Hepatic and Vascular Vitamin K Status in Patients with High Cardiovascular Risk.

Vitamin K dependent proteins (VKDP), such as hepatic coagulation factors and vascular matrix Gla protein (MGP), play key roles in maintaining physiological functions. Vitamin K deficiency results in inactive VKDP and is strongly linked to vascular calcification (VC), one of the major risk factors for cardiovascular morbidity and mortality. In this study we investigated how two vitamin K surrogate markers, dephosphorylated-undercarboxylated MGP (dp-ucMGP) and protein induced by vitamin K absence II (PIVKA-II), reflect vitamin K status in patients on hemodialysis or with calcific uremic arteriolopathy (CUA) and patients with atrial fibrillation or aortic valve stenosis. Through inter- and intra-cohort comparisons, we assessed the influence of vitamin K antagonist (VKA) use, vitamin K supplementation and disease etiology on vitamin K status, as well as the correlation between both markers. Overall, VKA therapy was associated with 8.5-fold higher PIVKA-II (0.25 to 2.03 AU/mL) and 3-fold higher dp-ucMGP (843 to 2642 pM) levels. In the absence of VKA use, non-renal patients with established VC have dp-ucMGP levels similar to controls (460 vs. 380 pM), while in HD and CUA patients, levels were strongly elevated (977 pM). Vitamin K supplementation significantly reduced dp-ucMGP levels within 12 months (440 to 221 pM). Overall, PIVKA-II and dp-ucMGP showed only weak correlation (r2 ≤ 0.26) and distinct distribution pattern in renal and non-renal patients. In conclusion, VKA use exacerbated vitamin K deficiency across all etiologies, while vitamin K supplementation resulted in a vascular VKDP status better than that of the general population. Weak correlation of vitamin K biomarkers calls for thoughtful selection lead by the research question. Vitamin K status in non-renal deficient patients was not anomalous and may question the role of vitamin K deficiency in the pathogenesis of VC in these patients.

Development of the BioHybrid Assay: Combining Primary Human Vascular Smooth Muscle Cells and Blood to Measure Vascular Calcification Propensity.

BACKGROUND: Vascular calcification is an active process that increases cardiovascular disease (CVD) risk. There is still no consensus on an appropriate biomarker for vascular calcification. We reasoned that the biomarker for vascular calcification is the collection of all blood components that can be sensed and integrated into a calcification response by human vascular smooth muscle cells (hVSMCs). METHODS: We developed a new cell-based high-content assay, the BioHybrid assay, to measure in vitro calcification. The BioHybrid assay was compared with the o-Cresolphthalein assay and the T50 assay. Serum and plasma were derived from different cohort studies including chronic kidney disease (CKD) stages III, IV, V and VD (on dialysis), pseudoxanthoma elasticum (PXE) and other cardiovascular diseases including serum from participants with mild and extensive coronary artery calcification (CAC). hVSMCs were exposed to serum and plasma samples, and in vitro calcification was measured using AlexaFluor®-546 tagged fetuin-A as calcification sensor. RESULTS: The BioHybrid assay measured the kinetics of calcification in contrast to the endpoint o-Cresolphthalein assay. The BioHybrid assay was more sensitive to pick up differences in calcification propensity than the T50 assay as determined by measuring control as well as pre- and post-dialysis serum samples of CKD patients. The BioHybrid response increased with CKD severity. Further, the BioHybrid assay discriminated between calcification propensity of individuals with a high CAC index and individuals with a low CAC index. Patients with PXE had an increased calcification response in the BioHybrid assay as compared to both spouse and control plasma samples. Finally, vitamin K1 supplementation showed lower in vitro calcification, reflecting changes in delta Agatston scores. Lower progression within the BioHybrid and on Agatston scores was accompanied by lower dephosphorylated-uncarboxylated matrix Gla protein levels. CONCLUSION: The BioHybrid assay is a novel approach to determine the vascular calcification propensity of an individual and thus may add to personalised risk assessment for CVD.

Role of Vitamin K in CKD: Is Its Supplementation Advisable in CKD Patients?

