Mechanisms of Vascular Calcification

Mechanisms of Vascular Calcification Mohga El-Abbadi and Cecilia M. Giachelli Vascular calcification is highly prevalent and correlated with high rates of cardiovascular mortality in chronic kidney disease patients. Recent evidence suggests that mineral, hormonal, and metabolic imbalances that promote phenotype change in vascular cells as well as deficiencies in specific mineralization inhibitory pathways may be important contributory factors for vascular calcification in these patients. This article reviews current mechanisms proposed for the regulation of vascular calcification and data supporting their potential contribution to this process in chronic kidney disease. © 2007 by the National Kidney Foundation, Inc. Index Words: Vascular calcification; Phosphate; Calcium; Smooth muscle; Chronic kidney disease.

V

ascular calcification, the inappropriate deposition of calcium phosphate salts in cardiovascular tissues, is highly correlated with cardiovascular mortality in chronic kidney disease (CKD) patients.1 In these patients, calcification occurs in both the tunica media (associated with Monckeberg’s sclerosis, elastocalcinosis, and calcific uremic arteriolopathy) as well as the tunica intima (associated with atherosclerosis).2 Calcification at these sites often includes bone and cartilage-like tissues, suggesting that developmental processes similar to those in osseous tissues may be relevant. Regardless in which anatomic compartment it occurs, calcification of blood vessels leads to altered hemodynamics and mechanical properties, ultimately causing increased wall stiffness, pulse pressure, left ventricular stress, and decreased coronary perfusion.3 The underlying mechanisms that cause and/or regulate vascular calcification are beginning to be unraveled. Exciting research in this area has defined a wide array of circulating factors, ion transporters/metabolic enzymes, matrix molecules, and signaling molecules that appear to comprise a complex calcification regulatory network involved in the pathogenesis of vascular calcification. Table 1 summarizes, by category, the growing list of molecules that have been implicated as From the Department of Bioengineering, University of Washington, Seattle, WA. Supported in part by NIH grants HL62329 and HL081785. Address correspondence to Cecilia M. Giachelli, PhD, Box 355061, Bioengineering, University of Washington, Seattle WA 98195. E-mail: [email protected] © 2007 by the National Kidney Foundation, Inc. 1548-5595/07/1401-0009$32.00/0 doi:10.1053/j.ackd.2006.10.007

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regulators of vascular calcification and those that have been associated with CKD. Interestingly, common themes and pathways are arising that are leading to an increased understanding of this complex system.

Phosphate/Pyrophosphate Balance Hyperphosphatemia is prevalent in CKD and is a strong predictor of morbidity and cardiovascular mortality in end-stage renal disease (ESRD) patients.4-6 Furthermore, hyperphosphatemia is correlated with vascular calcification,7,8 and effective control of hyperphosphatemia without increasing total calcium load has been correlated with attenuated progression of vascular calcification in dialysis patients.9-11 In addition to elevating the calcium ⫻ phosphorus product (Ca ⫻ P), elevated phosphate has direct effects on vascular cells that promote matrix calcification.12-15 Several groups have shown that elevated phosphate induces a profound phenotypic change in smooth muscle cells (SMCs) in vitro exemplified by loss of smooth muscle lineage gene expression (SM alpha actin, SM22) and gain of osteochondrogenic gene expression (Runx2, osteopontin, tissue nonspecific alkaline phosphatase [TNAP]).12-15 Similar gene expression patterns have been observed in calcifying vessels in animals12,16 and humans.2,17 Recent studies have begun to elucidate the pathways by which elevated phosphate may signal SMC phenotypic modulation and calcification. By using RNA silencing techniques, Li et al18 found that Pit-1, a sodium dependent phosphate cotransporter, was required for both phenotypic modulation as well as calcification of human SMCs in vitro but did not promote cell death or matrix vesicle

Advances in Chronic Kidney Disease, Vol 14, No 1 (January), 2007: pp 54-66

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Vascular Calcification

Table 1. Vascular Calcification Regulatory Molecules Vascular Calcification Regulatory Molecules Circulating factors Phosphate Calcium Vitamin D PTH Fetuin A Pyrophosphate FGF-23 Magnesium HDL LDL OPG Ion transporters/homeostasis Pit-1 TNAP Ank PC-1/NPP-1 Klotho Matrix molecules MGP OPN Signaling molecules/pathways Runx2/Cbfa1 Msx2/Wnt BMP2/4 BMP7 TNF-␣

