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. 2012 Oct;181(4):1190-6.
doi: 10.1016/j.ajpath.2012.06.014. Epub 2012 Jul 28.

A Novel Animal Model for Pseudoxanthoma Elasticum: The KK/HlJ Mouse

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Free PMC article

A Novel Animal Model for Pseudoxanthoma Elasticum: The KK/HlJ Mouse

Qiaoli Li et al. Am J Pathol. .
Free PMC article

Abstract

Pseudoxanthoma elasticum is a multisystem ectopic mineralization disorder caused by mutations in the ABCC6 gene. A mouse model with targeted ablation of the corresponding gene (Abcc6(tm1JfK)) develops ectopic mineralization on the dermal sheath of vibrissae as biomarker of the progressive mineralization disorder. Survey of 31 mouse strains in a longitudinal aging study has identified three mouse strains with similar ectopic mineralization of the vibrissae, particularly the KK/HlJ strain. We report here that this mouse strain depicts, in addition to ectopic mineralization of the dermal sheath of vibrissae, mineral deposits in a number of internal organs. Energy dispersive X-ray analysis and topographic mapping found the presence of calcium and phosphate as the principal ions in the mineral deposits, similar to that in Abcc6(tm1JfK) mice, suggesting the presence of calcium hydroxyapatite. The mineralization was associated with a splice junction mutation at the 3' end of exon 14 of the Abcc6 gene, resulting in a 5-bp deletion from the coding region and causing frame-shift of translation. As a consequence, essentially no Abcc6 protein was detected in the liver of the KK/HlJ mice, similar to that in Abcc6(tm1JfK) mice. Collectively, our studies found that the KK/HlJ mouse strain is characterized by ectopic mineralization due to a mutation in the Abcc6 gene and therefore provides a novel model system to study pseudoxanthoma elasticum.

Figures

Figure 1
Figure 1
Ectopic mineralization of the connective tissue sheath of vibrissae in KK/HlJ mice at 6 months of age is shown in comparison with age-matched Abcc6tm1JfK mice. Parallel sections were stained with H&E (A and B), alizarin red (AR; C and D), or von Kossa (vK; E and F) stain. The dark red brownish color represents mineral deposits. The magnification in all frames is the same. Scale bar = 100 μm (A).
Figure 2
Figure 2
Mineral deposits in the dermal sheath of vibrissae in KK/HlJ mice consist of calcium and phosphate, similar to that in Abcc6tm1JfK mice. The ratio of calcium to phosphate is ∼2.1, similar to that noted in endochondral bone as a control (A). RADAR mapping shows topographic colocalization of calcium (Ca) and phosphate (P) in the area corresponding to the mineralization, as visualized by scanning electron microscopy (B).
Figure 3
Figure 3
Sequence comparison of the KK/HlJ mice that shows ectopic mineralization and C57BL/6J mice without mineralization. Sequencing of the Abcc6 gene at the 3′ end of exon 14 shows a C-to-T transition substitution (arrowheads), which in pre-mRNA is predicted to create a new splice donor site. Sequencing of the RT-PCR product (cDNA) shows a frame shift in the KK/HlJ mouse mRNA compared with the wild-type sequence in C57BL/6J. Wt, wild-type; Mu, mutant.
Figure 4
Figure 4
A 5-bp deletion in the Abcc6 mRNA in KK/HlJ mice is shown. RT-PCR amplification of Abcc6 mRNA with the use of primers on exon 14 upstream from the C-to-T transition mutation and on exon 15 resulted in two products, 144 bp and 139 bp, respectively, reflecting the deletion of 5 bp in the mutant mRNA. Mu, mutant; NC, negative control; Wt, wild type.
Figure 5
Figure 5
Quantitation of the Abcc6 mRNA in the liver of the KK/HlJ mice in comparison with Abcc6tm1JfK mice. Note that the transcript levels were reduced by ∼20% in KK/HlJ mice, whereas it was completely absent in the Abcc6tm1JfK mice compared with Abcc6+/+ controls (100%). The values are mean ± SE; n = 9. N.S., not significant. ***P < 0.001.
Figure 6
Figure 6
IF analysis of Abcc6 protein in the liver of KK/HlJ mice in comparison with Abcc6tm1JfK mouse and its wild-type counterpart (Abcc6+/+). The magnifications in frames are shown on the right side of the figure.

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