Profiles of von Willebrand factor antigen, von Willebrand factor propeptide and ADAMTS13 activity in patients with carotid stenosis and their relationship with cerebral micro-embolic signal status

Abstract

Aims: The aims of this thesis were to comprehensively investigate the profiles of von Willebrand Factor (VWF:Ag), von Willebrand Factor propeptide (VWFpp) and A Disintegrin And Metalloprotease with ThromboSpondin type 1 motif number 13 (ADAMTS13)activity in patients with asymptomatic and symptomatic carotid artery stenosis and their relationship with cerebral micro-embolic signals (MES) detected on Transcranial Doppler ultrasound(TCD). Methods:We performed an observational analytical case-case/control study to assess levels of VWF:AgandVWFpp, VWFpp/VWF:Ag ratio and ADAMTS13activity in patients with moderate or severe (≥50-99%) asymptomatic vs. early(≤4 weeks) and latephase symptomatic(≥3 months after TIA or ischaemic stroke) carotid stenosis,with nested longitudinal follow-up studiesin the symptomatic cohort. VWF:Ag and VWFpp levels were quantified with an enzyme-linked immunosorbent assay(ELISA)on thawed,double-spun platelet poor plasma. ADATMS13 activity was assessed with a Fluorescence Resonance Energy Transfer(FRET) assay in non-double spun platelet poor plasma. One hour, bilateral TCD of the middle cerebral arteries was performed in as many patients as possible to classify patients asMES+ve or MES-ve. Results: Data from 34 asymptomatic patients were compared with 43 symptomatic patients in the ‘early phase’ (≤4 weeks) and 37 patients in the ‘late phase’ (≥3 months) after TIA/ischaemic stroke. VWF:Ag levels were significantly higher in early phase symptomatic than asymptomatic patients (12.53 vs. 10.57μg/mL; P = 0.049), but were similar in the late symptomatic and ‘late symptomatic post-intervention’ groups to those seen in the asymptomatic group(P ≥ 0.66). There was no difference in VWFpp expression between groups (P ≥ 0.15). However, the VWFpp/VWF:Ag ratio was significantly lower in early symptomatic (0.094 vs. 0.118; P = 0.006), but not in late symptomatic or late symptomatic post-intervention patients compared with the asymptomatic cohort (P ≥ 0.37). After controlling for differences in age and the proportion of active smokers at the time of enrolment between groups, VWF:Ag levels were still significantly higher (P = 0.018) and the VWFpp/VWF:Ag ratio remained lower (P = 0.015) in early symptomatic vs. asymptomatic patients; however, there was still no significant difference in VWFpp expression between groups (P = 0.93). There were no significant differences in ADAMTS13 activity between early symptomatic (134.71%; P = 0.79) or late symptomatic (127.84%; P = 0.33) and asymptomatic patients (132.74%). In symptomatic patients followed up longitudinally from the early to the late phase, there was a significant reduction in VWF:Ag levels (11.36 →9.05 μg/mL; P = 0.048), a significant increase in the VWFpp/VWF:Ag ratio (0.095 →0.109; P = 0.032), and a significant reduction in ADAMTS13 activity (144.1% →129.1%; P = 0.015),but no differences in VWFpp levels (P = 0.52)over time. There was a significant positive correlation between VWF:Ag and VWFpp levels in the early symptomatic (R2= 40.7%; P < 0.0001), late symptomatic (R2= 41.3%; P < 0.001) and asymptomatic cohorts (R2= 26.5%; P = 0.002). There were no significant correlations between ADAMTS13 activity and VWF:Ag levels in symptomatic or asymptomatic patients (R2≤ 2.3%; P ≥ 0.35). In the overall cohort of early symptomatic and asymptomatic patients, patients with bloodgroup O had significantly lower median VWF:Ag levels (9.59μg/mL) compared with ‘non-bloodgroup O patients’(12.32μg/mL; P = 0.035). However,there were no significant differences in VWFpp levels,the VWF:Ag/VWFpp ratio or ADAMTS13 activity between blood-group O and non-bloodgroup O patients (P > 0.05).As reported previously, the proportion of patients who were MES+ve was higher in early symptomatic (28.5%, N = 10, P = 0.049) than in asymptomatic patients (7.1%, N = 2), but had fallen to similar levels to the asymptomatic group by the late phase after symptom onset (6.7%, N = 2; P = 0.996). Early symptomatic MES-ve patients had significantly higher VWF:Ag levels (12.72 vs. 10.12 μg/mL; P = 0.023) and a lower VWFpp/VWF:Ag ratio (0.094 vs. 0.115; P = 0.008) than asymptomatic MES-ve patients. There were no differences in VWFpp levels or ADAMTS13 activity in MES-ve early symptomatic vs. asymptomatic patients (P ≥ 0.05). Conclusions: VWF:Ag expression, a marker of endothelial and/or platelet activation, is enhanced in recently symptomatic compared with asymptomatic 50-99% carotid stenosis patients, including in the subset ofMES-vepatients, and decreases along with ADAMTS13 activity over time following atherosclerotic TIA/ischaemic stroke. VWF:Ag levels and the VWFpp/VWF:Ag ratio may represent more sensitive biomarkers of endothelial activation in symptomatic carotid stenosis than VWFpp levels alone. MES were more common in early symptomatic than asymptomatic carotid stenosis patients and reduced over time with successful interventional and optimal medical treatment. Combining clinical, endothelial and platelet biomarker data with data on MES status improves our understanding of the pathogenesis of cerebrovascular events and could assist in risk-stratification in this important population of cerebrovascular disease patients, including in the MES-ve carotid stenosis subgroup. Such comprehensive profiling has the potential to target patients who might benefit from earlier, more intensive medical or interventional treatment to prevent subsequent TIA or stroke.