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. Author manuscript; available in PMC: 2023 Mar 13.
Published in final edited form as: Nat Neurosci. 2022 Aug 1;25(8):1020–1033. doi: 10.1038/s41593-022-01127-0

Fig. 7 ∣. ApoE isoform-specific effects of young plasma on human endothelial barrier integrity and transcriptomic signature.

Fig. 7 ∣

a, Schematic diagram of human brain endothelial cell (iBMEC) differentiation from iPSCs and the experimental design. b, Characterization of brain endothelial cell differentiation at different time points. Similar results were observed in at least three independent experiments. The iPSC pluripotency and mesoderm lineage differentiation were confirmed by Nanog, Brachyury (a primitive streak marker), or PAX2/5/8 (an intermediate mesoderm marker) staining, respectively. The iPSC-derived iBMECs were stained for Glut1 (an endothelial marker) and ZO1 (a tight junction protein) at Day 12. Scale bars: 100 μm. c, The transendothelial electrical resistance (TEER) in iBMECs and human umbilical vein endothelial cells (HUVECs) was measured. **, P < 0.0001. d, The transwell setup in which the iBMECs were treated with control or young plasma, followed by the assessment of the endothelial cell integrity. e, Human iBMECs cultured in the transwell system were treated with PBS-citrate (control), apoE3 young plasma (2%) or apoE4 young plasma (2%) for 24 hr. The TEER values were measured and compared to the control. Data represent mean ± s.e.m. from three independent experiments. **, P=0.002; *, P=0.034; N.S., not significant. One-way ANOVA with a Tukey’s post-hoc test. f, Modules significantly correlated with apoE3 or apoE4 plasma treatment compared to control identified from RNA-Seq. The box in the plots displays 25th and 75th percentile values. The center line represents the median and the whiskers show minimum and maximum values. g, Network plots of the proteins with high intramodular connectivity in the turquoise module of apoE3 group, and in the green module of apoE4 group. h, Pathway analysis using gene ontology showing the pathways enriched in the gene-sets of the turquoise module and green module. i, The human iBMECs were treated with apoE3 or apoE4 young plasma, or apoE4 plasma together with Timp3 (0.5 μg/ml) for 24 hr, and the TEER values were measured. Data represent mean ± s.e.m. from three independent experiments. **, P=0.003; *, P=0.012; N.S., not significant. One-way ANOVA with a Tukey’s post-hoc test was used to determine the significance between groups.