BACKGROUND: Patients with CKD are at an increased risk of developing vascular calcification (VC) and bone complications which translate into a higher morbidity and mortality. The dephosphorylated and uncarboxylated matrix Gla protein (dp-ucMGP) is considered to be an indicator of vitamin K2 status and correlates with markers of VC. It is activated by γ-glutamyl carboxylase that converts inactive MGP into an active form, and vitamin K2 is a cofactor of this reaction. The active form of MGP is a known inhibitor of arterial wall calcification and plays an important role in bone turnover. Recent studies show poor vitamin K2 status in CKD patients. We aimed to review the literature for the association between vitamin K2 status and calcification and bone disease risk and the efficacy of vitamin K2 supplementation in CKD population. SUMMARY: Most CKD patients, including those on renal replacement therapy, have vitamin K2 deficiency. The dp-ucMGP level, a marker of vitamin K2 status, is decreased by vitamin K2 supplementation in CKD patients, but there is no unequivocal proof that it influences arterial calcification progression and bone complications. Key Messages: CKD population are at risk of vitamin K deficiency. Supplementation of vitamin K2 is safe and improves the serum markers of its deficiency. There is lack of strong evidence that vitamin K2 supplementation slows progression of calcification or reduces the frequency of bone complications. More prospective studies are needed.

Association of Zinc Deficiency with Development of CVD Events in Patients with CKD.

Deficiency of the micronutrient zinc is common in patients with chronic kidney disease (CKD). The aim of this review is to summarize evidence presented in literature for consolidation of current knowledge regarding zinc status in CKD patients, including those undergoing hemodialysis. Zinc deficiency is known to be associated with various risk factors for cardiovascular disease (CVD), such as increased blood pressure, dyslipidemia, type 2 diabetes mellitus, inflammation, and oxidative stress. Zinc may protect against phosphate-induced arterial calcification by suppressing activation of nuclear factor kappa light chain enhancer of activated B. Serum zinc levels have been shown to be positively correlated with T50 (shorter T50 indicates higher calcification propensity) in patients with type 2 diabetes mellitus as well as those with CKD. Additionally, higher intake of dietary zinc was associated with a lower risk of severe abdominal aortic calcification. In hemodialysis patients, the beneficial effects of zinc supplementation in relation to serum zinc and oxidative stress levels was demonstrated in a meta-analysis of 15 randomized controlled trials. Thus, evidence presented supports important roles of zinc regarding antioxidative stress and suppression of calcification and indicates that zinc intake/supplementation may help to ameliorate CVD risk factors in CKD patients.

Fortified phosphorus­lowering treatment through administration of lanthanum protects against vascular calcification via regulation of FGF23 in chronic kidney disease.

Phosphorus reduction can prevent against vascular calcification (VC) in chronic kidney disease (CKD), but the mechanisms underlying its actions remain unclear. The aim of the present study was to determine the effect of a fortified phosphorus­lowing treatment on VC in CKD. Serum levels of creatinine, blood urea nitrogen (BUN), fibroblast growth factor 23 (FGF23), calcium and phosphorus, and the plasma levels of parathyroid hormone (PTH) were determined in an animal model of CKD treated with or without lanthanum. Haematoxylin and eosin (H&E) staining was performed to examine the structure of kidney tissues. Western blot analysis was performed to compare the levels of total­ (t­) extracellular signal­related kinase (ERK) and phospho­ (p­)ERK among the different experimental groups to investigate the effect of FGF23 on p­ERK expression. In the animal model, administration of adenine increased the serum levels of creatinine, BUN, FGF23 and phosphorus but decreased the serum levels of calcium. In addition, adenine treatment increased the plasma levels of PTH. H&E staining showed that lanthanum treatment did not alter the severity of renal cortex injury. Furthermore, the levels of t­ERK levels did not notably differ between the Adenine­free, Adenine­vehicle and Adenine­lanthanum groups, whereas the levels of p­ERK and aortic calcium in the Adenine­vehicle group were significantly upregulated. In addition, ectopic overexpression of FGF23 increased the levels of p­ERK, Msx2 and Osx in a dose­dependent manner. Furthermore, a total of 48 patients were enrolled in the present study. In the fortified group, the serum levels of FGF23, phosphorus and PTH were significantly reduced, whereas the serum levels of calcium were significantly increased, indicating an enhanced preventative effect in the fortified group. The results of the present study suggest that FGF23 may be used as a therapeutic target in the management and prevention of VC in CKD.

