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Clostridioides difficile Flagellin Activates the Intracellular NLRC4 Inflammasome
Hiba Chebly1,2, Jean-Christophe Marvaud1, Layale Safa2
1Institut Micalis, Université Paris-Saclay, INRAE, AgroParisTech, 91400 Orsay, France.
International Journal of Molecular Sciences
|October 27, 2022
Summary
Clostridioides difficile flagellin (FliC) internalizes into intestinal cells and activates the NLRC4 inflammasome pathway. This FliC-mediated NLRC4 inflammasome activation contributes to inflammation during C. difficile infection.
Area of Science:
- Immunology
- Microbiology
- Cell Biology
Background:
- Clostridioides difficile (C. difficile) is a significant cause of hospital-acquired diarrhea and colitis.
- C. difficile flagellin (FliC) is known to activate Toll-like receptor 5 (TLR5), leading to NF-kB and MAPK signaling and gut inflammation.
- Intracellular pathogens' flagella can activate the NLRC4 inflammasome, but FliC's role in this pathway for extracellular bacteria was unclear.
Purpose of the Study:
- To investigate whether flagellin from the extracellular bacterium C. difficile internalizes into epithelial cells.
- To determine if internalized C. difficile flagellin activates the NLRC4 inflammasome pathway.
- To elucidate the role of FliC internalization and NLRC4 inflammasome activation in C. difficile-associated inflammation.
Main Methods:
- Confocal microscopy to visualize the internalization of recombinant green fluorescent protein (GFP)-tagged FliC into Caco-2/TC7 intestinal cells.
- Assessing NLRC4 inflammasome activation by detecting the cleavage of pro-caspase-1 and gasdermin D (GSDMD).
- Analyzing the expression of inflammasome-related cytokines (IL-1β, IL-18, IL-33) and the effect of NF-kB pathway inhibition (IKK-α) on inflammasome activation.
Main Results:
- Internalization of full-length GFP-FliC into Caco-2/TC7 cells was confirmed via confocal microscopy.
- Full-length FliC, but not truncated FliC, activated the NLRC4 inflammasome, evidenced by pro-caspase-1 and GSDMD cleavage.
- FliC stimulation led to increased expression of IL-1β, IL-18, and IL-33, with NF-kB pathway inhibition reducing inflammasome-related gene expression.
Conclusions:
- C. difficile flagellin (FliC) internalizes into intestinal epithelial cells.
- Internalized FliC activates the intracellular NLRC4 inflammasome pathway, involving caspase-1 and GSDMD.
- This FliC-mediated NLRC4 inflammasome activation contributes to the inflammatory response in C. difficile infections.
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