Inbred mouse strains provide phenotypic homogeneity between individual mice. However, stochastic morphogenetic events combined with epigenetic changes due to exposure to environmental factors and ontogenic experience result in variability among mice with virtually identical genotypes, reducing the reproducibility of experimental mouse models. Here we used microscopic and cytometric techniques to identify individual patterns in gut-associated lymphoid tissue (GALT) that are induced by exposure to microbiota. By comparing germ-free (GF), conventional (CV) and gnotobiotic mice colonized with a defined minimal mouse microbiota (oMM12) MHC II-EGFP knock-in mice we quantified antigen-presenting cells (APCs) in the lamina propria, cryptopatches (CP), isolated lymphoid follicles (ILFs), Peyer's patches (PPs) and specific sections of the mesenteric lymphoid complex. We found that GF mice had a significantly larger outer intestinal surface area compared to CV and oMM12-colonized mice, which partially compensated for their lower density of the villi in the distal ileum. GF mice also contained fewer APCs than oMM12 mice in the Iamina propria of the villi and had a significantly smaller volume of the solitary intestinal lymphoid tissue (SILT). In both GF and oMM12 mice, PP follicles were significantly smaller compared to CV mice, although number was similar. Concomitantly, the number of pDCs in PPs was significantly lower in GF mice than in CV mice. Moreover, the cecal patch was dispersed into small units in GF mice whereas it was compact in CV mice. Taken together, we here provide further evidence that microbiota regulates SILT differentiation, the size and morphology of PPs, the cellular composition of mesenteric lymph nodes (MLNs) and the morphology of cecal patch. As such, microbiota directly affect not only the functional configuration of the immune system but also the differentiation of lymphoid structures. These findings highlight how standardized microbiota, such as oMM12, can promote reproducibility in animal studies by enabling microbiologically controlled experiments across laboratories.

• Klíčová slova
• Germ-free and gnotobiotic models, Gut-associated lymphoid tissue (GALT), Lymphoid tissue morphogenesis, MHCII-EGFP knock-in mice, Microbiota-induced immunity, Phenotypic plasticity,
• Publikační typ
• časopisecké články MeSH

The organs of many female animals are remodeled by reproduction. Using the mouse intestine, a striking and tractable model of organ resizing, we find that reproductive remodeling is anticipatory and distinct from diet- or microbiota-induced resizing. Reproductive remodeling involves partially irreversible elongation of the small intestine and fully reversible growth of its epithelial villi, associated with an expansion of isthmus progenitors and accelerated enterocyte migration. We identify induction of the SGLT3a transporter in a subset of enterocytes as an early reproductive hallmark. Electrophysiological and genetic interrogations indicate that SGLT3a does not sustain digestive functions or enterocyte health; rather, it detects protons and sodium to extrinsically support the expansion of adjacent Fgfbp1-positive isthmus progenitors, promoting villus growth. Our findings reveal unanticipated specificity to physiological organ remodeling. We suggest that organ- and state-specific growth programs could be leveraged to improve pregnancy outcomes or prevent maladaptive consequences of such growth.

• Klíčová slova
• Fgfbp1, SGLT3a, adult organ remodeling, intestinal epithelium, isthmus progenitor, lactation, plasticity, pregnancy, reproduction, small intestine,
• MeSH
• enterocyty metabolismus   cytologie   MeSH
• myši inbrední C57BL MeSH
• myši MeSH
• pohyb buněk MeSH
• rozmnožování * fyziologie   MeSH
• střeva * růst a vývoj   MeSH
• střevní sliznice metabolismus   MeSH
• těhotenství MeSH
• tenké střevo * růst a vývoj   metabolismus   MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• těhotenství MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH

