title: α-Synuclein vaccination modulates regulatory T cell activation and microglia in the absence of brain pathology.
abstract: BACKGROUND
  Passive and active immunization with α-synuclein has been shown to be neuroprotective in animal models of Parkinson's disease. We have previously shown that vaccination with α-synuclein, long before α-synuclein-induced brain pathology, prevents striatal degeneration by inducing regulatory T cell infiltration in parenchyma and antibody deposition on α-synuclein overexpressing neurons. However, the effect of peripheral α-synuclein on the immune system is unknown, as are the mechanistic changes induced in the CD4 T cell population during successful neuroprotective animal studies. We have studied the changes induced by vaccination with α-synuclein in the CD4 T cell pool and its impact on brain microglia to understand the immune mechanisms behind successful vaccination strategies in Parkinson's disease animal models.
  
  
  METHODS
  Mice were immunized with WT or nitrated α-synuclein at a dose equivalent to the one used in our previous successful vaccination strategy and at a higher dose to determine potential dose-dependent effects. Animals were re-vaccinated 4 weeks after and sacrificed 5 days later. These studies were conducted in naive animals in the absence of human α-synuclein expression.
  
  
  RESULTS
  The CD4 T cell response was modulated by α-synuclein in a dose-dependent manner, in particular the regulatory T cell population. Low-dose α-synuclein induced expansion of naive (Foxp3 + CCR6-CD127lo/neg) and dopamine receptor type D3+ regulatory T cells, as well as an increase in Stat5 protein levels. On the other hand, high dose promoted activation of regulatory T cells (Foxp3CCR6 + CD127lo/neg), which were dopamine receptor D2+D3-, and induced up-regulation of Stat5 and production of anti-α-synuclein antibodies. These effects were specific to the variant of α-synuclein used as the pathology-associated nitrated form induced distinct effects at both doses. The changes observed in the periphery after vaccination with low-dose α-synuclein correlated with an increase in CD154+, CD103+, and CD54+ microglia and the reduction of CD200R+ microglia. This resulted in the induction of a polarized tolerogenic microglia population that was CD200R-CD54CD103CD172a+ (82 % of total microglia).
  
  
  CONCLUSIONS
  We have shown for the first time the mechanisms behind α-synuclein vaccination and, importantly, how we can modulate microglia's phenotype by regulating the CD4 T cell pool, thus shedding invaluable light on the design of neuroimmunoregulatory therapies for Parkinson's disease.
authors.affiliation: Neuroimmunology of Degenerative Diseases group, Department of Biomedicine, HEALTH, Aarhus University, Aarhus, Denmark.|CNS Disease Modeling group, Department of Biomedicine, HEALTH , Aarhus University, Aarhus, Denmark.|AU Ideas Pilot Center NEURODIN, Department of Biomedicine, HEALTH, Aarhus University, Aarhus, Denmark.
authors.forename: Josefine R
authors.initials: JR
authors.lastname: Christiansen
authors.affiliation: Neuroimmunology of Degenerative Diseases group, Department of Biomedicine, HEALTH, Aarhus University, Aarhus, Denmark.|AU Ideas Pilot Center NEURODIN, Department of Biomedicine, HEALTH, Aarhus University, Aarhus, Denmark.
authors.forename: Mads N
authors.initials: MN
authors.lastname: Olesen
authors.affiliation: Interdisciplinary Nanoscience Center - iNANO, Department of Molecular Biology and Genetics, Aarhus University, Aarhus, Denmark.
authors.forename: Daniel E
authors.initials: DE
authors.lastname: Otzen
authors.affiliation: CNS Disease Modeling group, Department of Biomedicine, HEALTH , Aarhus University, Aarhus, Denmark.|AU Ideas Pilot Center NEURODIN, Department of Biomedicine, HEALTH, Aarhus University, Aarhus, Denmark.
authors.forename: Marina
authors.initials: M
authors.lastname: Romero-Ramos
authors.affiliation: Neuroimmunology of Degenerative Diseases group, Department of Biomedicine, HEALTH, Aarhus University, Aarhus, Denmark. v.sanchez-guajardo@biomed.au.dk.|AU Ideas Pilot Center NEURODIN, Department of Biomedicine, HEALTH, Aarhus University, Aarhus, Denmark. v.sanchez-guajardo@biomed.au.dk.
authors.forename: Vanesa
authors.initials: V
authors.lastname: Sanchez-Guajardo
mesh_terms: D000818:Animals; D015153:Blotting, Western; D004797:Enzyme-Linked Immunosorbent Assay; D005434:Flow Cytometry; D006801:Humans; D007150:Immunohistochemistry; D008213:Lymphocyte Activation / Q000276:immunology; D051379:Mice; D008810:Mice, Inbred C57BL; D017628:Microglia / Q000276:immunology*; D020734:Parkinsonian Disorders / Q000276:immunology* / Q000517:prevention & control; D011994:Recombinant Proteins / Q000276:immunology; D050378:T-Lymphocytes, Regulatory / Q000276:immunology*; D014611:Vaccination; D051844:alpha-Synuclein / Q000008:administration & dosage / Q000276:immunology*
chemical_list: D011994:Recombinant Proteins; D051844:alpha-Synuclein
keywords: CCR6; CD103; Dopamine receptor D2 (DR-D2); Dopamine receptor D3 (DR-D3); Foxp3; Immunotherapy; Neuroinflammation; Parkinson’s disease; Stat5