) Integrin adhesion receptors mediate the interactions between endothelial cells and the extracellular matrix and thus are key determinants of angiogenesis. Modification of certain integrins, in particular, the endothelial cell victronectin receptions alpha-v-beta-3 and alpha-v-beta-5, has been shown to be successful in preventing angiogenesis. These integrin receptors can convey signals from the extracellular matrix to the cell interior; conversely, intracellular signal transduction can influence the affinity of integrins for their ligands. The two integrin receptors for victronectin appear to mediate separate pathways to angiogenesis, which may be regulate separately. Our laboratory has recently identified a new 20 kilodalton intracellular mediator, which we have designated as TAP20 this mediator is regulated by PKCO and modulates the function of the alpha-v-beta-5 integrin. Preliminary data presented in this proposal shows that overexpression of TAP20 in endothelial cells enhances their ability to migrate and form tubules in three dimensional cell culture. The primary focus of this proposal is to investigate whether TAP20 is an important mediator of pathological angiogenesis and is useful in therapeutic angiogenesis in ischemic cardiovascular disease. The molecular mechanism by which TAP20 mediate signal transduction and how that may affect endothelial cell migration and tube formation will be determined.
The specific aims for this proposal are first, to test the hypothesis the TAP20 enhances endothelial cell migration and tubule formation by altering release of known autologous agonist or antagonist and/or by affecting intercellular mediators resulting from """"""""outside in"""""""" integrin signaling. Whether endothelial cells that overexpress TAP20 release increased amounts of FGF2, VEGF or IL8 and whether the release of antiangiogenic substances such as thrombospondin 2 or TGF-beta is suppressed will be determined. We also will determine whether TAP20 affects cellular migration and tube formation by signaling events associated with alpha-v-beta-5 integrin function, such as activation of ERK, GTPase in the Rho family, Src kinase activity, and focal adhesion formation and kinase (FAK) activity. Antisense for TAP20 to prevent those events will be used whether TAP20 enhances migration specifically in endothelial cells by overexpressing it in non endothelial cells that express alpha-v-beta-5 and in non alpha-v-beta-5 containing cells will be determined. An antibody raised against TAP20 will be used in immunohistochemistry to determine whether the expression of TAP20 in a variety of angiogenic tissues is increased over that in similar tissues in which angiogenesis is not prominent. These tissues include various cancers and ischemic myocardium. Finally whether TAP20 enhances angiogenesis in vivo will be determined by utilizing a corneal model of angiogenesis with endothelial cell that overexpress TAP20 inserted and to inject the TAP20 adenovirus into ischemic rat myocardium to determine whether it enhances angiogenesis following ischemia. We anticipate that these studies should provide data that will be useful in designing new strategic for pharmacologic intervention in ischemic vascular disease and cancer.
| Wang, Aihong; Nomura, Motohiro; Patan, Sybill et al. (2002) Inhibition of protein kinase Calpha prevents endothelial cell migration and vascular tube formation in vitro and myocardial neovascularization in vivo. Circ Res 90:609-16 |
