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01-03-2018 | Methods Paper

Analysis of the vasculature by immunohistochemistry in paraffin-embedded brains

Authors: Yann Decker, Andreas Müller, Eszter Németh, Walter J. Schulz-Schaeffer, Marc Fatar, Michael D. Menger, Yang Liu, Klaus Fassbender

Published in: Brain Structure and Function | Issue 2/2018

Abstract

The brain vasculature can be investigated in different ways ranging from in vivo to biochemical analysis. Immunohistochemistry is a simple and powerful technique that can also be applied to archival tissues. However, staining of brain vessels on paraffin sections has been challenging. In this study, we developed an optimized method that can be used in paraffin-embedded mouse and human brain tissues derived from healthy controls and neurological disorders such as Alzheimer’s disease. We subsequently showed that this method is fully compatible with the detection of glial cells and key markers of Alzheimer’s disease including amyloid beta and phosphorylated Tau protein. Furthermore, we observed that the length of microvasculature in hippocampus of TgCRND8 Alzheimer’s disease mouse model is reduced, which is correlated with the decreased blood flow in hippocampus as determined by arterial spin labeling perfusion magnetic resonance imaging. Finally, we determined that the microvasculature length in two other Alzheimer’s disease mouse models, APP and PS1 double-transgenic mice and P301S Tau-transgenic mice, is also shortened in the dentate gyrus. Thus, we have established a new, simple and robust method to characterize the brain vasculature in the mouse and human brain.

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Asllani I, Habeck C, Scarmeas N, Borogovac A, Brown TR, Stern Y (2008) Multivariate and univariate analysis of continuous arterial spin labeling perfusion MRI in Alzheimer’s disease. J Cereb Blood Flow Metabol 28:725–736. https://doi.org/10.1038/sj.jcbfm.9600570 CrossRef

Attems J, Jellinger KA (2014) The overlap between vascular disease and Alzheimer’s disease—lessons from pathology. BMC Med 12:206. https://doi.org/10.1186/s12916-014-0206-2 CrossRefPubMedPubMedCentral

Binnewijzend MA et al (2013) Cerebral blood flow measured with 3D pseudocontinuous arterial spin-labeling MR imaging in Alzheimer disease and mild cognitive impairment: a marker for disease severity. Radiology 267:221–230. https://doi.org/10.1148/radiol.12120928 CrossRefPubMed

Blair LJ et al (2015) Tau depletion prevents progressive blood-brain barrier damage in a mouse model of tauopathy. Acta Neuropathol Commun 3:8. https://doi.org/10.1186/s40478-015-0186-2 CrossRefPubMedPubMedCentral

Bordeleau M, ElAli A, Rivest S (2016) Severe chronic cerebral hypoperfusion induces microglial dysfunction leading to memory loss in APPswe/PS 1 mice. Oncotarget 7:11864–11880. https://doi.org/10.18632/oncotarget.7689 CrossRefPubMedPubMedCentral

Bouras C et al (2006) Stereologic analysis of microvascular morphology in the elderly: Alzheimer disease pathology and cognitive status. J Neuropathol Exp Neurol 65:235–244. https://doi.org/10.1097/01.jnen.0000203077.53080.2c CrossRefPubMed

Buee L, Hof PR, Bouras C, Delacourte A, Perl DP, Morrison JH, Fillit HM (1994) Pathological alterations of the cerebral microvasculature in Alzheimer’s disease and related dementing disorders. Acta Neuropathol 87:469–480CrossRefPubMed

Chishti MA et al (2001) Early-onset amyloid deposition and cognitive deficits in transgenic mice expressing a double mutant form of amyloid precursor protein 695. J Biol Chem 276:21562–21570. https://doi.org/10.1074/jbc.M100710200 CrossRefPubMed

Clark PJ, Brzezinska WJ, Puchalski EK, Krone DA, Rhodes JS (2009) Functional analysis of neurovascular adaptations to exercise in the dentate gyrus of young adult mice associated with cognitive gain. Hippocampus 19:937–950. https://doi.org/10.1002/hipo.20543 CrossRefPubMedPubMedCentral

