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. 2019 Jun 27;10:2817. doi: 10.1038/s41467-019-10643-w

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© The Author(s) 2019

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 4 — Functional defects of Rbpj mutant mural cells. a Confocal images of P10 cortical arteries stained for isolectin B4 (IB4) or CDH5 (red), αSMA (green), and Desmin (Des, white). Note irregular SMC coverage of mutant arteries (asterisk) and lack of αSMA labelling in Des⁺ regions (white arrow). Scale bar, 50 μm (left panels) and 25 μm (right panel, high magnification). b SMC coverage in P10 brain cortex arteries. p-values, Student’s t-test, n = 6. c Confocal images of P10 cortical vasculature (tdT⁺, red) stained for GFP (green), SM22α (blue, white), and αSMA (white). Note the expression of αSMA and SM22α in the Rbpj^iPC microvasculature (white arrowheads). Scale bar, 100 μm. d–f Confocal images of brain cortex vasculature showing increased mural cell-specific staining of Vimentin (d, arrowheads), Desmin (Des, e), and Nestin (f) (all in white) in P10 Rbpj^iPC blood vessels (labelled with IB4, CD31, or tdT, red) relative to control. Scale bar, 50 μm (d) or 100 μm (e, f). g Gene expression fold change (RNA-seq fpkm) in Rbpj^iPC mutants relative to age-matched controls (set as 1). p-adjusted values < 0.0001 for all with exception of Des expression at P7. n = 3. n.s., not statistically significant. h Confocal images of P10 cortical blood vessels showing increased immunostaining for phospho-myosin light chain 2 (phMLC2, white) in pericytes (PDGFRβ⁺, red; GFP⁺, green). Scale bar, 50 μm. i Collagen gel contraction assay showing increased contractile ability of lentivirus-Cre transfected Rbpj^iPC relative to control primary mouse brain pericytes. Scale bar, 4 mm. j Contractility quantitation at 18 and 30 h after pericyte seeding. p-values, Kruskal–Wallis with Dunn’s test, n = 5. k RT-qPCR analysis of Acta2 and Tagln expression in cultured pericytes. p-values, one-way ANOVA with Sidak’s test, n = 3. l Confocal images of P10 brain cortex microvasculature stained for ICAM2 (red), GFP (green), and PDGFRβ (white). Cre-induced membrane-tagged GFP expression allows detailed imaging of abnormal protrusions emerging from Rbpj^iPC pericytes (yellow arrowheads). Scale bar, 25 μm. Error bars represent s.e.m. Source data are provided as a Source Data file