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

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.

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 (K_Ca1.1), 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 K_Ca1.1 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.

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 K_Ca1.1 in fibroblasts obtained from patients with FD. Gene expression (p < .05) and protein levels of K_Ca1.1 (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 K_Ca1.1 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).

Gb3 deposition in skin fibroblasts may impair K_Ca1.1 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.