June 2015
Volume 56, Issue 7
Free
ARVO Annual Meeting Abstract  |   June 2015
Mouse Model for Tamoxifen-inducible Cre Targeting of Retinal Microglia
Author Affiliations & Notes
  • Senthil Selvam
    Institute of Ophthalmology, University College London, London, United Kingdom
    Moorfields Eye Hospital, London, United Kingdom
  • Andrew Scott
    Moorfields Eye Hospital, London, United Kingdom
  • Sidath E Liyanage
    Institute of Ophthalmology, University College London, London, United Kingdom
  • Michael Powner
    Institute of Ophthalmology, University College London, London, United Kingdom
  • Marcus Fruttiger
    Institute of Ophthalmology, University College London, London, United Kingdom
  • Footnotes
    Commercial Relationships Senthil Selvam, Fight for Sight (F); Andrew Scott, None; Sidath Liyanage, None; Michael Powner, None; Marcus Fruttiger, Amakem (F), AstraZeneca (F), Fight for Sight (F), Novartis (C), Novartis (F)
  • Footnotes
    Support None
Investigative Ophthalmology & Visual Science June 2015, Vol.56, 3556. doi:https://doi.org/
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      Senthil Selvam, Andrew Scott, Sidath E Liyanage, Michael Powner, Marcus Fruttiger; Mouse Model for Tamoxifen-inducible Cre Targeting of Retinal Microglia. Invest. Ophthalmol. Vis. Sci. 2015;56(7 ):3556. doi: https://doi.org/.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract
 
Purpose
 

Microglia are a specialised population of the mononuclear phagocyte lineage. Resident microglia are distinct to invading populations of circulating monocyte-derived macrophages in their localisation to the central nervous system (CNS) and their homeostatic role in the neurogeneis and angiogenesis of the developing retina. Microglial activation is commonly seen in inflammatory and degenerative pathologies of the retina. Resident retinal microglia are known to specifically express the protein ionized calcium binding adaptor molecule 1 (Iba1; also known as allograft inflammatory factor-1, Aif1). Using the Aif1 gene we present a mouse model of tamoxifen inducible, cre-recombinase activation that allows the genetic targeting of resident central nervous system microglia.

 
Methods
 

A bacterial artificial chromosome (BAC), containing the Aif1 gene was obtained and a construct coding for a tamoxifen inducible form of Cre (CreERT2) was recombined into the open reading frame of the Aif1 gene. The modified BAC was then used for transgenesis by pronuclear injections and resulting offspring was screened for founders by PCR. In vivo recombination efficiency was tested in a ROSA-EGFP reporter strain (Gt(ROSA)26Sortm4(ACTB-tdTomato,-EGFP)Luo/J) using flow cytometry in both p7 neonatal mice and 6 week old adult mice. Using the same strain we demonstrate the use of this tool to label Iba1 expressing resident retinal microglia in murine models of oxygen induced retinopathy (OIR), laser induced choroidal neovasculairsation (CNV) and endotoxin induced uveitis (EIU).

 
Results
 

Two founders were obtained and a transgenic line was established from one of them (Aif1-CreER). Flow cytometry of the ROSA-EGFP reporter mice showed that recombination could be achieved in upto 72% of resident microglia in the p7 perinatal retina and upto 95% in the 6 week adult retina. The brain, choroid and spleen also showed GFP expression. Immunohistochemistry performed on retinal flat mounts of ROSA-EGFP reporter mice undergoing OIR, laser induced CNV and EIU demonstrated co-localisation of GFP expression and Iba1 antibody labelling.

 
Conclusions
 

This model of Aif1-CreER provides a valuable research tool to genetically target resident microglia in the central nervous system of mice in various murine models of retinal disease.  

 
GFP expression in resident retinal microglia.
 
GFP expression in resident retinal microglia.

 
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