Effect*

Reference†

P P P I/P I

12-16

I I I I P I

23-25,161

P P I I I

16,18

CKD‡

Reference§

Increase Increase Decrease Increase Decrease/no change Decrease Increase Increase Decrease Increase Increase

4-8,158

Increase

2,180,182

65

Decrease

175

I I

83-86

Increase Increase

80

P P P I P

12,14,181

Increase

180,181

Increase

184

Increase

186

37-39 37,44,45,49,50,168 53,56,59,60,170,171 80,106,163

64 128,129,131-134 146,152 20,147,173 164,165

2,19 24

5,6,9,159,160 40,41,169 41,52,157,172 80,100-104

26 67,68 133,134,162 146,157 157 166,167

24,28,174

176-179

74,75 76,77,115,183 116-118,185 21

2,180,181,182

*Role of regulatory molecule in vascular calcification. P, promoter; I, inhibitor. †References supporting the role of the regulatory molecule in calcification. ‡Directional change in the concentration of calcification regulatory molecules in CKD patients. §References reporting the prevalence of the regulatory molecule in the CKD patients.

phosphate loading. Thus, levels of Pit-1 expressed by SMCs appear to regulate their susceptibility to calcification. Interestingly, Pit-1 levels were recently shown to be upregulated in calcified arteries of uremic rats.16 In addition to increased systemic phosphate levels because of renal insufficiency, extracellular phosphate levels are regulated locally by the action of phosphatases, such as TNAP, on inorganic and organic phosphate containing molecules. TNAP is absolutely required for bone formation,19 and growing evidence indicates that it may play a similar role in mediating ectopic calcification. Alkaline phosphatase is strongly induced in SMCs by calcification inducers, such as elevated phosphate, tumor necrosis factor ␣, and oxidized lipids.2,15,20,21 One key substrate for TNAP is pyrophosphate, a potent inhibitor of apatite formation,22-25 that is reduced in the sera of

dialysis patients.26 In addition to generating phosphate, TNAP activity decreases pyrophosphate levels, thereby potentially promoting vascular calcification by 2 mechanisms: increased promineralizing phosphate levels and decreased antimineralizing pyrophosphate levels. Indeed, both the human disorder hypophosphatasia and mice deficient in TNAP gene are characterized by poorly mineralized bones and increased serum pyrophosphate levels.19,27 That loss of pyrophosphate predisposes to vascular calcification is clearly shown by the human genetic disease infantile arterial calcification. Patients with infantile arterial calcification have a deficiency of the ectonucleotide pyrophosphatase phosphodiesterase (eNPP-1/PC-1), the enzyme that generates pyrophosphate from nucleoside triphosphates, leading to pyrophosphate deficiency, widespread, medial arterial calci-

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fication, cardiac events, and premature death.28 Likewise, mice with a targeted deletion of eNPP1 have decreased pyrophosphate serum levels, show a vascular calcification phenotype, and develop hypermineralization in bone and cartilage.19 A similar phenotype is observed in mice deficient in a major pyrophosphate plasma membrane transporter, ANK. Indeed, mineralization disorders including arterial calcification were partially reversed by crossing TNAP⫺/⫺ mice with ank⫺/⫺ or eNPP1⫺/⫺ mice.19 In vitro, eNPP-1⫺/⫺ and ank⫺/⫺ calvarial osteoblasts have decreased extracellular pyrophosphate and increased mineralization. Transfection of eNPP-1⫺/⫺ osteoblasts with the eNPP-1 gene corrects both extracellular pyrophosphate levels and mineralization.29 Thus, the enzymes and transporters that maintain phosphate and pyrophosphate homeostasis have emerged as critical regulators of vascular calcification.

