Vitamin K-Dependent Carboxylation in Articular Chondrocytes

Abstract

A nonhepatic vitamin K-dependent protein, matrix Gla protein, has recently been identified in cartilage where it may play an important role in the control of mineralization or matrix development. We have investigated the vitamin K cycle in chondrocytes isolated from bovine and rabbit articular cartilage and examined these cells for their ability to synthesize vitamin K-dependent proteins. Chondrocytes were found to have an active vitamin K-dependent carboxylation system. Preincubation of the cells with warfarin resulted in a significant increase in the measured carboxylase activity. Both vitamin K epoxide reductase and DT-diaphorase (EC 1.6.99.2) activity were present indicating that chondrocytes are capable of producing reduced vitamin K1H2, the cofactor for the vitamin K-dependent carboxylase. Specific 14C-labeling of microsomal vitamin K-dependent protein precursors demonstrated synthesis of several vitamin K-dependent proteins by chondrocytes. 35S-labeling of chondrocyte proteins provided evidence that matrix Gla protein is synthesized by these cells.

Articular-cartilage matrix γ-carboxyglutamic acid-containing protein. Characterization and immunolocalization

“…Abnormal articular-cartilage calcification has been seen in conditions such as chondrocalcinosis, hydroxyapatite deposition and osteoarthritis [8][9][10] and could be due to a local defect in MGP production. Recently we have discovered that articular-cartilage chondrocytes contain the necessary enzymes for the synthesis of vitamin K-dependent proteins and that they do synthesize MGP [11]. Our present studies demonstrate for the first time the presence and localization of MGP in bovine, monkey and human articular cartilage.…”

“…MGP appears to represent about 0.05 % of the total amount of cartilage protein extracted with guanidinium chloride, and the concentration of 0.5 jig/mg of tissue protein is consistent with concentrations noted in rat cartilages [2]. It has been shown that articularcartilage chondrocytes are capable of synthesizing a family of vitamin K-dependent proteins including MGP [11]. MGP is the first of this family to be isolated and characterized.…”

“…Cartilage MGP was extracted from bovine and human articular cartilage with guanidinium chloride as follows. Bovine cartilage was shaved from fetlock joints as previously described [11]. Human cartilage was obtained from femoral heads at the time of hip replacement surgery for osteoarthritis or femoral neck fracture.…”

Biosynthesis of the vitamin K-dependent matrix Gla protein (MGP) in chondrocytes: a fetuin–MGP protein complex is assembled in vesicles shed from normal but not from osteoarthritic chondrocytes

“…g-Carboxylase activity g-Carboxylase activity was assayed in 24 h cultured chondrocytes from normal (n ¼ 2) and osteoarthritic tissues (n ¼ 2) as 14 CO 2 incorporation into the synthetic g-carboxylase peptide substrate FLEEL as described 9 . The reaction was triggered by adding chemically reduced vitamin K 1 H 2 (100 mg/ml) to the assay mixture.…”

“…This modification of coagulation factors made in the liver is essential for normal hemostasis. We have shown previously that chondrocytes synthesize MGP and possess an active post-translational g-carboxylation system 9,10 .…”