1
/ physiology
Wnt Signaling Pathway
beta Catenin
/ metabolism
Journal
Nephrology, dialysis, transplantation : official publication of the European Dialysis and Transplant Association - European Renal Association
ISSN: 1460-2385
Titre abrégé: Nephrol Dial Transplant
Pays: England
ID NLM: 8706402
Informations de publication
Date de publication:
01 01 2019
01 01 2019
Historique:
received:
28
12
2017
accepted:
18
03
2018
pubmed:
5
5
2018
medline:
28
12
2019
entrez:
5
5
2018
Statut:
ppublish
Résumé
Our laboratory has previously demonstrated that Sirt1endo-/- mice show endothelial dysfunction and exaggerated renal fibrosis, whereas mice with silenced endothelial transforming growth factor beta (TGF-β) signaling are resistant to fibrogenic signals. Considering the fact that the only difference between these mutant mice is confined to the vascular endothelium, this indicates that secreted substances contribute to these contrasting responses. We performed an unbiased proteomic analysis of the secretome of renal microvascular endothelial cells (RMVECs) isolated from these two mutants. We cultured renal fibroblasts and RMVECs and used microfluidic devices for coculturing. Dickkopf-3 (DKK3), a putative ligand of the Wnt/β-catenin pathway, was present exclusively in the fibrogenic secretome. In cultured fibroblasts, DKK3 potently induced myofibroblast activation. In addition, DKK3 antagonized effects of DKK1, a known inhibitor of the Wnt pathway, in conversion of fibroblasts to myofibroblasts. In RMVECs, DKK3 induced endothelial-mesenchymal transition and impaired their angiogenic competence. The inhibition of endothelial outgrowth, enhanced myofibroblast formation and endothelial-mesenchymal transition were confirmed in coculture. In reporter DKK3-eGFP × Col3.6-GFPcyan mice, DKK3 was marginally expressed under basal conditions. Adriamycin-induced nephropathy resulted in upregulation of DKK3 expression in tubular and, to a lesser degree, endothelial compartments. Sulindac sulfide was found to exhibit superior Wnt pathway-suppressive action and decreased DKK3 signals and the extent of renal fibrosis. In conclusion, this unbiased proteomic screen of the profibrogenic endothelial secretome revealed DKK3 acting as an agonist of the Wnt pathway, enhancing formation of myofibroblasts and endothelial-mesenchymal transition and impairing angiogenesis. A potent inhibitor of the Wnt pathway, sulindac sulfide, suppressed nephropathy-induced DKK3 expression and renal fibrosis.
Sections du résumé
Background
Our laboratory has previously demonstrated that Sirt1endo-/- mice show endothelial dysfunction and exaggerated renal fibrosis, whereas mice with silenced endothelial transforming growth factor beta (TGF-β) signaling are resistant to fibrogenic signals. Considering the fact that the only difference between these mutant mice is confined to the vascular endothelium, this indicates that secreted substances contribute to these contrasting responses.
Methods
We performed an unbiased proteomic analysis of the secretome of renal microvascular endothelial cells (RMVECs) isolated from these two mutants. We cultured renal fibroblasts and RMVECs and used microfluidic devices for coculturing.
Results
Dickkopf-3 (DKK3), a putative ligand of the Wnt/β-catenin pathway, was present exclusively in the fibrogenic secretome. In cultured fibroblasts, DKK3 potently induced myofibroblast activation. In addition, DKK3 antagonized effects of DKK1, a known inhibitor of the Wnt pathway, in conversion of fibroblasts to myofibroblasts. In RMVECs, DKK3 induced endothelial-mesenchymal transition and impaired their angiogenic competence. The inhibition of endothelial outgrowth, enhanced myofibroblast formation and endothelial-mesenchymal transition were confirmed in coculture. In reporter DKK3-eGFP × Col3.6-GFPcyan mice, DKK3 was marginally expressed under basal conditions. Adriamycin-induced nephropathy resulted in upregulation of DKK3 expression in tubular and, to a lesser degree, endothelial compartments. Sulindac sulfide was found to exhibit superior Wnt pathway-suppressive action and decreased DKK3 signals and the extent of renal fibrosis.
Conclusions
In conclusion, this unbiased proteomic screen of the profibrogenic endothelial secretome revealed DKK3 acting as an agonist of the Wnt pathway, enhancing formation of myofibroblasts and endothelial-mesenchymal transition and impairing angiogenesis. A potent inhibitor of the Wnt pathway, sulindac sulfide, suppressed nephropathy-induced DKK3 expression and renal fibrosis.
