Influence of phytostanol phosphoryl ascorbate, FM-VP4, on pancreatic lipase activity and cholesterol accumulation within Caco-2 cells.
OBJECTIVE
The objective of this study was to determine how a novel hydrophilic phytostanol (FM-VP4) affects the cellular accumulation of [3H]cholesterol in human colon carcinoma (Caco-2) cell monolayers grown in Transwell chambers.


METHODS
To determine cellular accumulation of cholesterol and FM-VP4, [3H]cholesterol- containing micelles (50 microM cholesterol containing 1.27x10 (-4)% [3H]cholesterol) or [3H]FM-VP4 (50 microM) was incubated on the apical side of differentiated Caco-2 cell monolayers for 1 to 4 h at 37 degrees C in the absence or presence of increasing concentrations (10-200 microM) of unlabeled FM-VP4 or cholesterol, respectively.


RESULTS
The accumulation of [3H]cholesterol (presented in micelles) into Caco-2 cell monolayers in the presence of 50 microM FM-VP4 was significantly lower (33.7 +/- 7.0%) compared to control (59.8 +/- 5.2%, p<0.05) following 4 h of incubation. Conversely, cholesterol inhibited the accumulation of [3H]FM-VP4, although to a lesser extent, suggesting competition for binding sites. The inhibitory effects of FM-VP4 and cholesterol on each other were detectable after 1 h of incubation and increased with time. The extent of FM-VP4 inhibition of [3H]cholesterol accumulation was consistent whether FM-VP4 was co-incorporated into micelles or added separately in solution, suggesting that FM-VP4 does not elicit its effects through inhibition of cholesterol incorporation into micelles. In addition, pancreatic lipase activity ([3H]triolein hydrolysis) and p-glycoprotein (rhodamine 123 fluorescence) activity, were not affected by FM-VP4.


CONCLUSIONS
In conclusion, FM-VP4 rapidly inhibits cholesterol accumulation within Caco-2 cell monolayers in a mode independent of pancreatic lipase activity, p-glycoprotein activity or cholesterol incorporation in micelles.
Ramaswamy
Manisha
M
Division of Pharmaceutics and Biopharmaceutics, Faculty of Pharmaceutical Sciences, University of British Columbia, B.C, Canada.
Yau
Edwin
E
Wasan
Kishor M
KM
Boulanger
Kathy D
KD
Li
Ming
M
Pritchard
P Haydn
PH
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000378:metabolism; D018528:ATP-Binding Cassette Transporters / Q000378:metabolism; D000924:Anticholesteremic Agents / Q000737:chemistry / Q000494:pharmacology*; D001205:Ascorbic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D001667:Binding, Competitive; D018938:Caco-2 Cells / Q000201:enzymology* / Q000378:metabolism; D002784:Cholesterol / Q000378:metabolism*; D016166:Free Radical Scavengers / Q000494:pharmacology; D006801:Humans; D008049:Lipase / Q000378:metabolism*; D008823:Micelles; D009943:Organophosphorus Compounds / Q000494:pharmacology; D010179:Pancreas / Q000187:drug effects* / Q000201:enzymology*; D010840:Phytosterols / Q000737:chemistry / Q000494:pharmacology*; D014316:Tritium / Q000378:metabolism
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D018528:ATP-Binding Cassette Transporters; D000924:Anticholesteremic Agents; C445524:FM-VP4 compound; D016166:Free Radical Scavengers; D008823:Micelles; D009943:Organophosphorus Compounds; D010840:Phytosterols; D014316:Tritium; D002784:Cholesterol; D008049:Lipase; D001205:Ascorbic Acid