Efficacy of Selected Live Biotherapeutic Candidates to Inhibit the Interaction of an Adhesive-Invasive Escherichia coli Strain with Caco-2, HT29-MTX Cells and Their Co-Culture

Abstract

Adherent-invasive Escherichia coli (AIEC) has been implicated as a microbiological factor in the pathogenesis of inflammatory bowel disease (IBD). We evaluated the ability of six live biotherapeutic products (LBPs) to inhibit the interaction of an AIEC strain to three cell lines representing human gut epithelium. Co-inoculation of LBPs with AIEC showed a reduction in adhesion (up to 73%) and invasion of AIEC (up to 89%). Pre-inoculation of LBPs in HT-29-MTX and Caco-2 cells before challenging with AIEC further reduced the adhesion and invasion of the AIEC, with three LBPs showing significantly (p < 0.0001) higher efficiency in reducing the adhesion of AIEC. In co-inoculation experiments, the highest reduction in adhesion (73%) of AIEC was observed in HT-29-MTX cells, whereas the highest reduction in invasion (89%) was seen in HT-29-MTX and the co-culture of cells. Pre-inoculation of LBPs further reduced the invasion of AIEC with highest reduction (97%) observed in co-culture of cells. Our results indicated that whilst there were differences in the efficacy of LBPs, they all reduced interaction of AIEC with cell lines representing gut epithelium. Their efficiency was higher when they were pre-inoculated onto the cells, suggesting their potential as candidates for alleviating pathogenesis of AIEC in patients with IBD.

The Efficacy of Selected Probiotic Strains and Their Combination to Inhibit the Interaction of Adherent-Invasive Escherichia coli (AIEC) with a Co-Culture of Caco-2:HT29-MTX Cells

“…This strain has also been shown to be very effective in inhibiting adhesion of other enteric pathogens to cell lines representing the intestinal epithelium [ 53 ]. However, we have previously shown that there was no correlation between the F44A-1 AIEC adhesion and invasion of the co-culture cell model, and that this also did not correlate with the adhesion capabilities of live biotherapeutics products tested in that study [ 54 ]. For translocation assays we used E. coli strain HMLN-1, a highly efficient translocating strain as shown before [ 56 , 57 , 71 ], as a positive control.…”

“…It is generally accepted that probiotics are beneficial bacteria, competing with pathogens for binding sites and resources, essentially inhibiting them from colonising the gut epithelium [ 17 , 20 ]. Previous studies have shown probiotic strains as both a valuable prevention strategy and a therapeutic treatment option, reducing bacterial interaction with the gut epithelium [ 54 , 61 , 62 ]. Whilst many studies use a single cell line such as Caco-2 or HT-29 to assess the efficacy of probiotic strains in vitro, the lack of mucin production in these cell lines renders them suboptimal as a representation of the cell model of the GI tract when measuring bacterial colonisation [ 63 ].…”

“…A wild type E. coli strain (F44A-1) initially isolated from a patient with colorectal cancer was used as a challenging strain. This strain was characterised for the presence of all virulence genes involved in the pathogenesis of AIEC strains, specifically their diffuse adhesion pattern to Caco-2 cells, as well as their intramacrophage survival and replication [ 52 , 54 ]. As a negative control for adhesion and invasion assays, we used the K12 E. coli strain 46-4 [ 55 ], and for translocation experiments, we used a highly translocating E. coli strain HMLN-1, as a positive control and a non-translocating E. coli strain JM109 [ 55 , 56 , 57 ].…”

“…The adhesion of AIEC in the presence and absence of four probiotics and their combination was tested in 8-well chamber slides (Nunc Lab-Tek II, Thermo Fisher Scientific, Waltham, MA, USA) using the method adapted from Smit et al [ 54 ]. Co-culture cells were grown to >70% confluence, washed three times with sterile PBS and the media replaced with 200 μL of antibiotic-free EMEM.…”

Cellular Uptake and Transport Mechanism Investigations of PEGylated Niosomes for Improving the Oral Delivery of Thymopentin

“…It can be seen that Caco-2 and HT-29 cells with ratios of 1:0, 7:1, and 9:1 met the requirement of tissue integrity. Because of the benefits of the co-culture of Caco-2 and HT-29 cells, it provides the presence of both absorptive and goblet cells, both of which have different culture requirements for optimal growth and function to mimic intestine tissue in vivo [ 38 , 39 , 40 , 41 , 42 ]. Therefore, the ratio of 9:1 was selected based on the presence of both cell types and highest TEER value for subsequent uptake and transport studies.…”