Transforming growth factor beta mediates the angiotensin-II-induced stimulation of collagen type IV synthesis in cultured murine proximal tubular cells

Wolf, G.; Zahner, G.; Schroeder, R.; Stahl, R.A.

Nephrology Dialysis Transplantation Official Publication of the European Dialysis and Transplant Association - European Renal Association 11(2): 263-269


ISSN/ISBN: 0931-0509
PMID: 8671777
DOI: 10.1093/oxfordjournals.ndt.a027251
Accession: 009664678

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Background: Angiotensin II (Ang II) stimulates synthesis of type IV collagen in a cultured murine proximal tubular cell line (MCT cells). In addition, Ang II also induces the expression of TGF-beta-1 in these cells. Since TGF-beta has well-known stimulatory effects on the transcription of various collagens, we tested whether the Ang-II-mediated stimulation of type IV collagen is due to induction of endogenous TGF-beta-1 synthesis in MCT cells. Results: A neutralizing monoclonal anti-TGF-beta-1-3 antibody abolished the Ang II-stimulated release of type IV collagen in culture supernatants. The anti-TGF-beta-1-3 antibody also partly blocked Ang-II-mediated incorporation of 3(H)proline into de novo synthesized collagens. Moreover, 5 mu-M TGF-beta-1 antisense oligonucleotides, but not the same concentration of sense oligonucleotides, completely blocked Ang-II-stimulated 3(H)proline incorporation. MCT cells incubated with TGF-beta-1 antisense phosphorothioate-modified oligonucleotides failed to synthesize TGF-beta-1 protein after Ang II treatment as measured by a sandwich ELISA in culture supernatants. SDS-polyacrylamide electrophoresis of 3(H)proline-labelled collagens and comparison with standard collagens also demonstrated that the neutralizing anti-TGF-beta-1-3 antibody abolished the Ang-II-mediated stimulation in type IV collagen. Semiquantitative cDNA amplification for collagen type alpha-1 (IV) transcripts revealed that the anti-TGF-beta-1-3 antibody abrogates the increase in mRNA after Ang II treatment. Transient transfection studies in MCT cells using murine collagen alpha-1 (IV) enhancer/promoter constructs also demonstrated the suppressive effect of the neutralizing antibody on Ang-II-stimulated gene transcription. Conclusions: Our data collectively suggest that the Ang-II-mediated increase in type IV collagen in MCT cells is mediated by endogenous synthesis and autocrine action of TGF-beta-1. These findings may be important in changes of the tubulointerstitial architecture during the progression of renal disease.