The aim of this study was to characterize an in vitro modulating effect of three commensal Lactobacillus strains on cellular differentiation of non-transformed crypt-like rat small intestinal cell line IEC-18. IEC-18 was grown on extracellular matrix, with or without presence of Lactobacillus strains. Gene expression of IEC-18 bacterial detection system - such as Toll-like receptors TLR-2, TLR-4, signal adapter MyD88, cytoplasmic NOD2 receptor, inflammatory cytokines IL-18, IL-1beta, chemokine IL-8 and enzyme caspase-1 - was evaluated using real-time PCR. Expression and localization of TLR-2, TLR-4, IL-18 and caspase-1 proteins was demonstrated by Western blotting and immunofluorescent staining. Secretion of IL-18 to apical and basolateral surfaces was assayed by ELISA. Our results suggested that L. casei LOCK0919 accelerated differentiation of IEC-18 by stimulating TLR-2, TLR-4, MyD88, IL-18, caspase-1 mRNAs and proteins. L. casei LOCK0919 increased expression and transfer of villin and beta-catenin from cytoplasm to cell membrane. Presence of L. rhamnosus LOCK0900 resulted in detachment of IEC-18 layer from extracellular matrix leading to induction of IL-1beta, of TLR-2 and IL-8 mRNAs and stimulation of MyD88, caspase-1 and cytosolic receptor NOD2 mRNAs. L. rhamnosus LOCK0908 was not recognized by TLR-2 or TLR-4 receptors. Lactobacilli-IEC-18 crosstalk enhanced immune and barrier mucosal functions.
- MeSH
- beta-katenin biosyntéza MeSH
- buněčná diferenciace účinky léků MeSH
- cytokiny biosyntéza MeSH
- epitelové buňky účinky léků MeSH
- interleukin-18 biosyntéza MeSH
- kaspasa 1 biosyntéza MeSH
- krysa rodu rattus MeSH
- Lacticaseibacillus rhamnosus * MeSH
- Lactobacillus casei * MeSH
- messenger RNA biosyntéza MeSH
- mikrofilamentové proteiny biosyntéza MeSH
- probiotika farmakologie MeSH
- regulace genové exprese účinky léků MeSH
- střevní sliznice cytologie účinky léků MeSH
- subcelulární frakce metabolismus MeSH
- toll-like receptory biosyntéza účinky léků MeSH
- zvířata MeSH
- Check Tag
- krysa rodu rattus MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
The multifunctional cytokine interleukin-18 (IL-18) is an important mediator in intestinal inflammatory processes. The aim of this study was to evaluate the constitutive expression of IL-18 and its receptors (IL-18Ralpha and IL-18Rbeta) in intestinal epithelial cells (IEC) stimulated by tumor necrosis factor-alpha (TNF-alpha) and interferon-gamma (IFN-gamma). In addition, cellular proliferation and evaluation of brush border enzymes as differentiation markers were studied. Nontransformed rat intestinal epithelial IEC-6 cells were grown on an extracellular matrix (ECM) in medium with or without TNF-alpha, IFN-gamma, or a combination of both. Gene expression of IL-18, its receptors and apoptotic markers was evaluated using real-time PCR. Expression of IL-18Ralpha protein was demonstrated by flow cytometry and Western blot. Enzymatic activities of brush border enzymes and caspase-1 were determined. The constitutive expression of IL-18, IL-18Ralpha and IL-18Rbeta mRNAs and proteins were detected in IEC-6 cells. The biologically active form of IL-18 was released in response to TNF-alpha and IFN-gamma treatment. Exogenous IL-18 had no effect on cellular proliferation, brush border enzyme activities, and gene expression of apoptotic markers. However, the addition of IL-18 stimulated production and release of the chemokine IL-8. These data suggest that IEC-6 cells may be not only a source of IL-18 but also a target for its action.
- MeSH
- apoptóza účinky léků MeSH
- biologické markery metabolismus MeSH
- buněčná diferenciace účinky léků MeSH
- buněčná membrána metabolismus účinky léků MeSH
- buněčné linie MeSH
- chemokin CCL2 metabolismus MeSH
- epitelové buňky MeSH
- financování organizované MeSH
- interferon gama farmakologie MeSH
- interleukin-18 genetika sekrece MeSH
- kaspasa 1 metabolismus MeSH
- krysa rodu rattus MeSH
- kultivační média MeSH
- messenger RNA genetika metabolismus MeSH
- počet buněk MeSH
- polymerázová řetězová reakce s reverzní transkripcí MeSH
- receptor interleukinu-18 - alfa-podjednotka genetika metabolismus MeSH
- receptor interleukinu-18 - beta-podjednotka genetika metabolismus MeSH
- regulace genové exprese účinky léků MeSH
- střeva cytologie MeSH
- TNF-alfa farmakologie MeSH
- zvířata MeSH
- Check Tag
- krysa rodu rattus MeSH
- zvířata MeSH
