Journal: Brain, behavior, & immunity - health

Article Title: Investigation in blood-brain barrier integrity and susceptibility to immune cell penetration in a mouse model of Dravet syndrome.

doi: 10.1016/j.bbih.2025.100955

Figure Lengend Snippet: Fig. 3. Evaluation of neutrophil infiltration into the brain parenchyma: Representative images of a double immunofluorescence of the endothelial marker laminin (red) and neutrophils (NIMP-R14, green) in the hippocampus of Syn1-Cre/Scn1aWT/A1783V mice and their control group (Scn1aWT/WT) at PND25. Positive control corresponds to an ischemic brain tissue with proven infiltration of neutrophils into the brain ischemic core. Scale bar = 50 μm.

Article Snippet: Sections were mounted on gelatin-coated slides and, once adhered, washed with TBS, permeabilized with TBS containing Triton X-100 0.2% for 30 min, and blocked with TBS containing Triton X-100 0.1% and BSA 5% for 1 h. After several washes with TBS, sections were incubated overnight at 4 ◦C with the following primary antibodies: (i) Analysis of neutrophils: antilaminin rabbit polyclonal antibody (ref. NB300-144; Novus Biologicals, Centennial, CO, USA) used at 1/200, and anti-NIMP-R14 rat monoclonal antibody (ref. ab2557; Abcam, Cambridge, UK) used at 1/200 for labelling neutrophils; and (ii) Analysis of B and T cells: antiCD31/ PECAM-1 rat monoclonal antibody (ref. 553370, BD-Biosciences, Madrid, Spain) used at 1/100 for labelling endothelial cells, antiB220/CD45R rabbit monoclonal antibody (ref. NBP3-11980; Novus Biological Centennial, CO, USA) used at 1/250 for labelling B-cells, and anti-CD3 rabbit polyclonal antibody (ref. ab5690, Abcam, Cambridge, UK) used at 1/250 for labelling T-cells.

Techniques: Immunofluorescence, Marker, Control, Positive Control

Journal: Acta Neuropathologica

Article Title: Microglial-mediated PDGF-CC activation increases cerebrovascular permeability during ischemic stroke

doi: 10.1007/s00401-017-1749-z

Figure Lengend Snippet: CD11b, LRP1, and PDGFRα are expressed in the NVU of wild-type (WT) mice. 50 μm vibratome sections from WT murine brains were analyzed by immunofluorescence staining and confocal microscopy for CD11b (the alpha chain of Mac-1), LRP1, PDGFRα, and the endothelial cell marker CD31 ( white ) to visualize vessels. a PDGFRα ( red ) is expressed both in nonvascular cells ( arrows ) and in cells associated with a subset of cerebral vessels, presumably arterioles ( closed arrowheads ), but not in capillaries ( double arrows ). CD11b ( green ) is seen throughout the field of view in resting microglia with their characteristic ramified appearance with occasional branching processes contacting both PDGFRα positive vessels, as well as PDGFRα negative microvasculature ( open arrowheads ). b LRP1 ( red ) and PDGFRα ( green ) are co-localized in both nonvascular cells ( arrows ) and vessel-associated cells ( closed arrowheads ). c CD11b ( green ) is not co-expressed by LRP1 positive cells ( red ), neither by nonvascular ( arrows ) or by vessel-associated LRP1 positive cells ( closed arrowheads ). However, several CD11b branching processes are contacting LRP1 positive vessels ( open arrowheads ). The images were captured in the cerebral cortex ( a , b ) and in the hippocampus ( c ) and display the maximum intensity projections generated from confocal Z-stacks. Scale bars 10 μm

Article Snippet: Goat anti-CD31 or goat anti-podocalyxin antibodies were used to visualize endothelial cells (1:250; R&D Systems, UK).

Techniques: Immunofluorescence, Staining, Confocal Microscopy, Marker, Generated

Journal: Acta Neuropathologica

Article Title: Microglial-mediated PDGF-CC activation increases cerebrovascular permeability during ischemic stroke

doi: 10.1007/s00401-017-1749-z

Figure Lengend Snippet: Time course of monocytes infiltration after MCAO in R/G mice. a R/G mice subjected to MCAO showed a clear increase in RFP + monocyte infiltration overtime. Representative time course images of the ischemic penumbra from RFP + (monocytes/macrophages) and GFP + (microglia) brain sections stained for the endothelial cell marker podocalyxin ( cyan ). At 0 (no MCAO) and 6 h post-MCAO, there are few detectable RFP + monocytes infiltrating the parenchyma near blood vessels in the ischemic penumbra. By 24 h, a significant number of RFP + monocytes can be observed not only in the blood vessels, but also in the parenchyma ( scale bars 25 μm). b Quantification of GFP + and RFP + cells ( n = 2–4; errors represent SEM). The significance for all comparisons was determined from a two-way ANOVA with a Fisher’s LSD test (Online Resource 10); * p < 0.05 vs 0 h microglia, ‡ p < 0.05 vs 0 h monocyte, δ p < 0.05 vs 6 h microglia, ϕ p < 0.05 vs 6 h monocyte, § p < 0.0001 vs 24 h microglia. c Mac-1-mediated BBB permeability is independent of infiltrating leukocytes. Wild-type (WT) and Mac-1 −/− mice received bone marrow transplant from either WT or Mac-1 −/− mice, and 8–10 weeks later, they were subjected to MCAO. BBB permeability was quantified by fluorescent microscopy and image analysis of 70-kDa rhodamine-dextran extravasation 24 h after MCAO. In each group, n = 7–8; errors represent SEM, ** p < 0.01, *** p < 0.001 (two-way ANOVA with Fisher’s LSD test)

Article Snippet: Goat anti-CD31 or goat anti-podocalyxin antibodies were used to visualize endothelial cells (1:250; R&D Systems, UK).

Techniques: Staining, Marker, Permeability, Microscopy