Terminal sialylation determines plasma VWF half-life. A role for macrophage galactose lectin (MGL) in regulating hyposialylated VWF clearance has recently been proposed. In this study, we show that MGL influences physiological plasma VWF clearance. MGL inhibition was associated with a significantly extended mean residence time and 3-fold increase in endogenous plasma VWF:Ag levels (p<0.05). Using a series of VWF truncations, we further demonstrate that the A1 domain of VWF is predominantly responsible for enabling MGL interaction. Binding of both full-length and VWF-A1-A2-A3 to MGL were significantly enhanced in the presence of ristocetin (p<0.05), suggesting that the MGL-binding site in A1 is not fully accessible in globular VWF. Additional studies using different VWF glycoforms demonstrated that VWF O-linked glycans, clustered at either end of the A1 domain play a key role in protecting VWF against MGL-mediated clearance. Reduced sialylation has been associated with pathological increased clearance in patients with VWD. Herein, we demonstrate that specific loss of α2-3 linked sialylation from Oglycans results in markedly increased MGL-binding in vitro, and markedly enhanced MGLmediated clearance in vivo. Our data further show that the Asialoglycoprotein receptor (ASGPR) has no significant role in mediating the increased clearance of VWF following loss of O-sialylation. Conversely however, we observed that loss of N-linked sialylation from VWF drives enhanced circulatory clearance predominantly via the ASGPR. Altogether, our data support the hypothesis that in addition to regulating physiological VWF clearance, the MGL receptor works in tandem with ASGPR to modulate enhanced clearance of aberrantly sialylated VWF in VWD pathogenesis.
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