Randomized controlled trial of aspirin and clopidogrel versus aspirin and placebo on markers of smooth muscle proliferation before and after peripheral angioplasty.
OBJECTIVE
In peripheral arterial disease (PAD) patients, a limiting factor in the success of percutaneous transluminal angioplasty (PTA) is the development of restenosis secondary to vascular smooth muscle cell (SMC) proliferation. Following endothelial damage and platelet activation, there is release of factors and adhesion molecules which affect SMC proliferation. The aim of this study was to determine the effect of combination antiplatelet therapy (clopidogrel and aspirin compared with aspirin and placebo) on the ability of plasma from PAD patients undergoing PTA to stimulate SMCs in vitro. We further aimed to investigate the effect of combination treatment on the levels of circulating adhesion molecules and factors, which are known to mediate SMC proliferation in experimental models.


METHODS
Fifty patients were randomized to receive blinded clopidogrel or placebo, for thirty days, in addition to their daily 75 mg aspirin. To measure proliferative capacity, diluted plasma was incubated for 15 minutes with 24 hour-growth-arrested rat vascular smooth muscle cells, and extracellular regulated kinase (ERK)1/2 activation was analyzed by Western blotting at baseline, one hour pre-PTA, one hour, 24 hours and 30 days post-PTA. Plasma platelet-derived growth factor (PDGF), sE-selectin, intracellular adhesion molecule-1 (sICAM-1), and von Willebrand factor (vWF) were measured by ELISA, at the same five timepoints. Platelet activation was measured by flow cytometry of ADP-stimulated platelet fibrinogen binding at baseline and one hour post-PTA.


RESULTS
ADP-stimulated platelet fibrinogen binding was significantly inhibited by clopidogrel before and after PTA. ERK 1/2 activation was significantly increased post-PTA in both the aspirin/clopidogrel and aspirin/placebo groups (P < .001). There was a statistically significant decrease in PDGF (P = .004), and increase in vWF (P = .026), following loading with clopidogrel. sICAM-1 levels significantly decreased (P = .016) in the aspirin/placebo group following PTA. There were no other significant changes and also there was no statistically significant difference between the two treatment groups for each of ERK 1/2, sICAM-1, sE-selectin, or vWF.


CONCLUSIONS
This is the first study to show in-vitro ERK 1/2 activation (a surrogate marker of SMC proliferation) increases post-PTA. Combination antiplatelet therapy had no significant effect on this, although it did reduce PDGF. Further work is required to evaluate potential therapeutic treatments, which may reduce peripheral PTA-induced smooth muscle cell activation.


CONCLUSIONS
High rates of restenosis remain the major limitation of peripheral arterial angioplasty and stenting.The restenotic lesion occurs secondary to platelet activation, released circulating factors, and subsequent smooth musclecell proliferation and migration into the intima. Methods to limit the restenotic lesion are poorly understood. This paperinvestigates the effect of PTA on smooth muscle cell activation and the release of factors in plasma which mediate SMCproliferation. It also examines the effect of combination antiplatelet therapy as a potential therapeutic strategy.
Wilson
Alasdair M
AM
Department of Surgery, University of Aberdeen, Aberdeen, United Kingdom. awilson78@doctors.net.uk
Brittenden
Julie
J
Bachoo
Paul
P
Ford
Isobel
I
Nixon
Graeme F
GF
D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D000800:Angioplasty, Balloon / Q000379:methods; D001241:Aspirin / Q000008:administration & dosage*; D015415:Biomarkers / Q000097:blood*; D015153:Blotting, Western; D049109:Cell Proliferation / Q000187:drug effects*; D002478:Cells, Cultured; D000077144:Clopidogrel; D004305:Dose-Response Relationship, Drug; D004311:Double-Blind Method; D004334:Drug Administration Schedule; D019040:E-Selectin / Q000032:analysis; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D005434:Flow Cytometry; D006083:Graft Occlusion, Vascular / Q000517:prevention & control; D006801:Humans; D018799:Intercellular Adhesion Molecule-1 / Q000032:analysis; D008297:Male; D008875:Middle Aged; D048052:Mitogen-Activated Protein Kinase 3 / Q000032:analysis; D015999:Multivariate Analysis; D009131:Muscle, Smooth, Vascular / Q000187:drug effects; D016491:Peripheral Vascular Diseases / Q000097:blood / Q000000981:diagnostic imaging / Q000188:drug therapy* / Q000628:therapy; D010975:Platelet Aggregation Inhibitors / Q000008:administration & dosage; D010982:Platelet-Derived Growth Factor / Q000032:analysis; D011336:Probability; D011859:Radiography; D012016:Reference Values; D018709:Statistics, Nonparametric; D013988:Ticlopidine / Q000008:administration & dosage / Q000031:analogs & derivatives*
D015415:Biomarkers; D019040:E-Selectin; D010975:Platelet Aggregation Inhibitors; D010982:Platelet-Derived Growth Factor; D018799:Intercellular Adhesion Molecule-1; D000077144:Clopidogrel; D048052:Mitogen-Activated Protein Kinase 3; D013988:Ticlopidine; D001241:Aspirin