title: 3D visualization and quantification of microvessels in the whole ischemic mouse brain using solvent-based clearing and light sheet microscopy.
abstract: The visualization of cerebral microvessels is essential for understanding brain remodeling after stroke. Injection of dyes allows for the evaluation of perfused vessels, but has limitations related either to incomplete microvascular filling or leakage. In conventional histochemistry, the analysis of microvessels is limited to 2D structures, with apparent limitations regarding the interpretation of vascular circuits. Herein, we developed a straight-forward technique to visualize microvessels in the whole ischemic mouse brain, combining the injection of a fluorescent-labeled low viscosity hydrogel conjugate with 3D solvent clearing followed by automated light sheet microscopy. We performed transient middle cerebral artery occlusion in C57Bl/6j mice and acquired detailed 3D vasculature images from whole brains. Subsequent image processing, rendering and fitting of blood vessels to a filament model was employed to calculate vessel length density, resulting in 0.922 ± 0.176 m/mm3in healthy tissue and 0.329 ± 0.131 m/mm3in ischemic tissue. This analysis showed a marked loss of capillaries with a diameter ≤ 10 µm and a more moderate loss of microvessels in the range > 10 and ≤ 20 µm, whereas vessels > 20 µm were unaffected by focal cerebral ischemia. We propose that this protocol is highly suitable for studying microvascular injury and remodeling post-stroke.
authors.affiliation: 1 Department of Neurology, University Hospital Essen, University of Duisburg-Essen, Essen, Germany.|2 Department of Cell Morphology and Molecular Neurobiology, Ruhr University Bochum, Bochum, Germany.|3 Department of Physiology and Biochemistry, School of Medicine, Faculty of Health Sciences, University of Carabobo, La Morita, Venezuela.
authors.forename: Erlen
authors.initials: E
authors.lastname: Lugo-Hernandez
authors.affiliation: 4 Institute for Experimental Immunology and Imaging and Imaging Center Essen (IMCES), University Hospital Essen, University of Duisburg-Essen, Essen, Germany.
authors.forename: Anthony
authors.initials: A
authors.lastname: Squire
authors.affiliation: 1 Department of Neurology, University Hospital Essen, University of Duisburg-Essen, Essen, Germany.
authors.forename: Nina
authors.initials: N
authors.lastname: Hagemann
authors.affiliation: 4 Institute for Experimental Immunology and Imaging and Imaging Center Essen (IMCES), University Hospital Essen, University of Duisburg-Essen, Essen, Germany.
authors.forename: Alexandra
authors.initials: A
authors.lastname: Brenzel
authors.affiliation: 1 Department of Neurology, University Hospital Essen, University of Duisburg-Essen, Essen, Germany.
authors.forename: Maryam
authors.initials: M
authors.lastname: Sardari
authors.affiliation: 1 Department of Neurology, University Hospital Essen, University of Duisburg-Essen, Essen, Germany.
authors.forename: Jana
authors.initials: J
authors.lastname: Schlechter
authors.affiliation: 1 Department of Neurology, University Hospital Essen, University of Duisburg-Essen, Essen, Germany.
authors.forename: Eduardo H
authors.initials: EH
authors.lastname: Sanchez-Mendoza
authors.affiliation: 4 Institute for Experimental Immunology and Imaging and Imaging Center Essen (IMCES), University Hospital Essen, University of Duisburg-Essen, Essen, Germany.
authors.forename: Matthias
authors.initials: M
authors.lastname: Gunzer
authors.affiliation: 2 Department of Cell Morphology and Molecular Neurobiology, Ruhr University Bochum, Bochum, Germany.
authors.forename: Andreas
authors.initials: A
authors.lastname: Faissner
authors.affiliation: 1 Department of Neurology, University Hospital Essen, University of Duisburg-Essen, Essen, Germany.
authors.forename: Dirk M
authors.initials: DM
authors.lastname: Hermann
mesh_terms: D000818:Animals; D001921:Brain / Q000098:blood supply / Q000000981:diagnostic imaging; D002545:Brain Ischemia / Q000000981:diagnostic imaging*; D002196:Capillaries / Q000503:physiopathology; D021621:Imaging, Three-Dimensional / Q000379:methods*; D020244:Infarction, Middle Cerebral Artery; D051379:Mice; D008853:Microscopy / Q000379:methods; D055806:Microvessels / Q000000981:diagnostic imaging*; D012997:Solvents; D020521:Stroke / Q000000981:diagnostic imaging / Q000473:pathology
chemical_list: D012997:Solvents
keywords: Angiography; brain imaging; capillaries; focal ischemia; microcirculation