FIG. 1.

Effects of 3,5,3′-triiodothyronine (T₃) on AKT and endothelial nitric oxide synthase (eNOS) phosphorylation. Human umbilical vein endothelial cells (HUVECs) were treated with or without T₃ (10⁻⁷ M) at different times as described in Materials and Methods. Equal amounts of protein were then resolved on 4%–12% sodium dodecyl sulfate-polyacrylamide gel electrophoresis, transferred onto a nitrocellulose membrane, probed with appropriate primary and secondary antibodies, and visualized by enhanced chemiluminescence. A representative western blot from three independent experiments indicates the increase of AKT phosphorylation at serine 473 and eNOS phosphorylation at serine1177 with different fold increases that were time dependent, with maximal activation levels at 20 minutes for phospho-AKT serine 473 (a) and at 60 minutes for phospho-eNOS serine 1177 (c). Quantification of bands was performed by densitometric analysis (b, d). Data are reported as mean±standard deviation (SD) of three independent experiments.