Everted intestinal sacs as an in-vitro model for testing efficacy of new hypocholesterolemic agents

ABSTRACT

In the current study we investigated the effects of five new Acyl-CoACholesterol acyltransferase enzyme [ACAT] inhibitors on the extent of absorption of tritiated cholesterol using the everted intestinal sacs obtained from naive or cholesterol-fed rats. The extent of absorption of cholesterol was determined in the presence of the five analogs. Everted intestinal segments from male Sprague-Dawley rats filled with serosal incubation medium were added to a mucosal solution incubation medium containing 750. [micro]M [3] H-cholesterol. The compounds were added to the mucosal solution at a concentration of 15 mM. Samples were taken from serosal fluid over a period of two hours and monitored for increases in radioactivity concentrations [[3] H-cholesterol] inside the everted sac. Comparisons were made between incubations with and without the drugs in order to assess inhibitory effects of the compounds on cholesterol absorption. All calculations were based on the extent of cholesterol accumulation at the end of two hours. Absorption of cholesterol increased considerably in the cholesterol-fed rats, relative to naive rats. This effect is thought to be due to increased ACAT enzyme activity following cholesterol pretreatment. In naive rats, the addition of the different analogs [I through V] decreased the absorption of cholesterol by 58, 49, 22, 0, and 0%, respectively. Decreases of 97, 67, 58,44 and 54% were obtained for the same compounds in cholesterol-fed rats. The parent compounds were not detected in the serosal fluids using HPLC assay, indicating poor absorption of the compounds through the intestinal wall. Significant concentrations of the drugs were detected within the intestinal tissues. The everted gut technique provided a good model for assessing the efficacy of the hypolipidemic agents based on inhibition of cholesterol absorption at a single incubation concentration