Figure 1.
APOE4 disrupts the endothelial BBB transcriptome. (A) Schematic of nuclei isolation and sampling workflow from mouse cortex for snRNA-seq. See Materials and methods for details. (B) UMAP space representing six distinct clusters obtained via unsupervised clustering analysis and subsequent definition of each cluster based on cell type–specific cell markers. OPC, oligodendrocyte precursor cells. (C) Dot plot reporting average expression of the cell-specific markers of ECs and PCs in the vascular cluster by in silico sorting (see Fig. S1 D). (D) Volcano plot showing DEGs in ECs in 2–3-mo-old (red) and 9–12-mo-old (cyan) E4F compared with E3F mice. (E) Plot comparing the average log2 fold-change of DEGs in ECs in 2–3-mo-old (y axis) and 9–12-mo-old (x axis) E4F compared with E3F mice (n = 140 DEGs with known function according to the UniProt Knowledgebase of 158 total). (F–H) Bar charts reporting the number of DEGs in EC-encoding proteins with known function in each functional class in 2–3-mo-old (F) and 9–12-mo-old (G) E4F compared with E3F mice, and the DEGs found in common in both age groups of E4F compared with E3F mice (H). All data in B–H are from four mice per group. (I) Log2 fold-change of 140 DEGs in ECs common to both 2–3- and 9–12-mo-old E4F compared with E3F mice. Box-and-whisker plots indicating median (dark horizonal line) and interquartile range (IQR; box representing 25th to 75th percentiles), and whiskers representing IQR upper and lower limits ±1.5 IQR; significance by Wilcoxon two-tailed paired test. (J and K) BBB permeability Ktrans maps in the cortex of 2-mo-old E3F and E4F mice by DCE-MRI (J) and Ktrans values in the cortex (Ctx; K) of 2–3-, 4–6-, and 9–12-mo-old E3F and E4F mice. (L and M) Fibrinogen (red) and lectin+ endothelial profiles (white) in the cortex of 6-mo-old E3F and E4F mice (L; bar = 25 µm) and quantification of fibrinogen perivascular deposits in 2–3-, 4–6-, and 9–12-mo-old E3F and E4F mice (M). Mean ± SEM; in K, n = 8 mice per group; in M, n = 5 mice per group. Significance by one-way ANOVA (K and M) with Bonferroni post hoc test. (N) Heatmap showing overlap between DEGs in ECs from 9–12-mo-old E4F compared with E3F mice (columns of the heatmap) and DEGs in ECs from the published mouse models of acute, subacute, and chronic EAE, epilepsy, stroke, and TBI (Munji et al., 2019; rows of the heatmap). Color scale represents –log10 P value. Significance by Fisher’s exact test. *, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001.

APOE4 disrupts the endothelial BBB transcriptome. (A) Schematic of nuclei isolation and sampling workflow from mouse cortex for snRNA-seq. See Materials and methods for details. (B) UMAP space representing six distinct clusters obtained via unsupervised clustering analysis and subsequent definition of each cluster based on cell type–specific cell markers. OPC, oligodendrocyte precursor cells. (C) Dot plot reporting average expression of the cell-specific markers of ECs and PCs in the vascular cluster by in silico sorting (see Fig. S1 D). (D) Volcano plot showing DEGs in ECs in 2–3-mo-old (red) and 9–12-mo-old (cyan) E4F compared with E3F mice. (E) Plot comparing the average log2 fold-change of DEGs in ECs in 2–3-mo-old (y axis) and 9–12-mo-old (x axis) E4F compared with E3F mice (n = 140 DEGs with known function according to the UniProt Knowledgebase of 158 total). (F–H) Bar charts reporting the number of DEGs in EC-encoding proteins with known function in each functional class in 2–3-mo-old (F) and 9–12-mo-old (G) E4F compared with E3F mice, and the DEGs found in common in both age groups of E4F compared with E3F mice (H). All data in B–H are from four mice per group. (I) Log2 fold-change of 140 DEGs in ECs common to both 2–3- and 9–12-mo-old E4F compared with E3F mice. Box-and-whisker plots indicating median (dark horizonal line) and interquartile range (IQR; box representing 25th to 75th percentiles), and whiskers representing IQR upper and lower limits ±1.5 IQR; significance by Wilcoxon two-tailed paired test. (J and K) BBB permeability Ktrans maps in the cortex of 2-mo-old E3F and E4F mice by DCE-MRI (J) and Ktrans values in the cortex (Ctx; K) of 2–3-, 4–6-, and 9–12-mo-old E3F and E4F mice. (L and M) Fibrinogen (red) and lectin+ endothelial profiles (white) in the cortex of 6-mo-old E3F and E4F mice (L; bar = 25 µm) and quantification of fibrinogen perivascular deposits in 2–3-, 4–6-, and 9–12-mo-old E3F and E4F mice (M). Mean ± SEM; in K, n = 8 mice per group; in M, n = 5 mice per group. Significance by one-way ANOVA (K and M) with Bonferroni post hoc test. (N) Heatmap showing overlap between DEGs in ECs from 9–12-mo-old E4F compared with E3F mice (columns of the heatmap) and DEGs in ECs from the published mouse models of acute, subacute, and chronic EAE, epilepsy, stroke, and TBI (Munji et al., 2019; rows of the heatmap). Color scale represents –log10 P value. Significance by Fisher’s exact test. *, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001.

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