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. 2012 Mar 20;287(20):16121–16131. doi: 10.1074/jbc.M112.348383

FIGURE 3.

FIGURE 3.

TLR4 activation by triacylated compounds using target TLR4-tranfected HEK293 cells. A, activation of TLR4/MD-2/CD14(+)HEK293 cells by a series of triacylated compounds. Each triacylated compound was tested with a dose response ranging from 10 μg/ml up to 1.37 ng/ml, diluted 1:3 in RPMI. LPS (starting concentration 1 μg/ml, diluted 1:3) served as a positive control (TLR4 agonist), and PAM3CSK4 (TLR2 agonist) was used as a negative control in this assay. IL-8 production was measured in conditioned supernatants after 24 h of incubation time. Only OM-197-MP-AC was a (weak) TLR4 agonist in this assay. Data are expressed as mean ± S.D. from triplicates for each point. This is a representative experiment of three experiments with similar results. B, same experiment but with TLR4/MD-2/CD14(+)HEK293 Bluetm cells expressing alkaline phosphatase under the control of TLR4 activation. Again, only OM-197-MP-AC was a (weak) TLR4 agonist in this assay. Alkaline phosphatase was measured in conditioned supernatants after 24 h of incubation time using an alkaline substrate in a colorimetric assay. This is a representative experiment of three experiments with similar results.