Resveratrol attenuates cigarette smoke induced endothelial apoptosis by activating Notch1 signaling mediated autophagy.

BACKGROUND: Increasing evidence shows that endothelial apoptosis contributes to cigarette smoke (CS)-induced disease progression, such as chronic obstructive pulmonary disease (COPD). Our previous studies have validated Notch1 as an anti-apoptotic signaling in CS-induced endothelial apoptosis. Resveratrol (RESV) is a naturally occurring polyphenol that exhibits an anti-apoptotic activity in endothelial cells that exposed to many kinds of destructive stimulus. However, the effects of resveratrol on Notch1 signaling in CS-induced endothelial apoptosis have not yet been fully elucidated. Therefore, the aim of this study was to examine whether RESV can protect endothelial cells from CS-induced apoptosis via regulating Notch1 signaling. METHODS: Human umbilical vein endothelial cells (HUVECs) were pretreated with RESV for 2 h, followed by cotreatment with 2.5%CSE for 24 h to explore the role of RESV in CSE induced endothelial apoptosis. 3-methyladenine (3-MA) or rapamycin was used to alter autophagic levels. Lentivirus Notch1 intracellular domain (LV-N1ICD), γ-secretase inhibitor (DAPT) and Notch1 siRNA were used to change Notch1 expression. The expression of Notch1, autophagic and apoptotic markers were examined by Western blot and the apoptosis rate was detected by Flow cytometry analysis. RESULTS: Our results showed that activating autophagy reduced CSE-induced endothelial apoptosis, while blocking autophagy promoted cell apoptosis in HUVECs. RESV pretreatment attenuated the CSE-induced endothelial apoptosis and activated Notch1 signaling. RESV pretreatment also increased LC3b-II and Beclin1 production, decreased p62 and mTOR expression. 3-MA treatment inhibited autophagy and aggravated CSE induced apoptosis, while rapamycin promoted autophagy, led to a decrease in cell apoptosis. LV-N1ICD transfection upregulated autophagy and reduced apoptosis. However, this protective effect was abolished by 3-MA treatment. In cells treated with DAPT or Notch1 siRNA, autophagy was decreased, while apoptosis was increased. RESV partly rescued the DAPT or Notch1 siRNA induced apoptosis by activating Notch1 signaling. CONCLUSION: In HUVECs, RESV attenuates CSE induced endothelial apoptosis by inducing autophagy in a Notch1-dependent manner.

miR-34a is involved in CSE-induced apoptosis of human pulmonary microvascular endothelial cells by targeting Notch-1 receptor protein.

BACKGROUND: Abnormal apoptosis of lung endothelial cells has been observed in emphysematous lung tissue and has been suggested to be an important upstream event in the pathogenesis of chronic obstructive pulmonary disease (COPD). Studies have shown that microRNAs (miRNAs) contribute to the pathogenesis of pulmonary diseases by regulating cell apoptosis. The present study was designed to investigate the expression of microRNA-34a (miR-34a) in human pulmonary microvascular endothelial cells (HPMECs) exposed to cigarette smoke extract (CSE), and the potential regulatory role of miR-34a in endothelial cell apoptosis. RESULTS: Our results showed that the expression of miR-34a was significantly increased in CSE-treated HPMECs, and inhibiting miR-34a attenuated CSE-induced HPMEC apoptosis. Furthermore, expression of Notch-1, a receptor protein in the Notch signalling pathway, was decreased and was inversely correlated with miR-34a expression in HPMECs treated with CSE. Computational miRNA target prediction confirmed that Notch-1 is a target of miR-34a. Luciferase reporter assay further confirmed the direct interaction between miR-34a and the 3'-untranslated region (UTR) of Notch-1. Restoration of Notch-1 pathway was able to partially block the effect of miR-34a on HPMEC apoptosis. These results indicate that Notch-1 is a critical downstream target of miR-34a in regulating the CSE-induced HPMEC apoptosis. CONCLUSIONS: Our results suggest that miR-34a plays a key role in CSE-induced endothelial cell apoptosis by directly regulating its target gene Notch-1 in endothelial cells.