Inhibition of Rho proteins blocks protein tyrosine phosphorylation in LPS-stimulated endothelial cells.

(A) Cells were preexposed to 10 ng/mL TcdB-10463 for 60 minutes as indicated and stimulated with 100 ng/mL LPS for 15 minutes. Cell lysates were obtained as described in “Materials and methods” and were subjected to Western blot analysis (7.5% SDS-PAGE). Membranes were labeled with antibodies against tyrosine-phosphorylated proteins. Note reduction of protein tyrosine phosphorylation in LPS-treated cells after inhibition of Rho proteins with TcdB-10463. (B) Endothelial cells were incubated with 25nM to 500nM geldanamycin 30 minutes prior to stimulation with 100 ng/mL LPS for 8 hours. Supernatants were subsequently analyzed by IL-8 ELISA. A representative gel (1 of 3 separate experiments) is shown in (A). Data presented in (B) are mean ± SEM of 3 separate experiments.