Fig. 5

Expression of inflammatory factors in cultured microglia. a ELISA. Mixed glia cells were prepared from 5 neonates and cultured for 2 weeks and then treated with 1, 10 or 100 ng/mL of LPS or PBS for an additional 24 or 48 h. Media were recovered and subjected to ELISA for IL-6, PGE2 and IL-10. WT and KI denote ERα wild type and KI mice, respectively. ONE Way ANOVA plus post hoc test with Tukey-Kramer’s multiple comparisons test (Version 5.0, Stat view-j) was used for statistical analysis. Values are presented as means ± S.D.. **, p < 0.01, n = 4. b FACS. Mixed glia cells were separately prepared from male and female neonates, cultured for 2 weeks and then treated with 100 ng/mL of LPS or PBS for an additional 2 or 24 h. IL-6 production was determined by intracellular staining, followed by flow cytometry analysis. Assay was triplicated with three independent. ONE Way ANOVA plus post hoc test with Tukey-Kramer’s multiple comparisons test (Version 5.0, Stat view-j) was used for statistical analysis. Values are presented as means ± S.D.. **, p < 0.01. c Cell death was assessed by Annexin V staining, following by flow cytometry analysis. Assay was performed in triplicate eight times with three independent samples. These were separately prepared from male and female neonates. Two Way ANOVA plus post hoc test with Tukey-Kramer’s multiple comparisons test (GraphPad Prism) was used for statistical analysis. Values are presented as means ± S.D.. *, p < 0.05. d The gating strategy and the representative FACS plots for b and c