Fig. 3

Impaired TLR-4 signaling in R6/1 BMMCs. WT and R6/1 BMMCs were treated with LPS (500 ng/mL), at 0, 15, 30, and 60 min after stimulus, the cells were isolated, and the phosphorylation of specific proteins was determined by Western blot. a Time-course of IKK, b ERK1/2, c p65, and d ELK-1 activation. e Time-course of c-fos mRNA accumulation in FR180204-treated WT BMMCs. Two-million WT BMMCs were pre-incubated for 15 min with FR180204 (10 μM) and stimulated with LPS (500 ng/mL). Total RNA was isolated at the indicated times and RT-PCR was performed. f Time-course of LPS-induced c-fos mRNA accumulation in WT and R6/1 BMMCs. The cells were treated with 500 ng/mL of LPS and total RNA was isolated at different times to perform RT-PCR. Data shown in the upper part of all panels are from a representative experiment, whereas the lower part shows the results of at least three independent experiments performed with different cultures. Data are presented as the mean ± SD, *p < 0.05, **p < 0.01, ***p < 0.001 vs. WT values; ^&p < 0.05, ^&&p < 0.01, ^&&&p < 0.001 vs. to the value at time zero or in non-stimulated cells; #p < 0.05, ##p < 0.01, ###p < 0.001 vs. vehicle-treated cells, as determined by two-way ANOVA followed by Tukey’s test