mTORC1 signaling regulates vascular endothelial function via reactive oxygen species and NFκB signaling
Authors: Reho John J. , Guo Deng‐Fu , Olson Andrew , Rahmouni Kamal 
Published: June 2021

The mechanistic target of rapamycin complex 1 (mTORC1) is an important intracellular energy sensor that regulates protein synthesis through its downstream signaling components the S6‐kinase and the ribosomal S6 protein. Recently, our laboratory has demonstrated a critical role of the mTORC1 signaling pathway in the cardiovascular regulation with implications for obesity and hypertension. In this study, we tested the hypothesis that the mTORC1 signaling pathway in the vasculature is involved in the regulation of vascular endothelial function and that dysregulation of this pathway contributes to the cardiovascular disorders associated with obesity. We began by assessing the effects of activating mTORC1 signaling in cultured mouse lung endothelial cells (MLECs) using leucine (10 mM; 16hrs) or a constitutively active S6‐kinase adenoviral construct (Ad‐S6KCA; 48hrs). Activated mTORC1 signaling was confirmed by the increased phosphorylated levels of the ribosomal S6 protein (2–4 fold; p<0.05). Increasing mTORC1 signaling elevated mRNA expression of oxidative stress markers (NOX1 and NOX2; 1.5–5 fold; p<0.05), decreased mRNA expression of superoxide dismutase 2 (0.5 fold; p<0.05) and increased reactive oxygen species (ROS) generation (via dihydroethidium staining; 1.5 fold; p<0.05) in MLECs demonstrating a pro‐oxidant gene environment evoked by activation of mTORC1 signaling. Blockade of the IKKβ subunit of the NFκB transcriptional complex (BMS‐345541; 300nM) prevented mTORC1 signaling induced ROS generation in MLECs demonstrating a critical role of NFκB signaling in the cellular response to mTORC1 activation. Next, we used leucine and Ad‐S6KCA to test the consequence of enhancing mTORC1 signaling on endothelial function in aortic rings isolated from wildtype mice. Both leucine and an adenovirus expressing a constitutively active form of S6K (Ad‐S6KCA) impaired endothelial dependent acetylcholine‐induced relaxation (~10–15%; p<0.05) without changing endothelial‐independent relaxation responses evoked by sodium nitroprusside indicating endothelial but not smooth muscle dysfunction in response to increased mTORC1 signaling. Blockade of mTORC1 signaling using a dominant negative S6K adenoviral construct (Ad‐S6KDN) as well as inhibition of ROS signaling by Tempol (non‐specific free radical scavenger) ameliorated mTORC1‐induced endothelial dysfunction (p<0.05). To determine the involvement of mTORC1 signaling in the endothelial dysfunction associated with obesity, we utilized diet‐induced obese mice that display vascular endothelial dysfunction as compared to lean controls. Obese mice exhibited increased mTORC1 signaling in the aorta and mesenteric artery indicated by the elevated phosphorylated levels of the ribosomal S6 protein (1.5–3 fold; p<0.05) and this was associated with increases in NOX2 expression (1.5 fold; p<0.05) further indicating ROS signaling in the vascular pathology associated with obesity. We conclude that mTORC1 signaling is a novel regulator of vascular endothelial function through its effects on the NFκB complex and ROS signaling. Our data also indicate that dysregulation of the mTORC1 signaling pathway may be involved in the endothelial dysfunction associated with obesity.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.