High Glucose Environment Regulates The Interaction Between VE-cadherin And Orais In Mouse Aortic Endothelial Cells And Its Mechanism

Background: Injury of vascular endothelial barrier function is an early sign of diabetic vascular lesions,and injury of intercellular adhesion junction（AJ）can cause increased endothelial permeability.VE-cadherin is a specific adhesion linker in vascular endothelium.it has been reported that abnormal phosphorylation of adhesion connexin alters the connectivity between VE-cadherin and leads to changes in the permeability between endothelial cells,which is an important factor affecting the endothelial cells barrier.Store operated Ca²⁺ entry（SOCE）is a hot topic in calcium signaling.Ca²⁺-release-activated Ca²⁺ channel proteins（Orais）are key molecular proteins involved in the SOCE process,which affects intercellular permeability.Objective: To study the role and molecular mechanism of Orais-VE-cadherin signaling complex and its downstream signaling pathway in diabetic endothelial injury using mouse aortic endothelial cells（MAECs）.Methods: The activity of SOCE was detected by calcium imaging after 7 days of high glucose and normal sugar medium,the expression of Orais after high glucose treatment was detected by western blotting.The effect of high glucose treatment on the expression of ve-cadherin and p-VE-cadherin on cell membrane was observed by immunofluorescence test.The interaction between Orais and VE-cadherin was detected by immunoblotting method.Immunohistochemical experiments were used to detect the expression changes of Orais p-VE-cadherin in diabetic mice aortic endothelial.Results:（1）The expression level of Orais proteins were significantly increased and SOCE activity was significantly increased in MAECs cultured at high glucose for 7 days.（2）In MAECs cultured at high glucose for 7 days,the ratio ofVE-cadherin/p-VE-cadherin expressed on the cell membrane increased,indicating that p-VE-cadherin expression increased and intercellular permeability increased.（3）Orai 1,2 or 3 and VE-cadherin can interact and enhance the interaction ratio through high-glucose stimulation.（4）In MAECs cultured with high glucose,the SOCE activator ATP enhanced the expression level of p-VE-cadherin protein,and the SOCE inhibitor BTP2 decreased the expression level of p-VE-cadherin protein.（5）Significantly increased expression of p-VE-cadherin and Orais proteins in the aortic endothelium in diabetic mice.Conclusions:（1）Orais internalized VE-cadherin phosphorylation and increased endothelial permeability by co-distribution of calcium signaling complexes with VE-cadherin in cells in contact with each other;（2）High glucose environment stimulated increased expression of Orais protein S in endothelial cells,and then increased VE-cadherin phosphorylation through Orais-VE-cadherin complex and a series of downstream signaling pathways,resulting in disruption of endothelial cell junctions and initiation of atherosclerosis;（3）The interaction and expression of VE-cadherin complexes in type 2 diabetes mellitus can regulate the function of VE-cadherin junctions in endothelial cells to slow down the permeability of atherosclerotic lesions.