• Analysis of factors contributing to variation in the C57BL/6J fecal microbiota across German animal facilities.

      Rausch, Philipp; Basic, Marijana; Batra, Arvind; Bischoff, Stephan C; Blaut, Michael; Clavel, Thomas; Gläsner, Joachim; Gopalakrishnan, Shreya; Grassl, Guntram A; Günther, Claudia; et al. (2016-08)
      The intestinal microbiota is involved in many physiological processes and it is increasingly recognized that differences in community composition can influence the outcome of a variety of murine models used in biomedical research. In an effort to describe and account for the variation in intestinal microbiota composition across the animal facilities of participating members of the DFG Priority Program 1656 "Intestinal Microbiota", we performed a survey of C57BL/6J mice from 21 different mouse rooms/facilities located at 13 different institutions across Germany. Fresh feces was sampled from five mice per room/facility using standardized procedures, followed by extraction and 16S rRNA gene profiling (V1-V2 region, Illumina MiSeq) at both the DNA and RNA (reverse transcribed to cDNA) level. In order to determine the variables contributing to bacterial community differences, we collected detailed questionnaires of animal husbandry practices and incorporated this information into our analyses. We identified considerable variation in a number of descriptive aspects including the proportions of major phyla, alpha- and beta diversity, all of which displayed significant associations to specific aspects of husbandry. Salient findings include a reduction in alpha diversity with the use of irradiated chow, an increase in inter-individual variability (beta diversity) with respect to barrier access and open cages and an increase in bacterial community divergence with time since importing from a vendor. We further observe a high degree of facility-level individuality, which is likely due to each facility harboring its own unique combination of multiple varying attributes of animal husbandry. While it is important to account and control for such differences between facilities, the documentation of such diversity may also serve as a valuable future resource for investigating the origins of microbial-driven host phenotypes.
    • c-FLIP is crucial for IL-7/IL-15-dependent NKp46 ILC development and protection from intestinal inflammation in mice.

      Bank, Ute; Deiser, Katrin; Plaza-Sirvent, Carlos; Osbelt, Lisa; Witte, Amelie; Knop, Laura; Labrenz, Rebecca; Jänsch, Robert; Richter, Felix; Biswas, Aindrila; et al. (Nature research, 2020-02-26)
      NKp46+ innate lymphoid cells (ILC) modulate tissue homeostasis and anti-microbial immune responses. ILC development and function are regulated by cytokines such as Interleukin (IL)-7 and IL-15. However, the ILC-intrinsic pathways translating cytokine signals into developmental programs are largely unknown. Here we show that the anti-apoptotic molecule cellular FLICE-like inhibitory protein (c-FLIP) is crucial for the generation of IL-7/IL-15-dependent NKp46+ ILC1, including conventional natural killer (cNK) cells, and ILC3. Cytokine-induced phosphorylation of signal transducer and activator of transcription 5 (STAT5) precedes up-regulation of c-FLIP, which protects developing NKp46+ ILC from TNF-induced apoptosis. NKp46+ ILC-specific inactivation of c-FLIP leads to the loss of all IL-7/IL-15-dependent NKp46+ ILC, thereby inducing early-onset chronic colitis and subsequently microbial dysbiosis; meanwhile, the depletion of cNK, but not NKp46+ ILC1/3, aggravates experimental colitis. In summary, our data demonstrate a non-redundant function of c-FLIP for the generation of NKp46+ ILC, which protect T/B lymphocyte-sufficient mice from intestinal inflammation.
    • Caecal Microbiota of Experimentally Infected Chickens at Different Ages.

