Figure 5.

TSAd deficiency attenuates phosphorylation of Src but not VEGFR2 and eNOS pathways in vivo. (A) WT and tsad^−/− mice were injected (i.v.) with VEGF and lungs were harvested 1 min later. Blotting was performed on total lung lysates using antibodies against VEGFR2 phosphorylation sites pY1175 and pY951, VEGFR2 protein, VE-cadherin and ZO-1. Control for equal loading was done by immunoblotting for actin. Molecular weights in kilodaltons (kD) are indicated to the right. Data are shown for one mouse injected with PBS and two mice injected with VEGF. Representative results of at least three independent experiments. n = 3 mice/genotype. (B) Mice were treated as in A, but lungs were harvested after 20 min after injection (i.v.) of PBS or VEGF. Blotting was performed on total lung lysates using antibodies against c-Src pY418 and c-Src protein. Immunoblotting for β2 microglobulin (β2M) was performed to control for equal loading. Molecular weights in kilodaltons (kD) are indicated to the right. Numbers above lanes indicate fold-change in c-Src pY418/ total c-Src protein, set to 1 for the unstimulated WT sample. Representative results of three separate experiments are shown. (C) WT and tsad^−/− mice were injected (i.v.) with PBS or VEGF, and lungs were harvested 1, 10, and 20 min later. Conversion of l-arginine to nitric oxide (NO) in lung tissue was determined using stoichiometric production of l-citrulline. *, P < 0.05. n = 3 mice/genotype for each time point from 1 experiment.