Rutaecarpine Prevents High Glucose-Induced Endothelial Cell Senescence Through Transient Receptor Potential Vanilloid Subtype 1/ SIRT1 Pathway.

ABSTRACT: SIRT1 functions as a longevity factor to counteract vascular aging induced by high glucose. Our previous study revealed that rutaecarpine, the natural agonist of transient receptor potential vanilloid subtype 1 (TRPV1), prevented high glucose-induced endothelial dysfunction. The present study aims to evaluate the effects of rutaecarpine on endothelial cell senescence induced by high glucose, and focus on the regulatory effect on SIRT1 expression. In cultured human umbilical vein endothelial cell (HUVEC), exposure to 33 mM high glucose for 72 hours induced cellular senescence, demonstrated as cell cycle arrest at G0/G1 phase, decreased cell viability, and increased number of senescence-associated ß-galactosidase positive senescence cells and ROS production, which were effectively attenuated by treatment with rutaecarpine (0.3, 1, and 3 µM). Furthermore, rutaecarpine upregulated longevity protein SIRT1 expression in HUVECs, accompanied by decreased level of senescence marker p21. In addition, rutaecarpine increased intracellular calcium level in HUVECs, and pretreatment with TRPV1 antagonist capsazepine, intracellular Ca2+ chelator BAPTA-AM or CaM antagonist W-7 abolished the effects of rutaecarpine on SIRT1 expression. In summary, this study shows that rutaecarpine upregulates SIRT1 expression and prevents high glucose-induced endothelial cell senescence, which is related to activation of TRPV1/[Ca2+]i/CaM signal pathway. Our findings provide evidence that rutaecarpine may be a promising candidate with a novel mechanism in prevention vascular aging in diabetes.

Capsaicin ameliorates intermittent high glucose-mediated endothelial senescence via the TRPV1/SIRT1 pathway.

BACKGROUND: Patients with diabetes have accelerated vascular aging when compared with healthy individuals. Hyperglycemia, especially intermittent high glucose (IHG), is the main cause of vascular endothelial senescence. Capsaicin, a major component of chili pepper is thought to contribute to cardiovascular protection by spicy food. OBJECTIVE: To investigate the pathway related with the effects of capsaicin on endothelial cell senescence induced by IHG. METHODS: HUVECs were exposed to IHG (5 mM or 33 mM glucose, alternating every 12 hours for 3 days) and treated with capsaicin at 0.3, 1 and 3 µM. To determine endothelial cell senescence, we examined the senescence-related ß-galactosidase staining, cell cycle arrest, cell viability, as well as production of reactive oxygen species (ROS). To evaluate the involvement of TRPV1/[Ca2+]i/CaMKII/AMPK/SIRT1 pathway in anti- senescence effects of capsaicin, HUVECs were treated with CAPZ (a TRPV1 antagonist), BAPTA-AM (an intracellular calcium chelator), KN62 (a CaMKII antagonist), compound C (an AMPK inhibitor), or EX527 (a SIRT1 inhibitor). To knockdown TRPV1, HUVECs were transfected with shRNA lentivirus targeting TRPV1. The levels of SIRT1, p21, TRPV1, AMPK and phospho-AMPK were evaluated by western blotting. RESULTS: IHG suppressed the levels of SIRT1 and enhanced endothelial senescence. Capsaicin upregulated SIRT1 expression and downregulated the senescence marker, p21, thereby protecting endothelial cells from IHG-induced senescence as indicated by relieved G0/G1 phase arrest, improved cell viabilities, and reduced counts of senescent cells and ROS production. Pre-treatment with CAPZ, BAPTA-AM, KN62 or compound C abrogated the anti-senescence effects of capsaicin. Capsaicin restored AMPK phosphorylation and IHG-inhibited TRPV1 expression. Moreover, TRPV1 silencing suppressed SIRT1 expression and abolished the anti-senescence effects of capsaicin. CONCLUSION: Capsaicin elevates SIRT1 levels through TRPV1/[Ca2+]i/CaMKII/AMPK pathway and suppresses IHG-mediated endothelial cell senescence. This study provides initial evidence that capsaicin is a potential candidate for the prevention of vascular aging in diabetes.