Abstract
ABSTRACTBroadly neutralizing antibodies that recognize the conserved hemagglutinin (HA) stalk have emerged as exciting new biotherapeutic tools to combat seasonal and pandemic influenza viruses. Our general understanding of the mechanisms by which stalk-specific antibodies achieve protection is rapidly evolving. It has recently been demonstrated that broadly neutralizing HA stalk-specific IgG antibodies require Fc-Fcγ receptor (FcγR) interactions for optimal protection in vivo. Here we examine the neutrophil effector functions induced by stalk-specific antibodies. As the most abundant subset of blood leukocytes, neutrophils represent a critical innate effector cell population and serve an instrumental role in orchestrating downstream adaptive responses to influenza virus infection. Yet, the interplay of HA stalk-specific IgG, Fc-FcγR engagement, and neutrophils has remained largely uncharacterized. Using an in vitro assay to detect the production of reactive oxygen species (ROS), we show that human and mouse monoclonal HA stalk-specific IgG antibodies are able to induce the production of ROS by neutrophils, while HA head-specific antibodies do not. Furthermore, our results indicate that the production of ROS is dependent on Fc receptor (FcR) engagement and phagocytosis. We went on to assess the ability of monoclonal HA stalk-specific IgA antibodies to induce ROS. Consistent with our findings for monoclonal IgGs, only HA stalk-specific IgA antibodies elicited ROS production by neutrophils. This induction is dependent on the engagement of FcαR1. Taken together, our findings describe a novel FcR-dependent effector function induced by HA stalk-specific IgG and IgA antibodies, and importantly, our studies shed light on the mechanisms by which HA stalk-specific antibodies achieve protection.
Highlights
Influenza virus vaccines, for decades, have been the mainstay approach to combat seasonal influenza and pandemic outbreaks
Neutralizing stalk-specific HA antibodies have been shown to require Fc-Fc␥ receptor (Fc␥R) interaction for optimal protection in vivo [17], yet the full scope of Fc-mediated effector functions elicited by these antibodies has not been fully described
Luminol reacts with reactive oxygen species (ROS) generated by neutrophils to produce an excited state intermediate that emits light as it relaxes to the ground state (Fig. 1)
Summary
For decades, have been the mainstay approach to combat seasonal influenza and pandemic outbreaks. The mechanisms by which bnAbs achieve protection are fundamentally different from strain-specific antibodies that target the globular head domain. While the latter prevent binding of HA to sialic acid on the host cell receptor, stalk-specific antibodies can prevent fusion of virus and endosomal membranes, interfere with viral egress, and inhibit cleavage of HA required for maturation [16]. Given the ubiquitous expression of Fc␥R on innate immune cells, this led us to hypothesize that additional Fc-mediated effector functions would be elicited by this class of antibodies. While several studies have illustrated the critical role of neutrophils in limiting viral replication [23, 24], there is a paucity of data regarding the antibody-mediated effector function of neutrophils in the context of influenza virus infection. Our findings reveal a previously unappreciated effector function of HA stalk-specific bnAbs and help to provide a more complete picture of the mechanisms induced by this class of antibodies
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