Effects of fetuin-A-containing calciprotein particles on posttranslational modifications of fetuin-A in HepG2 cells

AbstractFetuin-A is an inhibitor of ectopic calcification that is expressed mainly in hepatocytes and is secreted into the circulation after posttranslational processing, including glycosylation and phosphorylation. The molecular weight (MW) of fully modified fetuin-A (FM-fetuin-A) is approximately 60 kDa in an immunoblot, which is much higher than the estimated MW by amino acid sequence. Under conditions of calcification stress such as advanced stage chronic kidney disease, fetuin-A prevents calcification by forming colloidal complexes, which are referred to as calciprotein particles (CPP). Since the significance of CPP in this process is unclear, we investigated the effect of synthetic secondary CPP on the level of FM-fetuin-A in HepG2 cells. Secondary CPP increased the level of FM-fetuin-A in dose- and time-dependent manners, but did not affect expression of mRNA for fetuin-A. Treatment with O- and/or N-glycosidase caused a shift of the 60 kDa band of FM-fetuin-A to a lower MW. Preincubation with brefeldin A, an inhibitor of transport of newly synthesized proteins from the endoplasmic reticulum to the Golgi apparatus, completely blocked the secondary CPP-induced increase in FM-fetuin-A. Treatment with BAPTA-AM, an intracellular calcium chelating agent, also inhibited the CPP-induced increase in the FM-fetuin-A level. Secondary CPP accelerate posttranslational processing of fetuin-A in HepG2 cells.

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FP033Effects of fetuin-A-containing calciprotein particle (CPP) on posttranslational modifications of fetuin-A in HepG2 cells

Nephrology Dialysis Transplantation ◽

10.1093/ndt/gfz106.fp033 ◽

2019 ◽

Vol 34 (Supplement_1) ◽

Author(s):

Hideki Uedono ◽

Katsuhito Mori ◽

Akinobu Ochi ◽

Shinya Nakatani ◽

Akihiro Tsuda ◽

...

Keyword(s):

Posttranslational Modifications ◽

Hepg2 Cells ◽

Fetuin A

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Fetuin-A-Containing Calciprotein Particles Reduce Mineral Stress in the Macrophage

PLoS ONE ◽

10.1371/journal.pone.0060904 ◽

2013 ◽

Vol 8 (4) ◽

pp. e60904 ◽

Cited By ~ 85

Author(s):

Edward R. Smith ◽

Eric Hanssen ◽

Lawrence P. McMahon ◽

Stephen G. Holt

Keyword(s):

Fetuin A ◽

Calciprotein Particles ◽

Mineral Stress

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Microvasculopathy, Luminal Calcification and Premature Aging in Fetuin-A Deficient Mice

10.1101/577098 ◽

2019 ◽

Cited By ~ 2

Author(s):

Marietta Herrmann ◽

Anne Babler ◽

Irina Moshkova ◽

Felix Gremse ◽

Fabian Kiessling ◽

...

Keyword(s):

