Emergence of dysfunctional neutrophils with a defect in arginase-1 release in severe COVID-19
Amrita Dwivedi, Aisling Ui Mhaonaigh, Makala Carroll, Bahareh Khosravi, Isabella Batten, Robert Seán Ballantine, Stuart Hendricken Phelan, Laura O’Doherty, Angel Mary George, Jacklyn Sui, Heike C. Hawerkamp, Padraic G. Fallon, Elnè Noppe, Sabina Mason, Niall Conlon, Clíona Ni Cheallaigh, Conor M. Finlay, Mark A. Little, Bioresource on behalf of the St James’s and Tallaght Trinity Allied Researchers (STTAR)
Amrita Dwivedi, Aisling Ui Mhaonaigh, Makala Carroll, Bahareh Khosravi, Isabella Batten, Robert Seán Ballantine, Stuart Hendricken Phelan, Laura O’Doherty, Angel Mary George, Jacklyn Sui, Heike C. Hawerkamp, Padraic G. Fallon, Elnè Noppe, Sabina Mason, Niall Conlon, Clíona Ni Cheallaigh, Conor M. Finlay, Mark A. Little, Bioresource on behalf of the St James’s and Tallaght Trinity Allied Researchers (STTAR)
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Research Article COVID-19 Immunology

Emergence of dysfunctional neutrophils with a defect in arginase-1 release in severe COVID-19

Abstract

Neutrophilia occurs in patients infected with SARS-CoV-2 (COVID-19) and is predictive of poor outcomes. Here, we link heterogenous neutrophil populations to disease severity in COVID-19. We identified neutrophils with features of cellular aging and immunosuppressive capacity in mild COVID-19 and features of neutrophil immaturity and activation in severe disease. The low-density neutrophil (LDN) number in circulating blood correlated with COVID-19 severity. Many of the divergent neutrophil phenotypes in COVID-19 were overrepresented in the LDN fraction and were less detectable in normal-density neutrophils. Functionally, neutrophils from patients with severe COVID-19 displayed defects in neutrophil extracellular trap formation and reactive oxygen species production. Soluble factors secreted by neutrophils from these patients inhibited T cell proliferation. Neutrophils from patients with severe COVID-19 had increased expression of arginase-1 protein, a feature that was retained in convalescent patients. Despite this increase in intracellular expression, there was a reduction in arginase-1 release by neutrophils into serum and culture supernatants. Furthermore, neutrophil-mediated T cell suppression was independent of arginase-1. Our results indicate the presence of dysfunctional, activated, and immature neutrophils in severe COVID-19.

Authors

Amrita Dwivedi, Aisling Ui Mhaonaigh, Makala Carroll, Bahareh Khosravi, Isabella Batten, Robert Seán Ballantine, Stuart Hendricken Phelan, Laura O’Doherty, Angel Mary George, Jacklyn Sui, Heike C. Hawerkamp, Padraic G. Fallon, Elnè Noppe, Sabina Mason, Niall Conlon, Clíona Ni Cheallaigh, Conor M. Finlay, Mark A. Little, Bioresource on behalf of the St James’s and Tallaght Trinity Allied Researchers (STTAR)

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Figure 1

Phenotypic analysis of whole blood neutrophils reveals distinct neutrophil populations in mild and severe COVID-19.

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Phenotypic analysis of whole blood neutrophils reveals distinct neutroph...
Fresh whole blood from healthy controls and patients with COVID-19 was subjected to phenotypic analysis using a combination of markers identifying the activation, maturation, and immunomodulatory status of neutrophils. (A) Total neutrophil count from patients with mild (n = 41) and severe (n = 29) COVID-19. (B) Flow cytometric gating strategy for identification of neutrophils in whole blood. (C) A total of 63,000 cells concatenated from 6 healthy controls (HC) and 10 patients with mild COVID-19 and 5 patients with severe COVID-19 were visualized using uniform manifold approximation and projection (UMAP) for dimensionality reduction. The UMAPs of all samples and separate study groups are shown. (D) Median fluorescence intensity (MFI) heatmap of surface markers projected on UMAP. (E) Distribution of identified FlowSOM populations overlaid on the UMAPs. (F) Frequency of the FlowSOM populations within HC (n = 6) and patients with mild (n = 10) and severe (n = 5) COVID-19. (G) Heatmap of selected surface markers in the 8 FlowSOM populations (Pop). (H) The HyperFinder plugin in FlowJo was used to determine the shortest gating strategy for the identified FlowSOM populations. Fraction of mature-homeostatic (HC, n = 6; mild C-19, n = 10; sev C-19, n = 5), aged (HC, n = 6; mild C-19, n = 17; sev C-19, n = 6), immunosuppressive (HC, n = 6; mild C-19, n = 10; sev C-19, n = 8), immature non-degranulated (HC, n = 6; mild C-19, n = 10; sev C-19, n = 9), immature (HC, n = 6; mild C-19, n = 18; sev C-19, n = 8), immature-activated (HC, n = 6; mild C-19, n = 15; sev C-19, n = 7). Statistical analysis performed using 2-tailed Mann-Whitney test (A). Differences between HC and mild and severe groups were analyzed using ordinary 1-way ANOVA with Tukey’s multiple comparisons test. *P < 0.05; **P < 0.01. Median with IQR is shown.