Fibroblast involvement in soft connective tissue calcification

Seven Italian centers reported data on survival, causes of death and appearance of complications in patients with thalassemia major. The interactions between gender, birth cohort, complications, and ferritin on survival and complications were analyzed. Survival after the first decade was studied for 977 patients born since 1960 whereas survival since birth and complication appearance was studied for the 720 patients born after 1970. Better survival was demonstrated for patients born in more recent years (p<0.00005) and for females (p=0.0003); 68% of the patients are alive at the age of 35 years. In the entire population 67% of the deaths were due to heart disease. There was a significant association between birth cohort and complication-free survival (p<0.0005). The prevalence of complications was: heart failure 6.8%, arrhythmia 5.7%, hypogonadism 54.7%, hypothyroidism 10.8%, diabetes 6.4%, HIV infection 1.7%, and thrombosis 1.1%. Lower ferritin levels were associated with a lower probability of heart failure (hazard ratio =3.35, p<0.005) and with prolonged survival (hazard ratio = 2.45, p<0.005), using a cut-off as low as 1,000 ng/mL. Survival and complication-free survival of patients with thalassemia major continue to improve, especially for female patients born shortly before or after the availability of iron chelation.

Idiopathic infantile arterial calcification (IIAC; OMIM 208000) is characterized by calcification of the internal elastic lamina of muscular arteries and stenosis due to myointimal proliferation. We analyzed affected individuals from 11 unrelated kindreds and found that IIAC was associated with mutations that inactivated ecto-nucleotide pyrophosphatase/phosphodiesterase 1 (ENPP1). This cell surface enzyme generates inorganic pyrophosphate (PP(i)), a solute that regulates cell differentiation and serves as an essential physiologic inhibitor of calcification.

Arterial calcifications are associated with increased cardiovascular risk, but the genetic basis of this association is unclear. We performed clinical, radiographic, and genetic studies in three families with symptomatic arterial calcifications. Single-nucleotide-polymorphism analysis, targeted gene sequencing, quantitative polymerase-chain-reaction assays, Western blotting, enzyme measurements, transduction rescue experiments, and in vitro calcification assays were performed. We identified nine persons with calcifications of the lower-extremity arteries and hand and foot joint capsules: all five siblings in one family, three siblings in another, and one patient in a third family. Serum calcium, phosphate, and vitamin D levels were normal. Affected members of Family 1 shared a single 22.4-Mb region of homozygosity on chromosome 6 and had a homozygous nonsense mutation (c.662C→A, p.S221X) in NT5E, encoding CD73, which converts AMP to adenosine. Affected members of Family 2 had a homozygous missense mutation (c.1073G→A, p.C358Y) in NT5E. The proband of Family 3 was a compound heterozygote for c.662C→A and c.1609dupA (p.V537fsX7). All mutations found in the three families result in nonfunctional CD73. Cultured fibroblasts from affected members of Family 1 showed markedly reduced expression of NT5E messenger RNA, CD73 protein, and enzyme activity, as well as increased alkaline phosphatase levels and accumulated calcium phosphate crystals. Genetic rescue experiments normalized the CD73 and alkaline phosphatase activity in patients' cells, and adenosine treatment reduced the levels of alkaline phosphatase and calcification. We identified mutations in NT5E in members of three families with symptomatic arterial and joint calcifications. This gene encodes CD73, which converts AMP to adenosine, supporting a role for this metabolic pathway in inhibiting ectopic tissue calcification. (Funded by the National Human Genome Research Institute and the National Heart, Lung, and Blood Institute of the National Institutes of Health.).