Shed XXXX enterocytes as a tool for the study of expression and function of intestinal drug-metabolizing enzymes and transporters.

@entity1 @entity39 @entity5322 @entity1855 @entity4433 @entity1195 @entity460
OBJECTIVES: Intestinal metabolism and transport are now recognized as protective barriers against orally ingested xenobiotics, including drugs. However, in vitro studies of the expression and function of intestinal proteins are hampered by the limited availability of @entity1 intestinal tissues. Because enterocytes are constantly shed in large numbers into the gut lumen, this study investigated whether these cells could be collected with a multilumen perfusion catheter and whether they are functionally active. METHODS: In healthy volunteers, a 20-cm isolated jejunal segment was generated with the perfusion catheter by inflating 2 balloons with air. Shed cells were characterized by fluorescence-activated cell sorting analysis for leukocyte-specific @entity1195 and enterocyte-specific villin, as well as for apoptosis. Homogenates of the cells were used for reverse transcriptase polymerase chain reaction and Western blotting. Cytochrome P450 enzyme activity was determined with the @entity460 channel blocker @entity1855 as a substrate. RESULTS: On average, 4.83 mg protein and 56.23 million cells were collected from a 20-cm segment during 2 hours. A total of 84.2% of the cells were positive for enterocyte-specific villin, and only 1.6% of the collected cells were positive for @entity1195 . The majority of cells (65.3%) were not in early or late apoptosis or @entity39 . In all volunteers, drug-metabolizing enzymes (such as members of the cytochrome P450 family) could be detected as both messenger ribonucleic acid and proteins. Consistent with expression data, formation of @entity1855 metabolites catalyzed by @entity5322 and @entity4433 was shown. CONCLUSIONS: The majority of shed @entity1 enterocytes collected with a multilumen perfusion catheter were still functionally active and not apoptotic. Harvesting of spontaneously shed enterocytes provides a new tool for studies on expression and function of intestinal proteins.
