Supplementary Figure 13: Physical presence of OPC clusters on vessels is required for effects on vascular and BBB integrity. | Nature Neuroscience

Supplementary Figure 13: Physical presence of OPC clusters on vessels is required for effects on vascular and BBB integrity.

From: Aberrant oligodendroglial–vascular interactions disrupt the blood–brain barrier, triggering CNS inflammation

Supplementary Figure 13

(ac) PDGFRα+ OPCs aggregate on blood vessels to form clusters in Olig2-cre:APC fl/fl mice P10 spinal cord (b, -AMD, arrow) but are not seen in APCfl/fl controls (a). Treatment of Olig2-cre:APCfl/fl mice with AMD3100 (+AMD) between P3-P10 leads to a reversal of OPC clustering on blood vessels (b), quantified in (c)(n=6 animals; APCfl/fl vs. Olig2-cre:APCfl/fl ****p = 6.29 E-8; Olig2-cre:APCfl/fl vs. Olig2-cre:APCfl/fl+AMD3100 ####p = 1.04 E-5). Data were analyzed by one-way ANOVA. (df) Astrocytic endfeet are displaced from blood vessels in P10 spinal cord of Olig2cre:APCfl/fl:Aldh1L1-GFP mice (e, -AMD, arrow), but AMD3100 treatment (+AMD) significantly reduces this loss of Aldh1L1-GFP+ astrocytic endfeet, quantified in (f)(n=6 animals; APCfl/fl vs. Olig2-cre:APCfl/fl ****p = 4.81 E-8; Olig2-cre:APCfl/fl vs. Olig2-cre:APCfl/fl+AMD3100 ###p = 0.0008). Data were analyzed by one-way ANOVA. (gi) OPC perivascular clustering in P10 spinal cord of Olig2-cre:APCfl/fl leads to deficits in astrocyte endfoot AQP4 staining at cluster sights (h, -AMD, arrows), but AMD3100 treatment (+AMD) significantly reverses this phenotype, quantified in (i)(n=6 animals; APCfl/fl vs. Olig2-cre:APCfl/fl ****p = 7.85 E-6; Olig2-cre:APCfl/fl vs. Olig2-cre:APCfl/fl+AMD3100 ###p = 0.0001). Data were analyzed by one-way ANOVA. (j-l) Staining for fibrinogen in P10 APCfl/fl, Olig2-cre:APCfl/fl (-AMD) and Olig2-cre:APC fl/fl+AMD3100 spinal cord shows that there are fibrinogen positive areas outside blood vessels in Olig2-cre:APCfl/fl mice (arrow, k, -AMD) but this extravasation is significantly reversed by AMD3100 treatment (l; n=6 animals; APCfl/fl vs. Olig2-cre:APCfl/fl ***p = 0.0008; Olig2-cre:APCfl/fl vs. Olig2-cre:APCfl/fl+AMD3100 ##p = 0.0023). Data were analyzed by one-way ANOVA. (m-o) Staining for CD11c positive activated microglial cells (green) in P10 APCfl/fl, Olig2-cre:APCfl/fl (-AMD) and Olig2-cre:APCfl/fl+AMD3100 spinal cord shows that there are significantly increased activated microglia around clusters in Olig2-cre:APCfl/fl mouse (-AMD, n), but this microglia activation is reduced by AMD3100 (+AMD), quantified in (o)(n=6 animals; APCfl/fl vs. Olig2-cre:APCfl/fl **p = 0.0032; Olig2 cre:APCfl/fl vs. Olig2-cre:APCfl/fl+AMD3100 ##p = 0.0044). Data were analyzed by one-way ANOVA. Scale bars, 20 μm in all panels. * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001. Values are mean ± s.d. For all staining results, experiments were repeated at least three times independently with similar results.

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