Fig. 6

AM-NPs suppress Aβ-mediated microglial activation. a–c BV2 microglia were treated with 20 µM Aβ in the presence or absence of NPs or 10 ng/ml lipopolysaccharide (LPS). After 24 h, supernatants were collected and assayed for inflammatory markers. Cells were fixed with 4% PFA and then permeabilized in 0.5% Triton-X-100 in potassium-buffered saline (PBS-T). After blocking, cells were incubated with anti-iNOS antibody overnight at 4 °C, then washed with PBS-T. Cells were incubated with secondary antibody Alex 594 for 1 h. a Confocal images representing the expression of inducible nitric oxide synthase (iNOS) in BV2 cells after 24-h co-incubation with fAβ and NPs. Scale bar, 25 µm. b Quantitative analysis of fluorescence intensity of iNOS in BV2 cells after 24-h co-incubation with fAβ and NPs. Data are presented as mean ± SEM; n = 3; ***P = 0.0002 for T₁₂P₅ versus fAβ, ***P = 0.0001 for M₁₂P₅ versus fAβ for Dunnett’s multiple comparisons shown on graph by one-way ANOVA. c Nitrite concentration measured by Griess reagent in the conditioned media harvested 24 h after cell co-incubation with Aβ and NPs. Data are presented as mean ± SEM; n = 3; **P = 0.0057 for T₁₂P₅ versus fAβ, **P = 0.0048 for M₁₂P₅ versus fAβ for Dunnett’s multiple comparisons shown on graph by one-way ANOVA. d Concentration of TNFα measured by ELISA in the conditioned media harvested 24 h after cell co-incubation with fAβ and NPs. Data are presented as mean ± SEM; n = 3; **P = 0.042 for T₁₂P₅ versus fAβ, **P = 0.0068 for M₁₂P₅ versus fAβ for Dunnett’s multiple comparisons shown on graph by one-way ANOVA. e SH-SY5Y cells were treated with conditioned media of  fAβ- or LPS-stimulated BV2 cells in the presence or absence of NPs. Cytotoxicity in response to stimulated BV2 conditioned media was quantified in SH-SY5Y media using LDH assay. Data are presented as mean ± SEM; n = 3; ***P = 0.003 for T₁₂P₅ versus fAβ and **P = 0.0027 for M₁₂P₅ versus fAβ for Dunnett’s multiple comparisons shown on graph by one-way ANOVA