Background/Purpose: Hidradenitis suppurativa (HS) is a chronic inflammatory skin condition characterized by painful nodules and abscesses in intertriginous areas. Despite its increasing prevalence, the etiology of HS remains unclear. A minority of HS cases are familial, involving germline mutations in one of the subunits of the membrane protease complex gamma-secretase (GS). Mutations are most frequent in the nicastrin (NCSTN) subunit. NCSTN regulates access to the proteolytic site of GS, thereby regulating cleavage of its many substrates including Amyloid Precursor Protein (APP). Given the biologic importance of GS and its known association to familial HS, we hypothesized that NCSTN levels may be low in common non-familial HS and lead to GS enzyme dysregulation, driving HS pathogenesis. We therefore sought to explore NCSTN expression in HS skin and the activity of GS in HS-derived fibroblasts. 

Methods: Immunofluorescence (IF) was used to quantify NCSTN protein in HS and healthy skin samples. For in vitro experiments, fibroblasts were derived from HS surgical specimens or axillary biopsies from healthy volunteers. Fibroblasts were maintained in culture for RNA, protein, and enzymatic analysis. TNFα treatment was used to induce inflammatory signaling. As a positive control for NCSTN loss, normal fibroblasts were treated with NCSTN siRNA (siNCSTN). To explore changes in GS kinetics, we used a Förster Resonance Energy Transfer (FRET) assay with an artificial APP-like FRET peptide. Fibroblast membrane fractions containing GS were incubated with FRET peptide for 6 hours in a microplate reader, with fluorescence recorded every 5 minutes. 

Results: Acquired HS patients had lower levels of NCSTN protein in dermal fibroblasts but not epidermal keratinocytes by IF (fibroblast p=0.0007, keratinocyte p=0.854; n=8 HS and 3 control). In normal fibroblasts in vitro, expression of NCSTN mRNA increased upon treatment with TNFα (n=6, p< 0.001). NCSTN protein was also increased (n=3, p< 0.01). GS-mediated cleavage of FRET peptide showed no difference in activity compared to normal fibroblasts (n=8). The knockdown of NCSTN expression did not modify FRET peptide cleavage in normal fibroblasts. Notably, siNCSTN fibroblasts had higher IL-8 expression (p < .0001, n = 10 over 4 experiments).

Conclusion: HS fibroblasts have lower NCSTN levels than healthy controls by IF and Western blot. Normal fibroblasts have increased NCSTN expression when stimulated with TNFα. Interestingly, while GS activity for an APP substrate was not altered in HS fibroblasts or in normal fibroblasts after NCSTN knockdown, increases in the inflammatory chemokine IL-8 were observed. These findings suggest that GS dysfunction in dermal fibroblasts may contribute to inflammation in familial and non-familial HS. Future studies should assess the etiology of NCSTN loss in HS and alterations in GS activity for key substrates including full-length proteins.

ACR Meeting Abstracts - https://acrabstracts.org/abstract/the-common-form-of-the-inflammatory-skin-disease-hidradenitis-suppurativa-is-associated-with-low-nicastrin-expression-in-dermal-fibroblasts/