Abstract-High doses of warfarin cause focal calcification of the elastic lamellae in the media of major arteries and in aortic heart valves in the rat. Aortic calcification was first seen after 2 weeks of warfarin treatment and progressively increased in density at 3, 4, and 5 weeks of treatment. By 5 weeks, the highly focal calcification of major arteries could be seen on radiographs and by visual inspection of the artery. The calcification of arteries induced by warfarin is similar to that seen in the matrix Gla protein (MGP)-deficient mouse, which suggests that warfarin induces artery calcification by inhibiting ␥-carboxylation of MGP and thereby inactivating the putative calcification-inhibitory activity of the protein. Warfarin treatment markedly increased the levels of MGP mRNA and protein in calcifying arteries and decreased the level of MGP in serum. Warfarin treatment did not affect bone growth, overall weight gain, or serum calcium and phosphorus levels, and, because of the concurrent administration of vitamin K, prothrombin times and hematocrits were normal. The results indicate that the improved warfarin plus vitamin K treatment protocol developed in this study should provide a useful model to investigate the role of MGP in preventing calcification of arteries and heart valves.

Abstract-The present experiments were carried out to test the hypothesis that arterial calcification is linked to bone resorption by determining whether the selective inhibition of bone resorption with osteoprotegerin will inhibit arterial calcification. In the first test, arterial calcification was induced by treating 22-day-old male rats with warfarin, a procedure that inhibits the ␥-carboxylation of matrix Gla protein and causes extensive calcification of the arterial media.Compared with rats treated for 1 week with warfarin alone, rats treated with warfarin plus osteoprotegerin at a dose of 1 mg/kg per day had dramatically reduced alizarin red staining for calcification in the aorta and in the carotid, hepatic, mesenteric, renal, and femoral arteries, and they had 90% lower levels of calcium and phosphate in the abdominal aorta (PϽ0.001) and in tracheal ring cartilage (PϽ0.01). More rapid arterial calcification was induced by treating 49-day-old male rats with toxic doses of vitamin D. Treatment for 96 hours with vitamin D caused widespread alizarin red staining for calcification in the aorta and the femoral, mesenteric, hepatic, renal, and carotid arteries, and osteoprotegerin completely prevented calcification in each of these arteries and reduced the levels of calcium and phosphate in the abdominal aorta to control levels (PϽ0.001). Treatment with vitamin D also caused extensive calcification in the lungs, trachea, kidneys, stomach, and small intestine, and treatment with osteoprotegerin reduced or prevented calcification in each of these sites. Measurement of serum levels of cross-linked N-teleopeptides showed that osteoprotegerin dramatically reduced bone resorption activity in each of these experiments (PϽ0.001). Therefore, we conclude that doses of osteoprotegerin that inhibit bone resorption are able to potently inhibit the calcification of arteries that is induced by warfarin treatment and by vitamin D treatment. 1,2 to explain the association between the increased bone resorption and increased arterial calcification that has been seen in the vitamin D-treated rat, 1 in the osteoprotegerin-deficient mouse, 3 and in patients with postmenopausal osteoporosis. 4 -12 One prediction of the hypothesis that arterial calcification is linked to bone resorption is that inhibitors of bone resorption should inhibit arterial calcification. 2 In previous studies, we have shown that arterial calcification induced by treatment with warfarin and by treatment with high doses of vitamin D is indeed inhibited by 2 potent inhibitors of bone resorption, the amino bisphosphonates alendronate and ibandronate, at doses of these drugs known to inhibit bone resorption in the rat. 2,13 In the present investigations, we have determined the effect of another potent inhibitor of bone resorption, osteoprotegerin, on arterial calcification by using subcutaneous doses of this protein that have been shown to inhibit bone resorption in the rat.Osteoprotegerin is a secreted protein of the tumor necrosis factor family, which regulat...

Phylloquinone supplementation slows the progression of CAC in healthy older adults with preexisting CAC, independent of its effect on total MGP concentrations. Because our data are hypothesis-generating, further studies are warranted to clarify this mechanism. This trial was registered at clinicaltrials.gov as NCT00183001.