Fig 2: Mechanism map was shown. Choroidal vascular endothelial cells (CVECs) release CSF1 under hypoxia. CSF1 binds to CSF1R on macrophages to activate the PI3K/AKT/FOXO1 pathway, which facilitates macrophage migration and M2 polarization. M2-type macrophages enhance the proliferation, migration and tube formation of CVECs in a CSF1/CSF1R-dependent manner to promote the formation of choroidal neovascularization.

Fig 3: CSF1 released from human choroidal vascular endothelial cells (HCVECs) promotes macrophage migration and M2 polarization via the CSF1R/PI3K/AKT/FOXO1 pathway. Macrophages were divided into normal control, human CSF1 recombinant protein, human CSF1 recombinant protein + pexidartinib (PLX3397; CSF1R inhibitor; 10 µM for 24 hours), coculture with HCVECs under normoxia, coculture with HCVECs under hypoxia, co-culture with HCVECs under hypoxia + CSF1 neutralizing antibody (0.2 µg/mL for 24 hours), coculture with HCVECs under hypoxia + PLX3397, coculture with HCVECs under hypoxia +LY294002 (PI3K/AKT inhibitor; 10 µM for 24 hours), and coculture with HCVECs under hypoxia + AS1842856 (FOXO1 inhibitor; 0.1 µM for 24 hours) groups. (A) Transwell assay was performed to measure macrophage migration. Scale bar: 50 µm. (B) The average number of migrated macrophages was analyzed. The qRT-PCR was performed to measure the mRNA levels of the M1-type macrophage marker iNOS (C) and the M2-type macrophage marker Arg1 (D). (D) Western blotting was performed to measure the protein levels of iNOS and Arg1. (E) The relative protein levels of iNOS and Arg1 were analyzed. *P < 0.05 versus normal group. #P < 0.05 versus CSF1 group. &P < 0.05 versus coculture under normoxia group. %P < 0.05 versus coculture under hypoxia group in Figures 3B to 3D, 3Fm and 3G. n = 4/each group.

Fig 4: The effect of propofol on PI3K/AKT signaling in ADSCs by western blot and the quantitative analysis, including p-FAK, FAK, p-PI3K, PI3K, p-AKT and AKT. The quantitative results were standardized according to GAPDH level. The low, medium and high concentrations were 1.7 µM, 2.8 µM and 4.2 µM respectively. Values are expressed as mean ± SD (n = 3). *p < 0.05, **P < 0.01 vs. control

Fig 5: SPAG5 reverses miR-133a-3p-inactivated PI3K/AKT pathway in PC9/GR cell. (A) Protein expression of phosphorylated PI3K, total PI3K, phosphorylated AKT, total AKT in PC9/GR cells co-transfected with SPAG5 or vector plasmids and miR-101-3p mimics was determined by western blotting. (B) Protein expression of phosphorylated PI3K, total PI3K, phosphorylated AKT, total AKT in PC9 cells co-transfected with si-SPAG5 or si-NC plasmids and miR-101-3p inhibitor was determined by western blotting.