Identifiants
pubmed: 29726981
pii: 4991926
doi: 10.1093/ndt/gfy100
pmc: PMC6322445
doi:
Substances chimiques
Adaptor Proteins, Signal Transducing
0
CTNNB1 protein, mouse
0
Dkk3 protein, mouse
0
Proteome
0
beta Catenin
0
Receptor, Transforming Growth Factor-beta Type II
EC 2.7.11.30
Tgfbr2 protein, mouse
EC 2.7.11.30
Sirt1 protein, mouse
EC 3.5.1.-
Sirtuin 1
EC 3.5.1.-
Types de publication
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Langues
eng
Sous-ensembles de citation
IM
Pagination
49-62Subventions
Organisme : NIDDK NIH HHS
ID : R01 DK045462
Pays : United States
Organisme : NIDDK NIH HHS
ID : R01 DK052783
Pays : United States
Organisme : NIDDK NIH HHS
ID : R01 DK054602
Pays : United States
Organisme : NIAMS NIH HHS
ID : R21 AR055750
Pays : United States
Références
Can J Cardiol. 2016 May;32(5):634-41
pubmed: 26948035
Nat Med. 2010 Dec;16(12):1400-6
pubmed: 21102460
Circ Res. 2015 Jan 16;116(2):312-22
pubmed: 25593276
Biochim Biophys Acta. 2012 Jan;1825(1):18-28
pubmed: 21982838
Stem Cells. 2006 Jan;24(1):13-22
pubmed: 16099999
Proc Natl Acad Sci U S A. 2000 Jul 5;97(14):8015-20
pubmed: 10859350
Biochimie. 2013 Dec;95(12):2326-35
pubmed: 24036368
Nat Rev Cancer. 2010 Feb;10(2):138-46
pubmed: 20094048
Angiogenesis. 2011 May;14(2):163-72
pubmed: 21234671
Nature. 2016 Jan 21;529(7586):316-25
pubmed: 26791722
J Am Soc Nephrol. 2017 Aug;28(8):2322-2336
pubmed: 28336721
Kidney Int. 2017 Sep;92(3):558-568
pubmed: 28476555
Cancer Res. 2007 Nov 1;67(21):10123-8
pubmed: 17974953
Growth Factors. 2010 Aug;28(4):232-42
pubmed: 20370576
J Clin Invest. 2015 Oct 26;125(12):4514-28
pubmed: 26517696
J Am Soc Nephrol. 2014 Feb;25(2):276-91
pubmed: 24136919
J Biol Chem. 2015 Aug 7;290(32):19844-52
pubmed: 26105053
JCI Insight. 2016 Jan 21;1(1):e84916
pubmed: 27699213
Nature. 2013 Jun 27;498(7455):492-6
pubmed: 23748444
Eur Heart J. 2017 May 7;38(18):1413-1425
pubmed: 27099262
J Fluoresc. 2014 Jul;24(4):1041-53
pubmed: 24728974
Stem Cells Transl Med. 2017 Mar;6(3):992-1005
pubmed: 28297566
J Bone Miner Metab. 2016 Nov;34(6):606-614
pubmed: 26369320
Mol Cell Proteomics. 2013 Apr;12(4):956-78
pubmed: 23345538
Proteomics. 2010 Feb;10(3):394-405
pubmed: 19953544
Nat Commun. 2017 Feb 09;8:14361
pubmed: 28181491
Science. 2014 Oct 3;346(6205):1248012
pubmed: 25278615
Am J Pathol. 2011 Sep;179(3):1074-80
pubmed: 21763673
Circ Res. 2009 Jan 2;104(1):32-40
pubmed: 19023133
Nat Med. 2007 Aug;13(8):952-61
pubmed: 17660828
Genes Dev. 2007 Oct 15;21(20):2644-58
pubmed: 17938244
Pflugers Arch. 2017 Aug;469(7-8):899-906
pubmed: 28685176
Bone. 2008 Sep;43(3):501-10
pubmed: 18571490
Am J Physiol Heart Circ Physiol. 2007 Jan;292(1):H285-94
pubmed: 16963618
J Am Soc Nephrol. 2016 Mar;27(3):781-90
pubmed: 26204899
Am J Physiol Heart Circ Physiol. 2018 Mar 1;314(3):H484-H496
pubmed: 29101181
Proc Natl Acad Sci U S A. 2015 Jul 7;112(27):8421-6
pubmed: 26109568
J Am Soc Nephrol. 2015 Apr;26(4):817-29
pubmed: 25535303
Lab Chip. 2013 Apr 21;13(8):1489-500
pubmed: 23440068
Biochem Biophys Res Commun. 2016 Sep 23;478(3):1074-9
pubmed: 27524235
Gene. 1999 Oct 1;238(2):301-13
pubmed: 10570958
Am J Pathol. 2012 Mar;180(3):973-83
pubmed: 22234173
Structure. 2011 Oct 12;19(10):1433-42
pubmed: 21944579
Cell Signal. 2015 Nov;27(11):2150-9
pubmed: 26278164
Circ Res. 2013 May 24;112(11):1433-43
pubmed: 23529184