Evidence for an intracellular mechanism contributing to cholesterol lowering activity of phytosterols

Background:

Phytosterols are enriched in functional food products to lower plasma LDL cholesterol by reducing fractional intestinal cholesterol absorption. Beside micellar intraluminal effects, intracellular mechanisms may be operative but direct evidence is still lacking. The ABC sterol transporters ABCG5/8 involved in phytosterol re-excretion and ABCA1 involved in cellular cholesterol export are expressed in the human enterocyte and carry a Liver X Receptor (LXR) response element. Ligands for LXR are side chain hydroxylated cholesterol metabolites. We hypothesized that phytosterols might in analogy to cholesterol undergo metabolization to oxy-phytosterols that act as LXR agonists.

Methods:

Caco-2 cells served as human enterocyte model and were incubated with 50µM 3H-labelled campesterol, sitosterol or cholesterol. Cellular uptake of the sterols was enhanced by 2-hydroxypropyl-beta-cyclodextrin (beta-CD). After incubation media and cells were extracted. The extracts were analysed by TLC with 3H-scanning and structural identification was done by GC/MS.

Results:

Incubation of enterocytes with 50µM cholesterol resulted in secretion of 27-hydroxycholesterol (27-OH-Chol) and cholestenoic acid (27-COOH-Chol) into media. 27-OH-Chol was also detected intracellularely. Sterol mass uptake mediated by beta-CD was similar for cholesterol and campesterol, but lower for sitosterol. After incubation with 50µM sitosterol no conversion to oxidation products was detected. In contrast, campesterol was metabolized to 27-hydroxycampesterol, although less efficiently than cholesterol.

Conclusion:

The human enterocyte is capable of converting cholesterol into 27-hydroxycholesterol and cholestenoic acid, both recognized as endogenous ligands for the transcription factor LXR. In this human enterocyte model sitosterol was not detectably metabolized, even in a cholesterol-depleted scenario and cyclodextrin-enhanced delivery. However, campesterol which is structurally closer related to cholesterol is converted into 27-hydroxycampesterol. Natural mixtures of phytosterols do not act in the enterocyte via conversion of sitosterol but campesterol may be metabolized into a possible LXR agonist.