Three-Dimensional Culture Enhances the Antimicrobial Activity of Mesenchymal Stem Cells against Shiga Toxin-Producing Escherichia Coli O157:H7 in Vitro

Abstract

Aims This study examines the in vitro antibacterial activity of palatal adipose tissue-derived mesenchymal stem cells (PAT-MSCs) and the expression of antimicrobial peptide LL-37, with a particular focus on the effect of three-dimensional (3D) nanofibrillar cellulose-based hydrogel against Shiga toxin-producing Escherichia coli (STEC) harboring stx1 and/or stx2 genes isolated from mastitis milk in Turkey.Methods and results The antibacterial activity of conventionally cultured PAT-MSCs and 3D-cultured PAT-MSCs (PAT-MSCs-3D) was evaluated against STEC isolates and Escherichia coli ATCC 35150 using quantitative colony-forming unit (CFU) assay. The expression levels of antimicrobial peptide (AMP)-encoding genes were evaluated by quantitative real-time PCR, and AMP production was further validated by immunocytochemical staining. The results indicated that PAT-MSCs-3D exhibited significantly enhanced antibacterial efficacy, resulting in marked bacterial inhibition of all tested STEC strains, with bacterial reductions reaching up to 6-7 log under specific experimental conditions. Molecular and immunocytochemical analyses demonstrated increased expression of the antimicrobial peptide LL-37 in PAT-MSCs-3D compared to 2D cultures.Conclusions Our results show that culturing PAT-MSCs in 3D conditions leads to a significant enhancement in their antimicrobial properties, which could be linked to the upregulation of LL-37.