Assessment of targeted therapies for inhibition of vascular smooth muscle cell invasion and proliferation

Cardiovascular surgical procedures, such as angioplasty, stenting and vascular grafts, have a high incidence of intimal hyperplasia, which often leads to restenosis or occlusion of the artery. Intimal hyperplasia is characterized by a hyperplastic response of arterial smooth muscle cells (SMCs). In a non-pathologic response, the SMCs heal a damaged artery wall and return to a quiescent state. In small diameter arteries and grafts (ID < 6 mm), this response tends to be pathologic when the SMCs proliferate, invade to the arterial lumen and deposit extracellular matrix (ECM) proteins in excess (hyperplasia) leading to reduced flow or occlusion.; Many proposed therapies targeting different mechanisms of SMC behavior are being studied, though few studies focus on quantitatively comparing and combining therapies to engineer a clinical solution. This study focuses on choosing which therapy or combination provides a superior solution to SMC hyperplastic behavior. The three potential therapies examined in the study are: tissue inhibitor of metalloproteinase 1 (TIMP-1) in solution, cyclic-RGD peptide fixed and in solution and over expression of hyperfunctional α _vβ₃ integrins (Hyp-int) using a gene therapy delivery vehicle. The experiments were separated into two main groups: (a) 2-D proliferation and spreading and (b) 1-D invasion behavior.; SMC behavior is affected by adhesion strength between the cell-surface vs. cell-cell interactions, cell phenotype, integrin expression, and time. Surface fixed cyclic-GpenGRGDSPCA (C-RGD) was compared with linear-GRGDSP (L-RGD), tissue culture polystyrene, and a laminin nonapeptide CAPGYIGSR. A dynamic flow environmental chamber system (DFLECTS) was designed to attempt a critical shear stress measurement, but the study revealed higher than expected SMC adhesion strength. Normal SMC behavior was compared with cells expressing Hyp-β₃ with optical fluorescent microscopy and custom image analysis programs.; Invasive behavior is not only affected by the factors involved in surface behavior, but also by ECM density and composition, proteinase activity and chemoattractor concentration gradients. All three potential therapies were quantitatively compared using an optimally designed fluorescent invasion assay to test for SMC invasion using platelet derived growth factor-BB (PDGF-BB) as a potent chemoattractor. TIMP-1/C-RGD and TIMP-1/Hyp-β ₃ combinations were also analyzed for synergistic, constructive or destructive interactions using general factorial design models.