The overgrowth of vascular smooth muscle cells is a hallmark of the late stages of vascular disease and contributes to the eventual blocking of affected arteries. Heparin has been shown to slow smooth muscle cell growth in vitro and in selected in vivo studies. Heparin effects on vascular smooth muscle cells have been explained by theories including heparin receptors, heparin interaction with growth factors and endocytosis of heparin. Data from the P.I.'s laboratory implicate a heparin receptor and signal transduction involving cGMP production and specific MAPK phosphatases. The proposed research aims to test the hypothesis that heparin treatment of cultured heparin-sensitive vascular smooth muscle cells will result in decreased activity of upstream signal steps in the MAPK pathway and decreased SAPK pathway activity through heparin's interaction with the heparin receptor previously identified in the P.l.'s laboratory. The proposed research will evaluate heparin and heparin receptor effects on upstream signaling from the PDGF receptor through SOS, Ras, and Raf using heparin, anti-heparin receptor antibodies, cGMP analogs, and PKG blockers. Studies of heparin effects on MAPK activity throughout the cell cycle will evaluate whether additional MAPK activity seen throughout the cycle is decreased, or whether the heparin effects are only at the Gi release from GO check point. The proposed research will evaluate signaling in response to Angiotensin II activation to determine whether the heparin effects on smooth muscle cell MAPK signal transduction are limited to MAPK activation in response to growth factors, or can modulate MAPK signaling regardless of activation system. These studies will also include evaluation of stress kinase activity in heparin treated smooth muscle cells to examine whether these parallel pathways are altered by heparin as might be expected based on our information about the mechanism by which MAPK activity is decreased. Finally, this study will continue work aimed at identifying a clone for the heparin receptor to enable further studies regarding the mechanism of heparin action. Together these studies will confirm the importance of the heparin receptor in heparin induced changes in vascular smooth muscle cells and will provide a significant advancement in understanding of how heparin alters sensitive vascular smooth muscle cells.
