Major clinical problems caused by unchecked vascular smooth muscle (VSM) proliferation and migration (intimal hyperplasia) can occur following surgical by-pass leading to vein-graft failure or restenosis following arterial angioplasty. Several agents present in serum have been shown to stimulate mitogenesis via dissociated heterotrimeric G proteins (Gq and Gi) appear to be critically involved in the pathological process of intimal hyperplasia since several serum mitogens, such as angiotensin II and thrombin, can activate cellular receptors that can couple to Gq and/or Gi. Peptide inhibitors targeted specifically against Galphaq (GqI) and Gbetagamma (BetaARKct) signaling have recently been developed and thus, molecular inhibitors of Galphaq and Gbetagamma-mediated signaling may represent a novel therapeutic approach to limit vein-graft failure and arterial restenosis after angioplasty. The Overall Hypothesis of this proposal is that G proteins are a critical regulator of VSM cellular proliferation and migration, especially in injured vessels. Furthermore, the targeted inhibition of intracellular G protein signaling, including Gbetagamma inhibition, will lead to a broader understanding of the molecular mechanisms involved in these processes, which may lead to novel molecular therapies to limit the pathological process of intimal hyperplasia. Several in vitro and in vivo model systems are available to test this hypothesis including a rabbit vein-graft model and a porcine coronary artery angioplasty and stent restenosis model. To delivery the GqI and betaArkct peptides to VSM cells in culture and in vivo to the vascular wall, we will utilize adenoviral vectors. The associated Specific Aims are: (1) To identify the specific role of Galphaq and Gbetagamma in the activation of cellular migration and proliferation in VSM cells using in vitro model systems; (2) To determine if in vivo inhibition of Galphaq or Gbetagamma signaling in VSM via adenoviral-mediated gene transfer will limit intimal hyperplasia during vein-graft failure; (3) To determine if adenoviral-mediated delivery of the GqI and betaArkct transgenes to the arterial wall will prevent restenosis is clinically relevant animal models; (4) To evaluate gene transfer to VSM cells within the vessel wall using """"""""advanced"""""""" adenovirus and AAV vectors in attempts to more efficiently deliver the peptide inhibitors of Galphaq and Gbetagamma.