BACKGROUND: Animals form complex symbiotic associations with their gut microbes, whose evolution is determined by an intricate network of host and environmental factors. In many insects, such as Drosophila melanogaster, the microbiome is flexible, environmentally determined, and less diverse than in mammals. In contrast, mammals maintain complex multispecies consortia that are able to colonize and persist in the gastrointestinal tract. Understanding the evolutionary and ecological dynamics of gut microbes in different hosts is challenging. This requires disentangling the ecological factors of selection, determining the timescales over which evolution occurs, and elucidating the architecture of such evolutionary patterns. RESULTS: We employ experimental evolution to track the pace of the evolution of a common gut commensal, Lactiplantibacillus plantarum, within invertebrate (Drosophila melanogaster) and vertebrate (Mus musculus) hosts and their respective diets. We show that in Drosophila, the nutritional environment dictates microbial evolution, while the host benefits L. plantarum growth only over short ecological timescales. By contrast, in a mammalian animal model, L. plantarum evolution results to be divergent between the host intestine and its diet, both phenotypically (i.e., host-evolved populations show higher adaptation to the host intestinal environment) and genomically. Here, both the emergence of hypermutators and the high persistence of mutated genes within the host's environment strongly differed from the low variation observed in the host's nutritional environment alone. CONCLUSIONS: Our results demonstrate that L. plantarum evolution diverges between insects and mammals. While the symbiosis between Drosophila and L. plantarum is mainly determined by the host diet, in mammals, the host and its intrinsic factors play a critical role in selection and influence both the phenotypic and genomic evolution of its gut microbes, as well as the outcome of their symbiosis.

• Klíčová slova
• Drosophila melanogaster, Experimental evolution, Gut microbiota evolution, Host–microbe symbiosis, Lactiplantibacillus plantarum, Mouse, Whole genome sequencing,
• MeSH
• Drosophila melanogaster genetika   MeSH
• Drosophila MeSH
• mikrobiota * MeSH
• myši MeSH
• savci MeSH
• střevní mikroflóra * MeSH
• symbióza MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, U.S. Gov't, Non-P.H.S. MeSH

Hundreds of studies in last decades have aimed to compare the microbiome of patients suffering from diverse diseases with that of healthy controls. The microbiome-related component was additionally identified in pathophysiology of many diseases formerly considered to depend only on the host physiology. This, however, opens important questions like: "What is the healthy microbiome?" or "Is it possible to define it unequivocally?". In this review, we describe the main hindrances complicating the definition of "healthy microbiome" in terms of microbiota composition. We discuss the human microbiome from the perspective of classical ecology and we advocate for the shift from the stress on microbiota composition to the functions that microbiome ensures for the host. Finally, we propose to leave the concept of ideal healthy microbiome and replace it by focus on microbiome advantageous for the host, which always depends on the specific context like the age, genetics, dietary habits, body site or physiological state.

• MeSH
• lidé MeSH
• mikrobiota * fyziologie   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• přehledy MeSH

Mature T cells are selected for recognizing self-antigens with low to intermediate affinity in the thymus. Recently, the relative differences in self-reactivity among individual T-cell clones were appreciated as important factors regulating their fate and immune response, but the role of self-reactivity in T-cell biology is incompletely understood. We addressed the role of self-reactivity in T-cell diversity by generating an atlas of mouse peripheral CD8+ T cells, which revealed two unconventional populations of antigen-inexperienced T cells. In the next step, we examined the steady-state phenotype of monoclonal T cells with various levels of self-reactivity. Highly self-reactive clones preferentially differentiate into antigen-inexperienced memory-like cells, but do not form a population expressing type I interferon-induced genes, showing that these two subsets have unrelated origins. The functional comparison of naïve monoclonal CD8+ T cells specific to the identical model antigen did not show any correlation between the level of self-reactivity and the magnitude of the immune response.

• Klíčová slova
• T cell, T-cell diversity, antigen-inexperienced memory-like CD8 T cells, interferon response, self-reactivity,
• MeSH
• autoantigeny MeSH
• buněčné klony MeSH
• CD8-pozitivní T-lymfocyty * MeSH
• interferon typ I * MeSH
• myši MeSH
• thymus MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• autoantigeny MeSH
• interferon typ I * MeSH

The development of inflammatory bowel disease (IBD) is associated with alterations in the gut microbiota. There is currently no universal treatment for this disease, thus emphasizing the importance of developing innovative therapeutic approaches. Gut microbiome-derived metabolite butyrate with its well-known anti-inflammatory effect in the gut is a promising candidate. Due to increased intestinal permeability during IBD, butyrate may also reach the liver and influence liver physiology, including hepatic drug metabolism. To get an insight into this reason, the aim of this study was set to clarify not only the protective effects of the sodium butyrate (SB) administration on colonic inflammation but also the effects of SB on hepatic drug metabolism in experimental colitis induced by dextran sodium sulfate (DSS) in mice. It has been shown here that the butyrate pre-treatment can alleviate gut inflammation and reduce the leakiness of colonic epithelium by restoration of the assembly of tight-junction protein Zonula occludens-1 (ZO-1) in mice with DSS-induced colitis. In this article, butyrate along with inflammation has also been shown to affect the expression and enzyme activity of selected cytochromes P450 (CYPs) in the liver of mice. In this respect, CYP3A enzymes may be very sensitive to gut microbiome-targeted interventions, as significant changes in CYP3A expression and activity in response to DSS-induced colitis and/or butyrate treatment have also been observed. With regard to medications used in IBD and microbiota-targeted therapeutic approaches, it is important to deepen our knowledge of the effect of gut inflammation, and therapeutic interventions were followed concerning the ability of the organism to metabolize drugs. This gut-liver axis, mediated through inflammation as well as microbiome-derived metabolites, may affect the response to IBD therapy.