Soft Tissue ◽

Early Stage ◽

Analytical Electron Microscopy ◽

Premature Aging ◽

Fetuin A ◽

Genome Wide ◽

Ectopic Mineralization ◽

Brown Adipose ◽

Calciprotein Particles ◽

Deficient Mice

AbstractObjectiveThe plasma protein fetuin-A mediates the formation of protein-mineral colloids known as calciprotein particles (CPP) – rapid clearance of these CPP by the reticuloendothelial system prevents errant mineral precipitation and therefore ectopic mineralization (calcification). The mutant mouse strain D2,Ahsg-/- combines fetuin-A deficiency with the mineralization-prone DBA/2 genetic background, having a particularly severe compound phenotype of microvascular and soft tissue mineralization. Here we studied mechanisms leading to soft tissue mineralization, organ damage and premature aging in these mice.Approach and ResultsWe analyzed mice longitudinally by echocardiography, X-ray-computed tomography, analytical electron microscopy, histology, mass spectrometry proteomics, and genome-wide microarray-based expression analyses of D2 wildtype and Ahsg-/- mice.Fetuin-A deficient mice had calcified lesions in myocardium, lung, brown adipose tissue, reproductive organs, spleen, pancreas, kidney and the skin, associated with reduced growth, cardiac output and premature aging. Importantly, early stage calcified lesions presented in the lumen of the microvasculature suggesting precipitation of mineral containing complexes from the fluid phase of blood. Genome-wide expression analysis of calcified lesions and surrounding (not calcified) tissue, together with morphological observations, indicated that the ectopic mineralization was not associated with osteochondrogenic cell differentiation, but rather with thrombosis and fibrosis.ConclusionsCollectively, these results demonstrate that pathological mineralization can start by intravascular mineral deposition causing microvasculopathy, which impacts on growth, organ function and survival. Our study underscores the importance of fetuin-A and related systemic regulators of mineralized matrix metabolism to prevent cardiovascular disease, especially in dysregulated mineral homeostasis.

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Phosphorylated fetuin-A-containing calciprotein particles are associated with aortic stiffness and a procalcific milieu in patients with pre-dialysis CKD

Nephrology Dialysis Transplantation ◽

10.1093/ndt/gfr609 ◽

2012 ◽

Vol 27 (5) ◽

pp. 1957-1966 ◽

Cited By ~ 101

Author(s):

Edward R. Smith ◽

Martin L. Ford ◽

Laurie A. Tomlinson ◽

Chakravarthi Rajkumar ◽

Lawrence P. McMahon ◽

...

Keyword(s):

Aortic Stiffness ◽

Fetuin A ◽

Calciprotein Particles

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The effect of increasing dialysate magnesium on calciprotein particles, inflammation and bone markers: post hoc analysis from a randomized controlled clinical trial

Nephrology Dialysis Transplantation ◽

10.1093/ndt/gfz234 ◽

2019 ◽

Cited By ~ 6

Author(s):

Iain Bressendorff ◽

Ditte Hansen ◽

Andreas Pasch ◽

Stephen G Holt ◽

Morten Schou ◽

...

Keyword(s):

Clinical Trial ◽

Bone Formation ◽

Bone Turnover ◽

Systemic Inflammation ◽

Maintenance Hemodialysis ◽

Controlled Clinical Trial ◽

Ectopic Calcification ◽

Tartrate Resistant Acid Phosphatase ◽

Calciprotein Particles ◽

Post Hoc

Abstract Background The formation of calciprotein particles (CPPs) may be an important component of the humoral defences against ectopic calcification. Although magnesium (Mg) has been shown to delay the transition of amorphous calcium-/phosphate-containing primary CPP (CPP-1) to crystalline apatite-containing secondary CPP (CPP-2) ex vivo, effects on the endogenous CPP pool are unknown. Methods We used post hoc analyses from a randomized double-blind parallel-group controlled clinical trial of 28 days treatment with high dialysate Mg of 2.0 mEq/L versus standard dialysate Mg of 1.0 mEq/L in 57 subjects undergoing maintenance hemodialysis for end-stage kidney disease. CPP load, markers of systemic inflammation and bone turnover were measured at baseline and follow-up. Results After 28 days of treatment with high dialysate Mg, serum total CPP (−52%), CPP-1 (−42%) and CPP-2 (−68%) were lower in the high Mg group (all P < 0.001) but were unchanged in the standard dialysate Mg group. Tumour necrosis factor-α (−20%) and interleukin-6 (−22%) were also reduced with high dialysate Mg treatment (both P < 0.01). High dialysate Mg resulted in higher levels of bone-specific alkaline phosphatase (a marker of bone formation) (+17%) but lower levels of tartrate-resistant acid phosphatase 5 b (a marker of bone resorption; −33%) (both P < 0.01). Inflammatory cytokines and bone turnover markers were unchanged in the standard dialysate Mg group over the same period. Conclusions In this exploratory analysis, increasing dialysate Mg was associated with reduced CPP load and systemic inflammation and divergent changes in markers of bone formation and resorption.

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