Determining how heparin interferes with the MAPK/ERK pathway: An attempt to understand the atherosclerotic process.

摘要

Necessary events for the development of atherosclerosis are the migration and subsequent proliferation of vascular smooth muscle cells (VSMCs) to the sites of vascular endothelial cell injury. The antiproliferative effects of heparin on VSMCS have been described for over thirty years, and still the mechanism(s) of heparin's effects remains poorly understood. It is most likely that heparin exerts its effects through the interaction of a cell surface receptor that at least in part functions to inhibit mitogenic growth factor signaling events. Previously, this laboratory identified a protein with an approximate molecular weight of 45 kDa that bound to heparin on the cell surface of both vascular endothelial and smooth muscle cells. Antibodies against this receptor protein mimic the effects of heparin on VSMCs. Both heparin and anti-receptor antibodies have been observed and reported to decrease DNA synthesis stimulated by platelet-derived growth factor (PDGF) and mitogen activated protein kinase (MAPK). In this report, both heparin and these anti-receptor antibodies induced the expression of mitogen activated protein kinase phosphatase 1 (MKP-1) and subsequent decrease in MAPK activity in VSMCs. Blocking the synthesis of MKP-1 blocked both heparin's and the anti-receptor antibodies' ability to decrease MAPK activity in VSMCs. The findings support a role for the heparin receptor protein in mediating heparin's effects on VSMCs. Herein we suggest a potential candidate protein as the putative heparin receptor namely transmembrane protein 184A, based on peptide mapping and sequencing analysis with matrix-assisted laser desorption ionization -- time of flight mass spectrometry (MALDI-TOF MS). The suggested protein has characteristics similar to the putative heparin receptors and little is known about the function of the protein. Previously, this laboratory suggested that the second messenger, cyclic guanidine monophosphate (cGMP), and cGMP-dependent protein kinase (PKG) are involved in the mediation of heparin effects. Decreasing the levels of PKG resulted in short term inhibition of heparin's ability to induce MKP-1 and decrease MAPK activity. These results support the hypothesis that PKG is involved in mediating heparin's effects on VSMCs. Heparin was found to have effects upstream of the MAPK pathway, as heparin altered phosphorylation levels on Raf-1 resulting in the overall decrease in Raf-1 activity. However, inhibiting the activity levels of PKG did not diminish heparin's ability to decrease Raf-1 activity. These findings suggest that heparin mediates its effects on VSMCs through a mechanism that involves cGMP and PKG-dependent induction of MKP-1 and PKG-independent decreases of Raf-1 activity.