October 31, 2004
American Society of Nephrology (ASN) Renal Week 2004, St. Louis, Missouri
Abstract SU-PO669
Poster: Neoplasia, Growth, Apoptosis, Injury Response: Tubule Epithelium and Renal Endothelium

Connective Tissue Growth Factor (CTGF) Activates Nuclear Factor-kB (NF-kB) in Tubulo Epithelial Cells. Andres F. Lopez, Vanesa Esteban, Elsa Sanchez-Lopez, Juan Rodriguez-Vita, Monica Ruperez, Noelynn Oliver, Leon Xu, Jesus Egido, Marta Ruiz-Ortega. Vascular and Renal Laboratory, Fundacion Jimenez Diaz. UAM., Madrid, MD, Spain; Fibrogen, CA.

The connective tissue growth factor (CTGF) has been described as a novel fibrotic mediator. CTGF is overexpressed in several kidney diseases and it is induced by different factors involved in renal injury. There are few investigations about the intracellular mechanisms involved in CTGF response. In kidney diseases elevated tissue activity of the nuclear factor-kB (NF-kB) has been observed. This transcription factor regulates many proinflammatory genes and products involved in proliferation/apoptosis. Our aim was to investigate whether CTGF could activate NF-kB pathway in tubulo epithelial cells (MCT cell line). In growth-arrested cells, recombinant CTGF (100 to 1 ng/mL) increased NF-kB DNA binding activity, as early as 15 minutes, decreasing after 2 hours. This response was dose-dependent, and maximal after 1 hour with 10 ng/mL CTGF (3-fold over control, n=6, p<0.05, gel mobility shift assays). This effect was similar to that observed with proinflammatory cytokines (TNF-alpha and AngII). By immunofluorescence we have located NF-kB subunits. In control cells a diffuse cytoplasmic fluorescence was seen with p65 antibody. Treatment with CTGF for 1 hour caused nuclear staining of p65, showing translocation of NF-kB to the nuclei. We have also investigated whether CTGF regulates NF-kB-mediated gene expression, by using a luciferase reporter plasmid (NF-kB/luc) that contains five copies of the recognition site for the NF-kB sequence. MCT cells were transiently transfected with NF-kB/luc and renilla as internal control. CTGF potently and significantly increased NF-kB promoter activity. This activation was similar to that observed with the cytokines IL-1-beta and TNF-alpha. Our data demonstrate that CTGF activates NF-kB pathway in tubuloepithelial cells. Future studies are needed to evaluate the biological significance of the activation of this transcription factor.