D’Amico F, Skarmoutsou E, Stivala F (2009) State of the art in antigen retrieval for immunohistochemistry. J Immunol Methods 341:1–18. https://doi.org/10.1016/j.jim.2008.11.007 CrossRefPubMed

De Strooper B, Karran E (2016) The cellular phase of Alzheimer’s. Dis Cell 164:603–615. https://doi.org/10.1016/j.cell.2015.12.056 CrossRef

Dobre MC, Ugurbil K, Marjanska M (2007) Determination of blood longitudinal relaxation time (T1) at high magnetic field strengths. Magn Reson Imaging 25:733–735. https://doi.org/10.1016/j.mri.2006.10.020 CrossRefPubMed

Dorr A et al (2012) Amyloid-beta-dependent compromise of microvascular structure and function in a model of Alzheimer’s disease. Brain 135:3039–3050. https://doi.org/10.1093/brain/aws243 CrossRefPubMed

Dudvarski Stankovic N, Teodorczyk M, Ploen R, Zipp F, Schmidt MH (2016) Microglia-blood vessel interactions: a double-edged sword in brain pathologies. Acta Neuropathol 131:347–363. https://doi.org/10.1007/s00401-015-1524-y CrossRefPubMed

Faure A et al (2011) Impaired neurogenesis, neuronal loss, and brain functional deficits in the APPxPS1-Ki mouse model of Alzheimer’s disease. Neurobiol Aging 32:407–418. https://doi.org/10.1016/j.neurobiolaging.2009.03.009 CrossRefPubMed

Fischer VW, Siddiqi A, Yusufaly Y (1990) Altered angioarchitecture in selected areas of brains with Alzheimer’s disease. Acta Neuropathol 79:672–679CrossRefPubMed

Franciosi S et al (2007) Pepsin pretreatment allows collagen IV immunostaining of blood vessels in adult mouse brain. J Neurosci Methods 163:76–82. https://doi.org/10.1016/j.jneumeth.2007.02.020 CrossRefPubMedPubMedCentral

Fries P et al (2012) Comparison of retrospectively self-gated and prospectively triggered FLASH sequences for cine imaging of the aorta in mice at 9.4T. Investig Radiol 47:259–266. https://doi.org/10.1097/RLI.0b013e31823d3eb6 CrossRef

Gama Sosa MA et al (2010) Age-related vascular pathology in transgenic mice expressing presenilin 1-associated familial Alzheimer’s disease mutations. Am J Pathol 176:353–368. https://doi.org/10.2353/ajpath.2010.090482 CrossRefPubMedPubMedCentral

Gama Sosa MA et al. (2014) Selective vulnerability of the cerebral vasculature to blast injury in a rat model of mild traumatic brain injury. Acta Neuropathol Commun 2:67 https://doi.org/10.1186/2051-5960-2-67 CrossRefPubMedPubMedCentral

Hebert F, Grand’maison M, Ho MK, Lerch JP, Hamel E, Bedell BJ (2013) Cortical atrophy and hypoperfusion in a transgenic mouse model of Alzheimer’s disease. Neurobiol Aging 34:1644–1652. https://doi.org/10.1016/j.neurobiolaging.2012.11.022 CrossRefPubMed

Herscovitch P, Raichle ME (1985) What is the correct value for the brain–blood partition coefficient for water? J Cereb Blood Flow Metabol 5:65–69. https://doi.org/10.1038/jcbfm.1985.9 CrossRef

Huang SN, Minassian H, More JD (1976) Application of immunofluorescent staining on paraffin sections improved by trypsin digestion Laboratory investigation. J Tech Methods Pathol 35:383–390

Huang W et al (2014) Novel NG2-CreERT2 knock-in mice demonstrate heterogeneous differentiation potential of NG2 glia during development. Glia 62:896–913. https://doi.org/10.1002/glia.22648 CrossRefPubMed

