Pseudoxanthoma elasticum is a metabolic disease

Abstract

Pseudoxanthoma elasticum (PXE) is a pleiotropic multisystem disorder affecting skin, eyes, and the cardiovascular system with progressive pathological mineralization. It is caused by mutations in the ABCC6 gene expressed primarily in the liver and kidneys, and at very low levels, if at all, in tissues affected by PXE. A question has arisen regarding the pathomechanism of PXE, particularly the "metabolic" versus the "PXE cell" hypotheses. We examined a murine PXE model (Abcc6(-/-)) by transplanting muzzle skin from knockout (KO) and wild-type (WT) mice onto the back of WT and KO mice using mineralization of the connective tissue capsule surrounding the vibrissae as an early phenotypic biomarker. Grafting of WT mouse muzzle skin onto the back of KO mice resulted in mineralization of vibrissae, whereas grafting KO mouse muzzle skin onto WT mice did not. Thus, these findings implicate circulatory factors as a critical component of the mineralization process. This mouse grafting model supports the notion that PXE is a systemic metabolic disorder with secondary mineralization of connective tissues and that the mineralization process can be countered or even reversed by changes in the homeostatic milieu.

Figure 1. Grafting of muzzle skin to the back of mice

Hair on the back of the recipient mice was clipped and a full thickness skin wound bed (0.5–1.0 cm in diameter) was prepared. The similar size sample of muzzle skin was obtained from the donor mice and grafted into the wound bed on the back of the recipient mice. The grafts were examined at 2 months after surgery by morphologic observations (a), by histology using H&E staining (b), and by genomic PCR (c). In (b), the arrows point to cross-sections of vibrissae in the graft of the donor mouse muzzle skin while they are not present in the adjacent back skin of the recipient mouse. In (c), the 430-bp band represents the wild-type allele, while the 320-bp PCR product is derived from amplification of knock-out (Abcc6^−/−) allele.

Figure 2. Progressive mineralization of connective tissue capsule of vibrissae in Abcc6^−/− mice with age

Sections of muzzle skin from 24-week old Abcc6^+/+ mice as well as from Abcc6^−/− mice at 4, 12 and 24 weeks of age were stained with H&E (top panel), Alizarin Red (middle panel) or von Kossa (bottom panel). No mineralization was noted in the 24-week old Abcc6^+/+ mice (a,e and i), nor was there any mineralization in the 4-week old Abcc6^−/− mice (b,f and j). In contrast, mineralization, as reflected by dark purple (top, c and d, arrows), red color (middle, g and h, arrows) or dark brown (bottom, k and l, arrows), was noted in the 12-week and 24-week old Abcc6^−/− mice which demonstrated a progressive increase in mineralization with advancing age. Scale bar = 100 μm

Figure 3. Aberrant mineralization of the connective tissue capsule of vibrissae in grafted skin

Two months after transplantation of grafts from 4-week old donor mice onto the back of recipient mice, the grafts were harvested and the mineralization was examined by H&E (a, d and g), Alizarin Red (b, e and h) and von Kossa (c, f and i) stains and by transmission electron microscopy (j, k). The connective tissue capsule of vibrissae grafted from the muzzle skin of Abcc6^+/+ mice onto the back of Abcc6^−/− mice developed characteristic mineralization (a–c, arrows), while no mineralization was observed in the grafts either from Abcc6^−/− (d–f) or Abcc6^+/+ (g–i) mice when transplanted onto the back of WT mice. TEM revealed mineralization (asterisks) of both collagen fibers (j) and elastic structures (k) in the vibrissae of Abcc6^+/+ mice at 2 months after grafting onto Abcc6^−/− mice. Scale bar = 100 μm (a–i); 1 μm (j and k).