Tumor necrosis factor alpha increases epithelial barrier permeability by disrupting tight junctions in Caco-2 cells.
The objectives of this study were to determine the effect of tumor necrosis factor alpha (TNF-alpha) on intestinal epithelial cell permeability and the expression of tight junction proteins. Caco-2 cells were plated onto Transwell microporous filters and treated with TNF-alpha (10 or 100 ng/mL) for 0, 4, 8, 16, or 24 h. The transepithelial electrical resistance and the mucosal-to-serosal flux rates of the established paracellular marker Lucifer yellow were measured in filter-grown monolayers of Caco-2 intestinal cells. The localization and expression of the tight junction protein occludin were detected by immunofluorescence and Western blot analysis, respectively. SYBR-Green-based real-time PCR was used to measure the expression of occludin mRNA. TNF-alpha treatment produced concentration- and time-dependent decreases in Caco-2 transepithelial resistance and increases in transepithelial permeability to the paracellular marker Lucifer yellow. Western blot results indicated that TNF-alpha decreased the expression of phosphorylated occludin in detergent-insoluble fractions but did not affect the expression of non-phosphorylated occludin protein. Real-time RT-PCR data showed that TNF-alpha did not affect the expression of occludin mRNA. Taken together, our data demonstrate that TNF-alpha increases Caco-2 monolayer permeability, decreases occludin protein expression and disturbs intercellular junctions.
Cui
W
W
Department of Infectious Diseases, the First Affiliated Hospital, China Medical University, Shenyang, Liaoning, China.
Li
L X
LX
Sun
C M
CM
Wen
Y
Y
Zhou
Y
Y
Dong
Y L
YL
Liu
P
P
D015153:Blotting, Western; D018938:Caco-2 Cells; D002463:Cell Membrane Permeability / Q000187:drug effects*; D004847:Epithelial Cells / Q000187:drug effects* / Q000378:metabolism; D006801:Humans; D007413:Intestinal Mucosa / Q000166:cytology*; D008565:Membrane Proteins / Q000187:drug effects* / Q000378:metabolism; D062793:Occludin; D016133:Polymerase Chain Reaction / Q000379:methods; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D019108:Tight Junctions / Q000187:drug effects* / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*
D008565:Membrane Proteins; C568985:OCLN protein, human; D062793:Occludin; D012333:RNA, Messenger; D014409:Tumor Necrosis Factor-alpha