Fig 2: Apelin induces the expression of cytoskeleton and tight junction proteins in C57/BL6 mice via PI-3K/Akt and MAPK/Erk signaling pathways. (a–e) The western blot results found that the expression of APJ and the phosphorylation of cytoskeleton (VE-Cadherin and FAK) and tight junction proteins (occludin and ZO-1) increased significantly in apelin-treated (10 and 100 ng/ml) C57BL/6J mice. (f–h) The phosphorylation of PI-3K/Akt and MEK/Erk signaling pathways, p38, Akt, and Erk also increased significantly. Protein intensity was quantified by Image J software and expressed as fold of change relative to control (mean ± SD, n = 3). ∗∗∗P < 0.001, ∗∗P < 0.01 versus control one, Student's t-test.

Fig 3: Inhibition of apelin receptor APJ using F13A decreased cell proliferation, migration, and the expression of cytoskeleton and tight junction proteins in HRMECs under high glucose condition. (a–c) According to the MTS and transwell assay results, the HRMECs proliferation and migration capability were decreased in a dose-dependent manner, significantly lower in the F13A group, compared with the control one, and the optimum concentration was 20 ng/ml. (d-e) IF staining showed that the phosphorylation of occludin and ZO-1 was downregulated after being treated with F13A in HRMECs under high glucose condition. Cells were stained with DAPI for visualization of nuclei (blue). Scale bar = 50 μm for (d and e). (f-g) The western blot results showed that the phosphorylation of cytoskeleton and tight junction were also downregulated in a dose-dependent way after being stimulated with F13A. Protein intensity was quantified by Image J software and expressed as fold of change relative to control (mean ± SD, n = 3). ∗∗∗P < 0.001, ∗∗P < 0.01 versus control one, Student's t-test.