3.5 The effect of Twist1 inhibitor on the distribution and expression of VE-cadherin in primary HUVECs under normal or hyperoxia conditions
There are three types of connections between endothelial cells: adhesive connections, tight connections and gap connections; however, adhesive connection are the most important structure with which to control vascular permeability24. Adhesive connections are mainly composed of vascular endothelial cadherin (VE-cadherin) complex and serve to maintain the barrier function and permeability of vascular endothelial cells25,26. Therefore, after reducing the expression of Twist1, the expression and distribution of VE-cadherin protein in HUVECs was detected by immunofluorescence cytochemistry. Laser confocal immunofluorescence (Figure 4A) showed that VE-cadherin was mainly expressed in the cell membranes of HUVECs, although a small amount was detected in the cytoplasm. The expression of this protein increased significantly at the adhesion junction between cells. As shown in Figure 4A, 4B, and 4C, there was high levels of VE-cadherin expression on the cell membranes of HUVECs in the control group. Furthermore, the staining of cell boundaries was clear, the intercellular connections were close, and the immunofluorescence intensity was( 9.21±2.11 A.U.). Compared with the control group, the expression of VE-cadherin in the cell membranes of HUVECs in the normoxic+harmine group and hyperoxia group was decreased, the cell boundary showed a discontinuous distribution, there was a weak connection between cells, and the immunofluorescence intensities respectively were (2.54±0.31 A.U.) and (2.19±0.46 A.U.). In contrast, compared with the hyperoxia group, the expression of VE-cadherin in the cell membranes of HUVECS in the hyperoxia+ harmine group was increased and red fluorescence was distributed continuously along the cell membrane, the cell boundary was clear and tightly connected, and the immunofluorescence intensity was (8.51±1.46 A.U.).