N-3 AND N-6 PUFAs SUPPRESS SREBP ACTIVITY AND INCREASE FLOW OF FREE CHOLESTEROL IN HEPG2 CELLS

The plasma lipid lowering effect of PUFAs, one of their main beneficial effects, is considered to be related to the regulation of lipid biosynthesis through transcription factors including sterol regulatory element binding proteins (SREBPs). In this study, we compared the effect of different PUFAs on SREBP activity in HepG2 cells, using a SRE-luciferase reporter construct as probe. Supplementation with different fatty acids reduced SREBP activity in the order 20 : 5n-3 = 18 : 2n-6 = 20 : 4n-6 >> 18 : 3n-3 = 22 : 6n-3 = 22 : 5n-6 >> 18 : 1n-9. The suppression of SREBP activity greatly depended on the degree of incorporation of the supplemented PUFA into cellular lipids, and correlated positively with the unsaturation index (r = 0·831; p<0·01) of total cell lipids. Supplemented PUFAs were also metabolized to longer and more unsaturated species. These processing activities were higher for n-3 than n-6 PUFAs (p<0·01). We studied the effect of PUFAs on the intracellular distribution of free cholesterol (FC), using filipin staining and fluorescence microscopy with or without the cholesterol traffic blocker U18666A. The data show that the incorporation of PUFAs increases FC flow from the plasma membrane to intracellular membranes. We conclude that suppression of SREBP activity by PUFAs depends on the degree of incorporation into cellular lipids, and is associated with increased flow of FC between the plasma membrane and intracellular membranes.