Fibroblasts from patients affected by Pseudoxanthoma elasticum exhibit an altered PPi metabolism and are more responsive to pro-calcifying stimuli

Background: Pseudoxanthoma elasticum (PXE) is a genetic disorder characterized by progressive calcification of soft connective tissues. The pathogenesis is still hard to pin down. In PXE dermal fibroblasts, in addition to impaired carboxylation of the vitamin K-dependent inhibitor matrix Gla protein (MGP), we have also demonstrated an up-regulation of alkaline phosphatase activity. In the light of these data we have suggested that both calcium and phosphate metabolism might be locally altered, both pathways acting in synergy on the occurrence of matrix calcification. Objective: This study aims to better explore if cultured PXE fibroblasts, compared to control cells, exhibit a modified inorganic pyrophosphate (PPi) metabolism and are more responsive to pro-calcifying stimuli. Methods: Primary human dermal fibroblasts isolated from healthy individuals and from PXE patients were cultured for different time points in standard and in pro-calcifying media. The expression of ANKH/ANKH, ENPP1/PC1, ALPL/TNAP, SPP1/OPN was evaluated by qRT-PCR and Western blot, respectively. TNAP activity was measured by spectrophotometric analyses, whereas calcification was investigated by light and electron microscopy as well as by micro-analytical techniques. Results: In the presence of pro-calcifying stimuli, dermal fibroblasts alter their phenotype favouring matrix mineralization. In particular, ENPP1/PC1 and SPP1/OPN expression, as well as TNAP activity, was differently expressed in control and in PXE fibroblasts. Moreover, in pathologic cells the ratio between factors favouring and reducing PPi availability exhibits a more pronounced shift towards a pro-calcifying balance. Conclusion: PXE fibroblasts are more susceptible to pro-calcifying stimuli and in these cells an altered PPi metabolism contributes to matrix calcification. (c) 2014 Japanese Society for Investigative Dermatology. Published by Elsevier Ireland Ltd. All rights reserved.