ROLE OF BILE SALT-DEPENDENT CHOLESTERYL ESTER HYDROLASE IN THE UPTAKEOF MICELLAR CHOLESTEROL BY INTESTINAL-CELLS

The bile salt-dependent cholesteryl ester hydrolase (CEH; EC 3.1.1.13) has been proposed to promote the intestinal absorption of both the fr ee and esterified (FC, CE) forms of dietary cholesterol. For example, it was recently reported that in the human intestinal cell line CaCo2, addition of bovine CEH to the medium increased the uptake and intrace llular esterification of micellar FC supplied at subphysiological conc entrations [Lopez-Candales et al. (1993) Biochemistry 32, 12085-12089] . To test the ability of CEH to promote micellar cholesterol uptake in a CaCo2 system under more physiological conditions, an in vitro model was developed. Cells stably expressing rat CEH were created by DNA tr ansfection (Tr cells), and the uptake of micellar FC and its intracell ular esterification were measured using isotopic methods in Tr and con trol cells. Experimental parameters that were varied included micellar composition (monoolein or egg PC; FC, CE, or both), the final concent ration of micellar cholesterol (1 nM to 50 mu M), the origin of CEH (e ndogenously synthesized vs exogenously added), and the species source of enzyme (rat, pig, man). The uptake of cholesterol that was derived from micellar CE was significantly increased 5-10-fold (p < 0.001) in Tr vs control cells as a result of the hydrolysis of the CE by the CEH and subsequent uptake of the liberated free cholesterol. In contrast, the uptake of micellar FC was not increased by the presence of CEH, w hether it was endogenous or exogenous. In addition, based on TLC analy sis of extracted cellular lipids, there was no evidence that CEH promo ted the esterification of the FC that was taken up. These results were independent of cholesterol concentration and the non-sterol compositi on of the micelles, Although in the presence and absence of CEH there was comparable uptake of cholesterol by cells after a 4 h incubation w ith a particular type of micelle, micelles containing egg PC were not as effective FC donors as those containing monoolein. Overall, the dat a support a role of CEH in modulating the absorption of CE present in the intestinal lumen by a mechanism involving the hydrolysis of CE by CEH, thereby increasing the FC concentration gradient between the mice llar and plasma membrane pools and enhancing the passive cellular upta ke of free cholesterol.