Jucker M, Bialobok P, Hagg T, Ingram DK (1992) Laminin immunohistochemistry in brain is dependent on method of tissue fixation. Brain Res 586:166–170CrossRefPubMed

Kai H, Shin RW, Ogino K, Hatsuta H, Murayama S, Kitamoto T (2012) Enhanced antigen retrieval of amyloid beta immunohistochemistry: re-evaluation of amyloid beta pathology in Alzheimer disease and its mouse model. J Histochem Cytochem 60:761–769. https://doi.org/10.1369/0022155412456379 CrossRefPubMedPubMedCentral

Kim SG, Tsekos NV, Ashe J (1997) Multi-slice perfusion-based functional MRI using the FAIR technique: comparison of CBF and BOLD effects. NMR Biomed 10:191–196CrossRefPubMed

Kloepper J et al (2016) Ang-2/VEGF bispecific antibody reprograms macrophages and resident microglia to anti-tumor phenotype and prolongs glioblastoma survival. Proc Natl Acad Sci USA 113:4476–4481. https://doi.org/10.1073/pnas.1525360113 CrossRefPubMedPubMedCentral

Kober F et al (2007) MRI follow-up of TNF-dependent differential progression of atherosclerotic wall-thickening in mouse aortic arch from early to. advanced stages. Atherosclerosis 195:e93–e99. https://doi.org/10.1016/j.atherosclerosis.2007.06.015 CrossRefPubMed

Kouznetsova E et al (2006) Developmental and amyloid plaque-related changes in cerebral cortical capillaries in transgenic Tg2576 Alzheimer mice. Int J Dev Neurosci 24:187–193. https://doi.org/10.1016/j.ijdevneu.2005.11.011 CrossRefPubMed

Lamoke F et al (2015) Amyloid beta peptide-induced inhibition of endothelial nitric oxide production involves oxidative stress-mediated constitutive eNOS/HSP90 interaction and disruption of agonist-mediated Akt activation. J Neuroinflamm 12:84. https://doi.org/10.1186/s12974-015-0304-x CrossRef

Lee GD, Aruna JH, Barrett PM, Lei DL, Ingram DK, Mouton PR (2005) Stereological analysis of microvascular parameters in a double transgenic model of Alzheimer’s disease. Brain Res Bull 65:317–322. https://doi.org/10.1016/j.brainresbull.2004.11.024 CrossRefPubMed

Love S, Miners JS (2016) Cerebrovascular disease in ageing and Alzheimer’s disease. Acta Neuropathol 131:645–658. https://doi.org/10.1007/s00401-015-1522-0 CrossRefPubMed

Massaad CA, Amin SK, Hu L, Mei Y, Klann E, Pautler RG (2010) Mitochondrial superoxide contributes to blood flow and axonal transport deficits in the Tg2576 mouse model of Alzheimer’s disease. PloS one 5:e10561 https://doi.org/10.1371/journal.pone.0010561 CrossRefPubMedPubMedCentral

Mauro A, Bertolotto A, Germano I, Giaccone G, Giordana MT, Migheli A, Schiffer D (1984) Collagenase in the immunohistochemical demonstration of laminin, fibronectin and factor VIII/RAg in nervous tissue after fixation. Histochemistry 80:157–163CrossRefPubMed

Montagne A, Nation DA, Pa J, Sweeney MD, Toga AW, Zlokovic BV (2016) Brain imaging of neurovascular dysfunction in Alzheimer’s disease. Acta Neuropathol 131:687–707. https://doi.org/10.1007/s00401-016-1570-0 CrossRefPubMedPubMedCentral

Mori S, Sternberger NH, Herman MM, Sternberger LA (1992) Variability of laminin immunoreactivity in human autopsy brain. Histochemistry 97:237–241CrossRefPubMed

Muneton-Gomez VC et al (2012) Neural differentiation of transplanted neural stem cells in a rat model of striatal lacunar infarction: light and electron microscopic observations. Front Cell Neurosci 6:30. https://doi.org/10.3389/fncel.2012.00030 CrossRefPubMedPubMedCentral

