Elastin was isolated from human aortic tissues by extraction with formic acid. The aortas came from individuals of both sexes with ages ranging from 18 to 67 years. The elastin was treated to remove pre-existing nuclei for hydroxyapatite and mineralizedin vitro by incubation in a solution metastable to hydroxyapatite. The results showed an increased rate of mineralizationin vitro as the age of the individual from whom the aorta was obtained increased. The lag period in mineral formation seen with elastin samples obtained from individuals below the age 40, disappeared with older elastin. Amino-acid analysis of the elastin samples confirmed earlier results by indicating an increase in the more polar amino acids in the older elastin samples. Exposure of the extracted elastin samples to several proteolytic enzymes including elastase confirmed the identity of the material as elastin.

Although invertebrate cartilage tissues do not mineralize in nature, it is now reported for the first time that when excised gill cartilage tissue fromLimulus (horse shoe crab) is placed in an appropriate incubation medium metastable to hydroxyapatite, mineralization will occur. The mineralization is temperature dependent, and takes place at 37° but not at 20°. Incubations in media metastable to calcite have not produced mineralization. Histologic examination of mineralized tissues showed mineral deposits predominantly within cells, and to a lesser extent in the matrix. X-ray diffraction of the deposited mineral revealed a typical biological hydroxyapatite pattern.

Twenty-four hour plaque samples from humans were tested for mineralizing activity. Dental calculus samples also were taken from the same individuals and data for each individual were correlated. Results showed a positive correlation between the mineralizing activity of a 24-hour plaque sample and the amount of dental calculus formed in the same individual's mouth.

WE have previously described the in vitro calcification of human aortic tissue with the formation of hydroxy-apatite1. We now wish to report the results of an investigation of the mechanism of this calcification.

CALCIFICATION in vitro of excised rat and rabbit aorta has been described previously1–5, but no comparable studies have been reported for the in vitro calcification of human aortic tissue. This communication reports the in vitro calcification of human aortic tissue using a procedure which measures the simultaneous deposition of calcium-45 and phosphorus-32. The mineral formed in the tissue was identified by X-ray diffraction analysis as hydroxy-apatite.