COVID-19, the disease caused by the SARS-CoV-2 virus, can progress to multi-organ failure characterized by respiratory insufficiency, arrhythmias, thromboembolic complications and shock. The mortality of patients hospitalized with COVID-19 is unacceptably high and new strategies are urgently needed to rapidly identify and treat patients at risk for organ failure. Clinical epidemiologic studies demonstrate that vulnerability to organ failure is greatest after viral clearance from the upper airway, which suggests that dysregulation of the host immune response is a critical mediator of clinical deterioration and death. Autopsy and pre-clinical evidence implicate aberrant complement activation in endothelial injury and organ failure. A potential therapeutic strategy warranting investigation is to inhibit complement, with case reports of successful treatment of COVID-19 with inhibitors of complement. However, this approach requires careful balance between the host protective and potential injurious effects of complement activation, and biomarkers to identify the optimal timing and candidates for therapy are lacking. Here we report the presence of complement activation products on circulating erythrocytes from hospitalized COVID-19 patients using flow cytometry. These findings suggest that novel erythrocyte-based diagnostics provide a method to identify patients with dysregulated complement activation.

RBC transfusion is associated with increased morbidity and mortality in critically ill patients. Endothelial cell necroptosis and subsequent damage-associated molecular pattern (DAMP) release has been identified as a mechanism of injury following RBC transfusion. Mounting evidence implicates the pro-inflammatory pattern recognition receptor, Receptor for Advanced Glycation End Products (RAGE), in initiating cell death programmes such as necroptosis. Here, we demonstrate the role of RAGE in endothelial necroptosis, as deletion of RAGE attenuates necroptotic cell death in response to TNFa, LPS or CpG-DNA. We show direct interaction of RAGE with the critical mediator of necroptosis, Receptor Interacting Protein Kinase 3 (RIPK3), during necroptosis. Furthermore, we observe decreased plasma High Mobility Group Box 1 (HMGB1) and RIPK3 levels in RAGE deficient mice compared to WT mice post-transfusion, substantiating the role for RAGE in transfusion-induced DAMP release in vivo. Collectively, these findings underscore RAGE as an essential mediator of regulated necrosis and post-transfusion DAMP release. Further studies to understand the role of RAGE and the necroptotic pathway in transfusion-induced organ injury may offer key targets to mitigate transfusion-related risks, including the risk of ARDS, in susceptible hosts.

Red blood cells (RBCs) express the nucleic acid-sensing toll-like receptor 9 (TLR9) and bind CpG-containing DNA. However, whether human RBCs express other nucleic acid-sensing TLRs and bind RNA is unknown. Here we show that human RBCs express the RNA sensor, TLR7. TLR7 is present on the red cell membrane and associates with the RBC membrane protein Band 3. RBCs bind synthetic single-stranded RNA and RNA from pathogenic single-stranded RNA viruses. RNA acquisition by RBCs is attenuated by recombinant TLR7 and inhibitory oligonucleotides. Thus, RBCs may represent a previously unrecognized reservoir for RNA, although how RNA-binding by RBCs modulates the immune response has yet to be elucidated. These findings add to the growing list of non-gas exchanging RBC immune functions.

Red blood cells (RBCs) demonstrate immunomodulatory capabilities through the expression of nucleic acid sensors. Little is known about bat RBCs, and no studies have examined the immune function of bat erythrocytes. Here we show that bat RBCs express the nucleic acid-sensing Toll-like receptors TLR7 and TLR9 and bind the nucleic acid ligands, single-stranded RNA, and CpG DNA. Collectively, these data suggest that, like human RBCs, bat erythrocytes possess immune function and may be reservoirs for nucleic acids. These findings provide unique insight into bat immunity and may uncover potential mechanisms by which virulent pathogens in humans are concealed in bats.

Following cell stress, injury, or infections, potentially injurious host-derived damageassociated molecular patterns (DAMPs) trigger the innate immune response. Among these DAMPs, the importance of extracellular nucleic acids, such as mitochondrial DNA and vesicleassociated non-coding RNA, in the pathogenesis of acute lung injury (ALI) and acute respiratory distress syndrome (ARDS), is increasingly recognized. Extracellular RNA is primarily composed of microRNA (miRNA) and ribosomal (rRNA) and can be vesicle-associated, protein-bound, or free form. These different forms of exRNA play roles in endothelial permeability and inflammation via recognition by receptors such as vascular endothelial growth factor receptor 2 (VEGFR-2) (1), receptor for advanced glycation end products (RAGE) (2), and toll-like receptors (TLRs). Plasma and alveolar extracellular RNA (exRNA) profiles are altered during lung injury and sepsis (3), and plasma microRNA-146-5p (miR146-5p) is elevated during sepsis (4). Given the diverse sequences and forms of exRNA and increasing interest in RNA-based therapeutics, understanding how some of these exRNAs contribute to lung injury or other diseases will advance our technology in applying RNA biology to mitigate inflammatory diseases and clinical syndromes including sepsis and acute lung injury.Here, Huang and colleagues report that extracellular miR146a-5p activates TLR7 in macrophages and induces vascular permeability via TNFα (5). miR146a-5p is a miRNA that negatively regulates TLR signaling intracellularly by silencing IRAK1/TRAF6 expression (6).However, the extracellular form of miR146a-5p is vesicle-associated and is proinflammatory in macrophages (7). Thus miR146a-5p fulfills the definition of a DAMP. In a cecal ligation and puncture-induced model of sepsis, miR146a-5p is elevated in both plasma and bronchoalveolar lavage. Intra-tracheal administration of miR146a-5p induced TLR7-dependent proinflammatory cytokine expression, endothelial disruption, and neutrophil recruitment. In vitro, conditioned