Deficiency of Socs3 leads to brain-targeted experimental autoimmune encephalomyelitis via enhanced neutrophil activation and ROS production
Zhaoqi Yan, Wei Yang, Luke Parkitny, Sara A. Gibson, Kevin S. Lee, Forrest Collins, Jessy S. Deshane, Wayne Cheng, Amy S. Weinmann, Hairong Wei, Hongwei Qin, Etty N. Benveniste
Zhaoqi Yan, Wei Yang, Luke Parkitny, Sara A. Gibson, Kevin S. Lee, Forrest Collins, Jessy S. Deshane, Wayne Cheng, Amy S. Weinmann, Hairong Wei, Hongwei Qin, Etty N. Benveniste
View: Text | PDF
Research Article Inflammation

Deficiency of Socs3 leads to brain-targeted experimental autoimmune encephalomyelitis via enhanced neutrophil activation and ROS production

Abstract

Dysregulation of the JAK/STAT signaling pathway is associated with multiple sclerosis (MS) and its mouse model, experimental autoimmune encephalomyelitis (EAE). Suppressors of cytokine signaling (SOCS) negatively regulate the JAK/STAT pathway. We previously reported a severe, brain-targeted, atypical form of EAE in mice lacking Socs3 in myeloid cells (Socs3ΔLysM), and that this atypical EAE is associated with cerebellar neutrophil infiltration. There is emerging evidence that neutrophils are detrimental in the pathology of MS/EAE; however, their exact function is unclear. Here we demonstrate that neutrophils from the cerebellum of Socs3ΔLysM mice show a hyperactivated phenotype with excessive production of reactive oxygen species (ROS) at the peak of EAE. Neutralization of ROS in vivo delayed the onset and reduced severity of atypical EAE. Mechanistically, Socs3-deficient neutrophils exhibited enhanced signal transducer and activator of transcription 3 (STAT3) activation, a hyperactivated phenotype in response to granulocyte colony–stimulating factor (G-CSF), and upon G-CSF priming, increased ROS production. Neutralization of G-CSF in vivo significantly reduced the incidence and severity of the atypical EAE phenotype. Overall, our work elucidates that hypersensitivity of G-CSF/STAT3 signaling in Socs3ΔLysM mice leads to atypical EAE by enhanced neutrophil activation and increased oxidative stress, which may explain the detrimental role of G-CSF in MS patients.

Authors

Zhaoqi Yan, Wei Yang, Luke Parkitny, Sara A. Gibson, Kevin S. Lee, Forrest Collins, Jessy S. Deshane, Wayne Cheng, Amy S. Weinmann, Hairong Wei, Hongwei Qin, Etty N. Benveniste

×

Figure 3

Socs3 deficiency promotes G-CSF hypersensitivity in neutrophils via JAK1 activation.

Options: View larger image (or click on image) Download as PowerPoint
Socs3 deficiency promotes G-CSF hypersensitivity in neutrophils via JAK...
(A) Bone marrow neutrophils were isolated from C57BL/6 mice and stimulated with G-CSF (10 ng/ml), IL-6 (100 ng/ml), IL-23 (10 ng/ml), or IFN-γ (10 ng/ml) for 2 hours. Expression of Socs3 mRNA was analyzed by qRT-PCR (n = 4). (B–E) Bone marrow neutrophils were isolated from Socs3fl/fl or Socs3ΔLysM mice (n = 3). (B and C) Neutrophils were stimulated with G-CSF (10 ng/ml) or IL-6 (100 ng/ml) for 2 hours followed by intracellular staining for phosphorylated STAT3 (p-STAT3) (Y705). (D and E) Neutrophils were stimulated with G-CSF (10 ng/ml) for 8 hours, and expression of surface markers was analyzed. (F) Bone marrow neutrophils isolated from Socs3ΔLysM mice were pretreated with AZD1480 (25 μM) or PF8041 (25 μM) for 2 hours and then stimulated with G-CSF (10 ng/ml) for 8 hours, followed by surface staining. Plots are representative of 3 independent experiments. All flow cytometry plots (B–F) were gated on live, single CD45+CD11b+Ly6G+ neutrophils. All error bars represent ± SEM. *P < 0.05, **P < 0.01, and ***P < 0.001 by 1-way ANOVA (A) or 2-tailed Student’s t test (C and E). MFI, mean fluorescence intensity.