      Hankel, Julia; Jung, Klaus; Kuder, Henrike; Keller, Birgit; Keller, Christoph; Galvez, Eric; Strowig, Till; Visscher, Christian; HZI,Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7,38124 Braunschweig, Germany. (Frontiers, 2019-01-01)
      Campylobacter jejuni is the most common bacterial cause of foodborne zoonosis in the European Union. Infections are often linked to the consumption and handling of poultry meat. The aim of the present study was to investigate the caecal microbiota of birds infected with C. jejuni at different ages. Therefore, a total of 180 birds of the laying hybrid Lohmann Brown-Classic were housed in 12 subgroups of 15 animals each in three performed repetitions. Three birds per subgroup were experimentally infected with C. jejuni at an age of about 21 days and about 78 days (4.46 ± 0.35 log10 CFU/bird). Twenty-one days after experimental infection, microbiome studies were performed on 72 caecal samples of dissected birds (three primary infected and three further birds/subgroup). Amplification within the hypervariable region V 4 of the 16S rRNA gene was performed and sequenced with the Illumina MiSeq platform. Statistical analyses were performed using SAS® Enterprise Guide® (version 7.1) and R (version 3.5.2). Both factors, the experimental replication (p < 0.001) and the chickens' age at infection (p < 0.001) contributed significantly to the differences in microbial composition of the caecal samples. The factor experimental replication explained 24% of the sample's variability, whereas the factor age at infection explained 14% thereof. Twelve of 32 families showed a significantly different count profile between the two age groups, whereby strongest differences were seen for seven families, among them the family Campylobacteraceae (adjusted p = 0.003). The strongest difference between age groups was seen for a bacterial species that is assigned to the genus Turicibacter which in turn belongs to the family Erysipelotrichaceae (adjusted p < 0.0001). Correlation analyses revealed a common relationship in both chicken ages at infection between the absolute abundance of Campylobacteraceae and Alcaligenaceae, which consists of the genus Parasutterella. In general, concentrations of particular volatile fatty acids (VFA) demonstrated a negative correlation to absolute abundance of Campylobacteraceae, whereby the strongest link was seen for n-butyrate (-0.51141; p < 0.0001). Despite performing consecutive repetitions, the factor experimental replication contributed more to the differences of microbial composition in comparison to the factor age at infection.
    • Chronic d-serine supplementation impairs insulin secretion.

      Suwandhi, Lisa; Hausmann, Simone; Braun, Alexander; Gruber, Tim; Heinzmann, Silke S; Gálvez, Eric J C; Buck, Achim; Legutko, Beata; Israel, Andreas; Feuchtinger, Annette; et al. (2018-07-25)
      The metabolic role of d-serine, a non-proteinogenic NMDA receptor co-agonist, is poorly understood. Conversely, inhibition of pancreatic NMDA receptors as well as loss of the d-serine producing enzyme serine racemase have been shown to modulate insulin secretion. Thus, we aim to study the impact of chronic and acute d-serine supplementation on insulin secretion and other parameters of glucose homeostasis. We apply MALDI FT-ICR mass spectrometry imaging, NMR based metabolomics, 16s rRNA gene sequencing of gut microbiota in combination with a detailed physiological characterization to unravel the metabolic action of d-serine in mice acutely and chronically treated with 1% d-serine in drinking water in combination with either chow or high fat diet feeding. Moreover, we identify SNPs in SRR, the enzyme converting L-to d-serine and two subunits of the NMDA receptor to associate with insulin secretion in humans, based on the analysis of 2760 non-diabetic Caucasian individuals. We show that chronic elevation of d-serine results in reduced high fat diet intake. In addition, d-serine leads to diet-independent hyperglycemia due to blunted insulin secretion from pancreatic beta cells. Inhibition of alpha 2-adrenergic receptors rapidly restores glycemia and glucose tolerance in d-serine supplemented mice. Moreover, we show that single nucleotide polymorphisms (SNPs) in SRR as well as in individual NMDAR subunits are associated with insulin secretion in humans. Thus, we identify a novel role of d-serine in regulating systemic glucose metabolism through modulating insulin secretion.
    • Distinct Microbial Communities Trigger Colitis Development upon Intestinal Barrier Damage via Innate or Adaptive Immune Cells.