Calcium/Vitamin D/Parathyroid Hormone/Fibroblast Growth Factor (FGF)23 Axis Elevated calcium burden, secondary hyperparathyroidism, and therapeutic vitamin D use are common in CKD patients together with underlying renal osteodystrophy. How these complex factors might affect vascular calcification is not entirely clear, but some evidence for their potential regulatory roles has been obtained recently. Derangements in calcium metabolism and elevated Ca ⫻ P have emerged as nontraditional risk factor for cardiovascular mortality in CKD patients.5,6 Calcium ingestion, in the form of phosphate binders, has been linked to the progression of vascular calcification in hemodialysis and ESRD patients8,30,31 and the use of sevelamer, a calcium-free phosphate binder attenuates the progression of arterial calcification in these patients.32-35 Furthermore, hypercalcemia has been linked to vascular calcification in humans and experimental animals.30,33,36,37 Elevated calcium levels upregulate the expression of the major sodium-dependent phosphate cotransporter Pit-1 in vascular SMCs, and adding phosphonoformic acid, a Pit-1 inhibitor, inhibits calciuminduced mineralization in these cells.38 Fi-

nally, elevated calcium increases matrix vesicle production,39 messenger RNA levels of Cbfa-1 and alkaline phosphatase,38 and is synergistic with elevated phosphate in promoting mineral deposition in vascular SMCs. Active vitamin D3 (1,25[OH]2 D3 or calcitriol) is primarily synthesized in the kidneys. Its production is impaired when there are low levels of the precursor form calcidiol or in the setting of decreased renal function.40,41 As a result, supplementation with calcitriol or its analogs is typically prescribed to control secondary hyperparathyroidism in CKD.42 Calcitriol overload, with subsequent hypercalcemia, has been implicated in vascular calcification in CKD patients.37,43 High doses of calcitriol have been used in several studies to promote vascular calcification in rats.44,45 In these studies, very high levels of calcitriol were used and were associated with hypercalcemia. In contrast, less calcemic vitamin D analogs have been developed that, although equipotent with calcitriol in managing secondary hyperparathyroidism, do not cause vascular calcification in animals46,47 and have a survival advantage in dialysis patients.48 In vitro, some studies have shown that high concentrations of calcitriol promote vascular SMC matrix calcification, induce the expression of proteins that are involved in calcification such as alkaline phosphatase, and downregulate the expression of proteins that inhibit calcification such as parathyroid hormone (PTH)-related peptide.49,50 On the other hand, other studies have detected no effect of calcitriol on vascular SMC calcification (Taniwaki and Giachelli, unpublished data, June 2005).51 Secondary hyperparathyroidism is characteristic in CKD and is mainly a consequence of defective calcitriol production, decreased serum calcium, and hyperphosphatemia.41,52 It is marked by parathyroid hyperplasia and enhanced synthesis and secretion of PTH. This ultimately leads to disregulated mineral metabolism, bone remodeling, and renal osteodystrophy.43 The role of PTH in vascular calcification is not conclusive. Some studies have linked elevated PTH53-55 with higher rates of vascular calcification, whereas others have not.1,8,56,57 In addition, calcific uremic arteriolopathy, a particularly lethal form of medial calcification of the small arterioles, has

Vascular Calcification

been linked with secondary hyperparathyroidism.58 On the other hand, administration of teriperatide (human PTH 1-24) inhibited arterial calcification in diabetic LDLR-null mice.59 Likewise, PTH and PTH-related peptide inhibited calcification of vascular SMCs in vitro.60 Thus, it is possible that an optimal range of PTH is required for maintenance of vessel health and that PTH concentration outside this range may exacerbate vascular calcification. In addition to PTH and vitamin D3, FGF23 has been identified recently as a regulator of mineral balance. FGF23 functions to suppress phosphate reabsorption and inhibit vitamin D3 production in the kidney.61,62 Indeed, mutations in FGF23 cause familial tumoral calcinosis in people,63 and FGF23-null mice develop hyperphosphatemia, elevated 1,25 (OH)2D3, and a premature aging syndrome that includes vascular calcification.62,64 Interestingly, the klotho-null mouse shows a similar phenotype,65 and klotho was recently shown to enhance FGF receptor binding to FGF23, thereby suggesting a potential link between these molecules.66 In CKD patients, circulating FGF23 is significantly elevated, and its concentration increases as kidney function decreases and serum phosphorus levels increase.67,68 Serum vitamin D3 levels, as expected, correlate negatively with serum FGF23 levels in these patients, potentially contributing to the development of secondary hyperparathyroidism.69 Finally, the relationship between vascular calcification, calcium burden, vitamin D3, PTH, FGF23, and renal osteodystrophy remains unclear. An inverse relationship has been documented between arterial calcification and bone density/turnover in uremic patients70 as well as the general population.71 Thus, abnormalities renal osteodystrophy may predispose to vascular calcification in ways that are not yet defined. Bone Morphogenetic Protein-2/Msx2/Wnt Pathway Many CKD patients are diabetic, and vascular calcification is highly correlated with cardiovascular mortality in diabetic patients.72,73 Studies of vascular calcification in diabetic animals have suggested that a signaling path-

