Abstract Rationale Sickle cell disease (SCD) is an inherited blood disorder affecting ∼100,000 African Americans and over 8 million individuals worldwide. In SCD, polymerization of mutant hemoglobin drives erythrocyte sickling, vaso-occlusion, and hemolysis, leading to a chronic thrombo-inflammatory milieu. Although influenza A virus (IAV) infection typically causes a self-resolving upper airway inflammation in healthy humans, SCD patients exhibit ∼50-fold higher risk of ICU admission and mortality following flu. However, mechanism linking viral infection to development of severe acute lung injury (ALI) in SCD remains poorly understood. The current treatment in SCD patients is limited to flu vaccine and antiviral therapy, however, these therapies need to be given at early stage of infection. Understanding the mechanism driving flu severity in SCD will enable new therapies to halt development of ALI in SCD patients with flu. Methods Murine model of IAV-induced ALI was developed in knock-in humanized-Townes SCD (SS) or control (AS) mice inoculated intranasally with a mild dose of A/PR/8/34 (H1N1) IAV. Lung injury severity was assessed 14 days post-infection based on histological scoring. Oxygen saturation and body weight were measured every alternate day. Real-time multi-photon-excitation-enabled intravital lung microscopy was performed at days-10 and 14 to visualize thrombo-inflammation, vascular leakage, and neutrophil extracellular traps (NETs) generation in the pulmonary microcirculation. Platelets, neutrophils and plasma were collected for western blotting, co-immunoprecipitation, imaging flow cytometry, transcriptomics, and assessment of circulating NETs and activation of type-I interferon signaling. Results At day-14 post flu infection, SCD mice manifested significant drop in oxygen saturation (90%), body weight (≥20%) and severe ALI indicated by the presence of hemorrhage, vascular congestion and edema. Severe ALI in SCD mice was associated with pulmonary thrombosis leading to microvasculature occlusion by neutrophil-platelet aggregates (NPAs), resulting in pulmonary ischemia and loss of blood-air barrier. Unlike SCD mice, control mice developed only a mild pulmonary inflammation at day 10, which was completely resolved by day 14-post flu infection, leading to absence of ALI and neutrophil-platelet aggregates in the pulmonary microcirculation. Pulmonary thrombosis in SCD mice was secondary to activation of viral-RNA sensing RIG-I-MAVS pathway, leading to sequestration of NPAs, upregulation of type-I interferon signaling in neutrophils and NETs generation in the lung microcirculation. Conclusions These findings reveal that mild flu infection promotes pulmonary thrombosis and ALI in SCD mice, which is secondary to activation of anti-viral-RIG1-signaling in platelets. Inhibition of platelet-RIG1 may offer novel anti-platelet therapy for preventing severe outcome in SCD patients with flu. This abstract is funded by: Versiti Foundation
Kaminski et al. (Fri,) studied this question.