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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.

Staphylococcus aureus (Sa), as one of the major human pathogens, has very effective strategies to subvert the human immune system. Virulence of the emerging community-associated methicillin-resistant Sa (CA-MRSA) depends on the secretion of phenol-soluble modulin (PSM) peptide toxins e.g., by binding to and modulation of innate immune cells. Previously, by using mouse bone marrow-derived dendritic cells we demonstrated that PSMs in combination with various Toll-like receptor (TLR) ligands induce a tolerogenic DC phenotype (tDC) characterized by the production of IL-10 and impaired secretion of pro-inflammatory cytokines. Consequently, PSM-induced tDCs favored priming of CD4+CD25+FoxP3+ Tregs with suppressor function while impairing the Th1 response. However, the relevance of these findings for the human system remained elusive. Here, we analyzed the impact of PSMα3 on the maturation, cytokine production, antigen uptake, and T cell stimulatory capacity of human monocyte-derived DCs (moDCs) treated simultaneously with either LPS (TLR4 ligand) or Sa cell lysate (TLR2 ligand). Herein, we demonstrate that PSMs indeed modulate human moDCs upon treatment with TLR2/4 ligands via multiple mechanisms, such as transient pore formation, impaired DC maturation, inhibited pro- and anti-inflammatory cytokine secretion, as well as reduced antigen uptake. As a result, the adaptive immune response was altered shown by an increased differentiation of naïve and even CD4+ T cells from patients with Th1/Th17-induced diseases (spondyloarthritis and rheumatoid arthritis) into CD4+CD127−CD25hiCD45RA−FoxP3hi regulatory T cells (Tregs) with suppressor function. This Treg induction was mediated most predominantly by direct DC-T-cell interaction. Thus, PSMs from highly virulent Sa strains affect DC functions not only in the mouse, but also in the human system, thereby modulating the adaptive immune response and probably increasing the tolerance toward the bacteria. Moreover, PSMα3 might be a novel peptide for tolerogenic DC induction that may be used for DC vaccination strategies.

DCs are professional APCs playing a crucial role in the initiation of T-cell responses to combat infection. However, systemic bacterial infection with various pathogens leads to DC-depletion in humans and mice. The mechanisms of pathogen-induced DC-depletion remain poorly understood. Previously, we showed that mice infected with Yersinia enterocolitica (Ye) had impaired de novo DC-development, one reason for DC-depletion. Here, we extend these studies to gain insight into the molecular mechanisms of DC-depletion and the impact of different bacteria on DC-development. We show that the number of bone marrow (BM) hematopoietic progenitors committed to the DC lineage is reduced following systemic infection with different Gram-positive and Gram-negative bacteria. This is associated with a TLR4- and IFN-γ-signaling dependent increase of committed monocyte progenitors in the BM and mature monocytes in the spleen upon Ye-infection. Adoptive transfer experiments revealed that infection-induced monopoiesis occurs at the expense of DC-development. Our data provide evidence for a general response of hematopoietic progenitors upon systemic bacterial infections to enhance monocyte production, thereby increasing the availability of innate immune cells for pathogen control, whereas impaired DC-development leads to DC-depletion, possibly driving transient immunosuppression in bacterial sepsis. Fil: Pasquevich, Karina Alejandra. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Biotecnológicas. Instituto de Investigaciones Biotecnológicas ; Argentina. University of Tübingen; Alemania Fil: Bieber, Kristin. University of Tübingen; Alemania Fil: Günter, Manina. University of Tübingen; Alemania Fil: Grauer, Matthias. University of Tübingen; Alemania Fil: Pötz, Oliver. University of Tübingen; Alemania Fil: Schleicher, Ulrike. Universität Erlangen; Alemania Fil: Biedermann, Tilo. University of Tübingen; Alemania Fil: Beer-Hammer, Sandra. University of Tübingen; Alemania Fil: Bühring, Hans-Jörg. University of Tübingen; Alemania Fil: Rammensee, Hans-Georg. University of Tübingen; Alemania Fil: Zender, Lars. University of Tübingen; Alemania Fil: Autenrieth, Ingo B.. University of Tübingen; Alemania Fil: Lengerke, Claudia. University of Tübingen; Alemania Fil: Autenrieth, Stella E.. University of Tübingen; Alemania