The Inhibitory Roles of Vitamin K in Progression of Vascular Calcification.

Vitamin K is a fat-soluble vitamin that is indispensable for the activation of vitamin K-dependent proteins (VKDPs) and may be implicated in cardiovascular disease (CVD). Vascular calcification is intimately associated with CV events and mortality and is a chronic inflammatory process in which activated macrophages promote osteoblastic differentiation of vascular smooth muscle cells (VSMCs) through the production of proinflammatory cytokines such as IL-1ß, IL-6, TNF-α, and oncostatin M (OSM) in both intimal and medial layers of arterial walls. This process may be mainly mediated through NF-κB signaling pathway. Vitamin K has been demonstrated to exert anti-inflammatory effects through antagonizing NF-κB signaling in both in vitro and in vivo studies, suggesting that vitamin K may prevent vascular calcification via anti-inflammatory mechanisms. Matrix Gla protein (MGP) is a major inhibitor of soft tissue calcification and contributes to preventing both intimal and medial vascular calcification. Vitamin K may also inhibit progression of vascular calcification by enhancing the activity of MGP through facilitating its γ-carboxylation. In support of this hypothesis, the procalcific effects of warfarin, an antagonist of vitamin K, on arterial calcification have been demonstrated in several clinical studies. Among the inactive MGP forms, dephospho-uncarboxylated MGP (dp-ucMGP) may be regarded as the most useful biomarker of not only vitamin K deficiency, but also vascular calcification and CVD. There have been several studies showing the association of circulating levels of dp-ucMGP with vitamin K intake, vascular calcification, mortality, and CVD. However, additional larger prospective studies including randomized controlled trials are necessary to confirm the beneficial effects of vitamin K supplementation on CV health.

Association between dietary zinc intake and abdominal aortic calcification in US adults.

BACKGROUND: In animal studies, zinc supplementation inhibited phosphate-induced arterial calcification. We tested the hypothesis that higher intake of dietary zinc was associated with lower abdominal aortic calcification (AAC) among adults in the USA. We also explored the associations of AAC with supplemental zinc intake, total zinc intake and serum zinc level. METHODS: We performed cross-sectional analyses of 2535 participants from the National Health and Nutrition Examination Survey 2013-14. Dietary and supplemental zinc intakes were obtained from two 24-h dietary recall interviews. Total zinc intake was the sum of dietary and supplemental zinc. AAC was measured using dual-energy X-ray absorptiometry in adults ≥40 years of age and quantified using the Kauppila score system. AAC scores were categorized into three groups: no AAC (AAC = 0, reference group), mild-moderate (AAC >0-≤6) and severe AAC (AAC >6). RESULTS: Dietary zinc intake (mean ± SE) was 10.5 ± 0.1 mg/day; 28% had AAC (20% mild-moderate and 8% severe), 17% had diabetes mellitus and 51% had hypertension. Higher intake of dietary zinc was associated with lower odds of having severe AAC. Per 1 mg/day higher intake of dietary zinc, the odds of having severe AAC were 8% lower [adjusted odds ratio 0.92 (95% confidence interval 0.86-0.98), P = 0.01] compared with those without AAC, after adjusting for demographics, comorbidities and laboratory measurements. Supplemental zinc intake, total zinc intake and serum zinc level were not associated with AAC. CONCLUSIONS: Higher intake of dietary zinc was independently associated with lower odds of having severe AAC among noninstitutionalized US adults.

Impact of hepatitis virus infection on arterial calcification among incident hemodialysis patients.

