Fig 1: Siponimod modulates microglial cytokine expression and secretion in an inflammatory milieu. (A,C,E,G) quantitative RT-PCR analysis of Tnf (A), Ifnb (C), Il1b (E) and Il10 (G) gene expression after stimulation for 1 day with +/- 50 µM siponimod +/- LPS. (B,D,F,H) Quantification of TNF-a (B), IFNß (D), IL-1ß (F) and IL-10 (H) protein concentration in the cell culture medium of microglial cells stimulated for 3 days with +/- 50 µM siponimod +/- LPS using respective quantitative sandwich ELISA assays. Grey-dotted lines indicate the lowest standard of the ELISA kit: TNFa = 82.3 pg/mL, IFNß = 15.63 pg/mL, IL-1ß = 68.59 pg/mL and IL-10 = 8.23 pg/mL. Data are presented as mean values ± SEM. Grey dots represent individual data points. Significance of gene expression analysis as well as ELISA was assessed by 1-way analysis of variance (ANOVA) followed by Tukey’s post hoc test using Graph-Pad Prism 8.4.3 (GraphPad Software, San Diego, CA, USA). The experimental groups were considered significantly different at ** p < 0.01, *** p < 0.001.