      Roy, Urmi; Gálvez, Eric J C; Iljazovic, Aida; Lesker, Till Robin; Błażejewski, Adrian J; Pils, Marina C; Heise, Ulrike; Huber, Samuel; Flavell, Richard A; Strowig, Till; et al. (2017-10-24)
      Inflammatory bowel disease comprises a group of heterogeneous diseases characterized by chronic and relapsing mucosal inflammation. Alterations in microbiota composition have been proposed to contribute to disease development, but no uniform signatures have yet been identified. Here, we compare the ability of a diverse set of microbial communities to exacerbate intestinal inflammation after chemical damage to the intestinal barrier. Strikingly, genetically identical wild-type mice differing only in their microbiota composition varied strongly in their colitis susceptibility. Transfer of distinct colitogenic communities in gene-deficient mice revealed that they triggered disease via opposing pathways either independent or dependent on adaptive immunity, specifically requiring antigen-specific CD4+ T cells. Our data provide evidence for the concept that microbial communities may alter disease susceptibility via different immune pathways despite eventually resulting in similar host pathology. This suggests a potential benefit for personalizing IBD therapies according to patient-specific microbiota signatures.
    • The DNA-sensing AIM2 inflammasome controls radiation-induced cell death and tissue injury.

      Hu, Bo; Jin, Chengcheng; Li, Hua-Bing; Tong, Jiyu; Ouyang, Xinshou; Cetinbas, Naniye Malli; Zhu, Shu; Strowig, Till; Lam, Fred C; Zhao, Chen; et al. (2016)
      Acute exposure to ionizing radiation induces massive cell death and severe damage to tissues containing actively proliferating cells, including bone marrow and the gastrointestinal tract. However, the cellular and molecular mechanisms underlying this pathology remain controversial. Here, we show that mice deficient in the double-stranded DNA sensor AIM2 are protected from both subtotal body irradiation-induced gastrointestinal syndrome and total body irradiation-induced hematopoietic failure. AIM2 mediates the caspase-1-dependent death of intestinal epithelial cells and bone marrow cells in response to double-strand DNA breaks caused by ionizing radiation and chemotherapeutic agents. Mechanistically, we found that AIM2 senses radiation-induced DNA damage in the nucleus to mediate inflammasome activation and cell death. Our results suggest that AIM2 may be a new therapeutic target for ionizing radiation exposure.
    • A flagellum-specific chaperone facilitates assembly of the core type III export apparatus of the bacterial flagellum.

      Fabiani, Florian D; Renault, Thibaud T; Peters, Britta; Dietsche, Tobias; Gálvez, Eric J C; Guse, Alina; Freier, Karen; Charpentier, Emmanuelle; Strowig, Till; Franz-Wachtel, Mirita; et al. (2017-08)
      Many bacteria move using a complex, self-assembling nanomachine, the bacterial flagellum. Biosynthesis of the flagellum depends on a flagellar-specific type III secretion system (T3SS), a protein export machine homologous to the export machinery of the virulence-associated injectisome. Six cytoplasmic (FliH/I/J/G/M/N) and seven integral-membrane proteins (FlhA/B FliF/O/P/Q/R) form the flagellar basal body and are involved in the transport of flagellar building blocks across the inner membrane in a proton motive force-dependent manner. However, how the large, multi-component transmembrane export gate complex assembles in a coordinated manner remains enigmatic. Specific for most flagellar T3SSs is the presence of FliO, a small bitopic membrane protein with a large cytoplasmic domain. The function of FliO is unknown, but homologs of FliO are found in >80% of all flagellated bacteria. Here, we demonstrate that FliO protects FliP from proteolytic degradation and promotes the formation of a stable FliP-FliR complex required for the assembly of a functional core export apparatus. We further reveal the subcellular localization of FliO by super-resolution microscopy and show that FliO is not part of the assembled flagellar basal body. In summary, our results suggest that FliO functions as a novel, flagellar T3SS-specific chaperone, which facilitates quality control and productive assembly of the core T3SS export machinery.
    • The gut microbiota drives the impact of bile acids and fat source in diet on mouse metabolism.

