β1-Integrin– and KV1.3 channel–dependent signaling stimulates glutamate release from Th17 cells
Katharina Birkner, … , Frauke Zipp, Stefan Bittner
Katharina Birkner, … , Frauke Zipp, Stefan Bittner
Published October 29, 2019
Citation Information: J Clin Invest. 2020;130(2):715-732. https://doi.org/10.1172/JCI126381.
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Research Article Autoimmunity Neuroscience Article has an altmetric score of 14

β1-Integrin– and KV1.3 channel–dependent signaling stimulates glutamate release from Th17 cells

Abstract

Although the impact of Th17 cells on autoimmunity is undisputable, their pathogenic effector mechanism is still enigmatic. We discovered soluble N-ethylmaleimide–sensitive factor attachment receptor (SNARE) complex proteins in Th17 cells that enable a vesicular glutamate release pathway that induces local intracytoplasmic calcium release and subsequent damage in neurons. This pathway is glutamine dependent and triggered by binding of β1-integrin to vascular cell adhesion molecule 1 (VCAM-1) on neurons in the inflammatory context. Glutamate secretion could be blocked by inhibiting either glutaminase or KV1.3 channels, which are known to be linked to integrin expression and highly expressed on stimulated T cells. Although KV1.3 is not expressed in CNS tissue, intrathecal administration of a KV1.3 channel blocker or a glutaminase inhibitor ameliorated disability in experimental neuroinflammation. In humans, T cells from patients with multiple sclerosis secreted higher levels of glutamate, and cerebrospinal fluid glutamine levels were increased. Altogether, our findings demonstrate that β1-integrin– and KV1.3 channel–dependent signaling stimulates glutamate release from Th17 cells upon direct cell-cell contact between Th17 cells and neurons.

Authors

Katharina Birkner, Beatrice Wasser, Tobias Ruck, Carine Thalman, Dirk Luchtman, Katrin Pape, Samantha Schmaul, Lynn Bitar, Eva-Maria Krämer-Albers, Albrecht Stroh, Sven G. Meuth, Frauke Zipp, Stefan Bittner

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Figure 7

KV1.3-mediated glutamate release is enhanced by activation of β1-integrin (CD29) and upon VCAM-1 binding.

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KV1.3-mediated glutamate release is enhanced by activation of β1-integri...
(A) Naive, Th17, and Th1 cells were stimulated for 4 hours and stained for CD29. Mean fluorescence intensity (MFI) is shown (n = 3–6). (B) Flow cytometric staining for KCNA3 expression (n = 6) and IL-7 production (n = 6) was performed on Th17 cells, comparing CD29hi- and CD29lo-expressing cell populations. (C) Glutamate levels in unstimulated or TCR-stimulated Th1 and Th17 cells with additional CD29 stimulation or VCAM-1 coating (n = 10–12 per group), 24 hours after culturing in HEPES complete. (D) Vcam1 mRNA expression levels in cortical neurons that were unstimulated, treated with IFN-γ, or stimulated with LPS, and after application of splenocyte supernatant (n = 5–9). (E) VCAM-1 protein analysis and quantification of cortical neurons with or without LPS stimulation or splenocyte supernatant addition. Western blot analysis showed protein levels in kDa (n = 3–4). (F) Immunohistochemical staining for VCAM-1 in infiltrated EAE lesions. Scale bars: 6 μm and 2 μm (zoom, enlarged inset). n = 8 animals from 2 independent EAE experiments; cells were isolated at the maximum disease stage. Costaining for NeuN and CD4 was performed. Graph shows quantification of VCAM-1–positive and –negative cells with or without T cell contact. (G) CD29 extracellular staining for CD29 in human MS Th17 cells (n = 6) compared with cells from healthy donors (n = 5). (H) VCAM-1–positive neurons were identified in brain tissue specimens from patients with MS (n = 5 patients with MS; 1 representative staining is shown). Scale bar: 10 μm. Data indicate the mean ± SEM. *P < 0.05, by unpaired Student’s t test (B), 1-way ANOVA with Tukey’s (A, C, and D) or Dunnett’s (E) post hoc test.