Glutamine and Alanyl-Glutamine Increase RhoA Expression and Reduce Clostridium difficile Toxin-A-Induced Intestinal Epithelial Cell Damage

Abstract EN

Clostridium difficile is a major cause of antibiotic-associated colitis and is associated with significant morbidity and mortality.

Glutamine (Gln) is a major fuel for the intestinal cell population.

Alanyl-glutamine (Ala-Gln) is a dipeptide that is highly soluble and well tolerated.

IEC-6 cells were used in the in vitro experiments.

Cell morphology was evaluated by atomic force microscopy (AFM) and scanning electron microscopy (SEM).

Cell proliferation was assessed by WST-1 and Ki-67 and apoptosis was assessed by TUNEL.

Cytoskeleton was evaluated by immunofluorescence for RhoA and F-actin.

RhoA was quantified by immunoblotting.

TcdA induced cell shrinkage as observed by AFM, SEM, and fluorescent microscopy.

Additionally, collapse of the F-actin cytoskeleton was demonstrated by immunofluorescence.

TcdA decreased cell volume and area and increased cell height by 79%, 66.2%, and 58.9%, respectively.

Following TcdA treatment, Ala-Gln and Gln supplementation, significantly increased RhoA by 65.5% and 89.7%, respectively at 24 h.

Ala-Gln supplementation increased cell proliferation by 137.5% at 24 h and decreased cell apoptosis by 61.4% at 24 h following TcdA treatment.

In conclusion, TcdA altered intestinal cell morphology and cytoskeleton organization, decreased cell proliferation, and increased cell apoptosis.

Ala-Gln and Gln supplementation reduced intestinal epithelial cell damage and increased RhoA expression.