Activation of TLR4 by LPS or cyclic stretch increases intracellular IL-33.

MLE-12 cells were transfected with 50 nM of TLR4-specific siRNA (TLR4siRNA) or nonspecific siRNA control (consiRNA) using Lipofectamine 2000 for 48h before stretch. (A) Transfected MLE-12 cells were further treated with 100 ng/ml LPS or control solution for 24h and IL-33 production were measured in cell lysate by ELISA. (B) IL-33 production after 6h cyclic stretch in transfected cells treated with or without TLR4-specific siRNA. (C) Total TLR4 expression in whole cell lysate and cytoplasm in MLE-12 cells with (cyclic S) or without (con) stretching were measured by Western blot and normalized by β-actin. ***P<0.001 when compared between groups denoted by horizontal lines.