BACKGROUND: The impact of hepatitis virus infection on arterial calcification (AC) was not studied. OBJECTIVE: To study the prevalence, severity and distribution of AC in incident hemodialysis patients with hepatitis B and C viral infection. CASES AND METHODS: 172 stage 5 CKD adults (98 male and 74 female) were included; 58 of them were seronegative for both hepatitis B and C (SN group), 48 were positive for hepatitis B virus infection (HBV group) and 66 were hepatitis C virus positive (HCV group). Beside histopathology of the obtained arterial samples, all these cases were examined for body mass index (BMI), serum calcium (Ca), phosphorus (P), alkaline phosphatase (AP), serum albumin, uric acid (UA), alanine transaminase (ALT), parathormone (PTH), fibroblast growth factor 23(FGF23), interleukin 6 (IL6), and 25 hydroxy vitamin D (25 (OH) vit D), hemoglobin concentration, and serum ferritin. RESULTS: 86 (50%) of the cases had AC; 11 of them were in SN group (19%), 9 in HBV group (18.8%) and all the 66 HCV group (100%). In SN group, 4 had intimal calcification, 5 had medial calcification, and 2 had both intimal and medial calcification. In HBV group, 9 had intimal calcification, while no cases were encountered with either medial or both site calcifications. In HCV group, 16 had intimal calcification, 31 had medial calcification, and 19 had both intimal and medial calcification. Calcification was in the form of spots in one case in SN group, and 6 cases in HBV group, a single plaque of calcification in 5 cases of SN group, 3 cases of HBV group, and 16 cases of HCV group, multiple plaques were detected in 4 cases in SN group, and 31 cases in HCV group, and diffuse calcification in one case in SN group, and 19 cases in HCV group. In HBV group, calcification was only detected in patients with high viremia, while all patients with low or moderate viremia were devoid of calcification. In HCV group, all patients with low viremia had intimal solitary plaque of calcification, all patients with moderate viremia had multiple plaques of medial calcification, while all patients with high viremia had diffuse intimal and medial calcification. Both groups of viral hepatitis were significantly different in comparison to SN group in either distribution or calcification score (P<0.001 in all). HBV group had significantly lower serum P, CaxP and PTH in comparison to SN group (4.6±0.66 vs. 5.45±0.77mg/dL, 36.4±7.2 vs. 44.1±8.69, and 348±65.4 vs. 405.9±83.2pg/mL, P<0.001, <0.001, and 0.035 respectively). On the other hand, HCV group did not show any significant difference in any of the studied parameters compared to SN group. CONCLUSION: HCV positive patients are more prone to develop AC that is more extensive. HBV positive patients were less likely to have arterial medial calcification, probably related to lower serum phosphorus, CaxP product and PTH. HCV infection should be added as risk factor for AC among CKD patients. Further studies are needed to confirm these findings.

FGF23 and Phosphate-Cardiovascular Toxins in CKD.

Elevated levels of fibroblast growth factor 23 (FGF23) and phosphate are highly associated with increased cardiovascular disease and mortality in patients suffering from chronic kidney disease (CKD). As the kidney function declines, serum phosphate levels rise and subsequently induce the secretion of the phosphaturic hormone FGF23. In early stages of CKD, FGF23 prevents the increase of serum phosphate levels and thereby attenuates phosphate-induced vascular calcification, whereas in end-stage kidney disease, FGF23 fails to maintain phosphate homeostasis. Both hyperphosphatemia and elevated FGF23 levels promote the development of hypertension, vascular calcification, and left ventricular hypertrophy by distinct mechanisms. Therefore, FGF23 and phosphate are considered promising therapeutic targets to improve the cardiovascular outcome in CKD patients. Previous therapeutic strategies are based on dietary and pharmacological reduction of serum phosphate, and consequently FGF23 levels. However, clinical trials proving the effects on the cardiovascular outcome are lacking. Recent publications provide evidence for new promising therapeutic interventions, such as magnesium supplementation and direct targeting of phosphate and FGF receptors to prevent toxicity of FGF23 and hyperphosphatemia in CKD patients.

The effect of vitamin K2 supplementation on vascular calcification in haemodialysis patients: a 1-year follow-up randomized trial.

