Rapamycin inhibits ox-LDL-induced inflammation in human endothelial cells in vitro by inhibiting the mTORC2/PKC/c-Fos pathway
Abstract
Abstract
Rapamycin and its derivative possess anti-atherosclerosis activity, but its effects on adhesion molecule expression and macrophage adhesion to endothelial cells during atherosclerosis remain unclear. In this study we explored the effects of rapamycin on ox-LDLinduced adhesion molecule expression and macrophage adhesion to endothelial cells in vitro and the underlying mechanisms. Ox-LDL (6–48 μg/mL) dose-dependently increased the protein levels of two adhesion molecules, intercellular adhesion molecule-1 (ICAM- 1) and E-selectin, in human umbilical vein endothelial cells (HUVECs), whereas pretreatment with rapamycin (1–10 μmol/L) dosedependently inhibited ox-LDL-induced increase in the adhesion molecule expression and macrophage adhesion to endothelial cells. Knockdown of mTOR or rictor, rather than raptor, mimicked the effects of rapamycin. Ox-LDL (100 μg/mL) time-dependently increased PKC phosphorylation in HUVECs, which was abolished by rapamycin or rictor siRNA. Pretreatment with PKC inhibitor staurosporine significantly reduced ox-LDL-stimulated adhesion molecule expression and macrophage adhesion to endothelial cells, whereas pretreatment with PKC activator PMA/TPA attenuated the inhibitory effect of rapamycin on adhesion molecule expression. Ox-LDL (100 μg/mL) time-dependently increased c-Fos levels in HUVECs, and pretreatment with rapamycin or rictor siRNA significantly decreased expression of c-Fos. Knockdown of c-Fos antagonized ox-LDL-induced adhesion molecule expression and macrophage adhesion to endothelial cells. Our results demonstrate that rapamycin reduces ox-LDL-stimulated adhesion molecule expression and macrophage adhesion to endothelial cells by inhibiting mTORC2, but not mTORC1, and mTORC2 acts through the PKC/c-Fos signaling pathway.
Keywords:
atherosclerosis; rapamycin; endothelial cells; ox-LDL; ICAM-1; E-selectin; mTORC2; PKC; c-Fos; HUVECs
Rapamycin and its derivative possess anti-atherosclerosis activity, but its effects on adhesion molecule expression and macrophage adhesion to endothelial cells during atherosclerosis remain unclear. In this study we explored the effects of rapamycin on ox-LDLinduced adhesion molecule expression and macrophage adhesion to endothelial cells in vitro and the underlying mechanisms. Ox-LDL (6–48 μg/mL) dose-dependently increased the protein levels of two adhesion molecules, intercellular adhesion molecule-1 (ICAM- 1) and E-selectin, in human umbilical vein endothelial cells (HUVECs), whereas pretreatment with rapamycin (1–10 μmol/L) dosedependently inhibited ox-LDL-induced increase in the adhesion molecule expression and macrophage adhesion to endothelial cells. Knockdown of mTOR or rictor, rather than raptor, mimicked the effects of rapamycin. Ox-LDL (100 μg/mL) time-dependently increased PKC phosphorylation in HUVECs, which was abolished by rapamycin or rictor siRNA. Pretreatment with PKC inhibitor staurosporine significantly reduced ox-LDL-stimulated adhesion molecule expression and macrophage adhesion to endothelial cells, whereas pretreatment with PKC activator PMA/TPA attenuated the inhibitory effect of rapamycin on adhesion molecule expression. Ox-LDL (100 μg/mL) time-dependently increased c-Fos levels in HUVECs, and pretreatment with rapamycin or rictor siRNA significantly decreased expression of c-Fos. Knockdown of c-Fos antagonized ox-LDL-induced adhesion molecule expression and macrophage adhesion to endothelial cells. Our results demonstrate that rapamycin reduces ox-LDL-stimulated adhesion molecule expression and macrophage adhesion to endothelial cells by inhibiting mTORC2, but not mTORC1, and mTORC2 acts through the PKC/c-Fos signaling pathway.