Dendritische Zellen (DC) sind von entscheidender Relevanz für das Zusammenspiel von angeborenem und erworbenem Immunsystem. Hierbei wird die Funktionalität dieses System unter anderem durch die Homöostase des DC-Netzwerkes gewährleistet, die sich auf einem schmalen Grat zwischen Immundefizienz und Autoimmunität bewegt. Vorarbeiten zeigten, dass sich der promiskuitive und G-Protein-gekoppelte FPR2 auf die Anzahl Dendritischer Zellen im lymphatischen Gewebe auswirkt. Dieser mögliche Einfluss auf die DC-Homöostase sollte hier untersucht werden. Hierzu wurden die Zellzahl und Verteilung von DCs, sowie ihren Subpopulationen und Vorläufern in Knochenmark und Milz von C57BL/6- und FPR2-/--Mäusen durchflusszytometrisch analysiert. Ebenso wurde das Apoptose-Verhalten der DCs, sowie die intrazelluläre Menge des anti-apoptotischen Proteins Bcl-2 per Durchflusszytometrie untersucht. Außerdem wurden in Kultur generierte C57BL/6- und FPR2-/--BMDCs in Bezug auf ihre Maturation und die Sezernierung von Zytokinen nach Stimulation mit LPS analysiert. Höhere DC Zahlen in der Milz von FPR2-/--Mäusen, wie in den vorherigen Versuchen, konnten nicht gezeigt werden, jedoch wurden veränderte prozentuale Anteile von DC-Subpopulationen beobachtet. Die FPR2-/--DCs in der Milz zeigten ein besseres Überleben und höhere intrazelluläre Mengen des anti-apoptotischen Protein Bcl-2. Die FPR2-/--BMDCs sezernierten nach Stimulation mit LPS signifikant geringere Konzentrationen des anti-apoptotischen Zytokins IL-10. Der FPR2 zeigt also einen Einfluss auf die Apoptose der Dendritischen Zellen, sowie auf ihre anti-inflammatorische Funktion. Zur abschließenden Klärung seines Einflusses auf die DC-Homöostase sind jedoch noch weitere Untersuchungen angezeigt.

Sleep has been linked to several vital body functions from memory and metabolism to immunity. The results in this work – showing that sleep in mice has a profound effect on innate immune cells and their ability to fight bacterial infection – support these findings. For the experiments two mice groups were used, sleep mice with 6 h of normal sleep were compared to mice with 6 h of enhanced wakefulness. 6 h of sleep increases the overall cellularity in the blood and spleen of WT mice, in particular the number and frequency of monocytes. This is described in this work as the sleep phenotype, a strong increase in frequency and count of monocytes due to sleep. This phenotype was found for classical monocytes and non-classical monocytes. The increase of monocytes is not due to stress or the progression into macrophages or DCs. Furthermore, no effect of sleep was found on monocytes precursors or their release from the BM. Interestingly, the increase of monocytes in blood and spleen did not lead to a reduction of monocytes in Lymph Nodes, the LP, or the lung. On the contrary, monocyte numbers as well increase in the lung after 6 h of sleep. Furthermore, the sleep phenotype is independent of the ICAM-1-LFA-1 axis and just partially dependent on CCR2. Both are very important factors for the migration of classical monocytes and their contribution to the marginal pool. However, it could be shown that for non-classical monocytes Cx3CR1 is involved in the sleep phenotype. Beyond this, it was demonstrated that the sleep phenotype for classical monocytes is dependent on the Clock gene Arntl, indicating a strong connection between the circadian system and sleep. With regards to immune cell function, sleep increases the ROS production of PMNs and classical monocytes. Consequential this leads to better bacterial clearance and improved survival time upon infection. Taken together sleep is imperative for immune functions and monocyte numbers are highly affected by sleep as well as their ability to fight a bacterial infection. Even though the exact mechanism is not yet fully understood there are strong indications for a connection to the circadian system.