Intermittent Hypoxia Aggravates Endothelial Cell Injury Induced By CSE Through The Nrf2/HO-1 Pathway

BackgroundObstructive sleep apnea syndrome(OSAS)is a sleep-related hypoxic disease.It is a clinical syndrome of complete or incomplete obstruction of the upper airway during sleep,resulting in snoring accompanied by apnea and / or hypoventilation,and recurrent hypoxia/ reoxygenation.The disease is a systemic disease,which causes multiple systems and organs damage,especially in cardiovascular complications.Some studies have confirmed that smoking,just like OSAS,is also an independent risk factor for atherosclerosis and death.The damage of cigarette smoking extract(CSE)on vascular endothelial cells initiates the process of atherosclerotic plaque formation,and it can lead to plaque rupture and thrombosis.The incidence of cardiovascular disease in patients with overlap syndrome(OS)caused by the combination of smoking with OSAS has increased compared with patients with simple OSAS or COPD.It has been reported that intermittent hypoxia and CSE can cause damage to the function of endothelial cells.However,whether the overlap of OSAS and COPD will cause aggravation of vascular endothelial function damage is rarely reported,and the mechanism is unknown.Oxidative stress is one of the most important pathophysiological mechanisms of endothelial dysfunction.According to the researches,the Nrf2 / HO-1 pathway plays an important role in the anti-oxidative stress of endothelial cells.Therefore,we hypothesize that intermittent hypoxia aggravates the damage of CSE to endothelial cell function,and is regulated by the Nrf2 / HO-1 pathway.Based on the hypothesis,we established the model of HUVECs exposed to intermittent hypoxia and CSE at the same time to determine whether IH aggravates the endothelial function damage caused by CSE.Simultaneously,we clarified the effect of the Nrf2 / HO-1pathway on endothelial cell damage under the overlap effect,so as to provide a theoretical basis for the diagnosis and treatment of cardiovascular loss in overlapping syndrome.Purpose:1.To study the effects of different exposure time and different concentrations of CSE on the apoptosis of HUVECs.2.To determine whether intermittent hypoxia aggravates CSE and induces apoptosis of HUVECs.3.To clarify the effect of the antioxidant pathway Nrf2 / HO-1 pathway on the apoptosis of HUVECs under the overlapping effect.Methods.1.We established a model of human umbilical vein endothelial cells exposed to CSE at different concentrations and at different durations to clarify the effect of CSE on the apoptosis rate and survival rate of HUVECs through cell CCK-8 experiment and flow cytometry experiment.2.We established a model of human umbilical vein endothelial cells exposed to intermittent hypoxia combined with tobacco to determine whether intermittent hypoxia aggravates the effect of tobacco extracts on the apoptosis rate and survival rate of human umbilical vein endothelial cells by cell CCK-8 experiment and flow cytometry experiment.3.We constructed a human umbilical vein endothelial cell model with Nrf2 knockdown and overexpression.Through flow cytometry and CCK-8 experiments,we clearly defined the effect of the Nrf2 / HO-1pathway on apoptosis and proliferation under overlapping effects.Meanwhile,we used Western Bolt Western blotting experiments and real-time Q-PCR to clarify the changes in the Nrf2 / HO-1 pathway.Results1.CSE increased the apoptosis rate of human umbilical vein endothelial cells and decreased its survival rate(p<0.05).At 2.5%,5%,and10% concentrations,the effect of CSE on the apoptosis rate of endothelial cells was most significant at 8 hours(p<0.05).2.Intermittent hypoxia significantly increased the apoptosis rate(p<0.05)of HUVECs exposed to CSE,and reduced its survival rate(p<0.05).3.Under the overlapping effect,Nrf2 of human umbilical vein endothelial cells was further reduced than that of pure intermittent hypoxia and tobacco exposure(p <0.05),and HO-1 increased(p <0.05).In the HUVECs model with Nrf2 knockdown,the cell survival rate was lower than that of normal cells(p <0.05),and the apoptosis rate was increased(p <0.05).The expression level of downstream protein HO-1 was normal to that of Nrf2.The overlap effect group was significantly decreased(p <0.05),and there was no difference in expression from the non-intervention group(p> 0.05).In the Nrf2 overexpression model,cell survival rate increased(p <0.05),and apoptosis rate decreased(p <0.05).Conclusion1.CSE has a significant damaging effect on endothelial cells,which is time-dependent.2.Intermittent hypoxia can significantly increase the damage of CSE to endothelial cells.3.Intermittent hypoxia and tobacco cause endothelial cell damage by inhibiting the Nrf2 / HO-1 pathway.Activation of the Nrf2 / HO-1pathway can alleviate endothelial cell damage.