Niwa K, Porter VA, Kazama K, Cornfield D, Carlson GA, Iadecola C (2001) A beta-peptides enhance vasoconstriction in cerebral circulation. Am J Physiol Heart Circ Physiol 281:H2417–H2424CrossRefPubMed

Paul J, Strickland S, Melchor JP (2007) Fibrin deposition accelerates neurovascular damage and neuroinflammation in mouse models of Alzheimer’s disease. J Exp Med 204:1999–2008. https://doi.org/10.1084/jem.20070304 CrossRefPubMedPubMedCentral

Poisnel G et al (2012) Increased regional cerebral glucose uptake in an APP/PS1 model of Alzheimer’s disease. Neurobiol Aging 33:1995–2005. https://doi.org/10.1016/j.neurobiolaging.2011.09.026 CrossRefPubMed

Radde R et al (2006) Abeta42-driven cerebral amyloidosis in transgenic mice reveals early and robust pathology. EMBO Rep 7:940–946. https://doi.org/10.1038/sj.embor.7400784 CrossRefPubMedPubMedCentral

Shi SR, Key ME, Kalra KL (1991) Antigen retrieval in formalin-fixed, paraffin-embedded tissues: an enhancement method for immunohistochemical staining based on microwave oven heating of tissue sections. J Histochem Cytochem 39:741–748CrossRefPubMed

Soto I et al (2015) APOE stabilization by exercise prevents aging neurovascular dysfunction and complement induction. PLoS Biol 13:e1002279. https://doi.org/10.1371/journal.pbio.1002279 CrossRefPubMedPubMedCentral

Spitzer M, Wildenhain J, Rappsilber J, Tyers M (2014) BoxPlotR: a web tool for generation of box plots. Nat Methods 11:121–122. https://doi.org/10.1038/nmeth.2811 CrossRefPubMedPubMedCentral

Vollert CT, Moree WJ, Gregory S, Bark SJ, Eriksen JL (2015) Formaldehyde scavengers function as novel antigen retrieval agents. Sci Rep 5:17322. https://doi.org/10.1038/srep17322 CrossRefPubMedPubMedCentral

Walchli T et al (2015) Quantitative assessment of angiogenesis, perfused blood vessels and endothelial tip cells in the postnatal mouse brain. Nat Protoc 10:53–74. https://doi.org/10.1038/nprot.2015.002 CrossRefPubMed

Whiteus C, Freitas C, Grutzendler J (2014) Perturbed neural activity disrupts cerebral angiogenesis during a postnatal critical period. Nature 505:407–411. https://doi.org/10.1038/nature12821 CrossRefPubMed

Yamashita S (2007) Heat-induced antigen retrieval: mechanisms and application to histochemistry. Prog Histochem Cytochem 41:141–200. https://doi.org/10.1016/j.proghi.2006.09.001 CrossRefPubMed

Yoshiyama Y et al (2007) Synapse loss and microglial activation precede tangles in a P301S tauopathy mouse model. Neuron 53:337–351. https://doi.org/10.1016/j.neuron.2007.01.010 CrossRefPubMed

Zerbi V et al (2013) Microvascular cerebral blood volume changes in aging APP(swe)/PS1(dE9) AD mouse model: a voxel-wise approach. Brain Struct Funct 218:1085–1098. https://doi.org/10.1007/s00429-012-0448-8 CrossRefPubMed

Title: Analysis of the vasculature by immunohistochemistry in paraffin-embedded brains
Authors: Yann Decker
Andreas Müller
Eszter Németh
Walter J. Schulz-Schaeffer
Marc Fatar
Michael D. Menger
Yang Liu
Klaus Fassbender
Publication date: 01-03-2018
Publisher: Springer Berlin Heidelberg
Published in: Brain Structure and Function / Issue 2/2018
Print ISSN: 1863-2653
Electronic ISSN: 1863-2661
DOI: https://doi.org/10.1007/s00429-017-1595-8

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Analysis of the vasculature by immunohistochemistry in paraffin-embedded brains

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