Soluble fibronectin induces chemokine gene expression in renal tubular epithelial cells.
BACKGROUND
Increasing proteinuria in kidney disease is associated with an increased risk of renal failure. Urinary proteins such as albumin induce inflammatory signaling and gene expression in tubular epithelial cells (TECs). Fibronectin is an extracellular matrix protein that can exist in soluble form and is excreted in the urine of patients with glomerular disease.


METHODS
To explore the impact of soluble fibronectin on tubular epithelium, murine TECs were stimulated with soluble fibronectin and chemokine mRNA was determined by RNase protection assay.


RESULTS
Fibronectin induced the expression of inflammatory chemokine genes, including monocyte chemoattractant protein-1 (MCP-1) (CCL2) and macrophage inflammatory protein-2 (MIP-2) within 2 hours in a dose-dependent manner. Phosphorylation of Src family tyrosine kinases was also increased in TECs following exposure to fibronectin. Src tyrosine kinases were involved in the fibronectin activation of MCP-1 since the Src inhibitors SU6656 and PP2 effectively reduced the induction of this chemokine. Fibronectin also induced the phosphorylation of extracellular signal-regulated protein kinase (ERK1/2) within minutes in TECs. The ERK kinase (MEK1/2) inhibitor U0126 inhibited the fibronectin induction of MCP-1 mRNA suggesting that ERK1/2 was also involved in this inflammatory pathway. Furthermore, fibronectin also induced phosphorylation of IkappaBalpha within 20 minutes in TECs. The nuclear factor-kappaB (NF-kappaB) inhibitors N-acetyl-L-cysteine (NAC) and pyrrolidinecarbodithioic acid (PDTC) effectively blocked fibronectin induction of MCP-1 mRNA.


CONCLUSIONS
Soluble fibronectin activates MCP-1 gene expression in TECs via Src tyrosine kinases, ERK1/2 and NF-kappaB. These data provide further support to the concept that proteinuria per se contributes to the tubulointerstitial injury observed in glomerular disease.
Ren
Li
L
Division of Nephrology, Department of Medicine, University of Calgary, Alberta, Canada.
Blanchette
Jason B
JB
White
Lindsay R
LR
Clark
Sharon A
SA
Heffner
Daniel J
DJ
Tibbles
Lee Anne
LA
Muruve
Daniel A
DA
D000818:Animals; D002478:Cells, Cultured; D018932:Chemokine CCL2 / Q000235:genetics; D054426:Chemokine CXCL2; D018925:Chemokines / Q000235:genetics*; D004847:Epithelial Cells / Q000166:cytology / Q000187:drug effects* / Q000276:immunology; D005353:Fibronectins / Q000494:pharmacology*; D015870:Gene Expression / Q000187:drug effects / Q000276:immunology; D007684:Kidney Tubules / Q000166:cytology*; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism; D016328:NF-kappa B / Q000378:metabolism; D012995:Solubility; D019061:src-Family Kinases / Q000378:metabolism
C491768:Ccl2 protein, mouse; D018932:Chemokine CCL2; D054426:Chemokine CXCL2; D018925:Chemokines; C404915:Cxcl2 protein, mouse; D005353:Fibronectins; D016328:NF-kappa B; D019061:src-Family Kinases; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3