Supplementary Material for: Determinants of Platelet-Leukocyte Aggregation and Platelet Activation in Stroke

<b><i>Background:</i></b> Platelet-leukocyte aggregation (PLA) and platelet activation are found to be on the higher side in ischemic stroke patients. The correlation of PLA with clinical features has not been intensively investigated and the influence of genetic factors on PLA is still unexplored. The interaction of platelets with leukocytes is mainly determined by the proteins encoded by six genes: P-Selectin (<i>SELP </i>encodes CD62P) on the thrombocyte binding to P-Selectin-Glycoprotein-Ligand-1 <i>(PSGL1)</i> on the leukocyte, intracellular-adhesion-molecule 2 <i>(ICAM2)</i> interacting with Integrin alpha M (<i>ITGAM</i>) and Glycoprotein 1b-alpha <i>(GP1BA)</i> binding to Integrin alpha L <i>(ITGAL)</i>. <b><i>Methods:</i></b> Seventy-nine patients with acute ischemic stroke and 151 controls without vascular disease from a single German center were enrolled. A neurologist and a neuroradiologist ascertained clinical and radiological features. PLA and platelet activation were analyzed using flow cytometry with various antibodies. Coding as well as tagging SNPs in six genes determining PLA were genotyped. Three groups of parameters were correlated with each other: (i) clinical and radiological parameters, (ii) laboratory parameters, (iii) genetic parameters. For the comparisons, robust nonparametric statistical tests were applicable. <b><i>Results:</i></b> PLA and platelet activation were higher in ischemic stroke patients compared to controls. Both, anticoagulant and antiplatelet treatment in the patient group affected platelet activation but not PLA. PLA correlated weakly with measures of stroke severity but not with thrombus length or stroke etiology. The association of SNP rs2228315 in the P-Selectin Glycoprotein Ligand-1-gene (PSGL1) with ischemic stroke and platelet activation was significant before correction for multiple testing while a trend was observed for the association with PLA. Regression analysis revealed that (i) platelet activation was an independent determinant of stroke, (ii) that PLA correlated with stroke, sex, age and platelet activation and (iii) that platelet activation correlated only with stroke. None of the SNPs survived in the regression analysis for stroke, PLA or platelet activation as dependent variables. <b><i>Conclusions:</i></b> The most important result of our study is that PLA and platelet activation are independent of other vascular risk factors correlated with stroke in our sample. In addition, we identified the missense SNP rs2228315 in the <i>PSGL1</i>-gene as a candidate polymorphism for ischemic stroke-related PLA. Association between this SNP and stroke as well as coronary artery disease has also been shown by two other studies.