Warfarin-induced artery calcification is accelerated by growth and vitaminD

The present studies demonstrate that growth and vitamin D treatment enhance the extent of artery calcification in rats given sufficient doses of Warfa rin to inhibit gamma-carboxylation of matrix Gla protein, a calcification i nhibitor known to be expressed by smooth muscle cells and macrophages in th e artery wall. The first series of experiments examined the influence of ag e and growth status on artery calcification in Warfarin-treated rats. Treat ment for 2 weeks with Warfarin caused massive focal calcification of the ar tery media in 20-day-old rats and less extensive focal calcification in 42- day-old rats. In contrast, no artery calcification could be detected in 10- month-old adult rats even after 4 weeks of Warfarin treatment. To directly examine the importance of growth to Warfarin-induced artery calcification i n animals of the same age, 20-day-old rats were fed for 2 weeks either an a d libitum diet or a 6-g/d restricted diet that maintains weight but prevent s growth. Concurrent treatment of both dietary groups with Warfarin produce d massive focal calcification of the artery media in the ad libitum-fed rat s but no detectable artery calcification in the restricted-diet, growth-inh ibited group. Although the explanation for the association between artery c alcification and growth status cannot be determined from the present study, there was a relationship between higher serum phosphate and susceptibility to artery calcification, with 30% higher levels of serum phosphate in youn g, ad libitum-fed rats compared with either of the groups that was resistan t to Warfarin-induced artery calcification, ie, the 10-month-old rats and t he restricted-diet, growth-inhibited young rats. This observation suggests that increased susceptibility to Warfarin-induced artery calcification coul d be related to higher serum phosphate levels. The second set of experiment s examined the possible synergy between vitamin D and Warfarin in artery ca lcification. High doses of vitamin D are known to cause calcification of th e artery media in as little as 3 to 4 days. High doses of the vitamin K ant agonist Warfarin are also known to cause calcification of the artery media, but at treatment times of 2 weeks or longer yet not at 1 week. In the curr ent study, we investigated the synergy between these 2 treatments and found that concurrent Warfarin administration dramatically increased the extent of calcification in the media of vitamin D-treated rats at 3 and 4 days. Th ere was a close parallel between the effect of vitamin D dose on artery cal cification and the effect of vitamin D dose on the elevation of serum calci um, which suggests that vitamin D may induce artery calcification through i ts effect on serum calcium. Because Warfarin treatment had no effect on the elevation in serum calcium produced by vitamin D, the synergy between Warf arin and vitamin D is probably best explained by the hypothesis that Warfar in inhibits the activity of matrix Gla protein as a calcification inhibitor . High levels of matrix Gla protein are found at sites of artery calcificat ion in rats treated with vitamin D plus Warfarin, and chemical analysis sho wed that the protein that accumulated was indeed not gamma-carboxylated. Th ese observations indicate that although the gamma-carboxyglutamate residues of matrix Gla protein are apparently required for its function as a calcif ication inhibitor, they are not required for its accumulation at calcificat ion sites.