Mass Cytometry Phenotyping of Human Granulocytes Reveals Novel Basophil Functional Heterogeneity

Abstract

Summary Basophils, the rarest granulocyte, play critical roles in parasite- and allergen-induced inflammation. We applied mass cytometry (CyTOF) to simultaneously asses 44 proteins to phenotype and functionally characterize neutrophils, eosinophils, and basophils from 19 healthy donors. There was minimal heterogeneity seen in eosinophils and neutrophils, but data-driven analyses revealed four unique subpopulations within phenotypically basophilic granulocytes (PBG; CD45 + HLA-DR − CD123 + ). Through CyTOF and fluorescence-activated cell sorting (FACS), we classified these four PBG subpopulations as (I) CD16 low FcεRI high CD244 high (88.5 ± 1.2%), (II) CD16 high FcεRI high CD244 high (9.1 ± 0.4%), (III) CD16 low FcεRI low CD244 low (2.3 ± 1.3), and (IV) CD16 high FcεRI low CD244 low (0.4 ± 0.1%). Prospective isolation confirmed basophilic-morphology of PBG I–III, but neutrophilic-morphology of PBG IV. Functional interrogation via IgE-crosslinking or IL-3 stimulation demonstrated that PBG I–II had significant increases in CD203c expression, whereas PBG III–IV remained unchanged compared with media-alone conditions. Thus, PBG III–IV could serve roles in non-IgE-mediated immunity. Our findings offer new perspectives in human basophil heterogeneity and the varying functional potential of these new subsets in health and disease.

Effect of intranasal corticosteroid treatment on allergen‐induced changes in group 2 innate lymphoid cells in allergic rhinitis with mild asthma

“…Due to the low numbers of ILC2s in the nasal curettage samples detected by flow cytometry, the current data should be interpreted with caution. Supportive data from more rigorous techniques such as CyTOF (cytometry by time of flight) 28 and multiplexed ion beam imaging 29 will be essential to confirm the current findings and to exclude the possibility of ILC2s from blood vessels contaminating the mucosal scraping taken in this study. IL‐2 maintains ILC2 survival and cytokine production, 30,31 whereas IL‐4 is important for ILC2 proliferation 31,32 .…”

A census of the lung: CellCards from LungMAP

“…Function: Basophils produce antimicrobial extracellular traps and regulate group 2 innate lymphoid cell (ILC2) responses (Lee and McGarry, 2007;Siracusa, 2016;Cohen et al, 2018;Kabashima et al, 2018;Inclan-Rico et al, 2020;Marone et al, 2020;Vivanco Gonzalez et al, 2020). In addition, they produce effector molecules in response to stimuli such as inflammatory mediators (histamine, serotonin, platelet-activating factor, and leukotriene), cytokines , and chemokines (CCL3, CCL4, CCL12, and CXCL12) (Siracusa, 2016;Inclan-Rico et al, 2020).…”

“…Heterogeneity and cellular states: Unknown in lung. Four subpopulations have been characterized in blood (Vivanco Gonzalez et al, 2020) ll Regenerative potential: No self-renewal potential (Yamanishi and Karasuyama, 2016).…”

Differential Effects of Alarmins on Human and Mouse Basophils

“…We used flow cytometry to purify mBaso and there is a possibility that this procedure might have activated murine basophils. Finally, Gonzales and collaborators have recently identified four phenotypically distinct subpopulations of human peripheral blood basophils ( 90 ) and Pellefigues and coworkers have provided some evidence of the existence of mBaso functional heterogeneity ( 50 ). In our study, we did not address the emerging issue of possible existence of human and mouse basophil heterogeneity.…”