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Exp Neurol. 2020 Feb;324:113134. doi: 10.1016/j.expneurol.2019.113134. Epub 2019 Nov 25.

Globotriaosylceramide-induced reduction of KCa1.1 channel activity and activation of the Notch1 signaling pathway in skin fibroblasts of male Fabry patients with pain.

Author information

1
Department of Neurology, University of Würzburg, Germany.
2
Department of Neurology, University of Würzburg, Germany; Fabry Center for Interdisciplinary Therapy Würzburg (FAZIT), University of Würzburg, Germany.
3
Molecular Electrophysiology, Institute of Physiology, Center of Mental Health, University of Würzburg, 97080 Würzburg, Germany.
4
Department of Neurology, University of Würzburg, Germany; Fabry Center for Interdisciplinary Therapy Würzburg (FAZIT), University of Würzburg, Germany. Electronic address: ueceyler_n@ukw.de.

Abstract

BACKGROUND:

Fabry disease (FD) is an X-linked lysosomal storage disorder that leads to cellular globotriaosylceramide (Gb3) accumulation due to mutations in the gene encoding α-galactosidase A. Trigger-induced acral burning pain is an early FD symptom of unknown pathophysiology. We aimed at investigating the potential role of skin fibroblasts in nociceptor sensitization.

PATIENTS AND METHODS:

We enrolled 40 adult FD patients and ten healthy controls, who underwent a 6-mm skin punch biopsy at the lower leg. Dermal fibroblasts were cultivated and analyzed for Gb3 load. Fibroblast electrical activity was assessed using patch-clamp analysis at baseline and upon incubation with agalsidase-α for 24 h. We investigated gene expression of CC motif chemokine ligand 2 (CCL2), Ca2+activated K+-channel 1.1 (KCa1.1), interferone-γ (IFN-γ), transforming growth factor-β1 (TGF-β1), and transmembrane receptor notch homolog 1 (Notch1) using quantitative real-time-PCR, and protein levels of KCa1.1 by ELISA. Gene expression was determined at baseline and after fibroblast stimulation with tumor necrosis factor-α (TNF), modeling inflammation as a common pain trigger in FD.

RESULTS:

Total Gb3 load was higher in FD fibroblasts than in control fibroblasts (p < .01). Upon increase of intracellular Ca2+ concentrations, we detected differential electrical activity of KCa1.1 in fibroblasts obtained from patients with FD. Gene expression (p < .05) and protein levels of KCa1.1 (p < .05) were higher in fibroblasts from FD patients compared to control fibroblasts, whereas electric channel activity was lower in FD fibroblasts. After incubation with agalsidase-α, we observed an over-proportionate increase of KCa1.1 activity in FD fibroblasts reaching 7-fold the currents of control cells (p < .01). Gene expression studies revealed higher mRNA levels of CCL2, INF-γ, and Notch1 in FD fibroblasts compared to controls at baseline and after TNF incubation (p < .05 each), while TGF-β1 was higher in FD fibroblasts only after incubation with TNF (p < .05).

CONCLUSIONS:

Gb3 deposition in skin fibroblasts may impair KCa1.1 activity and activate the Notch1 signaling pathway. The resulting increase in pro-inflammatory mediator expression may contribute to cutaneous nociceptor sensitization as a potential mechanism of FD-associated pain.

KEYWORDS:

Fabry disease; Fabry pain; Fibroblast; KCa1.1 channel; Patch clamp;Notch1; Skin punch biopsy

PMID:
31778662
DOI:
10.1016/j.expneurol.2019.113134
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