Insights into the association of <scp>G</scp>la‐rich protein and osteoarthritis, novel splice variants and γ‐carboxylation status

Rafael, Marta S.; Cavaco, Sofia; Viegas, Carla S. B.; Santos, Sofia; Ramos, Acácio; Willems, Brecht A. G.; Herfs, Marjolein; Theuwissen, Elke; Vermeer, Cees; Simes, Dina C.

doi:10.1002/mnfr.201300941

Cited by 46 publications

(56 citation statements)

References 40 publications

(73 reference statements)

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“…Western blot detection of tGRP, cGRP, tMGP, cMGP and GAPDH was performed using specific primary antibodies CTerm-GRP (5 μg/ml), cGRP (1:1000 v/v) (GenoGla Diagnostics, Faro, Portugal), tMGP and cMGP (1:1000 v/v, both from VitaK BV, Maastricht, The Netherlands) previously validated as described [16, 30, 31, 33, 34, 36], and anti-GAPDH (1:500 v/v) (Santa Cruz Biotechnology). Detection was achieved using species-specific secondary horseradish peroxidase-conjugated antibodies and Western Lightning Plus-ECL (PerkinElmer Inc., Waltham, MA, USA).…”

Section: Methodsmentioning

confidence: 99%

“…The pCRII-TOPO/GRP clone [34] was used as template to PCR amplify the complete GRP ORF using primers HSaGRPmMax_F, containing a T7 Polymerase promoter minimal sequence and a kozak sequence upstream ATG initiation codon, and HuGRP_Ex5R (Table 1). The resulting PCR product was used to in vitro synthetize capped and poly(A) tailed GRP mRNA using the mMESSAGE mMACHINE T7 Ultra kit (Ambion, Life Technologies), according to manufacturer’s recommendations.…”

Section: Methodsmentioning

confidence: 99%

“…Our previous studies also shown that down-regulation of mediators of inflammation in chondrocytes and synoviocytes treated with GRP is apparently independent of its γ-carboxylation status [16]. GRP calcium binding properties [29, 33] and association to calcification processes [16, 30, 31, 34] indicate that its function might be related with prevention of calcium-induced signaling pathways and to inhibition of crystal formation/maturation by direct mineral-binding. Moreover, GRP is also involved in the mineralization-competence of VSMCs-derived EVs possibly associated with the fetuin-A-MGP calcification inhibitory system [31].…”

Section: Introductionmentioning

confidence: 99%

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Gla-rich protein function as an anti-inflammatory agent in monocytes/macrophages: Implications for calcification-related chronic inflammatory diseases

et al. 2017

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Calcification-related chronic inflammatory diseases are multifactorial pathological processes, involving a complex interplay between inflammation and calcification events in a positive feed-back loop driving disease progression. Gla-rich protein (GRP) is a vitamin K dependent protein (VKDP) shown to function as a calcification inhibitor in cardiovascular and articular tissues, and proposed as an anti-inflammatory agent in chondrocytes and synoviocytes, acting as a new crosstalk factor between these two interconnected events in osteoarthritis. However, a possible function of GRP in the immune system has never been studied. Here we focused our investigation in the involvement of GRP in the cell inflammatory response mechanisms, using a combination of freshly isolated human leucocytes and undifferentiated/differentiated THP-1 cell line. Our results demonstrate that VKDPs such as GRP and matrix gla protein (MGP) are synthesized and γ-carboxylated in the majority of human immune system cells either involved in innate or adaptive immune responses. Stimulation of THP-1 monocytes/macrophages with LPS or hydroxyapatite (HA) up-regulated GRP expression, and treatments with GRP or GRP-coated basic calcium phosphate crystals resulted in the down-regulation of mediators of inflammation and inflammatory cytokines, independently of the protein γ-carboxylation status. Moreover, overexpression of GRP in THP-1 cells rescued the inflammation induced by LPS and HA, by down-regulation of the proinflammatory cytokines TNFα, IL-1β and NFkB. Interestingly, GRP was detected at protein and mRNA levels in extracellular vesicles released by macrophages, which may act as vehicles for extracellular trafficking and release. Our data indicate GRP as an endogenous mediator of inflammatory responses acting as an anti-inflammatory agent in monocytes/macrophages. We propose that in a context of chronic inflammation and calcification-related pathologies, GRP might act as a novel molecular mediator linking inflammation and calcification events, with potential therapeutic application.

