title: Diverse activation of microglia by chemokine (C-C motif) ligand 2 overexpression in brain.
abstract: BACKGROUND
  The chemokine (C-C motif) ligand 2 (CCL2) is a monocyte chemoattractant protein that mediates macrophage recruitment and migration during peripheral and central nervous system (CNS) inflammation.
  
  
  METHODS
  To determine the impact of CCL2 in inflammation in vivo and to elucidate the CCL2-induced polarization of activated brain microglia, we delivered CCL2 into the brains of wild-type mice via recombinant adeno-associated virus serotype 9 (rAAV-9) driven by the chicken β-actin promoter. We measured microglial activation using histological and chemical measurement and recruitment of monocytes using histology and flow cytometry.
  
  
  RESULTS
  The overexpression of CCL2 in the CNS induced significant activation of brain resident microglia. CD45 and major histocompatibility complex class II immunoreactivity significantly increased at the sites of CCL2 administration. Histological characterization of the microglial phenotype revealed the elevation of "classically activated" microglial markers, such as calgranulin B and IL-1β, as well as markers associated with "alternative activation" of microglia, including YM1 and arginase 1. The protein expression profile in the hippocampus demonstrated markedly increased levels of IL-6, GM-CSF and eotaxin (CCL-11) in response to CCL2, but no changes in the levels of other cytokines, including TNF-α and IFN-γ. Moreover, real-time PCR analysis confirmed increases in mRNA levels of gene transcripts associated with neuroinflammation following CCL2 overexpression. Finally, we investigated the chemotactic properties of CCL2 in vivo by performing adoptive transfer of bone marrow-derived cells (BMDCs) isolated from donor mice that ubiquitously expressed green fluorescent protein. Flow cytometry and histological analyses indicated that BMDCs extravasated into brain parenchyma and colabeled with microglial markers.
  
  
  CONCLUSIONS
  Taken together, our results suggest that CCL2 strongly activates resident microglia in the brain. Both pro- and anti-inflammatory activation of microglia were prominent, with no bias toward the M1 or M2 phenotype in the activated cells. As expected, CCL2 overexpression actively recruited circulating monocytes into the CNS. Thus, CCL2 expression in mouse brain induces microglial activation and represents an efficient method for recruitment of peripheral macrophages.
authors.affiliation: Department of Molecular Pharmacology & Physiology, Byrd Alzheimer Institute, University of South Florida, Tampa, FL, USA.
authors.forename: Maj-Linda B
authors.initials: ML
authors.lastname: Selenica
authors.forename: Jennifer A
authors.initials: JA
authors.lastname: Alvarez
authors.forename: Kevin R
authors.initials: KR
authors.lastname: Nash
authors.forename: Daniel C
authors.initials: DC
authors.lastname: Lee
authors.forename: Chuanhai
authors.initials: C
authors.lastname: Cao
authors.forename: Xiaoyang
authors.initials: X
authors.lastname: Lin
authors.forename: Patrick
authors.initials: P
authors.lastname: Reid
authors.forename: Peter R
authors.initials: PR
authors.lastname: Mouton
authors.forename: Dave
authors.initials: D
authors.lastname: Morgan
authors.forename: Marcia N
authors.initials: MN
authors.lastname: Gordon
mesh_terms: D019264:Adoptive Transfer; D000818:Animals; D001854:Bone Marrow Cells / Q000187:drug effects; D001923:Brain Chemistry / Q000502:physiology*; D002465:Cell Movement / Q000187:drug effects; D018932:Chemokine CCL2 / Q000096:biosynthesis / Q000502:physiology*; D016207:Cytokines / Q000096:biosynthesis; D000229:Dependovirus / Q000235:genetics; D015870:Gene Expression / Q000187:drug effects; D018014:Gene Transfer Techniques; D005822:Genetic Vectors; D049452:Green Fluorescent Proteins; D006801:Humans; D007091:Image Processing, Computer-Assisted; D007150:Immunohistochemistry; D008262:Macrophage Activation / Q000187:drug effects*; D051379:Mice; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic; D017628:Microglia / Q000187:drug effects*; D060888:Real-Time Polymerase Chain Reaction; D015398:Signal Transduction / Q000187:drug effects
chemical_list: D018932:Chemokine CCL2; D016207:Cytokines; D049452:Green Fluorescent Proteins