title: Expression pattern of Ccr2 and Cx3cr1 in inherited retinal degeneration.
abstract: BACKGROUND
  Though accumulating evidence suggests that microglia, resident macrophages in the retina, and bone marrow-derived macrophages can cause retinal inflammation which accelerates photoreceptor cell death, the details of how these cells are activated during retinal degeneration (RD) remain uncertain. Therefore, it is important to clarify which cells play a dominant role in fueling retinal inflammation. However, distinguishing between microglia and macrophages is difficult using conventional techniques such as cell markers (e.g., Iba-1). Recently, two mouse models for visualizing chemokine receptors were established, Cx3cr1 (GFP/GFP) and Ccr2 (RFP/RFP) mice. As Cx3cr1 is expressed in microglia and Ccr2 is reportedly expressed in activated macrophages, these mice have the potential to distinguish microglia and macrophages, yielding novel information about the activation of these inflammatory cells and their individual roles in retinal inflammation.
  
  
  METHODS
  In this study, c-mer proto-oncogene tyrosine kinase (Mertk) (-/-) mice, which show photoreceptor cell death due to defective retinal pigment epithelium phagocytosis, were employed as an animal model of RD. Mertk (-/-) Cx3cr1 (GFP/+) Ccr2 (RFP/+) mice were established by breeding Mertk (-/-) , Cx3cr1 (GFP/GFP) , and Ccr2 (RFP/RFP) mice. The retinal morphology and pattern of inflammatory cell activation and invasion of Mertk (-/-) Cx3cr1 (GFP/+) Ccr2 (RFP/+) mice were evaluated using retina and retinal pigment epithelium (RPE) flat mounts, retinal sections, and flow cytometry.
  
  
  RESULTS
  Four-week-old Mertk (-/-) Cx3cr1 (GFP/+) Ccr2 (RFP/+) mice showed Cx3cr1-GFP-positive microglia in the inner retina. Cx3cr1-GFP and Ccr2-RFP dual positive activated microglia were observed in the outer retina and subretinal space of 6- and 8-week-old animals. Ccr2-RFP single positive bone marrow-derived macrophages were observed to migrate into the retina of Mertk (-/-) Cx3cr1 (GFP/+) Ccr2 (RFP/+) mice. These invading cells were still observed in the subretinal space in 18-week-old animals.
  
  
  CONCLUSIONS
  Cx3cr1-GFP-positive microglia and Ccr2-RFP-positive macrophages were distinguishable in the retinas of Mertk (-/-) Cx3cr1 (GFP/+) Ccr2 (RFP/+) mice. In addition, Ccr2 expression in Cx3cr1 positive microglia is a feature of microglial activation in RD. Mertk (-/-) Cx3cr1 (GFP/+) Ccr2 (RFP/+) mice enabled observation of microglial activation over time during RD and may be useful for developing inflammation-targeted treatment strategies for RD in the future.
authors.affiliation: Department of Ophthalmology, The Jikei University School of Medicine, 105-8461, Tokyo, Japan. kohnohideo@jikei.ac.jp.|Tokyu Hospital, 145-0062, Tokyo, Japan. kohnohideo@jikei.ac.jp.
authors.forename: Hideo
authors.initials: H
authors.lastname: Kohno
authors.affiliation: Division of Molecular and Developmental Biology, The Institute of Medical Science, The University of Tokyo, 108-8639, Tokyo, Japan.
authors.forename: Hideto
authors.initials: H
authors.lastname: Koso
authors.affiliation: Department of Ophthalmology, The Jikei University School of Medicine, 105-8461, Tokyo, Japan.
authors.forename: Kiichiro
authors.initials: K
authors.lastname: Okano
authors.affiliation: Department of Pharmacology, School of Medicine, Case Western Reserve University, Cleveland, OH, USA.
authors.forename: Thomas R
authors.initials: TR
authors.lastname: Sundermeier
authors.affiliation: Department of Molecular Immunology, The Jikei University School of Medicine, 105-8461, Tokyo, Japan.
authors.forename: Saburo
authors.initials: S
authors.lastname: Saito
authors.affiliation: Division of Molecular and Developmental Biology, The Institute of Medical Science, The University of Tokyo, 108-8639, Tokyo, Japan.
authors.forename: Sumiko
authors.initials: S
authors.lastname: Watanabe
authors.affiliation: Department of Ophthalmology, The Jikei University School of Medicine, 105-8461, Tokyo, Japan.
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Tsuneoka
authors.affiliation: Department of Ophthalmology, The Jikei University School of Medicine, 105-8461, Tokyo, Japan. tstmski@jikei.ac.jp.
authors.forename: Tsutomu
authors.initials: T
authors.lastname: Sakai
mesh_terms: D000818:Animals; D000073883:CX3C Chemokine Receptor 1; D002465:Cell Movement / Q000235:genetics; D004195:Disease Models, Animal; D005260:Female; D005786:Gene Expression Regulation / Q000235:genetics*; D007962:Leukocytes / Q000378:metabolism / Q000473:pathology; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D008264:Macrophages / Q000378:metabolism / Q000473:pathology; D008297:Male; D051379:Mice; D008822:Mice, Transgenic; D017628:Microglia / Q000378:metabolism / Q000473:pathology; D009154:Mutation / Q000235:genetics; D009474:Neurons / Q000378:metabolism / Q000473:pathology; D011518:Proto-Oncogene Proteins / Q000172:deficiency / Q000235:genetics; D020794:Receptor Protein-Tyrosine Kinases / Q000172:deficiency / Q000235:genetics; D054390:Receptors, CCR2 / Q000235:genetics / Q000378:metabolism*; D019707:Receptors, Chemokine / Q000235:genetics / Q000378:metabolism*; D012160:Retina / Q000378:metabolism / Q000473:pathology; D012162:Retinal Degeneration / Q000235:genetics* / Q000378:metabolism*; D055213:Retinal Pigment Epithelium / Q000378:metabolism / Q000473:pathology; D013997:Time Factors; D000076202:c-Mer Tyrosine Kinase
chemical_list: D000073883:CX3C Chemokine Receptor 1; C515335:Ccr2 protein, mouse; C471885:Cx3cr1 protein, mouse; D008164:Luminescent Proteins; D011518:Proto-Oncogene Proteins; D054390:Receptors, CCR2; D019707:Receptors, Chemokine; C490767:Mertk protein, mouse; D020794:Receptor Protein-Tyrosine Kinases; D000076202:c-Mer Tyrosine Kinase