PURPOSE: Vascular calcification (VC) is an independent risk factor for cardiovascular disease in hemodialysis patients while Matrix GLA protein (MGP) is one of the most potent inhibitors of VC and its activation is vitamin K dependent. The aim of this study is to investigate the role of oral vitamin K2 supplementation in the prevention of VC progression in haemodialysis patients. METHODS: We conducted a prospective randomized interventional study in patients on hemodialysis. Patients were randomly assigned to either receiving orally 200 µgr of vitamin K2 (vitamin K2/MK-7, Solgar) every day for 1 year or no treatment. Uncarboxylated MGP (uc-MGP) concentrations were quantified using ELISA at randomization, at 3 and at 12 months. Aortic calcification was evaluated using Agatston score after an abdominal computed tomography scan that was performed at the beginning and at 12 months of follow-up. RESULTS: There were 102 patients that were randomized. After 1 year of follow-up, 22 patients from the vitamin K2 group and 30 patients from the control group were included in the analysis. After 3 months of treatment, uc-MGP values remained unchanged in the vitK2 group but after 1 year were reduced by 47% (p = 0.005). Furthermore, uc-MGP at 1 year was increased by 12% in the control group. At 1 year, vitK2 group had significantly lower values of uc-MGP in comparison to controls (p = 0.03). Agatston score was increased significantly both in vitamin K2 and control group at 1 year with no difference between groups. CONCLUSIONS: Oral administration of vitamin K2 in patients on haemodialysis reduced serum uc-MGP levels but did not have an effect in the progression of aortic calcification.

Calcification of abdominal aorta in patients recently starting hemodialysis: A single-center experience from Egypt.

Vascular calcification (VC) is a well-known complication in patients with chronic kidney disease (CKD). Keeping in mind, the end goal to assess the genuine effect of mineral bone disease in the pathogenesis of blood vessel calcification during the pre-dialysis course of CKD, we assessed the prevalence and extent of abdominal aortic calcification (AAC) in nondiabetic CKD patients recently starting hemodialysis (HD). Eighty-one patients with end-stage renal disease beginning HD over a one-month period were selected. They underwent a detailed clinical examination and laboratory evaluation, including serum calcium, phosphorus, parathyroid hormone, fibroblast growth factor (FGF-23), and alkaline phosphatase were measured, and spiral computed tomography was performed to evaluate AAC score. AAC was present in 64 patients (79%). There was a significant correlation between the AAC score and age (r = 0.609, P <0.001) and FGF-23 (r = 0.800, P <0.001). This study suggests that the prevalence and extent of AAC are critical in incident HD patients. Serum FGF-23 level is the sole statistically significant correlate of AAC in these patients.

Site-specific chelation therapy with EDTA-loaded albumin nanoparticles reverses arterial calcification in a rat model of chronic kidney disease.

Medial arterial calcification (MAC) is a common outcome in diabetes and chronic kidney disease (CKD). It occurs as linear mineral deposits along the degraded elastin lamellae and is responsible for increased aortic stiffness and subsequent cardiovascular events. Current treatments for calcification, particularly in CKD, are predominantly focused on regulating the mineral disturbance and other risk factors. Ethylene diamine tetraacetic acid (EDTA), a chelating agent, can resorb mineral deposits, but the systemic delivery of EDTA may cause side effects such as hypocalcemia and bone resorption. We have developed elastin antibody conjugated albumin nanoparticles that target only degraded elastin in vasculature while sparing healthy tissues. In this study, we tested a targeted nanoparticle-based EDTA chelation therapy to reverse CKD-associated MAC. Renal failure was induced in Sprague-Dawley rats by a high adenine diet supplemented by high P and Ca for 28 days that led to MAC. Intravenous delivery of DiR dye-loaded nanoparticles confirmed targeting to vascular degraded elastin and calcification sites within 24 hours. Next, EDTA-loaded albumin nanoparticles conjugated with an anti-elastin antibody were intravenously injected twice a week for two weeks. The targeted nanoparticles delivered EDTA at the site of vascular calcification and reversed mineral deposits without any untoward effects. Systemic EDTA injections or blank nanoparticles were ineffective in reversing MAC. Reversal of calcification seems to be stable as it did not return after the treatment was stopped for an additional four weeks. Targeted EDTA chelation therapy successfully reversed calcification in this adenine rat model of CKD. We consider that targeted NP therapy will provide an attractive option to reverse calcification and has a high potential for clinical translation.

Emerging Role of Vitamins D and K in Modulating Uremic Vascular Calcification: The Aspect of Passive Calcification.

