We investigated the evolutionary origin of a serum activity that induces calcification within a type I collagen matrix, an activity previously described in rat and bovine serum. Serum was obtained from vertebrates with calcified tissues (bony fish and shark), vertebrates without calcified tissues (lamprey and hagfish), and three invertebrates (marine worm, crab, and sea urchin). Serum from the bony fish and shark proved to contain a potent nucleator of collagen calcification; like the previously described calcifying activity in rat serum, the fish and shark activities are both able to recalcify a demineralized rat tibia when tested in Dulbecco's modified Eagle medium containing as little as 1.5% of the respective serum and have an apparent molecular weight of 50-150 kDa. No calcifying activity could be detected in any of several experimental tests of invertebrate or hagfish serum. Weak calcifying activity could be detected in lamprey serum, but calcification was restricted to the growth plate of the decalcified tibia, with no detectable calcification in the type I collagen of the midshaft. These studies reveal a correlation between the evolutionary timing of the appearance of calcified tissues in vertebrates and the appearance of the serum activity that initiates calcification within collagen and, therefore, support the hypothesis that this serum activity may play a role in normal calcification of bone.
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