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Fig. 4 | Acta Neuropathologica Communications

Fig. 4

From: Complement-independent bystander injury in AQP4-IgG seropositive neuromyelitis optica produced by antibody-dependent cellular cytotoxicity

Fig. 4

ADCC bystander killing in astrocyte-neuron cocultures. a Astrocyte-neuron cocultures following 30 min pre-coating with AQP4-IgG and 2 h incubation with NK cells at an effector:target cell ratio of 5:1, with fixable dead cell marker added for the final 30 min. GFAP (astrocyte) and MAP2 (neuron) immunofluorescence, with dead cells stained red, shown at high (left) and low (right) magnifications. Yellow filled arrows indicate dead astrocytes, yellow open arrows show dead neurons. b Fraction of dead neurons at different distances from the center of dead astrocytes (mean ± S.E.M., 3 slides with > 30 dead cells analyzed, ** P<0.01, *P<0.05 comparing AQP4-IgG + NK vs. control IgG + NK or AQP4-IgG without NK cells by two-way ANOVA). c Perforin immunofluorescence (red) of cocultures treated as in a. with GFAP (green) and MAP2 (gray) immunofluorescence. Yellow filled arrows indicate perforin on astrocytes, yellow open arrows show perforin on neurons. d AQP4 immunofluorescence (green) with dead cell stain (red) and DAPI (blue) at high (left) and low (center) magnifications in CHO-AQP4 and CHO-null cocultures following 5 μg/ml AQP4-IgG and neutrophils at an effector:target cell ratio of 5:1. Yellow filled arrows indicate dead CHO-AQP4 cells, yellow open arrows show dead CHO-null cells. (Right) Fraction of dead CHO-null cells at different distances from the center of dead CHO-AQP4 cells (mean ± S.E.M., 3 slides with > 50 dead cells analyzed, ** P<0.01, *P<0.05 comparing AQP4-IgG + neutrophils vs. neutrophils or pure CHO-null cells by two-way ANOVA)

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