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New and better protocols for a short-term Caco-2 cell culture system
Journal of pharmaceutical sciences, 2002-03, Vol.91 (3), p.669-679
Yamashita, Shinji
Konishi, Keisuke
Yamazaki, Yukako
Taki, Yoko
Sakane, Toshiyasu
Sezaki, Hitoshi
Furuyama, Yukihiro
2002
Volltextzugriff (PDF)
Details
Autor(en) / Beteiligte
Yamashita, Shinji
Konishi, Keisuke
Yamazaki, Yukako
Taki, Yoko
Sakane, Toshiyasu
Sezaki, Hitoshi
Furuyama, Yukihiro
Titel
New and better protocols for a short-term Caco-2 cell culture system
Ist Teil von
Journal of pharmaceutical sciences, 2002-03, Vol.91 (3), p.669-679
Ort / Verlag
New York: Elsevier Inc
Erscheinungsjahr
2002
Quelle
MEDLINE
Beschreibungen/Notizen
The aim of the present study was to develop new and better protocols for a short-term Caco-2 cell culture system for use in rapid screening of intestinal drug absorption. Caco-2 cells were cultured according to several protocols for short-term cell culture to obtain monolayers. The effects of serum (fetal bovine serum, FBS) in the culture medium and of the period of cell culture on the barrier function and transporter activities of the monolayers were examined. The barrier function was estimated both from the transepithelial electrical resistance (TEER) and the permeability of [14C]mannitol. Transporter activities were monitored by measuring the permeability of [14C]glycylsarcosine for oligopeptide transporter (PepT1) and of rhodamine 123 for P-glycoprotein (P-gp). Caco-2 monolayers obtained by 3-day culture in the BIOCOAT® HTS Caco-2 Assay System, developed by Becton Dickinson Bioscience, showed much higher permeability to hydrophilic compounds, such as mannitol, compared with those obtained by the standard 21-day culture system, due to the leaky structure of cell junctions. The newly developed 3-day protocol, which includes 10% FBS in the culture medium during the first day of culture, markedly enhanced TEER and lowered mannitol permeability of the monolayers. This protocol allowed us to better determine the rank order of permeability of compounds, giving results equivalent to those in the 21-day culture system. The longer culture period gave tighter monolayers, and the maximum value of TEER was obtained with 5 days in culture. However, after 5 days in culture, the integrity of monolayers decreased gradually. The highest activities of transporters, PepT1 and P-gp, in monolayers were obtained at days 5 or 6 of culture by the new protocol with FBS-containing medium. These results indicate that by a simple modification of the short-term culture protocol, it is possible to obtain Caco-2 monolayers with better barrier properties and higher activity of transporters that are equivalent to those found in the 21-day Caco-2 culture system. © 2002 Wiley-Liss, Inc. and the American Pharmaceutical Association J Pharm Sci 91:669–679, 2002
Sprache
Englisch
Identifikatoren
ISSN: 0022-3549
eISSN: 1520-6017
DOI: 10.1002/jps.10050
Titel-ID: cdi_proquest_miscellaneous_71568686
Format
–
Schlagworte
Algorithms
,
Animal cells
,
ATP Binding Cassette Transporter, Subfamily B, Member 1 - metabolism
,
barrier function
,
Biological and medical sciences
,
Caco-2 Cells - cytology
,
Caco-2 monolayers
,
Carrier Proteins - metabolism
,
Cell cultures. Hybridization. Fusion
,
Culture Media
,
Culture Media, Serum-Free
,
Cytological Techniques
,
drug permeability
,
Electric Conductivity
,
Fundamental and applied biological sciences. Psychology
,
General pharmacology
,
Humans
,
Hydrogen-Ion Concentration
,
Intestinal Absorption
,
Mannitol - metabolism
,
Medical sciences
,
Microscopy, Electron
,
Models, Biological
,
Molecular and cellular biology
,
Peptide Transporter 1
,
Permeability
,
Pharmacokinetics. Pharmacogenetics. Drug-receptor interactions
,
Pharmacology. Drug treatments
,
short-term cell culture
,
Symporters
,
transporter
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