Pyrophosphate Supplementation Prevents Chronic and Acute Calcification in ABCC6-Deficient Mice

Abstract

Soft tissue calcification occurs in several common acquired pathologies, such as diabetes and hypercholesterolemia, or can result from genetic disorders. ABCC6, a transmembrane transporter primarily expressed in liver and kidneys, initiates a molecular pathway inhibiting ectopic calcification. ABCC6 facilitates the cellular efflux of ATP, which is rapidly converted into pyrophosphate (PPi), a major calcification inhibitor. Heritable mutations in ABCC6 underlie the incurable calcification disorder pseudoxanthoma elasticum and some cases of generalized arterial calcification of infancy. Herein, we determined that the administration of PPi and the bisphosphonate etidronate to Abcc6 mice fully inhibited the acute dystrophic cardiac calcification phenotype, whereas alendronate had no significant effect. We also found that daily injection of PPi to Abcc6 mice over several months prevented the development of pseudoxanthoma elasticum-like spontaneous calcification, but failed to reverse already established lesions. Furthermore, we found that the expression of low amounts of the human ABCC6 in liver of transgenic Abcc6 mice, resulting in only a 27% increase in plasma PPi levels, led to a major reduction in acute and chronic calcification phenotypes. This proof-of-concept study shows that the development of both acute and chronic calcification associated with ABCC6 deficiency can be prevented by compensating PPi deficits, even partially. Our work indicates that PPi substitution represents a promising strategy to treat ABCC6-dependent calcification disorders.

Basic calcium phosphate crystal-associated musculoskeletal syndromes: an update

“…Although intravenously administered, PPi had a half-life of only 42 min, administration of a single dose after the initial injury that initiates calcification halted subsequent calcification. Similar results were seen with etidronate, but not with alendronate [21■■]. This fascinating work suggests that even transiently elevated levels of circulating PPi may reverse BCP crystal deposition in some settings.…”

“…The potential use of PPi as a therapeutic agent was recently highlighted [21■■]. For these studies, Pomozi et al used a mouse model of pathologic calcification based on mutations in ATP binding cassette subfamily C member 6 (ABCC6).…”

Oral administration of pyrophosphate inhibits connective tissue calcification

“…The observed transient elevation found in healthy volunteers (Fig A) indicates that in both GACI and in PXE patients, PPi levels may be transiently raised to the physiological level when 67 or 98 mg Na 4 PPi/kg of body weight is taken. Importantly, it has been shown that inhibition of calcification in uremic rats and in PXE mice can be achieved by daily intraperitoneal injections of PPi triggering transient increase in plasma PPi levels (O'Neill et al , ; Pomozi et al , ). We determined a t 1/2 = 44.7 min what is rather similar to that published in rat (34.1 min; O'Neill et al , ).…”

Inhibition of Tissue-Nonspecific Alkaline Phosphatase Attenuates Ectopic Mineralization in the Abcc6 Mouse Model of PXE but Not in the Enpp1 Mutant Mouse Models of GACI

“…Upstream of ENPP1, ABCC6-mediated ATP release from hepatocytes is the main source of circulating PPi (Jansen et al, 2014). The work by us and others suggests that reduced PPi levels is the major factor leading to ectopic mineralization in PXE, but there might be an alternative, as yet unknown mechanism, independent of PPi, by which ABCC6 prevents ectopic mineralization under physiologic conditions (Pomozi et al, 2017;Zhao et al, 2017). Recent observations suggest that ABCC6 is also involved in extracellular nucleotide metabolism, suggesting a central role for ABCC6 in purinergic signaling in the development of ectopic mineralization (Kauffenstein et al, 2018).…”

“…After centrifugation, plasma was depleted of platelets by filtration through a Centrisart I 300-kDa mass cutoff filter (Sartorius, New York, NY) and stored at e80 C until further processing. PPi in plasma was measured by an enzymatic reaction using ATP sulfurylase to convert PPi into ATP in the presence of excess adenosine 5 0 -phosphosulfate (Sigma-Aldrich), a methodology adopted worldwide as described previously (Bauer et al, 2018;Dedinszki et al, 2017;Jansen et al, 2014;Li et al, 2017;Pomozi et al, 2017).…”