Juxta-articular joint-capsule mineralization in CD73 deficient mice: similarities to patients with NT5E mutations

Abstract

Arterial calcification due to CD73 deficiency (ACDC), an autosomal recessive disorder, manifests with extensive mineralization of the lower-extremity arteries as well as of hand and foot joint-capsules. This disease is caused by mutations in the NT5E gene which encodes CD73, a membrane-bound ecto-5'-nucleotidase hydrolyzing 5'-AMP into adenosine and Pi. To gain insight into the pathophysiologic details of ACDC, we have characterized a Nt5e(-/-) knock out mouse (Nt5e(tm1Jgsc)) deficient in CD73. These mice, when maintained on appropriate strain background, demonstrated stiffening of the joints and micro CT revealed distinct changes in the thoracic skeletal structure with evidence of mineralization at the costochondral junctions. Mineralization was also noted in the juxta-articular spaces of the lower extremities as well as of ligaments and capsules adjacent to the bony structures. No evidence of vascular mineralization was noted either by CT or by microdissection of arteries in the thoracic area or in lower extremities. The Nt5e(-/-) mutant mice demonstrated significantly increased Pi levels in the serum and significantly reduced PPi concentration in the heparinized plasma, resulting in markedly increased Pi/PPi ratio, thus creating a pro-mineralization environment. In conclusion, the Nt5e(-/-) targeted mutant mice recapitulate some, but not all, features of ACDC and serve as a model system to study pharmacologic interventions for ectopic mineralization. Collectively, this mouse model deficient in CD73, with other targeted mutant mice with vascular mineralization, attests to the presence of a complex pro-mineralization/anti-mineralization network that under physiologic homeostatic conditions prevents ectopic tissue mineralization.

Keywords: ACDC, arterial calcification due to CD73 deficiency; CT, computed tomography; ENT1, equilibrative nucleoside transporter; Ectopic mineralization; GACI, generalized arterial calcification of infancy; PPi, inorganic pyrophosphate; PXE, pseudoxanthoma elasticum; Pi, inorganic phosphate; TNAP, tissue nonspecific alkaline phosphatase; animal models; mineralization networks.

Figure 1.

Genomic sequences of a SNP (rs32756904) in the Abcc6 gene of mice on different strain backgrounds. The original Nt5e^−/− mouse was developed on 129SvEv background (top line) which harbors a homozygous T nucleotide similar to 129S1/SvImJ (2nd row from top). The T allele is associated with ectopic mineralization. The Nt5e allele was separated from the mutant Abcc6 allele by crossing the mice on to C57BL/6J strain background which is homozygous for the C allele of the SNP (lower rows).

Figure 2.

Representative images of micro CT scans of Nt5e^−/− mice of 11–16 months of age. Note extensive ectopic mineralization of the sternocostal junctions (A, B), juxta-articular tissues in lower extremities (C), and knee (D) (arrows). Three-D renderings reveal skeletal deformities due to ectopic mineralization of thorax (E), foot (F), and knee (G). Note mineralization of the Achilles tendon in (F) (arrow).

Figure 3.

Schematic presentation of factors contributing to thepro-mineralization/anti-mineralization network preventing ectopicmineralization under physiologic conditions. Genetic defects in the genes encoding ABCC6, ENPP1 and CD73 result in ectopic mineralization in pseudoxanthoma elasticum (PXE), generalized arterial calcification of infancy (GACI) and arterial calcification due to CD73 deficiency (ACDC), respectively. (Modified from reference 12).