• Klíčová slova
• butyrate, cytochromes P450, drug metabolism, gut inflammation, gut–liver axis,
• Publikační typ
• časopisecké články MeSH

Mus musculus is the classic mammalian model for biomedical research. Despite global efforts to standardize breeding and experimental procedures, the undefined composition and interindividual diversity of the microbiota of laboratory mice remains a limitation. In an attempt to standardize the gut microbiome in preclinical mouse studies, here we report the development of a simplified mouse microbiota composed of 15 strains from 7 of the 20 most prevalent bacterial families representative of the fecal microbiota of C57BL/6J Specific (and Opportunistic) Pathogen-Free (SPF/SOPF) animals and the derivation of a standardized gnotobiotic mouse model called GM15. GM15 recapitulates extensively the functionalities found in the C57BL/6J SOPF microbiota metagenome, and GM15 animals are phenotypically similar to SOPF or SPF animals in two different facilities. They are also less sensitive to the deleterious effects of post-weaning malnutrition. In this work, we show that the GM15 model provides increased reproducibility and robustness of preclinical studies by limiting the confounding effect of fluctuation in microbiota composition, and offers opportunities for research focused on how the microbiota shapes host physiology in health and disease.

• MeSH
• Bacteria klasifikace   genetika   MeSH
• druhová specificita MeSH
• feces mikrobiologie   MeSH
• fenotyp MeSH
• gnotobiologické modely * MeSH
• metagenomika metody   MeSH
• myši inbrední C57BL MeSH
• myši MeSH
• organismy bez specifických patogenů * MeSH
• sekvenování celého genomu metody   MeSH
• střevní mikroflóra genetika   fyziologie   MeSH
• tělesná hmotnost genetika   fyziologie   MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Extramural MeSH

A steady rise in the number of poly-sensitized patients has increased the demand for effective prophylactic strategies against multi-sensitivities. Probiotic bacteria have been successfully used in clinics and experimental models to prevent allergic mono-sensitization. In the present study, we have investigated whether probiotic bacteria could prevent poly-sensitization by imprinting on the immune system early in life. We used two recombinant variants of probiotic Escherichia coli Nissle 1917 (EcN): i) EcN expressing birch and grass pollen, poly-allergen chimera construct (EcN-Chim), and ii) an "empty" EcN without allergen expression (EcN-Ctrl). Conventional mice (CV) were treated with either EcN-Chim or EcN-Ctrl in the last week of the gestation and lactation period. Gnotobiotic mice received one oral dose of either EcN-Chim or EcN-Ctrl before mating. The offspring from both models underwent systemic allergic poly-sensitization and intranasal challenge with recombinant birch and grass pollen allergens (rBet v 1, rPhl p 1, and rPhl p 5). In the CV setting, the colonization of offspring via treatment of mothers reduced allergic airway inflammation (AAI) in offspring compared to poly-sensitized controls. Similarly, in a gnotobiotic model, AAI was reduced in EcN-Chim and EcN-Ctrl mono-colonized offspring. However, allergy prevention was more pronounced in the EcN-Ctrl mono-colonized offspring as compared to EcN-Chim. Mono-colonization with EcN-Ctrl was associated with a shift toward mixed Th1/Treg immune responses, increased expression of TLR2 and TLR4 in the lung, and maintained levels of zonulin-1 in lung epithelial cells as compared to GF poly-sensitized and EcN-Chim mono-colonized mice. This study is the first one to establish the model of allergic poly-sensitization in gnotobiotic mice. Using two different settings, gnotobiotic and conventional mice, we demonstrated that an early life intervention with the EcN without expressing an allergen is a powerful strategy to prevent poly-sensitization later in life.