Vascular calcification is a critical complication in patients with chronic kidney disease (CKD) because it is predictive of cardiovascular events and mortality. In addition to the traditional mechanisms associated with endothelial dysfunction and the osteoblastic transformation of vascular smooth muscle cells (VSMCs), the regulation of calcification inhibitors, such as calciprotein particles (CPPs) and matrix vesicles plays a vital role in uremic vascular calcification in CKD patients because of the high prevalence of vitamin K deficiency. Vitamin K governs the gamma-carboxylation of matrix Gla protein (MGP) for inhibiting vascular calcification, and the vitamin D binding protein receptor is related to vitamin K gene expression. For patients with chronic kidney disease, adequate use of vitamin D supplements may play a role in vascular calcification through modulation of the calciprotein particles and matrix vesicles (MVs).

AT2 receptor stimulation inhibits phosphate-induced vascular calcification.

Vascular calcification is a common finding in atherosclerosis and in patients with chronic kidney disease. The renin-angiotensin system plays a role in the pathogenesis of cardiovascular remodeling. Here, we examined the hypothesis that angiotensin II type 2 receptor (AT2) stimulation has inhibitory effects on phosphate-induced vascular calcification. In vivo, calcification of the thoracic aorta induced by an adenine and high-phosphate diet was markedly attenuated in smooth muscle cell-specific AT2-overexpressing mice (smAT2-Tg) compared with wild-type and AT2-knockout mice (AT2KO). Similarly, mRNA levels of relevant osteogenic and vascular smooth muscle cell marker genes were unchanged in smAT2-Tg mice, while their expression was significantly altered in wild-type mice in response to high dietary phosphate. Ex vivo, sections of thoracic aorta were cultured in media supplemented with inorganic phosphate. Aortic rings from smAT2-Tg mice showed less vascular calcification compared with those from wild-type mice. In vitro, calcium deposition induced by high-phosphate media was markedly attenuated in primary vascular smooth muscle cells derived from smAT2-Tg mice compared with the two other mouse groups. To assess the underlying mechanism, we investigated the effect of PPAR-γ, which we previously reported as one of the possible downstream effectors of AT2 stimulation. Treatment with a PPAR-γ antagonist attenuated the inhibitory effects on vascular calcification observed in smAT2-Tg mice fed an adenine and high-phosphate diet. Our results suggest that AT2 activation represents an endogenous protective pathway against vascular calcification. Its stimulation may efficiently reduce adverse cardiovascular events in patients with chronic kidney disease.

Pseudoxanthoma Elasticum as a Paradigm of Heritable Ectopic Mineralization Disorders: Pathomechanisms and Treatment Development.

Ectopic mineralization is a global problem and leading cause of morbidity and mortality. The pathomechanisms of ectopic mineralization are poorly understood. Recent studies on heritable ectopic mineralization disorders with defined gene defects have been helpful in elucidation of the mechanisms of ectopic mineralization in general. The prototype of such disorders is pseudoxanthoma elasticum (PXE), a late-onset, slowly progressing disorder with multisystem clinical manifestations. Other conditions include generalized arterial calcification of infancy (GACI), characterized by severe, early-onset mineralization of the cardiovascular system, often with early postnatal demise. In addition, arterial calcification due to CD73 deficiency (ACDC) occurs late in life, mostly affecting arteries in the lower extremities in elderly individuals. These three conditions, PXE, GACI, and ACDC, caused by mutations in ABCC6, ENPP1, and NT5E, respectively, are characterized by reduced levels of inorganic pyrophosphate (PPi) in plasma. Because PPi is a powerful antimineralization factor, it has been postulated that reduced PPi is a major determinant for ectopic mineralization in these conditions. These and related observations on complementary mechanisms of ectopic mineralization have resulted in development of potential treatment modalities for PXE, including administration of bisphosphonates, stable PPi analogs with antimineralization activity. It is conceivable that efficient treatments may soon become available for heritable ectopic mineralization disorders with application to common calcification disorders.

Lanthanum carbonate, a phosphate binder, inhibits calcification of implanted aortic allografts in a rat model.