      Just, Sarah; Mondot, Stanislas; Ecker, Josef; Wegner, Katrin; Rath, Eva; Gau, Laura; Streidl, Theresa; Hery-Arnaud, Genevieve; Schmidt, Sinah; Lesker, Till Robin; et al. (2018-08-02)
      As the gut microbiota contributes to metabolic health, it is important to determine specific diet-microbiota interactions that influence host metabolism. Bile acids and dietary fat source can alter phenotypes of diet-induced obesity, but the interplay with intestinal microorganisms is unclear. Here, we investigated metabolic consequences of diets enriched in primary bile acids with or without addition of lard or palm oil, and studied gut microbiota structure and functions in mice. In combination with bile acids, dietary lard fed to male C57BL/6N mice for a period of 8 weeks enhanced fat mass accumulation in colonized, but not in germ-free mice when compared to palm oil. This was associated with impaired glucose tolerance, lower fasting insulin levels, lower counts of enteroendocrine cells, fatty liver, and elevated amounts of hepatic triglycerides, cholesteryl esters, and monounsaturated fatty acids. Lard- and bile acid-fed mice were characterized by shifts in dominant gut bacterial communities, including decreased relative abundances of Lachnospiraceae and increased occurrence of Desulfovibrionaceae and the species Clostridium lactatifermentans and Flintibacter butyricus. Metatranscriptomic analysis revealed shifts in microbial functions, including lipid and amino acid metabolism. Caution is required when interpreting data from diet-induced obesity models due to varying effects of dietary fat source. Detrimental metabolic consequences of a diet enriched with lard and primary bile acids were dependent on microbial colonization of the host and were linked to hepatic lipid rearrangements and to alterations of dominant bacterial communities in the cecum.
    • The gut microbiota promotes hepatic fatty acid desaturation and elongation in mice.

      Kindt, Alida; Liebisch, Gerhard; Clavel, Thomas; Haller, Dirk; Hörmannsperger, Gabriele; Yoon, Hongsup; Kolmeder, Daniela; Sigruener, Alexander; Krautbauer, Sabrina; Seeliger, Claudine; et al. (2018-09-14)
      Interactions between the gut microbial ecosystem and host lipid homeostasis are highly relevant to host physiology and metabolic diseases. We present a comprehensive multi-omics view of the effect of intestinal microbial colonization on hepatic lipid metabolism, integrating transcriptomic, proteomic, phosphoproteomic, and lipidomic analyses of liver and plasma samples from germfree and specific pathogen-free mice. Microbes induce monounsaturated fatty acid generation by stearoyl-CoA desaturase 1 and polyunsaturated fatty acid elongation by fatty acid elongase 5, leading to significant alterations in glycerophospholipid acyl-chain profiles. A composite classification score calculated from the observed alterations in fatty acid profiles in germfree mice clearly differentiates antibiotic-treated mice from untreated controls with high sensitivity. Mechanistic investigations reveal that acetate originating from gut microbial degradation of dietary fiber serves as precursor for hepatic synthesis of C16 and C18 fatty acids and their related glycerophospholipid species that are also released into the circulation.
    • Humanized mouse model supports development, function, and tissue residency of human natural killer cells.

      Herndler-Brandstetter, Dietmar; Shan, Liang; Yao, Yi; Stecher, Carmen; Plajer, Valerie; Lietzenmayer, Melanie; Strowig, Till; de Zoete, Marcel R; Palm, Noah W; Chen, Jie; et al. (2017-11-07)
      Immunodeficient mice reconstituted with a human immune system represent a promising tool for translational research as they may allow modeling and therapy of human diseases in vivo. However, insufficient development and function of human natural killer (NK) cells and T cell subsets limit the applicability of humanized mice for studying cancer biology and therapy. Here, we describe a human interleukin 15 (
    • IL22BP Mediates the Anti-Tumor Effects of Lymphotoxin Against Colorectal Tumors in Mice and Humans.

