title: Basic fibroblast growth factor stimulates the release of preformed transforming growth factor beta 1 from human proximal tubular cells in the absence of de novo gene transcription or mRNA translation.
abstract: Interstitial fibrosis is significantly correlated with the progression of renal impairment for most causes of renal insufficiency. Transforming growth factor beta 1 (TGF-beta 1) and basic fibroblast growth factor (bFGF) are two of a group of profibrotic cytokines that have been associated with the development of renal interstitial fibrosis. We have previously demonstrated that alterations in D-glucose concentrations modulate the synthesis of TGF-beta 1 by human renal proximal tubular cells (HPTC) in vitro. The aim of the present study was to examine the influence of bFGF on TGF-beta 1 synthesis by HPTC in culture and to examine any modulation of this response by changes in ambient glucose concentration. Incubation of growth-arrested HPTC (72 hours in serum-free medium) with bFGF resulted in a dose-dependent increase in latent TGF-beta 1 secretion. Maximal release of TGF-beta 1 was seen at a bFGF dose of 50 ng/ml in cells incubated in 5 mM D-glucose (7.48 +/- 2.5 ng/ml, mean +/- SEM; n = 3; p = 0.04). This release of TGF-beta 1 in response to bFGF was unaffected by increasing the concentration of glucose in the culture media to 25 mM (7.76 +/- 1.3, mean +/- SEM; n = 3; p < 0.02). It was also unaffected by pretreatment of cells with either actinomycin-D or cycloheximide. TGF-beta 1 secretion was, however, inhibited in a dose-dependent manner by the exposure of cells to the microtubule-disrupting agent vinblastine, indicating that the generation of TGF-beta 1 was dependent on the secretion of preformed, stored TGF-beta 1. In a separate series of experiments, exposure of HPTC to TGF-beta 1 (10 ng/ml) led to the induction of bFGF mRNA, which was first apparent at 12 hours and reached maximal levels 24 hours after stimulation (normalized bFGF/alpha-actin mRNA ratio was 1.5 times that of the control). This increase in bFGF mRNA was accompanied by a time-dependent increase in bFGF protein production, which was maximal after 24 hours (19.83 +/- 12.7 pg/ml versus 2.49 +/- 0.34 pg/ml, mean +/- SEM, stimulated versus control; n = 3; p = 0.03). These findings demonstrate that bFGF stimulates the secretion of preformed, latent TGF-beta 1 by HPTC but does not induce de novo TGF-beta 1 gene transcription or TGF-beta 1 protein synthesis. We have also demonstrated a positive-feedback loop involving TGF-beta 1 and bFGF and postulate that this may be involved in the progressive nature of renal fibrosis in vivo.
authors.affiliation: Institute of Nephrology, University of Wales College of Medicine, Cardiff Royal Infirmary, Wales, United Kingdom.
authors.forename: A O
authors.initials: AO
authors.lastname: Phillips
authors.forename: N
authors.initials: N
authors.lastname: Topley
authors.forename: K
authors.initials: K
authors.lastname: Morrisey
authors.forename: J D
authors.initials: JD
authors.lastname: Williams
authors.forename: R
authors.initials: R
authors.lastname: Steadman
mesh_terms: D002478:Cells, Cultured; D046508:Culture Techniques; D017931:DNA Primers / Q000737:chemistry; D004305:Dose-Response Relationship, Drug; D005109:Extracellular Matrix / Q000378:metabolism; D016222:Fibroblast Growth Factor 2 / Q000235:genetics / Q000494:pharmacology*; D006801:Humans; D007687:Kidney Tubules, Proximal / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D008870:Microtubules / Q000378:metabolism; D014176:Protein Biosynthesis / Q000502:physiology*; D012333:RNA, Messenger / Q000235:genetics*; D014158:Transcription, Genetic / Q000502:physiology*; D016212:Transforming Growth Factor beta / Q000096:biosynthesis* / Q000235:genetics
chemical_list: D017931:DNA Primers; D012333:RNA, Messenger; D016212:Transforming Growth Factor beta; D016222:Fibroblast Growth Factor 2