Strong tight junctions and curtailed inflammatory responses under stressful conditions are key for optimal digestive health. Bacillus -based probiotics are increasingly being used to maintain broilers' health, but their mode of action is often not well-defined. In the present study we used Caco-2 cells as a model for intestinal epithelia and assessed the effect of three Bacillus -based probiotics on intestinal barrier function and intestinal inflammation. Experimental results showed that one of the three tested strains, Bs 29784, significantly reinforced intestinal barrier integrity under basal conditions through an up-regulation of the expression of tight junction's proteins, whereas the others had no or detrimental effects. When Caco-2 cells were pre-treated with Bacillus subtilis strains, the subsequent IL-8 release to various pro-inflammatory signals (IL-1β, deoxynivalenol, or flagellin) was blunted compared to cells that had not been pretreated, but to a different extent depending on the strain of Bacillus used. Bs 29784, was able to significantly decrease IL-8 production in all stressed conditions tested. Mechanistically, Bs 29784 appeared to limit nuclear translocation of NF-κB during IL-1β exposure by preventing IκB degradation. The effects of Bs 29784 were observed independently with supernatant and cells but in a lesser extent than with the combination, indicating that they can thus likely be attributed to both secreted metabolites and cell-associated compounds. Moreover, under inflammatory conditions, Bs 29784 significantly reduced the upregulation of iNOS protein levels further underlining its intestinal anti-inflammatory potential. Our data show that Bacillus -based probiotics may indeed improve digestive health by strengthening intestinal barrier and limiting inflammatory responses and that these properties are strain-dependent.

Defensins represents an integral part of the innate immune system serving to ward off potential pathogens and to protect the intestinal barrier from microbial encroachment. In addition to their antimicrobial activities, defensins in general, and human β-defensin 2 (hBD2) in particular, also exhibit immunomodulatory capabilities. In this report, we assessed the therapeutic efficacy of systemically administered recombinant hBD2 to ameliorate intestinal inflammation in three distinct animal models of inflammatory bowel disease; i.e., chemically induced mucosal injury (DSS), loss of mucosal tolerance (TNBS), and T-cell transfer into immunodeficient recipient mice. Treatment efficacy was confirmed in all tested models, where systemically administered hBD2 mitigated inflammation, improved disease activity index, and hindered colitis-induced body weight loss on par with anti-TNF-α and steroids. Treatment of lipopolysaccharide (LPS)-activated human peripheral blood mononuclear cells with rhBD2 confirmed the immunomodulatory capacity in the circulatory compartment. Subsequent analyzes revealed dendritic cells (DCs) as the main target population. Suppression of LPS-induced inflammation was dependent on chemokine receptor 2 (CCR2) expression. Mechanistically, hBD2 engaged with CCR2 on its DC target cell to decrease NF-κB, and increase CREB phosphorylation, hence curbing inflammation. To our knowledge, this is the first study showing in vivo efficacy of a systemically administered defensin in experimental disease.

Antimicrobial therapy of infections with Staphylococcus aureus can pose a challenge due to slow response to therapy and recurrence of infection. These treatment difficulties can partly be explained by intracellular survival of staphylococci, which is why the intracellular activity of antistaphylococcal compounds has received increased attention within recent years. The intracellular activity of plectasin, an antimicrobial peptide, against S. aureus was determined both in vitro and in vivo. In vitro studies using THP-1 monocytes showed that some intracellular antibacterial activity of plectasin was maintained (maximal relative efficacy [E max ], 1.0-to 1.3-log reduction in CFU) even though efficacy was inferior to that of extracellular killing (E max , >4.5-log CFU reduction). Animal studies included a novel use of the mouse peritonitis model, exploiting extra-and intracellular differentiation assays, and assessment of the correlations between activity and pharmacokinetic (PK) parameters. The intracellular activity of plectasin was in accordance with the in vitro studies, with an E max of a 1.1-log CFU reduction. The parameter most important for activity was fC peak /MIC, where fC peak is the free peak concentration. These findings stress the importance of performing studies of extra-and intracellular activity since these features cannot be predicted from traditional MIC and killing kinetic studies. Application of both the THP-1 and the mouse peritonitis models showed that the in vitro results were similar to findings in the in vivo model with respect to demonstration of intracellular activity. Therefore the in vitro model was a good screening model for intracellular activity. However, animal models should be applied if further information on activity, PK/pharmacodynamic parameters, and optimal dosing regimens is required.

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NZ2114 displayed similar extracellular and intracellular activities as daptomycin, and was more effective than vancomycin against the intracellular forms of susceptible bacteria. However, the study also showed that the intracellular activities of NZ2114 and daptomycin are weaker than their extracellular activities.