IL-1β drives a cytokine/chemokine expression profile in mouse and human blood–brain barrier/BSCB ECs predicted to favor neutrophil and monocyte activities. (A) Cytokine levels in conditioned media collected from primary BMECs treated with IL-1β or TNF + IFN-γ as measured using a multiplex ELISA cytokine/chemokine array. Differences compared with the control condition, i.e., culture medium in the absence of cytokine treatment, are expressed in ng/ml for cells stimulated with either 10 ng/ml of recombinant mouse IL-1β (black bars; n = 3) or a combination of TNF and IFN-γ (gray bars; n = 3). (B–D) The combination of immunohistochemistry for the endothelial marker CD31 with ISH for the detection of G-CSF (B), GM-CSF (C), and Il-6 (D) mRNAs in the spinal cord of C57BL/6 mice at EAE onset is shown (n = 4). (E–I) Cytokines released by primary human BMECs after treatment with either 10 ng/ml of recombinant human IL-1β (black bars) or 100 U TNF + IFN-γ (gray bars). Levels of G-CSF (E) GM-CSF (F), IL-6 (G), CXCL8 (H), and CCL2 (I) are means ± SEM of experiments from two to five different primary cultures. (J and K) Immunohistochemistry against endothelial CD31 was combined with ISH for G-CSF (J) and Il-6 (K) mRNAs in spinal cord tissue sections from C57BL/6 mice (n = 4/group) that received a single subdural injection of recombinant IL-1β (J and K) or saline (not depicted). *, P < 0.05; **, P < 0.01; ***, P < 0.001 compared with unstimulated cells; ^&, P < 0.05; ^&&, P < 0.01 compared with TNF + IFN-γ treatment; one-way ANOVA followed by Dunnett’s post-hoc test (A and E–I). Bars, 25 µm.