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way comprising Msx2 and Wnt exists in the arterial adventitia and signals the arterial media to undergo osteogenic differentiation and mineralization.74,75 Diabetes may promote this process by upregulating BMP-2/4 production, and bone morphogenetic protein-2 (BMP2) injection promotes calcification as well as Wnt signaling in TOPGAL⫹ LDLR⫺/⫺ mice.76 BMP-2 may also be involved in the calcification of atherosclerotic lesions as first reported by the Demer group.77 These studies are very exciting and provide several signaling pathways as potential targets for therapeutic interventions aimed at blocking vascular calcification.

Matrix Gla Protein/Vitamin K Duo Matrix gla protein (MGP), a potent inhibitor of arterial calcification is a 10-kDa extracellular matrix protein that has 5 Gla residues, is normally synthesized in high concentrations by SMCs and chondrocytes, and is present in serum.78,79 Calcified arteries in CKD, diabetic, and cardiovascular disease patients strongly express MGP, particularly at the interface between normal tissue and regions of precipitated calcium salt crystals.80-82 In vitro, decreased MGP expression is observed in calcifying bovine vascular SMCs and inhibition of calcification, using bisphosphonates, restores MGP expression to the level of uncalcified controls.83 Clear evidence for the inhibitory role of MGP in vascular calcification is manifested in MGP knockout mice and human Keutel syndrome, where its deficiency causes spontaneous medial and cartilaginous calcification.84,85 Furthermore, restoring MGP expression in the arteries of MGP knockout mice rescues the hypermineralized arterial phenotype.86 Although the molecular mechanism for MGP action is still unclear, transgenic mouse studies have shown that posttranslational, vitamin K– dependent gamma carboxylation of the glutamate residues (gla) is required for MGP’s antimineralization function.86 This is supported by earlier studies showing that rats treated with the vitamin K–antagonist warfarin develop severe arterial calcification.87 Similar procalcific findings were obtained when human vascular SMCs were pretreated with warfarin.39 In the clinical

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setting, warfarin usage has been associated with the development of calciphylaxis in dialysis patients.88 The importance of vitamin K in MGP-mediated inhibition was recently underscored with the development of conformation-specific MGP antibodies that distinguish between carboxylated and undercarboxylated forms of the molecule. By using these antibodies, Schurgers’ group89 showed that impaired carboxylation of MGP is associated with intimal and medial calcification in human sclerotic arteries, whereas only active, carboxylated MGP is displayed in healthy arteries. Although a negative correlation between total serum MGP levels and the severity of coronary calcification has been detected in nonuremic patients,90,91 a recent study did not find any association between serum MGP levels and vascular calcification in CKD patients.80 Neither study, however, analyzed the carboxylation state of measured MGP and hence did not determine the proportions of inactive versus active MGP. The development of serumbased assays for undercarboxylated forms of MGP would be useful for this purpose.91 In addition, longitudinal versus cross-sectional population studies would be more appropriate for accurate assessment of serum changes during disease progression. The ability of MGP to inhibit vascular calcification is thought to be partly because of the calcium/hydroxylapatite chelating capacity of its gla residues. High molecular weight fetuinMGP-calcium mineral complexes were identified in the sera of rats challenged with active bisphosphonate. These complexes were cleared after 24 hours, suggesting that by chelating and eliminating nascent circulating mineral nuclei, MGP reduces their availability for ectopic calcification.92 Locally, direct binding of MGP to hydroxylapatite crystals could act as crystal poison, disrupting normal crystal growth. Next, MGP is thought to exert its anticalcific effects by binding and inhibiting BMP-2 activity.93 Furthermore, the discovery that mutations/polymorphisms in the promoter region of MGP and in the gene vitamin K epoxide reductase complex subunit 1 (VKORC1, a component of the vitamin K cycle) can influence arterial calcification suggests potential genetic-based regulation of MGP function.94,95 Finally, several in vitro

studies have shown the ability of some molecules/ions involved in CKD to modulate MGP expression. A single dose of 10⫺7 M PTH upregulates MGP expression by 10-fold in MC3T3-E1 osteoblast-like cells,96 elevated extracellular ionic calcium upregulates MGP in rat aortic vascular SMCs,97 and vitamin D3 as well as a cyclic adenosine monophosphate (cAMP) analog (forskolin) stimulate MGP messenger RNA transcription in rat transcriptional regulation studies.98