      Kempski, Jan; Giannou, Anastasios D; Riecken, Kristoffer; Zhao, Lilan; Steglich, Babett; Lücke, Jöran; Garcia-Perez, Laura; Karstens, Karl-Frederick; Wöstemeier, Anna; Nawrocki, Mikolaj; et al. (Elsevier, 2020-06-18)
      We obtained tumor and non-tumor tissues from patients with colorectal cancer (CRC) and measured levels of cytokines by quantitative PCR, flow cytometry, and immunohistochemistry. We measured levels of Il22bp mRNA in colon tissues from wild-type, Tnf-/-, Lta-/-, and Ltb-/- mice. Mice were given azoxymethane and dextran sodium sulfate, to induce colitis and associated cancer, or intra-caecal injections of MC38 tumor cells. Some mice were given inhibitors of lymphotoxin beta receptor (LTBR). Intestine tissues were analyzed by single-cell sequencing to identify cell sources of lymphotoxin. We performed immunohistochemistry analysis of colon tissue microarrays from patients with CRC (1475 tissue cores, contained tumor and non-tumor tissues) and correlated levels of IL22BP with patient survival times.
    • An Integrated Metagenome Catalog Reveals New Insights into the Murine Gut Microbiome.

      Lesker, Till R; Durairaj, Abilash C; Gálvez, Eric J C; Lagkouvardos, Ilias; Baines, John F; Clavel, Thomas; Sczyrba, Alexander; McHardy, Alice C; Strowig, Till; HZI,Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7,38124 Braunschweig, Germany.
      The complexity of host-associated microbial ecosystems requires host-specific reference catalogs to survey the functions and diversity of these communities. We generate a comprehensive resource, the integrated mouse gut metagenome catalog (iMGMC), comprising 4.6 million unique genes and 660 metagenome-assembled genomes (MAGs), many (485 MAGs, 73%) of which are linked to reconstructed full-length 16S rRNA gene sequences. iMGMC enables unprecedented coverage and taxonomic resolution of the mouse gut microbiota; i.e., more than 92% of MAGs lack species-level representatives in public repositories (<95% ANI match). The integration of MAGs and 16S rRNA gene data allows more accurate prediction of functional profiles of communities than predictions based on 16S rRNA amplicons alone. Accompanying iMGMC, we provide a set of MAGs representing 1,296 gut bacteria obtained through complementary assembly strategies. We envision that integrated resources such as iMGMC, together with MAG collections, will enhance the resolution of numerous existing and future sequencing-based studies.
    • Intestinal Microbiota of Fattening Pigs Offered Non-Fermented and Fermented Liquid Feed with and without the Supplementation of Non-Fermented Coarse Cereals.

      Bunte, Sebastian; Grone, Richard; Keller, Birgit; Keller, Christoph; Galvez, Eric; Strowig, Till; Kamphues, Josef; Hankel, Julia; HZI,Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7,38124 Braunschweig, Germany. (MDPI, 2020-04-27)
      Introducing high numbers of lactic acid bacteria into the gastrointestinal tract of pigs via fermented liquid feed (FLF) could have an impact on intestinal bacterial ecosystems. Twenty piglets were allocated into four groups and fed a botanically identical liquid diet that was offered either non-fermented (twice), fully fermented or partially fermented but supplemented with 40% of non-fermented coarse cereals. Microbiota studies were performed on the small and large intestine digesta and faecal samples. A 16S rRNA gene amplification was performed within the hypervariable region V4 and sequenced with the Illumina MiSeq platform. R (version 3.5.2) was used for the statistical analyses. The digesta of the small intestines of pigs fed FLF were dominated by Lactobacillaceae (relative abundance up to 95%). In the colonic contents, the abundance of Lactobacillaceae was significantly higher only in the pigs fed the FLF supplemented with non-fermented coarse cereals. Additionally, the digesta of the small and large intestines as well as in the faeces of the pigs fed the FLF supplemented with non-fermented coarse cereals were significantly enriched for two operational taxonomic units (OTUs) belonging to the genus Lactobacillus and Bifidobacterium. The FLF supplemented with non-fermented coarse cereals had probiotic and prebiotic-like impacts on the intestinal and faecal bacterial composition of pigs.
    • Loss of CNFY toxin-induced inflammation drives Yersinia pseudotuberculosis into persistency.

