Effects of High Dietary Levels of Cholesterol on the Metabolism of Tropoelastin and Proteolytic Enzymes in the Chick Aorta

Abstract

The synthesis of tropoelastin in chick arterial tissue was investigated using an in vitro system. Aortas from 2-week-old chicks were incubated in a synthetic medium containing l-[³H]valine and β-aminopropionitrile to inhibit cross-linking. Following incubation, the tissue was extracted with 0.1 m acetic acid, and nonradioactive tropoelastin was added to the extracts to facilitate the purification of radioactively labeled protein with the properties of tropoelastin. It was demonstrated by means of polyacrylamide gel systems employing either a cationic buffer or a neutral buffer containing 0.1% sodium dodecyl sulfate that l-[³H]valine is incorporated almost exclusively into tropoelastin. The incubation system also provided a means of measuring tropoelastin synthesis. When aortas from chicks fed diets containing 2% cholesterol were utilized for study, these aortas incorporated l-[³H]valine into tropoelastin at rates 2 to 4 times greater than that of aortas from control birds. In addition, the activities of four proteolytic enzymes in aorta were also examined in relationship to the effects of the high cholesterol diet. The levels of cathepsins A, B, and D and elastase were measured at 2, 4, 8, and 12 weeks from the day of hatching. The presence of cholesterol in the diet effected a marked increase in the levels of the four proteolytic enzymes compared with that in aorta from control chicks. The possible consequences of the observed alterations in protein synthesis and proteolytic enzymes in relationship to experimentally produced atherosclerosis are discussed.