• Klíčová slova
• BALB/c, Escherichia coli Nissle 1917, allergic poly-sensitization, germ-free, hygiene hypothesis, mouse model, mucosal tolerance,
• MeSH
• alergeny imunologie   MeSH
• alergie imunologie   MeSH
• antigeny rostlinné imunologie   MeSH
• bříza imunologie   MeSH
• epitelové buňky imunologie   MeSH
• Escherichia coli imunologie   MeSH
• gnotobiologické modely imunologie   MeSH
• homeostáza imunologie   MeSH
• imunitní systém imunologie   MeSH
• lipnicovité imunologie   MeSH
• myši inbrední BALB C MeSH
• myši MeSH
• probiotika aplikace a dávkování   MeSH
• pyl imunologie   MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• alergeny MeSH
• antigeny rostlinné MeSH

Our understanding of human gut microbiota in health and disease depends on accurate and reproducible microbial data acquisition. The critical step in this process is to apply an appropriate methodology to extract microbial DNA, since biases introduced during the DNA extraction process may result in inaccurate microbial representation. In this study, we attempted to find a DNA extraction protocol which could be effectively used to analyze both the bacterial and fungal community. We evaluated the effect of five DNA extraction methods (QIAamp DNA Stool Mini Kit, PureLinkTM Microbiome DNA Purification Kit, ZR Fecal DNA MiniPrepTM Kit, NucleoSpin® DNA Stool Kit, and IHMS protocol Q) on bacterial and fungal gut microbiome recovery using (i) a defined system of germ-free mice feces spiked with bacterial or fungal strains, and (ii) non-spiked human feces. In our experimental setup, we confirmed that the examined methods significantly differed in efficiency and quality, which affected the identified stool microbiome composition. In addition, our results indicated that fungal DNA extraction might be prone to be affected by reagent/kit contamination, and thus an appropriate blank control should be included in mycobiome research. Overall, standardized IHMS protocol Q, recommended by the International Human Microbiome Consortium, performed the best when considering all the parameters analyzed, and thus could be applied not only in bacterial, but also in fungal microbiome research.

• Klíčová slova
• 16S rDNA, DNA extraction method, ITS rDNA, fungal microbiota, gut microbiome, gut microbiota, gut mycobiome, gut mycobiota,
• Publikační typ
• časopisecké články MeSH

Background: Mucosal mast cells (MC) are key players in IgE-mediated food allergy (FA). The evidence on the interaction between gut microbiota, MC and susceptibility to FA is contradictory. Objective: We tested the hypothesis that commensal bacteria are essential for MC migration to the gut and their maturation impacting the susceptibility to FA. Methods: The development and severity of FA symptoms was studied in sensitized germ-free (GF), conventional (CV), and mice mono-colonized with L. plantarum WCFS1 or co-housed with CV mice. MC were phenotypically and functionally characterized. Results: Systemic sensitization and oral challenge of GF mice with ovalbumin led to increased levels of specific IgE in serum compared to CV mice. Remarkably, despite the high levels of sensitization, GF mice did not develop diarrhea or anaphylactic hypothermia, common symptoms of FA. In the gut, GF mice expressed low levels of the MC tissue-homing markers CXCL1 and CXCL2, and harbored fewer MC which exhibited lower levels of MC protease-1 after challenge. Additionally, MC in GF mice were less mature as confirmed by flow-cytometry and their functionality was impaired as shown by reduced edema formation after injection of degranulation-provoking compound 48/80. Co-housing of GF mice with CV mice fully restored their susceptibility to develop FA. However, this did not occur when mice were mono-colonized with L. plantarum. Conclusion: Our results demonstrate that microbiota-induced maturation and gut-homing of MC is a critical step for the development of symptoms of experimental FA. This new mechanistic insight into microbiota-MC-FA axis can be exploited in the prevention and treatment of FA in humans.

• Klíčová slova
• commensal bacteria, food allergy, germ-free, mast cells, mouse models,
• MeSH
• biologické markery MeSH
• buněčná diferenciace genetika   imunologie   MeSH
• cytokiny metabolismus   MeSH
• degranulace buněk genetika   imunologie   MeSH
• gnotobiologické modely MeSH
• mastocyty imunologie   metabolismus   MeSH
• metagenom MeSH
• metagenomika metody   MeSH
• mikrobiota * imunologie   MeSH
• modely nemocí na zvířatech MeSH
• myši MeSH
• náchylnost k nemoci MeSH
• potravinová alergie etiologie   metabolismus   patologie   MeSH
• RNA ribozomální 16S MeSH
• střevní mikroflóra MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• biologické markery MeSH
• cytokiny MeSH
• RNA ribozomální 16S MeSH