OBJECTIVES: Calcification is one of the major postoperative problems after aortic allograft implantation. We hypothesized that phosphate binders, lanthanum carbonate and calcium carbonate inhibit calcification of implanted aortic allografts and verified this hypothesis using a rat model. METHODS: Aortas were harvested from 4-week-old Brown Norway rats and implanted into the subdermal space of 4-week-old Lewis rats. Twenty-seven recipient Lewis rats were divided into Group N, Group L, and Group C (9 rats per group), which were fed a normal diet, a normal diet containing 3% lanthanum carbonate, and a normal diet containing 3% calcium carbonate, respectively. Implanted aortic allografts were explanted 2 weeks later. Calcification of aortic allografts was evaluated using von Kossa staining and calcium content assay. Calcification score was defined in von Kossa staining as 0 (none), 1 (mild), 2 (moderate), and 3 (severe). Serum calcium and phosphorus levels at euthanasia were measured. RESULTS: Calcification scores were 2.6, 1.2, and 0.8, and calcium content was 48.9, 15.8, and 8.9 mg/dry·g, in Groups N, L, and C, respectively. Calcification was significantly reduced in Groups L and C. Serum calcium level was 11.5, 12.2, and 13.5 mg/dl, and serum phosphorus level was 15.4, 12.5, and 11.7 mg/dl, in Groups N, L, and C, respectively. Serum calcium level in Group C was significantly higher than in the other two groups. CONCLUSIONS: Lanthanum carbonate and calcium carbonate significantly reduced calcification of implanted aortic allografts in young rats. Although calcium carbonate induced hypercalcemia, lanthanum carbonate has significant potential to inhibit calcification of implanted aortic allografts.

Vitamin K status, supplementation and vascular disease: a systematic review and meta-analysis.

OBJECTIVES: Vascular stiffness (VS) and vascular calcification (VC) are surrogate markers of vascular health associated with cardiovascular events. Vitamin K-dependent proteins (VKDP) are associated with VS and VC and require vitamin K for activity. We conducted a systematic review and meta-analysis of: (1) the effect of vitamin K supplementation on VS and VC and (2) association of inactive VKDP levels with incident cardiovascular disease and mortality. METHODS: Two authors searched MEDLINE and Embase databases and Cochrane and ISRCTN registries for studies of vitamin K clinical trials that measured effects on VC, VS or VKDP and longitudinal studies assessing effect of VKDP on incident CVD or mortality. Random effects meta-analyses were performed. RESULTS: Thirteen controlled clinical trials (n=2162) and 14 longitudinal studies (n=10 726) met prespecified inclusion criteria. Vitamin K supplementation was associated with significant reduction in VC (-9.1% (95% CI -17.7 to -0.5); p=0.04) and VKDP (desphospho-uncarboxylated matrix Gla protein; -44.7% (95% CI -65.1 to -24.3), p<0.0001) and uncarboxylated osteocalcin; -12.0% (95% CI -16.7 to -7.2), p<0.0001) compared with control, with a non-significant improvement in VS. In longitudinal studies with median follow-up of 7.8 (IQR 4.9-11.3) years, VKDP levels were associated with a combined endpoint of CVD or mortality (HR 0.45 (95% CI 0.07 to 0.83), p=0.02). CONCLUSIONS: Supplementation with vitamin K significantly reduced VC, but not VS, compared with control. The conclusions drawn are limited by small numbers of studies with substantial heterogeneity. VKDP was associated with combined endpoint of CVD or mortality. Larger clinical trials of effect of vitamin K supplementation to improve VC, VS and long-term cardiovascular health are warranted. TRIAL REGISTRATION NUMBER: CRD42017060344.

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.

