Fig 2: Physiological studies of ARPE-19 monolayers cultured on transwell inserts.Trans-epithelial Electrical Resistance (TEER) measurements were obtained from long-term cultures to evaluate effectiveness of the Retinal Pigment Epithelial barrier. [ A] Measurements were conducted from transwells (n=3) at weekly time intervals after seeding. Values were plotted as a percentage change from the previous measurement which show a gradual increase as junctions form and mature. [ B] Fluctuations between average weekly TEER were observed prior to week 6 (p=0.009 at 3 weeks, p=0.013 at 4 weeks and p=0.001 at 6 weeks, one-way ANOVA with Tukey’s multiple comparisons) after which a stable value of 40.72 Ω.cm 2 was achieved. Data is presented as mean ± SEM. Next, we quantified polarised secretion of Vascular Endothelial Growth Factor (VEGF) and Pigment Epithelium Derived Factor (PEDF) by ARPE-19 cells. Conditioned media was collected (n=3) after 72 hours and proteins quantified by ELISA. [ C] The apical compartment was found to contain 0.942 ± 0.035ng/ml of VEGF compared to 2.852 ± 0.145ng/ml in the basal chamber, which was statistically significant (p=0.0002). [ D] PEDF concentrations in the apical compartment was 16.95 ± 0.72ng/ml compared to 25.05 ± 3.93ng/ml in the basal chamber. There were no significant differences (p= 0.112) although more PEDF was secreted via the basolateral RPE surface. Data is presented as mean ± SEM with statistical comparisons made using the unpaired student’s t-test and sourced in part from material published previously 18 under the Creative Commons licence.

Fig 3: Receiver-operating characteristic curve for plasma PEDF levels at days 1–3 after subarachnoid hemorrhage (SAH) according to good and poor 3-month outcomes in elderly SAH patients. AUC, area under the curve; CI, confidence interval.

Fig 4: Box and jitter plots illustrating the relationships between (A) VEGFR2, (B) VEGF-A, (C) PIGF, (D) PDGF-BB, (E) FGF-2, (F) PEDF, and (G) ANG-1 and glycemic control (HBA1c cut-off at 8%). VEGF-A—vascular endothelial growth factor A, VEGF-R2—vascular endothelial growth factor receptor 2, FGF-2—fibroblast growth factor 2, PlGF—placental growth factor, PDGF-BB—platelet-derived growth factor-BB, PEDF—pigment epithelium-derived factor, and ANG-1—angiopoietin-1.

Fig 5: After maturation, iRPE cells present typical biological functions of RPE cells. (a) TER was measured on iRPE and fRPE monolayers at 14, 28 and 42 days in culture on transwell. Bars represents mean ± SD. (b) PEDF and VEGF secretion levels were measured by ELISA after 42 days in culture for both apical (blue) and basal (grey) compartments of n = 5 iRPE lines. Comparison between compartments was done by paired Wilcoxon match-pairs signed-rank test. Bars represent mean ± SD. (c–f) FITC-conjugated POS were fed to iRPE and fRPE cells for 16 h and the RHODOPSIN protein was immunolabelled without permeabilization (red labelling). As a result, the internalized POS fluoresce in green only, and microvilli-bound POS fluoresce in green and red. p: p-value. P-value significance threshold for all tests: 0.05.