Alterations in the gut microbiota composition play a crucial role in the pathogenesis of inflammatory bowel disease (IBD) as specific commensal bacterial species are underrepresented in the microbiota of IBD patients. In this study, we examined the therapeutic potential of three commensal bacterial species, Faecalibacterium prausnitzii (F. prausnitzii), Roseburia intestinalis (R. intestinalis) and Bacteroides faecis (B. faecis) in an in vitro model of intestinal inflammation, by using differentiated Caco-2 and HT29-MTX cells, stimulated with a pro-inflammatory cocktail consisting of interleukin-1β (IL-1β), tumor necrosis factor-α (TNFα), interferon-γ (IFNγ), and lipopolysaccharide (LPS). Results obtained in this work demonstrated that all three bacterial species are able to recover the impairment of the epithelial barrier function induced by the inflammatory stimulus, as determined by an amelioration of the transepithelial electrical resistance (TEER) and the paracellular permeability of the cell monolayer. Moreover, inflammatory stimulus increased claudin-2 expression and decreased occludin expression were improved in the cells treated with commensal bacteria. Furthermore, the commensals were able to counteract the increased release of interleukin-8 (IL-8) and monocyte chemoattractant protein-1 (MCP-1) induced by the inflammatory stimulus. These findings indicated that F. prausnitzii, R. intestinalis and B. faecis improve the epithelial barrier integrity and limit inflammatory responses.

Natural smectites have demonstrated efficacy in the treatment of diarrhea. The present study evaluated the prophylactic effect of a diosmectite (FI5pp) on the clinical course, colon damage, expression of tight junction (TJ) proteins and the composition of the gut microbiota in dextran sulfate sodium (DSS) colitis. Diosmectite was administered daily to Balb/c mice from day 1 to 7 by oral gavage, followed by induction of acute DSS-colitis from day 8 to 14 (“Control”, n = 6; “DSS”, n = 10; “FI5pp + DSS”, n = 11). Mice were sacrificed on day 21. Clinical symptoms (body weight, stool consistency and occult blood) were checked daily after colitis induction. Colon tissue was collected for histological damage scoring and quantification of tight junction protein expression. Stool samples were collected for microbiome analysis. Our study revealed prophylactic diosmectite treatment attenuated the severity of DSS colitis, which was apparent by significantly reduced weight loss (p = 0.022 vs. DSS), disease activity index (p = 0.0025 vs. DSS) and histological damage score (p = 0.023 vs. DSS). No significant effects were obtained for the expression of TJ proteins (claudin-2 and claudin-3) after diosmectite treatment. Characterization of the microbial composition by 16S amplicon NGS showed that diosmectite treatment modified the DSS-associated dysbiosis. Thus, diosmectites are promising candidates for therapeutic approaches to target intestinal inflammation and to identify possible underlying mechanisms of diosmectites in further studies.

Streptococcus pneumoniae remains a major global health threat and is listed by the by the World Health Organization as a pathogen in urgent need of new antimicrobial strategies. While primarily considered as an extracellular pathogen, S. pneumoniae can persist within splenic macrophages in severe disease, creating a protected intracellular niche that may contribute to fulminant sepsis. We recently demonstrated the concept of an mRNA-based therapeutic approach in which host cells produce the pneumococcal bacteriophage endolysin Cpl-1. Here, we investigated whether expression of Cpl-1 in macrophages can target S. pneumoniae residing within host cells. Using the human THP-1 macrophage line, we demonstrated successful translation and intracellular accumulation of bioactive Cpl-1 following IVT-mRNA transfection. Lysates from Cpl-1 mRNA-transfected cells exhibited bacteriolytic activity, and Western blotting as well as immunofluorescent staining confirmed cytosolic endolysin production. Phagocytosis assays using an encapsulated and unencapsulated pneumococcal strain showed a reduction of intracellular bacterial burden in Cpl-1 mRNA-transfected macrophages compared with control and inactive-mutant Cpl-1 mRNA groups, a flow cytometry-based assay further corroborated a decreased intracellular bacterial signal. Together, these findings suggest that mRNA-encoded Cpl-1 enhances intracellular killing of S. pneumoniae and support the feasibility of mRNA-based endolysin therapies to target intracellular pneumococcal reservoirs.

Streptococcus pneumoniae remains a major global health threat and is listed by the World Health Organization as a pathogen in urgent need of new antimicrobial strategies. While primarily considered an extracellular pathogen, S. pneumoniae can persist within splenic macrophages in severe disease, creating a protected intracellular niche that may contribute to fulminant sepsis. We recently demonstrated the concept of an mRNA-based therapeutic approach in which host cells produce the pneumococcal bacteriophage endolysin Cpl-1. Here, we investigated whether expression of Cpl-1 in macrophages can target S. pneumoniae residing within host cells. Using the human THP-1 macrophage line, we demonstrated successful translation and intracellular accumulation of bioactive Cpl-1 following IVT-mRNA transfection. Lysates from Cpl-1 mRNA-transfected cells exhibited bacteriolytic activity, and Western blotting as well as immunofluorescent staining confirmed cytosolic endolysin production. Phagocytosis assays using an encapsulated and unencapsulated pneumococcal strain showed a reduction in intracellular bacterial burden in Cpl-1 mRNA-transfected macrophages compared with control and inactive-mutant Cpl-1 mRNA groups, and a flow cytometry-based assay further corroborated a decreased intracellular bacterial signal. Together, these findings suggest that mRNA-encoded Cpl-1 enhances intracellular killing of S. pneumoniae and supports the feasibility of mRNA-based endolysin therapies to target intracellular pneumococcal reservoirs.