Pseudoxanthoma elasticum (PXE), a prototype of heritable ectopic mineralization disorders, is caused by mutations in the ABCC6 gene encoding a putative efflux transporter ABCC6. It was recently shown that the absence of ABCC6-mediated adenosine triphosphate release from the liver and, consequently, reduced inorganic pyrophosphate levels underlie the pathogenesis of PXE. Given that tissue-nonspecific alkaline phosphatase (TNAP), encoded by ALPL, is the enzyme responsible for degrading inorganic pyrophosphate, we hypothesized that reducing TNAP levels either by genetic or pharmacological means would lead to amelioration of the ectopic mineralization phenotype in the Abcc6-/- mouse model of PXE. Thus, we bred Abcc6-/- mice to heterozygous Alpl+/- mice that display approximately 50% plasma TNAP activity. The Abcc6-/-Alpl+/- double-mutant mice showed 52% reduction of mineralization in the muzzle skin compared with the Abcc6-/-Alpl+/+ mice. Subsequently, oral administration of SBI-425, a small molecule inhibitor of TNAP, resulted in 61% reduction of plasma TNAP activity and 58% reduction of mineralization in the muzzle skin of Abcc6-/- mice. By contrast, SBI-425 treatment of Enpp1 mutant mice, another model of ectopic mineralization associated with reduced inorganic pyrophosphate, failed to reduce muzzle skin mineralization. These results suggest that inhibition of TNAP might provide a promising treatment strategy for PXE, a currently intractable disease.

High calcium, phosphate and calcitriol supplementation leads to an osteocyte-like phenotype in calcified vessels and bone mineralisation defect in uremic rats.

A link between vascular calcification and bone anomalies has been suggested in chronic kidney disease (CKD) patients with low bone turnover disease. We investigated the vascular expression of osteocyte markers in relation to bone microarchitecture and mineralization defects in a model of low bone turnover CKD rats with vascular calcification. CKD with vascular calcification was induced by 5/6 nephrectomy followed by high calcium and phosphate diet, and vitamin D supplementation (Ca/P/VitD). CKD + Ca/P/VitD group (n = 12) was compared to CKD + normal diet (n = 12), control + normal diet (n = 8) and control + Ca/P/VitD supplementation (n = 8). At week 6, tibia, femurs and the thoracic aorta were analysed by Micro-Ct, histomorphometry and for expression of osteocyte markers. High Ca/P/VitD treatment induced vascular calcification only in CKD rats, suppressed serum parathyroid hormone levels and led to higher sclerostin, DKK1 and FGF23 serum levels. Expression of sclerostin, DKK1 and DMP1 but not FGF23 were increased in calcified vessels from CKD + Ca/P/VitD rats. Despite low parathyroid hormone levels, tibia bone cortical thickness was significantly lower in CKD + Ca/P/VitD rats as compared to control rats fed a normal diet, which is likely the result of radial growth impairment. Finally, Ca/P/VitD treatment in CKD rats induced a bone mineralization defect, which is likely explained by the high calcitriol dose. In conclusion, Ca/P/VitD supplementation in CKD rats induces expression of osteocyte markers in vessels and bone mineralisation anomalies. Further studies should evaluate the mechanisms of high dose calcitriol-induced bone mineralisation defects in CKD.

Impact of Blood or Erythrocyte Membrane Fatty Acids for Disease Risk Prediction: Focusing on Cardiovascular Disease and Chronic Kidney Disease.

Fatty acids (FAs) are essential nutrients and main constituents of cell membranes that are involved in the signaling pathway and associated with health conditions. We investigated if blood or erythrocyte membrane FAs can predict the risk of cardiovascular disease (CVD), chronic kidney disease (CKD), and related complications. Omega-3 (n-3) FAs are important predictors for metabolic syndrome, diabetes, CVD, and CKD risks, and the n-3 index is also a good biomarker for sudden cardiac death in coronary artery disease. Linoleic acid, which is one of the major n-6 FAs reflecting recent dietary FA intake, may predict CVD risk and mortality in the general population and patients with CKD. Monounsaturated FAs (MUFAs) are also related to diabetes or diabetic nephropathy. Oleic acid, a major MUFA, is an emerging marker that is related to acute coronary syndrome, low glomerular filtration rate, and vascular calcification in patients with CKD, and can be modified by n-3 FA supplementation. Saturated FAs, trans-FAs, and FA desaturation/elongation are associated with CVD risk; however, few studies have been conducted on patients with CKD. In summary, blood or erythrocyte membrane FA measurements are important for CVD and CKD risk prediction and management. Further studies are needed to elucidate the FAs for their risk predictions.