Live Faecalibacterium prausnitzii Does Not Enhance Epithelial Barrier Integrity in an Apical Anaerobic Co-Culture Model of the Large Intestine

Abstract

Appropriate intestinal barrier maturation during infancy largely depends on colonization with commensal bacteria. Faecalibacterium prausnitzii is an abundant obligate anaerobe that colonizes during weaning and is thought to maintain colonic health throughout life. We previously showed that F. prausnitzii induced Toll-like receptor 2 (TLR2) activation, which is linked to enhanced tight junction formation. Therefore, we hypothesized that F. prausnitzii enhances barrier integrity, an important factor in appropriate intestinal barrier maturation. In order to test metabolically active bacteria, we used a novel apical anaerobic co-culture system that allows the survival of both obligate anaerobic bacteria and oxygen-requiring intestinal epithelial cells (Caco-2). The first aim was to optimize the culture medium to enable growth and active metabolism of F. prausnitzii while maintaining the viability and barrier integrity, as measured by trans-epithelial electrical resistance (TEER), of the Caco-2 cells. This was achieved by supplementing the apical cell culture medium with bacterial culture medium. The second aim was to test the effect of F. prausnitzii on TEER across Caco-2 cell layers. Live F. prausnitzii did not improve TEER, which indicates that its benefits are not via altering tight junction integrity. The optimization of the novel dual-environment co-culturing system performed in this research will enable the investigation of new probiotics originating from indigenous beneficial bacteria.

Biotechnology of health-promoting bacteria

“…Recently, a comprehensive set of comparative studies were described for a dozen F. prausnitzii isolates showing considerable differences in oxygen sensitivity, antibiotic resistances and immunomodulatory properties (Martin et al, 2017). This is in line with another study that showed variability in improving barrier function of various F. prausnitzii strains and noted that strains A2-165 and HTF-F did not affect this parameter (Maier et al, 2017). All these findings are reminiscent of probiotic bacteria that in most cases show strain-specific effects and illustrate the heterogeneity of the F. prausnitzii group.…”

Primary human colonic mucosal barrier crosstalk with super oxygen-sensitiveFaecalibacterium prausnitziiin continuous culture

“…In vitro studies to test hypotheses about F. prausnitzii-mediated regulation of colon epithelial function or immune functions have been limited to <12-hr static culture by technical difficulties in maintaining an anaerobic apical environment supporting viable microbes in the context of an aerobic basal environment to maintain epithelial cell integrity. [22][23][24] As appreciated by these previous investigators, the influences of F. prausnitzii on cellular phenotype may arise from relatively long (>24 hr) time scale phenomena, such as accumulation of butyrate and other microbial metabolites, 25 in addition to other transcriptionally-controlled behaviors. Recent technological innovations to culture obligate anaerobes in modified Transwell formats are restricted to static cultures -where microbial nutrition become limiting -and have so far been used to culture anaerobes that are not as strictly oxygen sensitive as F. prausnitzii.…”

Epithelial-microbial diplomacy: escalating border tensions drive inflammation in inflammatory bowel disease

“…Interestingly, co-culture studies have demonstrated F. prausnitzii does not enhance barrier function but can induce activation of NF-κB signaling downstream of IL-1β in Caco-2 cells. 92,96 In contrast, Caco-2 cells treated with F. prausnitzii supernatant decreases basal and IL-1β-induced NF-κB activity. 92…”