Biological labeling of very low density lipoproteins with cholesteryl linoleyl ether and its fate in the intact rat

Abstract

In vitro labeling of very low density lipoproteins (VLDL) with radioactive cholesteryl linoleyl ether, an analog of cholesteryl linoleate, was studied. The protocol which gave the highest efficiency and seemed least injurious to the final product included: (1) sonication of the labeled cholesteryl ether with partially delipidated high density lipoproteins (HDL); (2) transfer of the labeled lipids to VLDL in the presence of lipoprotein-deficient human serum; (3) reisolation of the VLDL by ultracentrifugation. Under optimal conditions 70% of the added labeled lipid was recovered with HDL and 60% were transferred from HDL to VLDL. The labeled cholesteryl linoleyl ether was shown to comigrate with the protein of VLDL on agarose gel electrophoresis. In negatively stained preparations, the labeled VLDl and its unlabeled counterpart had similar appearance. The in vitro labeled VLDL was injected into rats and was cleared from the circulation with a t1/2 comparable to endogenously labeled VLDL. More than 80% of the injected dose was recovered in the liver between 3 and 48 h after injection of VLDL labeled with [3H]cholesteryl linoleyl ether of which 91-97% were in the ether form. On radioautography of fixed frozen sections of liver the bulk of the radioautographic reaction was associated with the cytoplasm of hepatocytes. When the VLDL had been labeled also with [14C]cholesteryl linoleate only 35% of injected dose was present in the liver at 3 h, of which 87% was in unesterified form. The distribution of the labeled cholesteryl linoleyl ether, 3-48 h after injection, expressed as per cent of injected dose per organ was 0.7-1.5 in spleen, 0.2-0.5 in lung, 0.1 in heart and 0.2-0.4 in adrenal. The main advantage of the presently described approach in which a nondegradable analog of cholesteryl ester was introduced into VLDL by a biological procedure is the possibility to study the role of various organs to take up circulating cholesteryl ester, especially in species in which LDL is produced from VLDL.