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Gram-negative Bacterial Protein Secretion Systems01:17

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Gram-negative bacteria utilize sophisticated protein secretion systems to transport proteins across their double-membrane envelope into the extracellular environment or host cells. Based on their mechanism of action, these systems are classified into one-step and two-step pathways.One-Step Secretion Systems (Types I, III, IV, and VI)One-step secretion systems bypass the periplasm entirely, forming a continuous channel that spans both the inner and outer membranes:Type I Secretion System (T1SS):...
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Detection of Toxin Translocation into the Host Cytosol by Surface Plasmon Resonance
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Shiga Toxin (Stx) Type 1a and Stx2a Translocate through a Three-Layer Intestinal Model.

Rebecca A Bova1,2,3, Andrew C Lamont2,3, Theodore J Picou2,3

  • 1Department of Microbiology and Immunology, Uniformed Services University, Bethesda, MD 20814, USA.

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|March 28, 2023
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Summary

A more complex, three-layer cell model better simulates Shiga toxin (Stx) translocation across the intestinal barrier than a single-layer model. This enhanced model is crucial for studying Stx-producing E. coli infections and developing inhibitors.

Keywords:
Escherichia coliShiga toxintissue modeltranslocation

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Area of Science:

  • Microbiology
  • Toxicology
  • Cell Biology

Background:

  • Shiga toxins (Stxs) from E. coli cause hemolytic uremic syndrome by crossing the intestinal barrier.
  • The precise mechanisms of Stx translocation into the bloodstream remain unclear.

Purpose of the Study:

  • To compare Stx translocation across single-layer and three-layer polarized cell models.
  • To evaluate the impact of model complexity on quantifying Stx movement.

Main Methods:

  • Utilized primary colonic epithelial cells (single-layer) and a co-culture model (epithelial cells, myofibroblasts, endothelial cells).
  • Measured Stx1a and Stx2a translocation by assessing Vero cell toxicity in apical and basolateral media.
  • Infected models with Shiga toxin-producing E. coli (STEC) strains to assess barrier function and toxin movement.

Main Results:

  • Stx1a and Stx2a translocated across both models, with significantly higher amounts (approx. 10-fold) in the three-layer model.
  • Stx2a translocated more efficiently (3-4 fold) than Stx1a in both models.
  • STEC O157:H7 reduced barrier function; STEC O26:H11 translocated Stx without compromising barrier integrity.

Conclusions:

  • Single-cell models may underestimate Stx translocation.
  • The biomimetic three-layer model provides a more accurate assessment of Stx translocation.
  • The three-layer model is suitable for evaluating Stx translocation inhibitors.