Dataset Information

Calciprotein particle inhibition explains magnesium-mediated protection against vascular calcification.

ABSTRACT:

Background

Phosphate (Pi) toxicity is a strong determinant of vascular calcification development in chronic kidney disease (CKD). Magnesium (Mg2+) may improve cardiovascular risk via vascular calcification. The mechanism by which Mg2+ counteracts vascular calcification remains incompletely described. Here we investigated the effects of Mg2+ on Pi and secondary crystalline calciprotein particles (CPP2)-induced calcification and crystal maturation.

Methods

Vascular smooth muscle cells (VSMCs) were treated with high Pi or CPP2 and supplemented with Mg2+ to study cellular calcification. The effect of Mg2+ on CPP maturation, morphology and composition was studied by medium absorbance, electron microscopy and energy dispersive spectroscopy. To translate our findings to CKD patients, the effects of Mg2+ on calcification propensity (T50) were measured in sera from CKD patients and healthy controls.

Results

Mg2+ supplementation prevented Pi-induced calcification in VSMCs. Mg2+ dose-dependently delayed the maturation of primary CPP1 to CPP2 in vitro. Mg2+ did not prevent calcification and associated gene and protein expression when added to already formed CPP2. Confirmatory experiments in human serum demonstrated that the addition of 0.2?mmol/L Mg2+ increased T50 from healthy controls by 51?±?15?min (P?ConclusionsOur results demonstrate that crystalline CPP2 mediates Pi-induced calcification in VSMCs. In vitro, Mg2+ delays crystalline CPP2 formation and thereby prevents Pi-induced calcification.

SUBMITTER: Ter Braake AD

PROVIDER: S-EPMC7203562 | biostudies-literature | 2020 May

REPOSITORIES: biostudies-literature

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Calciprotein particle inhibition explains magnesium-mediated protection against vascular calcification.

Ter Braake Anique D AD Eelderink Coby C Zeper Lara W LW Pasch Andreas A Bakker Stephan J L SJL de Borst Martin H MH Hoenderop Joost G J JGJ de Baaij Jeroen H F JHF

Nephrology, dialysis, transplantation : official publication of the European Dialysis and Transplant Association - European Renal Association 20200501 5

<h4>Background</h4>Phosphate (Pi) toxicity is a strong determinant of vascular calcification development in chronic kidney disease (CKD). Magnesium (Mg2+) may improve cardiovascular risk via vascular calcification. The mechanism by which Mg2+ counteracts vascular calcification remains incompletely described. Here we investigated the effects of Mg2+ on Pi and secondary crystalline calciprotein particles (CPP2)-induced calcification and crystal maturation.<h4>Methods</h4>Vascular smooth muscle cel ...[more]

PMID: 31605492

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Project description:BackgroundThe liver-derived plasma protein fetuin A is a systemic inhibitor of ectopic calcification. Fetuin-A stabilizes calcium phosphate mineral initially as ion clusters to form calciprotein monomers (CPM), and then as larger multimeric consolidations containing amorphous calcium phosphate (primary CPP, CPP 1) or more crystalline phases (secondary CPP, CPP 2). CPM and CPP mediate excess mineral stabilization, transport and clearance from circulation.MethodsWe injected i.v. synthetic fluorescent CPM and studied their clearance by live two-photon microscopy. We analyzed organ sections by fluorescence microscopy to assess CPM distribution. We studied cellular clearance and cytotoxicity by flow cytometry and live/dead staining, respectively, in cultured macrophages, liver sinusoidal endothelial cells (LSEC), and human proximal tubule epithelial HK-2 cells. Inflammasome activation was scored in macrophages. Fetuin A monomer and CPM charge were analyzed by ion exchange chromatography.ResultsLive mice cleared CPP in the liver as published previously. In contrast, CPM were filtered by kidney glomeruli into the Bowman space and the proximal tubules, suggesting tubular excretion of CPM-bound calcium phosphate and reabsorption of fetuin A. Fetuin-A monomer clearance was negligible in liver and low in kidney. Anion exchange chromatography revealed that fetuin A monomer was negatively charged, whereas CPM appeared neutral, suggesting electrochemical selectivity of CPM versus fetuin A. CPM were non-toxic in any of the investigated cell types, whereas CPP 1 were cytotoxic. Unlike CPP, CPM also did not activate the inflammasome.ConclusionsFetuin-A prevents calcium phosphate precipitation by forming CPM, which transform into CPP. Unlike CPP, CPM do not trigger inflammation. CPM are readily cleared in the kidneys, suggesting CPM as a physiological transporter of excess calcium and phosphate. Upon prolonged circulation, e.g., in chronic kidney disease, CPM will coalesce and form CPP, which cannot be cleared by the kidney, but will be endocytosed by liver sinusoidal endothelial cells and macrophages. Large amounts of CPP trigger inflammation. Chronic CPM and CPP clearance deficiency thus cause calcification by CPP deposition in blood vessels and soft tissues, as well as inflammation.

