Extended Data Fig. 8: IL-18 suppresses eNOS activity in RAEC cells.
From: Reversal of prolonged obesity-associated cerebrovascular dysfunction by inhibiting microglial Tak1
a, Quantitative RT-PCR analysis of the mRNA levels of cytokines and chemokines in the brainstem of chow- or HFD-fed mice. au, arbitrary unit. n = 6 (IL-1β, HFD or IL-23α, Chow) or 7 (all other groups) mice per group. b, Immunofluorescent staining for iNOS (red) and Iba1 (green). Tissue sections were counterstained with DAPI (blue) to visualize cell nuclei. Brainstem region close to basilar artery is shown. Scale bars, 20 µm. n = 3 mice per group. c, Percentage of iNOS+ microglia in chow- or HFD-fed mice. n = 3 mice per group. d, Rat aortic endothelial cells (RAECs) were incubated with vehicle (Ctrl) or IL-18 (20 ng ml-1) for 4 hrs. Total proteins were prepared and subjected to western blot analysis. β-Actin was used as a loading control. n = 3 cell cultures per group. e, RAECs were transfected with scramble RNA (Ctrl) or siRNA targeting IL-18Rα. The cells were further cultured for 4 days and then harvested for protein preparation. The proteins were subjected to western blot analysis. β-Actin was used as a loading control. n = 3 cell cultures per group. f, Quantification of the immunoblots shown in d and e. For p-eNOS/eNOS, *P = 0.005, t(4) = 5.55, Ctrl versus IL-18; or P = 0.03, t(4) = 3.3, Ctrl versus IL-18Rα siRNA. For IL-18Rα/β-Actin, *P = 0.0002, t(4) = 13.95. n = 3 cell cultures per group. Uncropped western blots can be found in Source Data Extended Data Fig. 8. Data are presented as mean ± SEM. Two-tailed Student’s t-test was used for statistical analysis (f).
