Figure 8.

FK506+minocycline treatment effectively leads to reduced NF-kB p65 and increased NRF2 nuclear translocation. (A) Representative slides stained with NF-kB p65 adopting immunohistochemistry technique. Top three slides are low power (50 μm) while bottom three slides are high power (20 μm). Graphical presentation at the bottom showing expression of NF-kB p65 in prion infected and non infected groups. Slides from 5 animals each per group were selected for statistical analysis. The data was analyzed using one way ANOVA test followed by post hoc test tukey’s multiple comparison. (P ≤ 0.05 = * and P < 0.0001 = ***). (MOD= mean optical density). (B) Representative pictures of confocal microscopy for nuclear translocation of NRF2 in different groups, First panel stained for nucleus with DAPI (blue), second panel stained for NRF2 (red), third panel is merge of first and second panel. (scale bar-40 μm). The data was analyzed using one way ANOVA test followed by post hoc test tukey’s multiple comparison. (P ≤ 0.01 = ** and P < 0.001 = ***) (AFU= arbitrary fluorescence units). (C) Representative western blot panel of two animals each per group for the protein expression level of NFKB p65 and NRF2 in cytoplasmic extracts (upper panel). (D,E) The data showing NF-kB p65 and NRF2 protein expression levels in cytoplasm and nucleus of different experimental groups based on 5 animals per group. Values were normalized using beta actin and nucleophosmin as loading controls. The data was analyzed by using 2 way ANOVA test followed by bonferroni post hoc test. (P ≤ 0.05 = *, P < 0.001 = *** and P > 0.05 = ns). (F) Representative blots of two animals each per group for the expression of HO-1 in prion infected and non infected animals. (G) Graphical presentation of HO-1in the brain homogenates of 5 animals each per group. Values were normalized using beta actin as loading control. The data was analyzed by using one way ANOVA test followed by post hoc test tukey’s multiple comparison. (P < 0.0001 = ***).