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Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Gla-rich protein function as an anti-inflammatory agent in monocytes/macrophages: Implications for calcification-related chronic inflammatory diseases

et al. 2017

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“…Gla rich protein (GRP), which is another vitamin K dependent implicated in calcification, has recently identified in human articular cartilage [68]. Similar to MGP, GRP from arthritic cartilage is primarily uncarboxylated, whereas GRP from healthy cartilage is primarily carboxylated [69]. …”

Section: Vitamin K and Osteoarthritismentioning

confidence: 99%

The Role of Vitamin K in Chronic Aging Diseases: Inflammation, Cardiovascular Disease, and Osteoarthritis

Harshman

Shea

2016

Curr Nutr Rep

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Vitamin K is an enzyme cofactor required for the carboxylation of vitamin K dependent proteins, several of which have been implicated in diseases of aging. Inflammation is recognized as a crucial component of many chronic aging diseases and evidence suggests vitamin K has an anti-inflammatory action that is independent of its role as an enzyme co-factor. Vitamin K-dependent proteins and inflammation have been implicated in cardiovascular disease and osteoarthritis, which are leading causes of disability and mortality in older adults. The purpose of this review is to summarize observational studies and randomized trials focused on vitamin K status and inflammation, cardiovascular disease, and osteoarthritis. Although mechanistic evidence suggests a protective role for vitamin K in these age-related conditions, the benefit of vitamin K supplementation is controversial because observational data are equivocal and the number of randomized trials is few.

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“…Interestingly, the Keutel syndrome in humans, which is characterized by loss-of-function mutations in the MGP gene results in non-lethal abnormal soft tissues calcification, which suggest additional or compensatory mechanisms of mineralization inhibition in humans. The discovery of an additional VKDP, Gla-rich protein (GRP), also expressed and accumulated in skeletal and vascular tissues [63][64][65], that we have recently shown to function as a calcification inhibitor in the cardiovascular [66] and articular systems [67], should open a new window of knowledge in the area of pathological calcification of connective tissues and increase the range of vitamin K action. GRP was initially identified in sturgeon calcified cartilage and characterized by the presence of an unprecedented 16 Gla residues within its 74 amino acid total protein sequence, and an impressive degree of evolutionary conservation [63].…”

Section: Vitamin K Mechanism Of Actionmentioning

confidence: 99%

New Perspectives for the Nutritional Value of Vitamin K in Human Health

Viegas¹,

Simes²

2016

J Nutr Disorders Ther

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Vitamin K is an essential micronutrient in the post-translational modification of specific glutamic acid residues (Glu) into γ-carboxyglutamic acid residues (Gla) in target proteins known as vitamin K-dependent proteins (VKDPs). In healthy conditions of sufficient vitamin K status, a vitamin K recycling system maintains sufficient vitamin K levels for proper γ-carboxylation of VKDPs, and vitamin K antagonists (VKAs) widely used as anticoagulants inhibit vitamin K recycling. Besides its well-known function in the maintenance of normal coagulation, vitamin K has been reported to have other diverse physiological functions with impact in human health. In extra-hepatic tissues vitamin K deficiency results in impairment of VKDPs γ-carboxylation with important implications in bone and cardiovascular health. Although most of the vitamin K effects have been associated with regulation of mineralization in connective tissues through the action of matrix Gla protein (MGP) and osteocalcin (OC), the discovery of Gla-rich protein (GRP) opens new perspectives on the potential therapeutic range of vitamin K.

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Insights into the association of Gla‐rich protein and osteoarthritis, novel splice variants and γ‐carboxylation status

Abstract: Overall, our results indicate the predominance of GRP-F1, and a clear association of ucGRP with OA cartilage and synovial membrane. Levels of vitamin K should be further assessed in these patients to determine its potential therapeutic use as a supplement in OA treatment.

Cited by 46 publications

References 40 publications

Gla-rich protein function as an anti-inflammatory agent in monocytes/macrophages: Implications for calcification-related chronic inflammatory diseases

Gla-rich protein function as an anti-inflammatory agent in monocytes/macrophages: Implications for calcification-related chronic inflammatory diseases

The Role of Vitamin K in Chronic Aging Diseases: Inflammation, Cardiovascular Disease, and Osteoarthritis

New Perspectives for the Nutritional Value of Vitamin K in Human Health

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