ES proteins elicit Th2-related chemokine production in the absence of protease inhibition.

<p>(A) ES proteins were treated with various protease inhibitors. The samples were incubated for 2 hr and were assayed by zymography on 0.1% gelatin SDS-PAGE gels (lane 1: 10 μg ES proteins; lane 2: with 1 mM PMSF; lane 3: with 5 mM PMSF; lane 4: with 10 μM E-64; lane 5: with 50 μM E-64; lane 6: with 10 μM MMP-9 inhibitor; lane 7: with 50 μM MMP-9 inhibitor; arrowhead: protease activity of ES proteins; arrow: protease activity inhibited by E64). (B) Th2-related chemokine gene expression (<i>TSLP</i>, <i>TARC</i>, <i>MDC</i>, eotaxin, and <i>IL-25</i>) was measured in MLE12 cells after incubation with 1 μg/ml of ES proteins (ES) for 2 hr or pre-treatment with 0.1 mM PMSF and 10 μg/ml polymyxin B (polymyxin) for 2 hr. (C) The fold-change in PAR mRNA levels in MLE12 cells treated with ES proteins relative to those treated with medium, as detected by real-time RT-PCR. (*<i>p</i><0.05, **<i>p</i><0.01, ***<i>p</i><0.001; three independent experiments).