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Organotypic Collagen I Assay: A Malleable Platform to Assess Cell Behaviour in a 3-Dimensional Context
Published on: October 13, 2011
Transcription factor Egr-1 activates collagen expression in immortalized fibroblasts or fibrosarcoma cells
Dorothea Alexander1, Martin Judex, Rotraud Meyringer
1Research Laboratories, Center for Orthopedic Surgery, University Hospital Tubingen, Pulvermühlstrasse 5, D-72070 Tübingen, Germany
Abstract:
Synovial fibroblasts from rheumatoid arthritis patients express elevated levels of the transcription factor Egr-1. The metabolic consequences of Egr-1 overexpression in fibroblasts are not known in detail. Therefore we searched for gene products that are differentially expressed in Egr-1(high) versus Egr-1(low) fibroblasts. Immortalized synovial fibroblasts were transfected with two different Egr-1 expression vectors. Expression of recombinant Egr-1 was confirmed by RT-PCR and immunoblots. Random arbitrarily primed PCR revealed that Egr-1 induces enhanced transcription levels of the alpha1 chain of type I collagen. Increased expression of the alpha2 (I) chain could also be observed. We found enhanced levels of type I collagen propeptide in supernatants and stronger signals of alpha2 (I) protein in extracts of the Egr-1(high) expressing clone versus controls. Additionally, Egr-1 was transiently expressed in fibrosarcoma cells. These cells showed a pronounced elevation of type I collagen (alpha1) transcripts as well. Moreover, we could demonstrate that Egr-1 induces transcription of other genes including type II collagen (alpha1) and plateled-derived growth factor beta1. These data suggest that upregulation of Egr-1 might contribute tofibrosis observed in rheumatoid arthritis synovium by activation of genes encoding the alpha1 and alpha2 chains of type I collagen.
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