CYCLIC-AMP STIMULATES EFFLUX OF INTRACELLULAR STEROL FROM CHOLESTEROL-LOADED CELLS

The interaction of high density lipoprotein with its putative receptor stimulates translocation and efflux of intracellular sterols by a pro cess involving activation of protein kinase C. This study shows that a ctivation of cAMP-dependent protein kinase also stimulates efflux of i ntracellular sterols. When intracellular sterol pools of cholesterol-l oaded cultured human skin fibroblasts and bovine aortic endothelial ce lls were radiolabeled with the biosynthetic precursor [H-3]mevalonolac tone, high density lipoprotein3 (HDL3)-mediated H-3-sterol efflux was enhanced by addition of the adenylylcyclase activator forskolin, the p hosphodiesterase inhibitors theophylline and 3-isobutyl-1-methylxanthi ne, and the cAMP analogues N-benzoyl-cAMP (N6-cAMP) and 8-thiomethyl-c AMP. The effect of N-cAMP was abolished by an inhibitor of cAMP-depend ent protein kinase (H8). The enhanced sterol efflux was independent of receptor binding of HDL3, as similar effects were observed in the pre sence of tetranitromethane-modified HDL3, which lacks receptor binding activity. N6-cAMP stimulated efflux of several subspecies of newly sy nthesized sterols, including cholesterol. Elevation of cAMP levels inc reased the proportion of radiosterols that were accessible to treatmen t of cells with the enzyme cholesterol oxidase, suggesting that activa tion of cAMP-dependent protein kinase stimulates translocation of ster ols from intracellular compartments to the plasma membrane where they desorb from the cell surface. Thus, at least two distinct protein kina se signalling pathways modulate transport of intracellular sterols in cholesterol-loaded cells.