Differential effects of transforming growth factor-beta on the synthesis of connective tissue growth factor and vascular endothelial growth factor by peritoneal mesothelial cell.
BACKGROUND
Previous studies found that transforming growth factor-beta (TGF-beta) plays a conflicting role in peritoneal fibrosis. We hypothesise that TGF-beta acts on peritoneal mesothelial cells (PMC) via VEGF and CTGF as downstream mediators.


METHODS
The effect of TGF-beta in primary culture of rat PMC was studied. VEGF and CTGF mRNA expression was examined by real time quantitative polymerase chain reaction (RT-QPCR), and VEGF antigen level in cell supernatant by ELISA.


RESULTS
Incubation of rat PMC with TGF-beta resulted in a time- (3-72 h) and concentration- (0-50 pg/ml) dependent increase in VEGF mRNA expression, and VEGF protein level in the cell supernatant. When stimulated with TGF-beta 100 pg/ml, there was a 20-fold up-regulation of VEGF mRNA expression (p < 0.001). The CTGF mRNA expression and protein level of PMC was slightly increased at low concentration of TGF-beta (50 pg/ml) but decreased at a higher concentration (100 pg/ml or above). The effect of TGF-beta on PMC CTGF, but not VEGF, gene expression was inhibited by Smad decoy oligodeoxynucleotide. The effect of TGF-beta on PMC VEGF gene expression and protein synthesis was inhibited by PD98059 (a specific MAP kinase inhibitor) and chelerythrine (a specific protein kinase C inhibitor), but not cholera toxin (activator of cyclic AMP) or herbimycin A (inhibitor of protein tyrosine kinase). The up-regulation of CTGF mRNA expression was inhibited by PD98059, but not chelerythrine, cholera toxin or herbimycin A. Furthermore, CTGF gene expression in TGF-beta-stimulated PMC was inhibited by co-administration of recombinant VEGF.


CONCLUSIONS
Our data demonstrate that TGF-beta induces PMC production of VEGF and CTGF via different signalling pathways. At high concentration of TGF-beta, VEGF production predominates and CTGF production was inhibited. Since CTGF and VEGF have different biologic effects, our results may explain the complex activity of TGF-beta in peritoneal physiology.
Szeto
Cheuk-Chun
CC
Department of Medicine and Therapeutics, Prince of Wales Hospital, The Chinese University of Hong Kong, Shatin, Hong Kong, China. ccszeto@cuhk.edu.hk
Lai
Ka-Bik
KB
Chow
Kai-Ming
KM
Szeto
Carol Yi-Ki
CY
Wong
Teresa Yuk-Hwa
TY
Li
Philip Kam-Tao
PK
D000818:Animals; D015153:Blotting, Western; D002478:Cells, Cultured; D055513:Connective Tissue Growth Factor; D004305:Dose-Response Relationship, Drug; D004347:Drug Interactions; D004797:Enzyme-Linked Immunosorbent Assay; D004847:Epithelial Cells / Q000187:drug effects / Q000378:metabolism; D017874:Immediate-Early Proteins / Q000037:antagonists & inhibitors / Q000096:biosynthesis*; D036341:Intercellular Signaling Peptides and Proteins / Q000096:biosynthesis*; D010537:Peritoneum / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D015398:Signal Transduction / Q000502:physiology; D051785:Smad Proteins / Q000502:physiology; D016212:Transforming Growth Factor beta / Q000008:administration & dosage / Q000494:pharmacology*; D015854:Up-Regulation / Q000502:physiology; D042461:Vascular Endothelial Growth Factor A / Q000096:biosynthesis*
C526316:CCN2 protein, rat; D017874:Immediate-Early Proteins; D036341:Intercellular Signaling Peptides and Proteins; D051785:Smad Proteins; D016212:Transforming Growth Factor beta; D042461:Vascular Endothelial Growth Factor A; D055513:Connective Tissue Growth Factor