title: CCR2 overexpression promotes the efficient recruitment of retinal microglia in vitro.
abstract: OBJECTIVE
  Retinal microglia can be activated and recruited by chemokines and play a protective role in early retinal degeneration. CC-chemokine ligand 2 (CCL2) and its receptor, CC-chemokine receptor 2 (CCR2), have been implicated as key mediators for the trafficking and accumulation of microglial cells in lesioned tissue. The current study investigates whether the overexpression of CCR2 allows microglia to migrate toward CCL2 more efficiently.
  
  
  METHODS
  Primary microglial cells were transduced with lentivirus carrying green fluorescent protein (GFP)-tagged CCR2 (CCR2-GFP). Overexpression of CCR2 was assessed by western blot analysis and fluorescence-assisted cell sorting. The chemotaxis of primary microglia transduced with lentivirus carrying CCR2-GFP was compared to either those transduced with GFP alone or those not transduced, using a chemotaxis chamber assay.
  
  
  RESULTS
  Primary microglia showed a high transduction rate following lentivirus application and maintained normal microglial morphology and a significant overexpression of CCR2 protein. We found that CCL2-mediated chemotaxis is concentration and time dependent in microglia. The chemotactic response of microglia cells overexpressing CCR2-GFP was significantly increased compared to that of nontransduced and GFP-expressing microglia.
  
  
  CONCLUSIONS
  These findings suggest that microglia can be efficiently transduced with CCR2-GFP lentiviral vectors and that the overexpression of CCR2 in retinal microglia promotes their chemotaxis in response to chemokines, suggesting that these cells may be promising targets for cell-based therapeutic manipulation in retinal disease.
authors.affiliation: Department of Ophthalmology, Eye & ENT Hospital of Fudan University, Shanghai, People's Republic of China.
authors.forename: Xiao-shuang
authors.initials: XS
authors.lastname: Jiang
authors.forename: Ying-qin
authors.initials: YQ
authors.lastname: Ni
authors.forename: Tian-jin
authors.initials: TJ
authors.lastname: Liu
authors.forename: Meng
authors.initials: M
authors.lastname: Zhang
authors.forename: Hui
authors.initials: H
authors.lastname: Ren
authors.forename: Rui
authors.initials: R
authors.lastname: Jiang
authors.forename: Xin
authors.initials: X
authors.lastname: Huang
authors.forename: Ge-zhi
authors.initials: GZ
authors.lastname: Xu
mesh_terms: D000818:Animals; D000831:Animals, Newborn; D015153:Blotting, Western; D018932:Chemokine CCL2 / Q000494:pharmacology*; D002633:Chemotaxis / Q000187:drug effects; D015194:Diffusion Chambers, Culture; D004305:Dose-Response Relationship, Drug; D015870:Gene Expression; D017930:Genes, Reporter; D005822:Genetic Vectors; D049452:Green Fluorescent Proteins; D016086:Lentivirus / Q000235:genetics; D017628:Microglia / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D061251:Primary Cell Culture; D051381:Rats; D017207:Rats, Sprague-Dawley; D054390:Receptors, CCR2 / Q000235:genetics* / Q000378:metabolism; D012160:Retina / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D014161:Transduction, Genetic; D019076:Transgenes
chemical_list: C491766:Ccl2 protein, rat; C515336:Ccr2 protein, rat; D018932:Chemokine CCL2; D054390:Receptors, CCR2; D049452:Green Fluorescent Proteins