Fig. 4

Immune functions of brain-resident astrocytes and microglia are not affected by direct FXR activation or supernatant of FXR-activated BMMs. The direct treatment with GW4064 (a, b, d, g, h, j) and the incubation with conditioned BMM supernatant of astrocytes (e, f) and ESdMs (k, l) do not suppress secretion of TNFα and NO. ESdMs and primary astrocytes were treated with different concentrations of GW4064 for 7 days. RNA was isolated after 6 h of stimulation with LPS and IFNγ. Differential expression of genes was assessed by performing RT² Profiler PCR Assay (c, d, i, j). After stimulation, a strong differential expression of genes in astrocytes (c) and ESdM (i) compared to unstimulated cells was detectable. However, treatment with GW4064 had no influence on the gene expression of either astrocytes (d) or ESdMs (j). Supernatants of pretreated BMMs were collected after 3 days of culturing and were transferred to ESdMs or astrocytes. Secreted amount of TNFα was determined by ELISA after stimulation with LPS and IFNγ for 24 h (g, k: ESdMs) or 72 h (a, e: astrocytes). Concentration of produced NO was detected by Griess assay after 48 h of stimulation of ESdMs (h, l) and after 72 h in the supernatant of astrocytes (b, f). Bar graphs: data are represented as mean ± SEM. Shown is one representative experiment of at least three independent experiments with triplets for each condition, two-tailed Student’s t test. Scatter plots: data are represented as logarithmic average fold change. Shown are data from two biological replicates