After a lag period, rat aortas incubated in rat serum in vitro accumulated substantial quantities of calcium and phosphate. Examination of the tissue by x-ray diffraction, microradiography, electron diffraction, and electron microscopy indicated that the calcium-phosphate phase which formed was hydroxyapatite and that the crystals were localized almost exclusively in elastin. Selective elimination of various components of the aorta with proteolytic enzymes indicated that the presence of elastin was required for mineralization. Collagen fibers did not appear to be required for the initiation of calcification, nor did they seem to undergo appreciable calcification in the time periods studied. Analysis of the initial lag period suggested that at least two changes occurred in serum prior to the mineralization of this tissue. Inhibitors of the reaction were destroyed, and the level of dialyzable calcium was elevated owing to its release from serum protein.

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