Figure 3.

MANF Is Protective against OGD in NSCs

(A) Timeline of the oxygen-glucose-deprivation/reperfusion experiment. (B) Representative photomicrographs of activated caspase-3 labeling in the NSCs after OGD or under normal conditions. (C) Ratio of activated caspase-3+ cells in relation to DAPI⁺ nuclei. Increase in activated caspase-3 density in Manf^−/− cells after OGD treatment compared to WT cells. rhMANF treatment before OGD decreased activated caspase-3 density in Manf^−/− cells (n = 6–11, two-way ANOVA, genotype X condition effect p = 0.042; **p < 0.01 genotype effect; *p < 0.05 Manf^−/− OGD versus OGD + rhMANF). (D) Analysis of cell death by PI labeling. The percentage of PI⁺Manf^−/− nuclei after OGD and reoxygenation was higher than in WT cells, indicating a greater apoptotic rate in Manf^−/− cells. OGD-induced Manf^−/− cell death was prevented by rhMANF treatment (n = 6 to 7, two-way ANOVA, genotype X condition effect p = 0.001; ***p < 0.001 genotype comparison; *p < 0.05 Manf^−/− OGD versus OGD + rhMANF). Scale bar, 100 μm (B). Data are presented as mean ± SEM.