Hypercalcemia and hyperphosphatemia associate with an elevated risk of cardiovascular events, yet the pathophysiological basis of this association is unclear. Disturbed mineral homeostasis and the associated hypercalcemia and hyperphosphatemia may result in the formation of circulating calciprotein particles (CPPs) that aggregate the excessive calcium and phosphate ions. If not counteracted, the initially formed harmless amorphous spherical complexes (primary CPPs) may mature into damaging crystalline complexes (secondary CPPs). Secondary CPPs are internalized by vascular cells, causing a massive influx of calcium ions into the cytosol, leading to a proinflammatory response, cellular dysfunction, and cell death. Although the pathophysiological effects induced by CPPs in vascular cells receive increasing attention, a complete picture of how these particles contribute to the development of atherosclerosis and vascular calcification remains elusive. We here discuss existing knowledge on CPP formation and function in atherosclerosis and vascular calcification, techniques for investigating CPPs, and models currently applied to assess CPP-induced cardiovascular pathogenesis. Lastly, we evaluate the potential diagnostic value of serum CPP measurements and the therapeutic potential of anti-CPP therapies currently under development.

BACKGROUND: Calciprotein particles (CPPs) are associated with the development of vascular calcifications in chronic kidney disease. The role of endothelial cells (ECs) in this process is unknown. Here, we investigated the interaction of CPPs and ECs, thereby focusing on endothelial nitric oxide metabolism and oxidative stress. METHODS: CPPs were generated in calcium- and phosphate-enriched medium. Human umbilical vein endothelial cells were exposed to different concentrations of CPPs (0–100 µg/mL) for 24 or 72 hours. Ex vivo porcine coronary artery rings were used to measure endothelial cell–dependent vascular smooth muscle cell relaxation after CPP exposure. Serum samples from an early chronic kidney disease cohort (n=245) were analyzed for calcification propensity (measure for CPP formation) and nitrate and nitrite levels (NO x ). RESULTS: CPP exposure for 24 hours reduced eNOS (endothelial nitric oxide synthase) mRNA expression and decreased nitrite production, indicating reduced nitric oxide bioavailability. Also, 24-hour CPP exposure caused increased mitochondria-derived superoxide generation, together with nitrotyrosine protein residue formation. Long-term (72 hours) exposure of human umbilical vein endothelial cells to CPPs induced eNOS uncoupling and decreased eNOS protein expression, indicating further impairment of the nitric oxide pathway. The ex vivo porcine coronary artery model showed a significant reduction in endothelial-dependent vascular smooth muscle cell relaxation after CPP exposure. A negative association was observed between NO x levels and calcification propensity (r=−0.136; P =0.049) in sera of (early) chronic kidney disease patients. CONCLUSIONS: CPPs cause endothelial cell dysfunction by impairing nitric oxide metabolism and generating oxidative stress. Our findings provide new evidence for direct effects of CPPs on ECs and pathways involved.

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