Background: Venous thromboembolism (VTE), comprising deep vein thrombosis (DVT) and pulmonary embolism (PE), affects nearly 1 of every 100 hospitalized children in the US each year and is associated with both increased mortality (up to 2%) and morbidity.
VTE development is multifactorial based upon Virchow's triad - vessel wall injury, stasis of blood flow, and hypercoagulability - to describe the pathophysiology and identify risk factors for VTE formation. A growing body of literature demonstrates that inflammation plays a key role in VTE formation. For example, in a risk-assessment model for hospital acquired-VTE, children with inflammatory/autoimmune disease had 4.32 increased odds of VTE development (95% CI 2.51-7.45).
Elevation of pro-inflammatory cytokines has been associated with risk for primary or recurrent VTE. Prior cohort studies demonstrate IL-6 elevation is independently associated with first occurrence of VTE. IL-6 is a cytokine synthesized in response to inflammation. IL-6 signaling requires formation of a 3-part complex including IL-6, one of two IL-6 receptor types (membrane-bound for classical signaling or soluble for trans-signaling) and the ubiquitously expressed cell surface receptor gp130 that acts as the signal transducer. Prior studies demonstrated that physiologic concentrations of IL-6 stimulation of platelet rich plasma (PRP) enhances agonist-induced platelet aggregation and secretion. Specifically, IL-6 signaling in platelets induces STAT3 phosphorylation and enhances collagen-induced platelet aggregation by influencing signaling through the GPVI platelet glycoprotein receptor.
Post-translational modification (such as glycosylation) of surface adhesion receptors affects platelet activity, localization, and adhesion. Inflammation-associated glycan alterations on platelets themselves have not been previously studied, thus representing a novel approach to exploring the mechanistic underpinnings of thromboinflammatory VTE.
Methods: Whole blood from healthy human donors was drawn into acid-citrate-dextrose and platelets were isolated by differential centrifugation, then incubated with various concentrations of hyper IL-6 (peptide of both recombinant IL-6 and soluble IL-6 receptor). Doses were comparable to patients receiving IL-6 immunotherapy (low-dose) and to those with meningococcal septic shock (high dose). Collagen response was assessed by addition of collagen-related peptide at low concentrations. Glycan changes were assessed via lectin array.
Results: In collagen-stimulated platelets, addition of hyper IL-6 increases α-linked fucosylation via increased binding of AAL and AOL lectins. Furthermore, the addition of hyper IL-6 leads to differential sialylation patterns of the collagen-stimulated platelets: α-2,3 linked sialylation was significantly decreased based on ACG lectin binding and α-2,6 linked sialylation was significantly increased based on binding of SNA and TJA-1 lectins.
Conclusions: Multiple glycosylation changes occur in collagen-stimulated platelets in the presence of IL-6. Our findings of increased fucosylation and differential patterns in sialylation linkages are consistent with those noted in other inflammatory conditions besides VTE. As such, this study lays the groundwork for exploration of the mechanisms of glycan alterations on platelet activation responses in the setting of inflammation.
No relevant conflicts of interest to declare.
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