␣2 Heremans-Schmid Glycoprotein (Fetuin A, Ahsg) Fetuin A is a calcium-binding, negative acutephase reactant glycoprotein that is synthesized by hepatocytes and secreted in high concentration into the circulation. Serum fetuin A is estimated to contribute about 50% of the precipitation inhibitory capacity of serum.99 In dialysis and ESRD patients, lowserum fetuin A levels are linked to the progression of atherosclerosis and increased cardiovascular mortality.100 Several studies have shown an inverse relationship between serum fetuin A levels and arterial calcification scores in dialysis patients, with calcific uremic arteriolopathy patients expressing particularly low-serum fetuin A levels.80,100-102 Patients with mild to moderate kidney dysfunction, including those with diabetic nephropathy, however, show no reduction in serum fetuin A with increased arterial calcification.103,104 This apparent contradiction is resolved once the acute reverse-phase nature of fetuin A is considered. The elevated state of inflammation in dialysis patients, featured by elevated inflammation biomarker CRP concentrations, suppresses fetuin A synthesis, allowing calcification to exacerbate, whereas earlier, less inflammatory CKD stages support the protective upregulation of fetuin A to fend off ectopic calcification.105 Evidence supporting the anticalcific role of fetuin A comes from studies with deficient mice showing their increased susceptibility to widespread soft-tissue calcification.106 In addition, fetuin A added to mineralizing vascular SMC inhibits calcification in a dose-dependent manner.80 In this context, Pseudoxanthoma elasticum, a he-

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reditary disorder characterized by progressive calcification in the cardiovascular system, skin, and retina, is associated with low serum fetuin A levels.107 Fetuin A inhibits vascular calcification partly by binding calcium ions and hydroxyapatite through its calcium-binding domain, D1.108 Increased fetuin A binding to calcified arterial lesions of dialysis patients is detected with immunohistochemical staining.80 As mentioned earlier, high molecular weight fetuin-MGPcalcium mineral complexes are detected in rat sera and are thought to clear early calcification nuclei.109 In addition, fetuin A binds directly to bone morphogenetic proteins (BMP-2, BMP-4, and BMP-6) and blocks their osteogenic activity in rat bone marrow culture assays.110,111 Furthermore, fetuin A is a natural antagonist of transforming growth factor-␤,110,111 and phosphorylated fetuin A impairs insulin signaling by inhibiting insulin receptor tyrosine kinase activity.112 Finally, rat fetuin appears to be susceptible to regulation by PTH,113 and native fetuin A undergoes several posttranslational modifications including phosphorylation, N-glycosylation, and O-glycosylation, allowing regulation of its diverse biological functions.114 In dialysis patients with particularly low-serum fetuin A levels, a fetuin A polymorphism was identified and associated with significantly poorer prognosis.105

BMP-7 BMP-7 is a newly emerging inhibitor of vascular calcification.115 It is a member of the transforming growth factor-␤ superfamily and is an important regulator of the skeletal remodeling. In human adult, BMP-7 is primarily expressed in the kidney and its synthesis is decreased by renal injury. In animals, BMP-7 expression is downregulated early in kidney failure,115 and as a treatment it prevents or reverses vascular calcification in LDLR-/-high-fat–fed CKD mice.116 The finding that BMP-7 concurrently reverses hyperphosphatemia in these mice led the Hruska group to suggest that BMP-7 in part exerted its vascular anticalcific effects by decreasing serum phosphorus and increasing skeletal deposition.117 Furthermore, BMP-7 stimulates

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the expression of SMC phenotype when added to human aorta-derived SMCs in culture.118 Finally, given BMP-7’s promising results in inhibiting vascular calcification in the uremic mouse model, it would be interesting to determine its efficacy in humans including its influence on vascular calcification-related morbidity and mortality in CKD patients.119

