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A2700 - Platelets Shed Extracellular Vesicles to Promote IL-1β Dependent Lung Vaso-Occlusion in Sickle Cell Disease
Author Block: P. Sundd, M. Jimenez, M. Bennewitz, T. Brzoska, M. T. Gladwin; Medicine-VMI-PACCM, University of Pittsburgh, Pittsburgh, PA, United States.
RATIONALE: Acute chest syndrome (ACS) is a type of acute lung injury and the leading cause of mortality in Sickle Cell Disease (SCD). Current treatments for ACS are primarily supportive, and there is a critical need for rescue therapies. ACS is often a sequela of acute systemic vaso-occlusive crisis and preceded by thrombocytopenia. However, the role of platelets in the pathogenesis of ACS remains largely unknown. METHODS: We used our validated model of vaso-occlusive crisis in transgenic, humanized SCD mice, which is triggered by intravenous challenge with nanogram levels of the TLR4 ligand, lipopolysaccharide (LPS). Platelet-neutrophil aggregates and blood flow in the lung microcirculation was visualized in real time in vivo, using multi-photon-excitation fluorescence microscopy of intact lung in live SCD mice. SCD or control human blood was perfused through microfluidic flow channels in vitro and neutrophil-platelet aggregation was visualized using fluorescence microscopy. Platelet derived extracellular vesicles were characterized using nanoparticle tracking and biochemical approaches. RESULTS: We have made a novel finding that the arrest of blood flow and injury in the lung is secondary to blockade of pulmonary arterioles by platelet-neutrophil aggregates. We found that platelets in SCD are hairy in appearance due to the presence of hair-like membrane tethers on the surface. The hair-like tethers shed IL-1β carrying extracellular vesicles and promoted platelet-neutrophil aggregation in lung arterioles in vivo. Remarkably, inhibition of TLR4 or NLRP3-inflammasome activated caspase-1 or IL-1β signaling abolished hair-like tethers from platelets, attenuated release of platelet-derived extracellular vesicles, and prevented platelet-neutrophil aggregation in the lung arterioles of SCD mice in vivo and SCD human blood in vitro. CONCLUSIONS: TLR4 and NLRP3-inflammasome-mediated caspase-1 activation in platelets promotes the release of IL-1β-containing extracellular vesicles in SCD. These circulating extracellular vesicles promote platelet-neutrophil aggregation in pulmonary arterioles, which results in arrest of blood flow in the lung, leading to ACS. Therapeutic inhibition of IL-1β dependent innate immune pathway can be a potential therapy for ACS in SCD patients.