      Heine, Wiebke; Beckstette, Michael; Heroven, Ann Kathrin; Thiemann, Sophie; Heise, Ulrike; Nuss, Aaron Mischa; Pisano, Fabio; Strowig, Till; Dersch, Petra; Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7, 38124 Braunschweig, Germany. (2018-02)
      Gastrointestinal infections caused by enteric yersiniae can become persistent and complicated by relapsing enteritis and severe autoimmune disorders. To establish a persistent infection, the bacteria have to cope with hostile surroundings when they transmigrate through the intestinal epithelium and colonize underlying gut-associated lymphatic tissues. How the bacteria gain a foothold in the face of host immune responses is poorly understood. Here, we show that the CNFY toxin, which enhances translocation of the antiphagocytic Yop effectors, induces inflammatory responses. This results in extensive tissue destruction, alteration of the intestinal microbiota and bacterial clearance. Suppression of CNFY function, however, increases interferon-γ-mediated responses, comprising non-inflammatory antimicrobial activities and tolerogenesis. This process is accompanied by a preterm reprogramming of the pathogen's transcriptional response towards persistence, which gives the bacteria a fitness edge against host responses and facilitates establishment of a commensal-type life style.
    • Microbiota Alters Urinary Bladder Weight and Gene Expression.

      Roje, Blanka; Elek, Anamaria; Palada, Vinko; Bom, Joana; Iljazović, Aida; Šimić, Ana; Sušak, Lana; Vilović, Katarina; Strowig, Till; Vlahoviček, Kristian; et al. (MDPI, 2020-03-17)
      We studied the effect of microbiota on the transcriptome and weight of the urinary bladder by comparing germ-free (GF) and specific pathogen-free (SPF) housed mice. In total, 97 genes were differently expressed (fold change > ±2; false discovery rate (FDR) p-value < 0.01) between the groups, including genes regulating circadian rhythm (Per1, Per2 and Per3), extracellular matrix (Spo1, Spon2), and neuromuscular synaptic transmission (Slc18a3, Slc5a7, Chrnb4, Chrna3, Snap25). The highest increase in expression was observed for immunoglobulin genes (Igkv1-122, Igkv4-68) of unknown function, but surprisingly the absence of microbiota did not change the expression of the genes responsible for recognizing microbes and their products. We found that urinary bladder weight was approximately 25% lighter in GF mice (p = 0.09 for males, p = 0.005 for females) and in mice treated with broad spectrum of antibiotics (p = 0.0002). In conclusion, our data indicate that microbiota is an important determinant of urinary bladder physiology controlling its gene expression and size.
    • Microbiota Normalization Reveals that Canonical Caspase-1 Activation Exacerbates Chemically Induced Intestinal Inflammation.

