Endoplasmic Reticulum Stress Contributes To Endothelial Dysfunction In Intermittent Hypoxia Mice

Background Obstructive sleep apnea syndrome(OSAS)is a highly prevalent sleep disorders that occurs recurrently obstructive hypoventilation or apnea during sleep.Intermittent hypoxia(IH)considered to be an important mechanism of the injury of OSAS is one of the key risks of cardiovascular diseases.Numerous pathophysiologic stimuli such as oxidative stress,ischemia and hypoxia may cause endoplasmic reticulum stress(ERS),which is involved in OSAS related diseases such as atherosclerosis,diabetes and ischemic cardiomyopathy.Vascular endothelial dysfunction induced by OSAS is associated with endothelial cell apoptosis.Besides the cell death receptors and mitochondria-dependent apoptotic pathways,ERS is thought to be the pathway that can induce endothelial cell apoptosis.Therefore,we hypothesized that ERS can lead to vascular dysfunction induced by IH.Aim1.To establish the IH mouse model,and to observe the effects of IH and ERS inhibitor on the blood pressure and vascular diastolic function in mice.2.To establish IH cell model and explore the effects of different degrees of IH on cell injury.To explore the mechanism of ERS in endothelial cell apoptosis induced by IH.To provide the evidence of the prevention and treatment of vascular endothelial dysfunction induced by IH.MethodsThe male healthy Kunming mice were randomly divided into control group,IH group and 4-phenylbutyric acid(4-PBA)treatment group.Tail artery systolic blood pressure was measured noninvasive by the tail cuff system.The changes of aorta endotheliumdependent relaxation was measured by isolated vascular tension.the expressions of GRP78 and CHOP protein in aortic endothelial cells was detected by Western Bolt.Human umbilical vein endothelial cells(HUVECs)were randomly divided into normal control(NC)group,IH 6 h group,IH 12 h group and IH 24 h group.Each group was treated with endoplasmic reticulum stress inhibitors(4-PBA).Cell viability was detected by Cell Counting Kit-8(CCK-8),and cell apoptosis was detected by flow cytometry.The content of e NOS was measured by ELISA.1.Role of ERS in the increased caudal artery systolic pressure induced by IH.The male Kunming mice were randomly divided into control group,IH group and IH+4-PBA group.Put IH and IH+4-PBA groups into IH chamber(8 h/d).4-PBA(100mg/kg/d)was injected into the abdominal cavity of IH+4-PBA mice in the last week.Control group was remaining the same conditions except IH.After 28 days of IH,tail artery systolic blood pressure of the three groups mice was measured by the tail cuff system.2.Role of ERS in aorta endothelial dysfunction of mice induced by IH.The changes of endothelium dependent and endothelium independent relaxation of three groups in thoracic aorta were detected by the method of isolated vascular tension.3.The effect of IH on the expression of ERS related protein in thoracic aorta of mice.The relative expression level of GRP78 and CHOP protein was detected by Westernblot in thoracic aorta endothelial cells of three groups.4.Role of ERS on the effect of IH on the viability of HUVECs.HUVECs after 12 h culture were divided into IH 0 h group,IH 6 h group,IH 12 h group and IH 24 h group in serum-free medium,and each group was treated with 4-PBA(2mmol/L).Cell viability was detected by CCK8 to observe the influence of IH and 4-PBA.5.Effect of ERS on the apoptosis of HUVECs induced by IH.The HUVECs were divided into IH 0 h group,IH 6 h group,IH 12 h group and IH 24 h group,and each group was treated with 4-PBA,and cell apoptosis was detected by flow cytometry.6.Role of ERS on effect of IH on the content of e NOS.The content of endothelial nitric oxide synthase(e NOS)in each cells group was measured by ELISA to observe the effects of IH and 4-PBA on the content of e NOS in endothelial cells.Results1.IH can significantly increase systolic blood pressure in mice,but the effect was inhibited by 4-PBA.2.IH can reduce the endothelium dependent relaxation of thoracic aorta in mice,and4-PBA can increase the endothelium dependent relaxation of thoracic aorta in IH mic.There is no significant change of the three groups in endothelium independent relaxation.3.The level of GRP78 in aorta tissue was decreased in IH and increased in 4-PBA compared with IH.The protein CHOP was increased in IH mice and decreased in4-PBA treated mice.4.IH could decrease the cell viability,but the effect was inhibited by 4-PBA.5.There is no change on cell apoptosis induced by IH 6 h.Prolonged IH(12 h and 24h)increased cell apoptosis,and 4-PBA could inhibit the apoptosis induced by IH.6.Prolonged IH(12 h and 24 h)could decrease the content of e NOS in endothelial cells,and the decrease of e NOS was inhibited by 4-PBA.Conclusion IH can cause blood pressure to rise induced by causing endothelial cells ERS,an increased level of apoptosis,the content of e NOS and vasodilatation function decline.