Figure 5.

Activation of IL-33/ST2 signaling protects against OGD in mixed N/G cultures but not in neuron-enriched cultures. A, B, Rat mixed N/G cultures with or without 3 h OGD were treated with a range of concentrations of IL-33 or PBS (control) for 24 h. A, Neuronal survival was quantified using MAP2 ELISA. n = 8 per group. Data represent three independent experiments. B, Representative images of Hoechst and NeuN double staining in N/G cultures treated with 50 ng/ml IL-33 or PBS (control) for 24 h. Scale bar, 30 μm. C, ELISA quantification of MAP2 expression at 24 h after OGD (3 h) treatments in N/G cultures prepared from WT and ST2 KO mice. Data represent three independent experiments. D, MTT assay in rat neuron-enriched cultures subjected to 1.5 h OGD or sham conditions followed by treatment with PBS or a range of concentrations of IL-33 for another 24 h. Data represent three independent experiments. E, F, MTT and LDH assays were performed at 24 h after OGD (1.5 h) in neuron-enriched cultures prepared from WT and ST2 KO mice. G, Quantification of RT-PCR for M2 markers IL-10, TGF-β, CD206, and M1 marker inducible nitric oxide synthase (iNOS) in microglial cultures treated with 50 ng/ml IL-33 or PBS (control) for 24 h. n = 8–9 per group. Data are mean ± SEM. *p < 0.05, **p < 0.01, ****p < 0.0001 (2-tailed Student's t test or 1-way ANOVA followed by the Bonferroni's post hoc).