      Błażejewski, Adrian J; Thiemann, Sophie; Schenk, Alexander; Pils, Marina C; Gálvez, Eric J C; Roy, Urmi; Heise, Ulrike; de Zoete, Marcel R; Flavell, Richard A; Strowig, Till; et al. (2017-06-13)
      Inflammasomes play a central role in regulating intestinal barrier function and immunity during steady state and disease. Because the discoveries of a passenger mutation and a colitogenic microbiota in the widely used caspase-1-deficient mouse strain have cast doubt on previously identified direct functions of caspase-1, we reassessed the role of caspase-1 in the intestine. To this end, we generated Casp1(-/-) and Casp11(-/-) mice and rederived them into an enhanced barrier facility to standardize the microbiota. We found that caspase-11 does not influence caspase-1-dependent processing of IL-18 in homeostasis and during DSS colitis. Deficiency of caspase-1, but not caspase-11, ameliorated the severity of DSS colitis independent of microbiota composition. Ablation of caspase-1 in intestinal epithelial cells was sufficient to protect mice against DSS colitis. Moreover, Casp1(-/-) mice developed fewer inflammation-induced intestinal tumors than control mice. These data show that canonical inflammasome activation controls caspase-1 activity, contributing to exacerbation of chemical-induced colitis.
    • MyD88 signaling in dendritic cells and the intestinal epithelium controls immunity against intestinal infection with C. rodentium.

      Friedrich, Christin; Mamareli, Panagiota; Thiemann, Sophie; Kruse, Friederike; Wang, Zuobai; Holzmann, Bernhard; Strowig, Till; Sparwasser, Tim; Lochner, Matthias; TWINCORE, Zentrum für experimentelle und klinische Infektionsforschung GmbH, Feodor-Lynen Str.7, 30625 Hannover, Germany. (2017-05)
      MyD88-mediated signaling downstream of Toll-like receptors and the IL-1 receptor family is critically involved in the induction of protective host responses upon infections. Although it is known that MyD88-deficient mice are highly susceptible to a wide range of bacterial infections, the cell type-specific contribution of MyD88 in protecting the host against intestinal bacterial infection is only poorly understood. In order to investigate the importance of MyD88 in specific immune and nonimmune cell types during intestinal infection, we employed a novel murine knock-in model for MyD88 that enables the cell type-specific reactivation of functional MyD88 expression in otherwise MyD88-deficient mice. We report here that functional MyD88 signaling in CD11c+ cells was sufficient to activate intestinal dendritic cells (DC) and to induce the early group 3 innate lymphoid cell (ILC3) response as well as the development of colonic Th17/Th1 cells in response to infection with the intestinal pathogen C. rodentium. In contrast, restricting MyD88 signaling to several other cell types, including macrophages (MO), T cells or ILC3 did not induce efficient intestinal immune responses upon infection. However, we observed that the functional expression of MyD88 in intestinal epithelial cells (IEC) also partially protected the mice during intestinal infection, which was associated with enhanced epithelial barrier integrity and increased expression of the antimicrobial peptide RegIIIγ and the acute phase protein SAA1 by epithelial cells. Together, our data suggest that MyD88 signaling in DC and IEC is both essential and sufficient to induce a full spectrum of host responses upon intestinal infection with C. rodentium.
    • Performance, Fermentation Characteristics and Composition of the Microbiome in the Digest of Piglets Kept on a Feed With Humic Acid-Rich Peat.

