FIGURE 8.

Treatment of hNSC with αC1q and αC3a Abs blocks PMN-CM–induced effects on astroglial fate and migration in vitro. (A–F) hNSC were treated with PMN-CM (orange bars) or DM control (green bars) with or without addition of αC1q and αC3a Abs for 7 DIV. Black bars indicate SDF1α (positive control for Transwell migration). Comparisons versus DM control (dashed line) were conducted using one-sample t tests (NS is p ≥ 0.05, ^#p < 0.05, ^##p < 0.01, ^###p < 0.001). Comparisons between treatment conditions were conducted using one-way ANOVA, followed by Tukey post hoc tests (NS is p ≥ 0.05, *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001). (A) hNSC treatment with PMN-CM plus αC1q/αC3a alone or in combination decreased GFAP⁺ cell number (one-way ANOVA, F = 6.394, p < 0.01). (B) hNSC treatment with DM control plus αC1q/αC3a alone or in combination did not alter GFAP⁺ cell number (one-way ANOVA, F = 0.0335, p > 0.05). (C) hNSC treatment with either DM control (green bars) or PMN-CM plus αTNF-α (orange bars) did not alter GFAP⁺ cell number (one-way ANOVA, F = 4.913, p < 0.01). (D) hNSC migration toward PMN-CM was decreased in the presence of αC1q/αC3a Ab alone or in combination (one-way ANOVA, F = 8.135, p < 0.0001). Treatment with both Abs prevented ∼76% hNSC chemotaxis. (E) hNSC migration was unaltered in the presence of αC1q plus αC3a Ab (one-way ANOVA, F = 22.38, p < 0.0001). (F) hNSC migration was unaltered for DM versus DM plus αSDF1 (green bar), and for PMN-CM versus PMN-CM plus αSDF1 (orange bars; one-way ANOVA, F = 13.44, p < 0.0001). αSDF1α Ab activity was verified by reversal of SDF1α-induced hNSC migration (black bar, one-sample t test SDF1α versus DM control ^###; gray bar, one-sample t test SDF1α plus αSDF1α versus DM control NS). (F′) RT-PCR did not detect mRNA expression of SDF1α by PMN. For (A–F), n = 2 technical replicates and n = 3–4 biological replicates/condition. Mean ± SEM.