Human blood neutrophils generate ROS through FcγR-signaling to mediate protection against febrile P. falciparum malaria

Human blood neutrophils generate ROS through FcγR-signaling to mediate protection against febrile P. falciparum malaria
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Here, we depict our study discerning the role of IgG-mediated generation of reactive oxygen species (ROS) by major human blood phagocytes (neutrophils and monocytes) against Plasmodium falciparum asexual blood stage parasites. We find that neutrophils play a dominant role in generation of ROS though Fc gamma receptor (FcγR) signaling stimulated by IgG-opsonized parasites, and neutrophil ROS was strongly correlated with protection against clinical malaria in immune-epidemiology studies.

Malaria, caused by P. falciparum parasites, poses a significant threat to global health, accounting for over 95% of malaria cases and 96% of deaths in Africa1. These obligate intracellular parasites undergo a major phase of their development within the human host involving several cycles of multiplication within red blood cells, causing severe symptoms and fatalities. Each round of this intra-erythrocytic cycle takes approximately 48 hours releasing short-lived extracellular progenies (merozoites) that quickly invade new red blood cells.

These blood parasite development stages are targets of IgG mediated naturally acquired immunity (NAI) leading to control of parasite load and conferring protection against clinical disease2,3. However, the specific ways in which these antibodies protect against the parasite remain uncertain. Different mechanisms of IgG mediated anti-parasite effects have been described. These include either the IgG acting on their own4,5 or in co-operation with other blood leukocytes. The Fc domains of these protective IgG engage with the cellular FcγR6 triggering a variety of potent parasite killing mechanisms (depending on the leukocyte subset and the combination of the FcγR-types involved) such as the antibody dependent cellular inhibition (ADCI)7; opsonic phagocytosis (OP)8-10; antibody-dependent respiratory burst (ADRB)11 and generation of ROS (as shown in this study). It is likely that combinations of these IgG mediated anti-parasite mechanisms may operate concurrently, and perhaps complement each other, in order to control the parasite load and accord protection against clinical malaria.

Our study delved into the mechanisms behind IgG-mediated protection using an ex vivo bioassay with human peripheral blood leukocyte samples (with normal distribution of the cell types). By examining different subsets of immune cells, we focused on their respective FcγR distribution and their ability to generate ROS when exposed to IgG-opsonized P. falciparum merozoites. Notably, we found that neutrophils play a dominant role in this process, primarily involving FcγRIIIB (exclusively expressed on neutrophils), as well as FcγRIIA, both acting in concert for IgG-mediated ROS generation by the neutrophils. Meanwhile, monocytes predominantly rely on FcγRIIA, with a minor involvement of FcγRIIIA for triggering ROS generation.

Our results unveiled that IgG-opsonized merozoites activate FcγRs, stimulate the assembly of the NOX212 system in blood phagocytes both on the cell membrane as well as within the phagosomal membrane. This process leads to ROS generation within these phagocytic cells themselves and in the surrounding vicinity. We also explored the signaling pathway linking the cell membrane FcγR to the downstream signals involved in the IgG-mediated ROS generation. Interestingly, inhibiting phosphoinositide 3-kinase (PI3K) using wortmannin (a specific PI3K inhibitor)13, but not the protein kinase C (PKC) using Gö6983 (a specific PKC inhibitor)14, significantly reduced FcγR-mediated ROS production. Notably, these inhibitors did not impact merozoite-phagocytosis, suggesting distinct downstream signaling pathways govern phagocytosis and ROS generation.

Furthermore, we explored the connection between IgG-dependent ROS generation in specific immune cell subsets and protection against febrile malaria. Two separate cohort studies conducted in geographically diverse malaria endemic regions from Ghana and India provided valuable insights. Remarkably, the ability of plasma samples to enhance neutrophil ROS generation strongly correlated with protection against febrile malaria in both locations. This finding highlights the critical role of neutrophil effector functions in naturally acquired immunity against malaria. To what extent the FcγR- mediated ROS generated by these phagocytes directly mediates parasite killing remains unclear. It is likely that the ROS generated by neutrophils could further trigger release of certain intracellular granule secretion to mediate the parasite killing.

In conclusion, our study sheds light on the vital role of neutrophils in FcγR-mediated protection against febrile P. falciparum malaria. Understanding the intricacies of these protective mechanisms is crucial for advancing our knowledge of naturally acquired immunity against malaria and developing effective immune-intervention strategies for combatting this devastating disease.

The full story can be read : 10.1038/s42003-023-05118-0

References:

  1. WHO. World Malaria Report 2022. (2022).
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  3. McGregor, I. & Carrington, S. Treatment of East African P. falciparum malaria with West African human gamma globulin. Transactions of the Royal Society of Tropical Medicine and Gygiene 57, 170-175 (1963).
  4. Hodder, A.N., Crewther, P.E. & Anders, R.F. Specificity of the protective antibody response to apical membrane antigen 1. Infect.Immun. 69, 3286-3294 (2001).
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  7. Bouharoun-Tayoun, H., Attanath, P., Sabchareon, A., Chongsuphajaisiddhi, T. & Druilhe, P. Antibodies that protect humans against Plasmodium falciparum blood stages do not on their own inhibit parasite growth and invasion in vitro, but act in cooperation with monocytes. J Exp Med 172, 1633-1641 (1990).
  8. Hill, D.L., et al. Opsonising antibodies to P. falciparum merozoites associated with immunity to clinical malaria. PLoS One 8, e74627 (2013).
  9. Osier, F.H., et al. Opsonic phagocytosis of Plasmodium falciparum merozoites: mechanism in human immunity and a correlate of protection against malaria. BMC medicine 12, 108 (2014).
  10. Kana, I.H., et al. Breadth of Functional Antibodies Is Associated With Plasmodium falciparum Merozoite Phagocytosis and Protection Against Febrile Malaria. J Infect Dis 220, 275-284 (2019).
  11. Joos, C., et al. Clinical protection from falciparum malaria correlates with neutrophil respiratory bursts induced by merozoites opsonized with human serum antibodies. PLoS One 5, e9871 (2010).
  12. Cross, A.R. & Jones, O.T. The effect of the inhibitor diphenylene iodonium on the superoxide-generating system of neutrophils. Specific labelling of a component polypeptide of the oxidase. Biochem J 237, 111-116 (1986).
  13. Rastogi, R., Geng, X., Li, F. & Ding, Y. NOX Activation by Subunit Interaction and Underlying Mechanisms in Disease. Front Cell Neurosci 10, 301 (2016).
  14. Chen, F., et al. Regulation of NADPH oxidase 5 by protein kinase C isoforms. PLoS One 9, e88405 (2014).

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