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Neuroscience

Trois dimensions d'analyse confocale de marqueurs microglies / macrophages de polarisation dans Experimental Brain Injury

Published: September 4, 2013 doi: 10.3791/50605
* These authors contributed equally

Materials

Name Company Catalog Number Comments
Materials
Rat Anti-mouse CD11b Kindly provided by Dr. A. Doni, Istituto Clinico Humanitas, Milan, Italy
Rat Anti-mouse CD68 AbD Serotec MCA 1957
Rabbit Anti-mouse Ym1 Stem Cell Technologies 1404
H–chst 33342 Life technologies H21492
Mouse Anti-Neural Nuclei (NeuN) CHEMICON MAB377
Biotinilated Goat Anti-Rat antibody Jackson Immuno Research 112-065-143
TSA Cyanine 5 System Perkin Elmer NEL705A001KT
Prolong Gold Invitrogen P36930
Anti-rat alexa 546 Invitrogen A-11081
Anti mouse Alexa 488 Invitrogen A-21121
Anti-rabbit Alexa 594 Invitrogen A-11037
Normal Goat Serum Vectors Laboratories S-1000
Tritin X-100 Sigma T8787
Phosphate Buffered Saline Sigma P4417-100
Equipment
Cryostat CM1850 Leica
Olympus IX81 confocal microscope Olympus
AnalySIS software Olympus
Imaris software 5.0 Bitplane
Photoshop cs2 Adobe Systems
Software packages GraphPad Prism version 4.0 GraphPad Software Inc.

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References

  1. Davalos, D., et al. ATP mediates rapid microglial response to local brain injury in vivo. Nat. Neurosci. 8, 752-758 (2005).
  2. Yenari, M. A., Kauppinen, T. M., Swanson, R. A. Microglial activation in stroke: therapeutic targets. Neurotherapeutics. 7, 378-391 (2010).
  3. Iadecola, C., Anrather, J. The immunology of stroke: from mechanisms to translation. Nat. Med. 17, 796-808 (2011).
  4. Jin, R., Yang, G., Li, G. Inflammatory mechanisms in ischemic stroke: role of inflammatory cells. J. Leukoc. Biol. 87, 779-789 (2010).
  5. Schilling, M., Besselmann, M., Muller, M., Strecker, J. K., Ringelstein, E. B., Kiefer, R. Predominant phagocytic activity of resident microglia over hematogenous macrophages following transient focal cerebral ischemia: an investigation using green fluorescent protein transgenic bone marrow chimeric mice. Exp. Neurol. 196, 290-297 (2005).
  6. Perego, C., Fumagalli, S., De Simoni, M. G. Temporal pattern of expression and colocalization of microglia/macrophage phenotype markers following brain ischemic injury in mice. Journal of Neuroinflammation. 8, 174-193 (2011).
  7. Zanier, E. R., et al. Human umbilical cord blood mesenchymal stem cells protect mice brain after trauma. Crit. CareMed. 39 (11), 2501-2510 (2011).
  8. Capone, C., et al. Neurosphere derived cells exert a neuroprotective action by changing the ischemic microenvironment. PLoS ONE. 2, e373 (2007).
  9. Bhatia, S., et al. Rapid host defense against Aspergillus fumigatus involves alveolar macrophages with a predominance ofalternatively activated phenotype. PLoS One. 6, e15943 (2011).
  10. Raes, G., Noel, W., Beschin, A., Brys, L., de Baetselier, P., Hassanzadeh, G. H. FIZZ1 and Ym as tools to discriminate between differentially activated macrophages. Dev. Immunol. 9, 151-159 (2002).
  11. Gesuete, R., et al. Recombinant C1 inhibitor in brain ischemic injury. Ann. Neurol. 66, 332-342 (2009).
  12. Sica, A., Mantovani, A. Macrophages plasticity and polarization: in vivo veritas. J. Clin. Invest. 122 (3), 787-795 (2012).
Trois dimensions d'analyse confocale de marqueurs microglies / macrophages de polarisation dans Experimental Brain Injury
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Cite this Article

Perego, C., Fumagalli, S., DeMore

Perego, C., Fumagalli, S., De Simoni, M. G. Three-dimensional Confocal Analysis of Microglia/macrophage Markers of Polarization in Experimental Brain Injury. J. Vis. Exp. (79), e50605, doi:10.3791/50605 (2013).

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