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. 2024 Mar 19;134(10):e169137. doi: 10.1172/JCI169137

Figure 8. Aberrant eNOS localization and PP2A expression in ISCLS-derived BOECs.

Figure 8

(A) BOECs stained with anti–p-eNOS (left, green) or anti-eNOS (middle, red) antibodies and DAPI. Arrows denote eNOS clusters. Scale bar: 20 μm. (B) Numbers of eNOS clusters/cell. Data indicate the mean ± SEM. n = 3 cell lines/group. *P = 0.03 and **P = 0.006, by 2-way ANOVA with Šidák’s multiple comparisons. (C) Representative immunoblot of PP2A subunits in BOEC lysates (n = 5 donors/group). (D) Quantification of relative protein expression. Data indicate the mean ± SEM. n = 7–9 donors/group. *P = 0.03, by 2-way ANOVA with Šidák’s multiple comparisons. (E) Relative PPP2R1B expression in BOECs evaluated by qPCR (normalized to GAPDH). Data indicate the mean ± SEM. n = 6–9 donors/group. NS, by unpaired, 2-tailed Student’s t test. (F) Representative immunoblot of FLAG-PP2A-Aβ in BOECs transfected with the respective lentiviruses. (G and H) Representative TER (G) and maximum decrease in VEGF-induced TER (H) from t = 0 (arrow) in BOECs infected with control or FLAG–PP2A-β–encoding lentivirus. Data indicate the mean ± SEM. n = 2 donors/group analyzed in 4 independent experiments. **P = 0.001, by unpaired, 2-tailed Student’s t test. (I and J). Representative blot (I) and quantification (J) of p-eNOS/eNOS in control versus FLAG–PP2A-β–overexpressing BOECs stimulated with VEGF (100 ng/mL). Data indicate the mean ± SEM. n = 5 independent experiments. **P = 0.004, by 2-way ANOVA with Šidák’s multiple comparisons.