cholesteryl-linoleyl-ether and dilactitol-tyramine

Earlier studies have shown that rat granulosa cells grown in serum-free medium are exquisitely responsive to exogenously provided lipoprotein cholesterol. In this study we compare the amount of cholesterol (cholesteryl ester) actually delivered from various homologous and heterologous cholesterol-rich lipoproteins and examine the intracellular pathways used in the delivery system. Granulosa cells were incubated for 5 or 24 h with 125I-labeled human (h) HDL3, rat (r) HDL or hLDL equipped with non-releasable apoprotein and cholesteryl ether tags which accumulate within cells, even after degradation. We show that all the tested lipoproteins were similarly efficient in cholesteryl ester delivery; i.e., based on cholesterol: protein ratios of the starting ligands, each delivered approximately the same cholesteryl ester mass and evoked a similar progestin response. However, each lipoprotein was processed quite differently by the granulosa cells: hHDL3-cholesteryl ester was taken up almost exclusively by an non-endocytic pathway, hLDL-cholesteryl ester almost exclusively by an endocytic pathway and rHDL-cholesteryl ester by both pathways. In general, there was no correlation between the total amount of lipoprotein bound or apoprotein internalized and/or degraded by the cells with the amount of cholesteryl ester received or the level of the progestin response. Hormone stimulation upregulated the preferred pathway for each lipoprotein.

Topics: Animals; Bucladesine; Cells, Cultured; Cholesterol; Cholesterol Esters; Endocytosis; Female; Granulosa Cells; Iodine Radioisotopes; Lipoproteins; Lipoproteins, HDL; Lipoproteins, HDL3; Lipoproteins, LDL; Microscopy, Electron; Rats; Rats, Inbred Strains; Tyramine

This study describes high density lipoprotein (HDL) uptake in the rat adrenal using a newly developed nonrecycling perfusion technique to control both the quality and quantity of the supplied lipoprotein. The aim of the study was to quantify a nonendocytic (alternative) pathway in the delivery of HDL-cholesterol. All experiments were conducted using an acute lipoprotein-deficient rat model (24 h 4-aminopyrazolo-[3, 4-d]-pyrimidine, 4-APP) in which circulating levels of cholesterol were reduced by one half, but various adrenal gland measurements of cholesterol metabolism were unchanged. Both rat HDL (rHDL) and affinity-purified human HDL3 (hHDL3) were used throughout the study. Microscopic autoradiographs (ARGs) indicate that both ligands bind avidly and exclusively to cells of the adrenal fasciculata and reticularis zones. Despite differences in binding affinity, both ligands deliver approximately the same total cholesterol to the cell interior as estimated by double-labeled residualizing tags on HDL (i.e., 125I-labeled dilactitol tyramine-[3H]cholesteryl linoleyl ether (DTT-CLE) HDL). The internalized cholesterol can account for much of the corticosterone produced during the 90-min time frame; however, only a small fraction of this cholesterol could have been provided via the endocytic pathway. Data obtained with the use of 125I-labeled DTT-[3H]CLE-HDL show that only 8.0% (or 0.7%) of corticosterone produced with rHDL (or hHDL3) could have come from cholesterol internalized as a component of intact HDL (i.e., via the endocytic pathway). These calculations strengthen the electron microscopy autoradiographic data that show that few exposed silver grains (representing the localization of the 125I-isotope) are found within the cell cytoplasm. Thus, despite differences in the uptake characteristics of the two ligands, most of the HDL-cholesterol internalized and used for corticosterone production during adrenal perfusion apparently comes from a pathway in which intact HDL are not internalized.

Topics: Adenine; Adrenal Glands; Animals; Autoradiography; Cell Membrane; Cholesterol; Cholesterol, HDL; Corticosterone; Humans; Male; Microscopy, Electron; Perfusion; Rats; Rats, Inbred Strains; Tissue Distribution; Tyramine