Gene Delivery of OP-1 in Rodent Model of Renal Fibrosis

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This application for Ro3 award is submitted as a supplement for the K08 award DK02546-02. Osteogenic Protein-1 (OP- 1) or Bone Morphogenetic Protein-7) (BMP-7) is a member of the transforming growth factor beta (TGF-beta) superfamily of secreted growth factors. The kidney is the primary site of OP-1 synthesis. Mice genetically deficient in OP-1 expression have markedly abnormal renal development, suggesting that this protein may be important in regulation of cell division and morphogenesis in the kidney. Furthermore, systematic administration of OP-1 has been shown to delay or halt progress to end stage renal failure in the 5/6 nephrectomy of renal fibrosis. To further explore this phenomenon, we evaluated the effects of OP-1 on the extracellular matrix in an in vitro model of renal fibrosis. Increasing concentrations of OP-1 0-300 ng/ml) resulted in a dose dependent increase in fibronectin levels in mouse messangial and proximal tubular cells. In duplicate samples processed for RNA revealed no significant changes in fibronectin RNA levels, suggesting a non- transcriptional mechanism. Concurrently, our laboratory has demonstrated that fibronectin is a target protein for ubiquitin dependent degradation. The research goals for the final two years of the proposal will focus on the observation that OP-1 decreases ubiquitin dependent degradation of fibronectin. These goals will be realized in the following ways. First we will determine id the effects of OP-1 are specific for fibronectin or if it has similar effects on other extracellular matrix proteins such as collagen I, III and IV. Second we will investigate the potential involvement of OP-1 on ubiquitination of fibronectin. Finally, utilizing confocal microscopy we will evaluate the effects of OP-1 on the subcellular distribution of fibronectin. This finding presented in this presented in this proposal represents a new paradigm in ECM homeostasis. Understanding the normal mechanism of action of this pathway will not only elucidate a novel mechanism of extracellular matrix homeostasis but may have major therapeutic implications.