Magnesium, a Mimic/Antagonist of Calcium Magnesium, the forth most abundant metal in living organisms and an essential bivalent cation, is a natural antagonist of calcium and has been implicated for some time in the inhibition of soft-tissue calcification.120-125 In vitro, magnesium inhibits the nucleation and crystal growth of apatite as well as collagen-induced mineralization.126,127 Furthermore, magnesium deficiency has been reported to induce arterial calcification in numerous animal studies.128-132 Although serum magnesium levels are typically increased during the earlier stages of CKD, decreased serum magnesium levels have been associated with increased vascular calcification in ESRD patients.133,134 Magnesium is thought to exert its anticalcific effects partly by competing with calcium for the physical binding with phosphate.124,127 In addition, magnesium has been extensively implicated in the regulation of PTH synthesis and/or secretion. Increased serum magnesium levels appear to mimic, although with lesser potency, the effects of increased serum calcium levels in suppressing PTH secretion.135-139 In dialysis patients, an inverse relationship between magnesium and PTH has been well documented.140-142 Under health conditions, serum magnesium levels are mainly controlled by the kidneys through renal tubular maximum reabsorption.143 In chronic dialysis patients, however, because of diminished kidney function, magnesium levels are primarily determined by the dialysate’s magnesium concentration.144 This presents an opportunity for control of serum magnesium levels in these patients. Finally, it appears that magnesium may play an important protective role in the pathogenesis and progression of vascular calcification in dialysis patients. Long-term, longitudinal studies addressing

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the role of magnesium in arterial calcification are warranted, particularly for ESRD patients. For those patients, identifying the optimal dialysate magnesium concentration that grants protection against pathological calcification while avoiding possible clinical implications of hypermagnesemia is critical. Also, studies investigating the efficacy of magnesium-containing phosphate binders in inhibiting vascular calcification seem justified.134

Low-Density Lipoprotein/High-Density Lipoprotein Balance Dyslipidemia is prevalent among CKD patients and includes a reduction in serum highdensity lipoprotein (HDL) and an elevation in the very low– density lipoprotein and triglycerides.145,146 There is a significant association between the progression of coronary artery calcification and dislipidemia in dialysis patients.146 In nonuremic patients, the extent of vascular calcification also correlates with serum low-density lipoprotein (LDL) exposure,147 and lipid-lowering medications reduce the progression of coronary calcification.148 LDL receptor–null mice fed a high-fat diabetogenic diet develop aortic calcification and are hyperlipidemic.149 Levi’s group has shown that murine dyslipidemia plays an important role, via sterol regulatory elementbinding proteins (SREBP)-dependent pathways, in glomerulosclerosis and the pathogenesis of kidney disease.150,151 In vitro, minimally oxidized LDL induces osteoblastic differentiation and calcification of calcifying vascular cells (CVCs),20 whereas HDL inhibits their calcification as well as any prior osteogenic activity induced by oxidized LDL or inflammatory cytokines.152 How disregulated lipid metabolism influences vascular calcification is still not fully understood. Demer et al153 proposed that HDL may exert its protective role by inhibiting the oxidation and/or the procalcific effects of oxidized LDL. In this context, it is interesting to include recent findings showing that omega-3 fatty acids (present in fish oil) inhibit CVCs calcification by activating osteoblastic differentiation inhibitors p38MAPK and peroxisome proliferator-activated receptor (PPAR)-␥.154 Furthermore, oxidized LDL-activated human monocytes/macro-

phages cocultured with CVCs cause an increase in their ALP activity compared with the nonactivated monocytes.155 Activation of the cAMP pathway by minimally oxidized LDL enhances CVCs osteogenic transdifferentiation.156 Finally, Zanos et al157 recently suggested that PTH regulates uremic dyslipidemia through its induction of intracellular calcium uptake. In their study, increased lipid profile abnormalities detected in dialysis patients with high-serum intact parathyroid hormone (iPTH) were corrected on administration of calcium-channel blockers, whereas patients with decreased iPTH levels had normal lipid profiles. Thus, it is evident that uremic dyslipidemia involves complex regulation and contributes to vascular calcification.

Conclusion It is becoming increasingly clear that the active process of vascular calcification is highly complex, involving the interplay between a large number of circulating factors, hormones, membrane transporters, and matrix and signaling molecules. For each metabolic state, a few pathways are pivotal and orchestrate the entire mineralization process. In chronic kidney disease, pathways directly influenced by the calcium-phosphorus balance, such as BMP-2, vitamin D, and PTH, are potential candidates. Identification of these critical modulators followed by the elucidation of the networks they stimulate and regulate under health and various disease conditions should aid in the development of effective therapies to control and potentially reverse vascular calcification.

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