Fig. 10

CXCR3 signaling negatively regulates NF-κB activation induced by IL-17 in a glioblastoma cell line. a U87MG cells were stained with an anti-CXCR3-APC (solid line) or isotype control (dashed line) antibody followed by FACS analysis. b U87MG cells were serum-starved for 6 h followed by stimulation with CXCL10 (100 ng/ml). Proteins in whole-cell lysates, prepared at the indicated times, were resolved by SDS-PAGE followed by immunoblotting with anti-p-ERK and anti-ERK antibodies. c U87MG cells were serum-starved for 6 h and preincubated with or without PD98059 (10 ng/ml) for 15 min followed by incubation with CXCL10 (100 ng/ml) for 60 min. Cells were then stimulated with IL-17 (100 ng/ml) for 15 min followed by cell lysis. Whole-cell lysates were subjected to SDS-PAGE followed by immunoblotting with antibodies to IκB and phospho-IκB. d Similar to b except that cells were stimulated with IL-17 for 30 min and cell lysates were fractionated into cytoplasmic and nuclear fractions. Proteins in whole-cell lysates, cytoplasmic fractions, and nuclear fractions were resolved by SDS-PAGE followed by immunoblotting with antibodies to NF-κB p65, α-tubulin, and PARP. PARP and α-tubulin were used as markers for the nuclear and the cytoplasmic fractions, respectively. e U87MG cells were cultured in complete medium containing IL-17 (100 ng/ml) or IL-17 (100 ng/ml) plus CXCL10 (100 ng/ml) for 48 h, after which CCL20 in culture supernatants was detected by ELISA. Data represent means ± SEM from four independent experiments. *P < 0.05