      Visscher, Christian; Hankel, Julia; Nies, Andrea; Keller, Birgit; Galvez, Eric; Strowig, Till; Keller, Christoph; Breves, Gerhard; HZI, Helmholtz Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7, 38124 Braunschweig Germany. (Frontiers, 2019-01-01)
      The transition from breast milk to solid feed is a dramatic change in the nutrition of piglets, frequently necessitating antibiotic treatment. In efforts to reduce the use of antibiotics, dietetic concepts based on natural feed additives are becoming more and more important. In the present study, experiments were carried out with 15 rearing piglets (days 28–56) divided into three groups that were offered different diets (Ctr [0% peat]; H1.5 [1.5% peat]; and H3.0 [3.0% peat] based on a commercial weaner recipe; all ~178 g CP, 13.7 MJ ME, 13.3 g Lys, as-fed). The contents of cecal and colon digesta were removed at necropsy. The gas formation (4 h) in colon digesta was measured using in vitro batch fermenters. For microbiome studies, 16S rRNA amplification was performed within the hypervariable region V 4 and sequenced with Illumina MiSeq platform. DNA read mapping and statistical analysis were performed using QIIME (version 1.8.0), MicrobiomeAnalyst, RStudio, and SAS Enterprise Guide. The mean body weight of the animals at the end of the trial did not show statistical differences (in kg: Ctr: 26.1 ± 4.85, H1.5: 28.5 ± 3.41, H3.0: 26.2 ± 4.92). The daily weight gains were high for this age (in g/day; Ctr: 607 ± 157; H1.5: 692 ± 101; H3.0: 615 ± 113) and the feed to gain ratio low (in kg/kg; Ctr: 1.538; H1.5: 1.462; H3.0: 1.462). Concentrations of short-chain fatty acids in the cecal content were significantly lower when peat was used (mmol/kg wet weight; Ctr: 173 ± 30.0; H1.5:134 ± 15.0; H3.0:133 ± 17.3). Numerical differences were found in the gas formation (in mL gas per 10 mL batch in 4 h; Ctr: 7.9 ± 2.2; H1.5: 7.4 ± 2.4; H3.0: 6.6 ± 1.1). The microbiome analyses in the cecal content showed significantly higher values for alpha diversity Chao 1 index for samples from the control group. Significant differences were found for bacterial relative abundance for Tenericutes at phylum level and Mollicutes at class level (p < 0.05) in cecal microbiota. Therefore, there was initial evidence that peat influences intestinal microflora causing a shift in the overall concentration of fermentation products in both, the cecal and the colon content.
    • Prdx4 limits caspase-1 activation and restricts inflammasome-mediated signaling by extracellular vesicles.

      Lipinski, Simone; Pfeuffer, Steffen; Arnold, Philipp; Treitz, Christian; Aden, Konrad; Ebsen, Henriette; Falk-Paulsen, Maren; Gisch, Nicolas; Fazio, Antonella; Kuiper, Jan; et al. (2019-09-23)
    • Prdx4 limits caspase‐1 activation and restricts inflammasome‐mediated signaling by extracellular vesicles

      Lipinski, Simone; Pfeuffer, Steffen; Arnold, Philipp; Treitz, Christian; Aden, Konrad; Ebsen, Henriette; Falk‐Paulsen, Maren; Gisch, Nicolas; Fazio, Antonella; Kuiper, Jan; et al. (EMBO Press, 2019-09-23)
      Inflammasomes are cytosolic protein complexes, which orchestrate the maturation of active IL-1β by proteolytic cleavage via caspase-1. Although many principles of inflammasome activation have been described, mechanisms that limit inflammasome-dependent immune responses remain poorly defined. Here, we show that the thiol-specific peroxidase peroxiredoxin-4 (Prdx4) directly regulates IL-1β generation by interfering with caspase-1 activity. We demonstrate that caspase-1 and Prdx4 form a redox-sensitive regulatory complex via caspase-1 cysteine 397 that leads to caspase-1 sequestration and inactivation. Mice lacking Prdx4 show an increased susceptibility to LPS-induced septic shock. This effect was phenocopied in mice carrying a conditional deletion of Prdx4 in the myeloid lineage (Prdx4-ΔLysMCre). Strikingly, we demonstrate that Prdx4 co-localizes with inflammasome components in extracellular vesicles (EVs) from inflammasome-activated macrophages. Purified EVs are able to transmit a robust IL-1β-dependent inflammatory response in vitro and also in recipient mice in vivo. Loss of Prdx4 boosts the pro-inflammatory potential of EVs. These findings identify Prdx4 as a critical regulator of inflammasome activity and provide new insights into remote cell-to-cell communication function of inflammasomes via macrophage-derived EVs