SARS-CoV-2 Spike Protein Stimulates Macropinocytosis in Murine and Human Macrophages via PKC-NADPH Oxidase Signaling

Coronavirus disease 2019 (COVID-19) is an infectious disease caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). While recent studies have demonstrated that SARS-CoV-2 may enter kidney and colon epithelial cells by inducing receptor-independent macropinocytosis, it remains unknown whether this process also occurs in cell types directly relevant to SARS-CoV-2-associated lung pneumonia, such as alveolar epithelial cells and macrophages. The goal of our study was to investigate the ability of SARS-CoV-2 spike protein subunits to stimulate macropinocytosis in human alveolar epithelial cells and primary human and murine macrophages. Flow cytometry analysis of fluid-phase marker internalization demonstrated that SARS-CoV-2 spike protein subunits S1, the receptor-binding domain (RBD) of S1, and S2 stimulate macropinocytosis in both human and murine macrophages in an angiotensin-converting enzyme 2 (ACE2)-independent manner. Pharmacological and genetic inhibition of macropinocytosis substantially decreased spike-protein-induced fluid-phase marker internalization in macrophages both in vitro and in vivo. High-resolution scanning electron microscopy (SEM) imaging confirmed that spike protein subunits promote the formation of membrane ruffles on the dorsal surface of macrophages. Mechanistic studies demonstrated that SARS-CoV-2 spike protein stimulated macropinocytosis via NADPH oxidase 2 (Nox2)-derived reactive oxygen species (ROS) generation. In addition, inhibition of protein kinase C (PKC) and phosphoinositide 3-kinase (PI3K) in macrophages blocked SARS-CoV-2 spike-protein-induced macropinocytosis. To our knowledge, these results demonstrate for the first time that SARS-CoV-2 spike protein subunits stimulate macropinocytosis in macrophages. These results may contribute to a better understanding of SARS-CoV-2 infection and COVID-19 pathogenesis

Grand Challenges in global eye health: a global prioritisation process using Delphi method

Background: We undertook a Grand Challenges in Global Eye Health prioritisation exercise to identify the key issues that must be addressed to improve eye health in the context of an ageing population, to eliminate persistent inequities in health-care access, and to mitigate widespread resource limitations. Methods: Drawing on methods used in previous Grand Challenges studies, we used a multi-step recruitment strategy to assemble a diverse panel of individuals from a range of disciplines relevant to global eye health from all regions globally to participate in a three-round, online, Delphi-like, prioritisation process to nominate and rank challenges in global eye health. Through this process, we developed both global and regional priority lists. Findings: Between Sept 1 and Dec 12, 2019, 470 individuals complete round 1 of the process, of whom 336 completed all three rounds (round 2 between Feb 26 and March 18, 2020, and round 3 between April 2 and April 25, 2020) 156 (46%) of 336 were women, 180 (54%) were men. The proportion of participants who worked in each region ranged from 104 (31%) in sub-Saharan Africa to 21 (6%) in central Europe, eastern Europe, and in central Asia. Of 85 unique challenges identified after round 1, 16 challenges were prioritised at the global level; six focused on detection and treatment of conditions (cataract, refractive error, glaucoma, diabetic retinopathy, services for children and screening for early detection), two focused on addressing shortages in human resource capacity, five on other health service and policy factors (including strengthening policies, integration, health information systems, and budget allocation), and three on improving access to care and promoting equity. Interpretation: This list of Grand Challenges serves as a starting point for immediate action by funders to guide investment in research and innovation in eye health. It challenges researchers, clinicians, and policy makers to build collaborations to address specific challenges. Funding: The Queen Elizabeth Diamond Jubilee Trust, Moorfields Eye Charity, National Institute for Health Research Moorfields Biomedical Research Centre, Wellcome Trust, Sightsavers, The Fred Hollows Foundation, The Seva Foundation, British Council for the Prevention of Blindness, and Christian Blind Mission. Translations: For the French, Spanish, Chinese, Portuguese, Arabic and Persian translations of the abstract see Supplementary Materials section.</p