Mba1049

We analysed formalin-fixed paraffin-embedded whole coronal sections at levels 6–8 [27 (link), 42 ] containing the frontal cortex (Brodmann area 9). We ensured to select the cortical regions without any obvious infarction. Unless otherwise stated, 2–5 adjacent or alternate whole or half-coronal sections were used for the morphological analyses. Immunohistochemistry was performed to examine different microvascular structures essentially as described before [13 (link), 22 (link)]. The following antibodies were used to assess various cellular features: collagen IV (COL4 at dilation 1:1000, C1926, Merck (Sigma-Aldrich), Branchburg, NJ, USA), a marker of basement membrane in the vessels, platelet-derived growth factor receptor-β (PDGFR-β at 1:200 dilution, clone 42G12, #AF385, R&D systems, Minneapolis, MN, USA), a marker for pericytes, bone morphogenetic protein 4 (BMP4 dilution at 1:100, MBA1049, Millipore, MA, USA), α-smooth muscle actin (αSMA at dilution 1:1000, Clone 1A4, Dako, Cambridge, UK), a marker for mural cells, and glucose transporter-1 (GLUT-1 at 1:200, PA1-21,041, Fisher Scientific, Waltham, MA, USA), a marker of endothelial cells. Vectastain ABC mouse kits (PK-6102, Vector Laboratories, Burlingame, CA, USA) and Diaminobenzidine were used for single or double immunohistochemistry. Haematoxylin counterstain was used for ease in localising regions of interest.

Formalin‐fixed paraffin‐embedded whole coronal sections at levels 6–8 (36, 48) containing the frontal lobe (Brodmann area 9) were analyzed. When sampling tissue, we ensured to select the WM regions free of any obvious infarcts and thus assessed effects of remote stroke injury. Immunohistochemistry was performed to examine various microvascular structures essentially as described before (16, 29). The following antibodies were used to assess various cellular features: collagen IV (COL4 at dilution 1:1000, C1926, Merck (Sigma‐Aldrich), Branchburg, NJ, USA), a marker of basement membrane in the vessels, platelet‐derived growth factor receptor‐β (PDGFR‐β at 1:200 dilution, clone 42G12, #AF385, R&D systems, Minneapolis, MN, USA), a marker for pericytes, bone morphogenetic protein 4 (BMP4 dilution at 1:100, MBA1049, Millipore, MA, USA), α‐smooth muscle actin (αSMA at dilution 1:1000, Clone 1A4, Dako, Cambridge, UK), a marker for mural cells and glucose transporter‐1 (GLUT‐1 at 1:200, PA1‐21041, Fisher Scientific, Waltham, MA, USA), a marker of endothelial cells. Vectastain ABC mouse kits (PK‐6102, Vector Laboratories, Burlingame, CA, USA) and DAB were used to localize single or double immunohistochemical stains. Tissue sections were then counter stained with hematoxylin to visualize the landmarks.