Fig 1: Inflammatory activation of HMC3 human microglia cell line by TNF-a and IFN-?. (a) Flow cytometric analyses demonstrate the increased protein expression of inflammatory microglia biomarkers (IBA-1, CD40, MHC-II, CD68, CD14, and CD86) in the HMC3 cells treated with IFN-? (0.3 µg/mL, 24 h) (red), but no alteration in the expression of the anti-inflammatory microglia biomarkers CD163 and CD206. TNF-a (0.3 µg/mL, 24 h) (blue) also increases the expression of inflammatory microglia biomarkers (IBA-1, CD40, and CD14). (b) q-PCR analysis revealed increased CD11b and TMEM119 expression by HMC3 microglia after addition of 0.3 µg/mL TNF-a or 0.3 µg/mL IFN-? for 24 h. Fold change is normalized to untreated control. GAPDH was used as an internal control to normalize mRNA level of the gene of measurement in each sample (n = 5 independent experiments). Quantitative analysis (c) and representative fluorescence micrographs (d, displayed at ×10 and ×4 magnification) of the cell viability assay show that both TNF-a and IFN-? induce cell death in a concentration-dependent manner (0, 0.01, 0.03, 0.1, 0.3 µg/mL) for 24 h in the HMC3 human microglial cells. Fluorescent green features are AO stains of live cells and fluorescent red features are PI stains of dying cells. *p < 0.1, **p < 0.01, ***p < 0.001, and NS (no significant difference).