Fig 2: Neutrophil phenotype, cytokines and NETs levels in children with COVID-19 according to disease severity. (a–g) Children with COVID-19 (asymptomatic, mild and moderate) and MIS-C were classified as described in Methods. (a–d) Expression of CD11b, CD66b, LAIR-1 and PD-L1 in gated neutrophils was analyzed by flow cytometry in whole blood samples. (a) CD11b (asymptomatic, n = 13; mild, n = 37; moderate, n = 15 and MIS-C, n = 15); (b) CD66b (asymptomatic, n = 10; mild, n = 17; moderate, n = 8 and MIS-C, n = 19); (c) LAIR-1 (asymptomatic, n = 7; mild, n = 9; moderate, n = 5 and MIS-C, n = 16); (d) PD-L1 (asymptomatic, n = 3; mild, n = 14; moderate, n = 4 and MIS-C, n = 8). (e) Spontaneous production of IL-1-ß (left) and IL-8 (right) evaluated in supernatants from unstimulated neutrophils (asymptomatic, n = 10; mild, n = 20; moderate, n = 10 and MIS-C, n = 10). (f) Plasma levels of cell-free DNA (left: asymptomatic, n = 13; mild, n = 24; moderate, n = 12 and MIS-C, n = 17) and citrullinated histones H3 (right: asymptomatic, n = 8; mild, n = 7; moderate, n = 4 and MIS-C, n = 8). (g) Levels of cell-free DNA in the supernatant of unstimulated neutrophils from asymptomatic, n = 7; mild, n = 16; moderate, n = 7 and MIS-C, n = 11. Dotted line depicts the median values found in healthy controls. Median and min to max of n donors are shown in a-g. P values were determined by Kruskal-Wallis test with Dunn's posttest and Mann-Whitney U test: * p < 0.05, ** p < 0.01, *** p < 0.001. Asymptomatic (light green circle), mild (dark green circle), moderate (yellow circle), MIS-C (red circle)(For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.) .

Fig 3: High expression of LAIR-1, PD-L1 and CD64 in neutrophils from children with COVID-19. (a) Frequency of LAIR-1 and PD-L1 positive cells in neutrophils from healthy (n = 20) and COVID-19 (n = 21). (b) Representative dot plots of LAIR-1 and PD-L1 expression in a donor from each cohort are shown. Numbers indicate the percentage of neutrophils positive for the expression of LAIR-1 or PD-L1. (c) Frequency of positive cells for CD16, CD32 and CD64 expression in neutrophils from healthy (n = 22) and COVID-19 (n = 56). (d) Representative dot plots of CD16 and CD64 expression are shown. Data show the percentages of positive cells in gated neutrophils. Median and min to max of n donors are shown in a and c. P values were determined by Kruskal-Wallis test with Dunn's posttest and Mann-Whitney U test: * p <0.05, ** p <0.01. Controls (white circle), COVID-19 (blue circle) (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.).