Wound healing requires the formation of new vasculature in a complex process that is dependent on stabilization and destabilization signals. The endothelial cell-specific receptor tyrosine kinase, Tie2, when stimulated by its primary ligand, angiopoietin-1 (Ang1), promotes blood vessel stability by stimulating cell viability, basement membrane integrity, and perivascular cell recruitment. Ang2, an antagonist of the Tie2 receptor, blocks these signals by competing for Ang1 binding, and is necessary for the locaiized vessel instability associated with angiogenesis. Thus, the ratio of Ang1 to Ang2 is a critical factor in the regulation of neovascularization; altering Ang1 or Ang2 expression can lead to profound vascular defects. This study focuses on evidence that Ang2 expression is regulated post-transcriptionally; endothelial and smooth muscle cell culture models demonstrate profound changes in Ang2 mRNA half-life in response to certain growth factors. This induced change in message stability is dependent on new transcription, presumably of factors associated with regulated mRNA turnover. It is hypothesized that such post-traiascriptional regulation is critical for the maintenance of appropriate Ang2 levels during tissue repair. To test this, the putative mRNA control elements within the human, mouse, and rat Ang2 MRNAs that regulate induced message stability/instability or translation will be identified and characterized. Their role in regulating Ang2 expression will then be tested in vivo by introducing these elements into reporter constructs; which will then be used to develop transgenic mice. The biological roles of these mRNA control elements will be further analyzed in mice in which these elements are ablated by conditional homologous recombination. These mouse models will be used to test the effects of dysfunctional Ang2 post-transcriptional regulation on wound healing. These studies will indicate the relative importance of post-transcriptional regulation of Ang2, and whether eliminating the ability of cells to regulate Ang2 mRNA stability or translation impacts vessel structure during the wound healing process.

R01GM063638

2002-06-01

2008-05-31