ISSN:
1573-904X
Keywords:
Caco-2
;
in vitro intestinal epithelial model
;
confocal laser scanning microscopy
;
tight junctions
;
cytoarchitecture
;
variability
Source:
Springer Online Journal Archives 1860-2000
Topics:
Chemistry and Pharmacology
Notes:
Abstract Purpose. To introduce confocal laser scanning microscopy (CLSM)combined with digital image restoration to characterise Caco-2 cellsunder different culture conditions, and thus to define additional validcriteria for the optimisation of culture models. Methods. Growth curves were established and transepithelial electricalresistance (TEER) measured for cells grown in EMEM or DMEMmedium on Cyclopore™ membranes. Cytoskeleton, cell nuclei and tightjunctions (TJ) were investigated by CLSM. Results. Cultures reached a plateau of ∼4.5 × 105 cells/cm2 after∼ 10 days. At the same time TEER reached 750 Ω cm2. An irregular,fairly complete network of TJ was present at confluence (∼2 d).Between 15 and 30 days a regular TJ network was established. Cellsformed mixed mono- and multilayers under most conditions with twoexceptions: flat monolayers were observed on polycarbonate filterswith EMEM and with the Biocoat™ intestinal epithelium differentiationenvironment system. In multilayers TJ were found in the upper aswell as in the lower cell layers although the regular vertical polaritywas disturbed. Conclusions. CLSM represents an important tool to investigate thecytoarchitecture of Caco-2 cells. 3D-analysis of confocal data givesimportant clues on the characteristics of cell layers and thus helps tovalidate optimisation strategies.
Type of Medium:
Electronic Resource
URL:
http://dx.doi.org/10.1023/A:1007585105753
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