title: TGF-beta1 blockade of microglial chemotaxis toward Abeta aggregates involves SMAD signaling and down-regulation of CCL5.
abstract: BACKGROUND
  Overactivated microglia that cluster at neuritic plaques constantly release neurotoxins, which actively contribute to progressive neurodegeneration in Alzheimer's disease (AD). Therefore, attenuating microglial clustering can reduce focal neuroinflammation at neuritic plaques. Previously, we identified CCL5 and CCL2 as prominent chemokines that mediate the chemotaxis of microglia toward beta-amyloid (Abeta)aggregates. Although transforming growth factor-beta1 (TGF-beta1) has been shown to down-regulate the expression of chemokines in activated microglia, whether TGF-beta1 can reduce the chemotaxis of microglia toward neuritic plaques in AD remains unclear.
  
  
  METHODS
  In the present study, we investigated the effects of TGF-beta1 on Abeta-induced chemotactic migration of BV-2 microglia using time-lapse recording, transwell assay, real-time PCR, ELISA, and western blotting.
  
  
  RESULTS
  The cell tracing results suggest that the morphological characteristics and migratory patterns of BV-2 microglia resemble those of microglia in slice cultures. Using this model system, we discovered that TGF-beta1 reduces Abeta-induced BV-2 microglial clustering in a dose-dependent manner. Chemotactic migration of these microglial cells toward Abeta aggregates was significantly attenuated by TGF-beta1. However, these microglia remained actively moving without any reduction in migration speed. Pharmacological blockade of TGF-beta1 receptor I (ALK5) by SB431542 treatment reduced the inhibitory effects of TGF-beta1 on Abeta-induced BV-2 microglial clustering, while preventing TGF-beta1-mediated cellular events, including SMAD2 phosphorylation and CCL5 down-regulation.
  
  
  CONCLUSIONS
  Our results suggest that TGF-beta1 reduces Abeta-induced microglial chemotaxis via the SMAD2 pathway. The down-regulation of CCL5 by TGF-beta1 at least partially contributes to the clustering of microglia at Abeta aggregates. The attenuating effects of SB431542 upon TGF-beta1-suppressed microglial clustering may be mediated by restoration of CCL5 to normal levels. TGF-beta1 may ameliorate microglia-mediated neuroinflammation in AD by preventing activated microglial clustering at neuritic plaques.
authors.affiliation: Institute of Neuroscience, Brain Research Center, National Yang-Ming University, Taipei 11221, Taiwan.
authors.forename: Wei-Chao
authors.initials: WC
authors.lastname: Huang
authors.forename: Feng-Chang
authors.initials: FC
authors.lastname: Yen
authors.forename: Feng-Shiun
authors.initials: FS
authors.lastname: Shie
authors.forename: Chih-Ming
authors.initials: CM
authors.lastname: Pan
authors.forename: Young-Ji
authors.initials: YJ
authors.lastname: Shiao
authors.forename: Cheng-Ning
authors.initials: CN
authors.lastname: Yang
authors.forename: Fong-Lee
authors.initials: FL
authors.lastname: Huang
authors.forename: Yen-Jen
authors.initials: YJ
authors.lastname: Sung
authors.forename: Huey-Jen
authors.initials: HJ
authors.lastname: Tsay
mesh_terms: D016229:Amyloid beta-Peptides / Q000276:immunology*; D000818:Animals; D001549:Benzamides / Q000494:pharmacology; D015153:Blotting, Western; D002460:Cell Line; D002465:Cell Movement / Q000187:drug effects; D018946:Chemokine CCL5 / Q000235:genetics / Q000502:physiology*; D002633:Chemotaxis / Q000187:drug effects*; D004149:Dioxoles / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D015536:Down-Regulation / Q000502:physiology; D004797:Enzyme-Linked Immunosorbent Assay; D017628:Microglia / Q000187:drug effects*; D010766:Phosphorylation; D051381:Rats; D018125:Receptors, Transforming Growth Factor beta / Q000037:antagonists & inhibitors; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology; D051785:Smad Proteins / Q000235:genetics / Q000502:physiology*; D053773:Transforming Growth Factor beta1 / Q000494:pharmacology*
chemical_list: C459179:4-(5-benzo(1,3)dioxol-5-yl-4-pyridin-2-yl-1H-imidazol-2-yl)benzamide; D016229:Amyloid beta-Peptides; D001549:Benzamides; D018946:Chemokine CCL5; D004149:Dioxoles; D018125:Receptors, Transforming Growth Factor beta; D051785:Smad Proteins; D053773:Transforming Growth Factor beta1