Project description:Rationale & objectiveCharacteristics of the transformation of primary to secondary calciprotein particles (CPPs) in serum, including the size of secondary CPP (CPP2) aggregates and the time of transformation (T50), may be markers for arterial calcification in patients undergoing hemodialysis (HD). We examined the associations of CPP2 aggregate size and T50 with arterial calcification in incident HD patients.Study designProspective cohort study.Setting & participantsIncident HD patients (n=402with available CPP2 measures and n=388with available T50 measures) from the Predictors of Arrhythmic and Cardiovascular Risk in End-Stage Renal Disease (PACE) Study PREDICTORS: Serum CPP2 size and T50 at baseline.OutcomesPrimary outcomes were baseline coronary artery and thoracic aorta calcifications. Exploratory outcomes included baseline arterial stiffness, measured by pulse wave velocity (PWV) and ankle brachial index, and longitudinally, repeat measures of PWV and all-cause mortality.Analytical approachTobit regression, multiple linear regression, Poisson regression, linear mixed-effects regression, and Cox proportional hazards regression.ResultsMean age was 55±13 years, 41% were women, 71% were Black, and 57% had diabetes mellitus. Baseline CPP2 size and T50 were correlated with baseline fetuin A level (r=-0.59 for CPP2 and 0.44 for T50; P<0.001 for both), but neither was associated with baseline measures of arterial calcification or arterial stiffness. Baseline CPP2 size and T50 were not associated with repeat measures of PWV. During a median follow-up of 3.5 (IQR, 1.7-6.2) years, larger CPP2 was associated with higher risk for mortality (HR, 1.17 [95% CI, 1.05-1.31] per 100nm larger CPP2 size) after adjusting for demographics and comorbid conditions, but there was no association between baseline T50 and risk for mortality.LimitationsPossible imprecision in assays, small sample size, limited generalizability to incident HD populations with different racial composition, and residual confounding.ConclusionsIn incident HD patients, neither CPP2 size nor T50 was associated with prevalent arterial calcification and stiffness. Larger CPP2 was associated with risk for mortality, but this finding needs to be confirmed in future studies.

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Project description:Sclerostin is a negative regulator of the Wnt/β-catenin signaling and is, therefore, an important inhibitor of bone formation and turnover. Because ectopic vascular calcification develops in a similar way to bone formation, one might reasonably attribute a role to sclerostin in this pathological process. Ectopic calcification, especially vascular calcification, importantly contributes to mortality in elderly and patients with diabetes, osteoporosis, chronic kidney disease (CKD), and hypertension. The central players in this ectopic calcification process are the vascular smooth muscle cells that undergo dedifferentiation and thereby acquire characteristics of bonelike cells. Therefore, we hypothesize that depletion/deactivation of the Wnt/β-catenin signaling inhibitor sclerostin may promote the development of ectopic calcifications through stimulation of bone-anabolic effects at the level of the arteries. We investigated the role of sclerostin (encoded by the Sost gene) during vascular calcification by using either Sost-/- mice or anti-sclerostin antibody. Sost-/- and wild-type (WT) mice (C57BL/6J background) were administered an adenine-containing diet to promote the development of CKD-induced vascular calcification. Calcifications developed more extensively in the cardiac vessels of adenine-exposed Sost-/- mice, compared to adenine-exposed WT mice. This could be concluded from the cardiac calcium content as well as from cardiac tissue sections on which calcifications were visualized histochemically. In a second experiment, DBA/2J mice were administered a warfarin-containing diet to induce vascular calcifications in the absence of CKD. Here, warfarin exposure led to significantly increased aortic and renal tissue calcium content. Calcifications, which were present in the aortic medial layer and renal vessels, were significantly more pronounced when warfarin treatment was combined with anti-sclerostin antibody treatment. This study demonstrates a protective effect of sclerostin during vascular calcification. © 2022 The Authors. Journal of Bone and Mineral Research published by Wiley Periodicals LLC on behalf of American Society for Bone and Mineral Research (ASBMR).

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Calciprotein particle inhibition explains magnesium-mediated protection against vascular calcification.

Background

Methods

Results

Publications

Calciprotein particle inhibition explains magnesium-mediated protection against vascular calcification.

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