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The effects of bacterial lysates on the gut barrier function and microbiota composition
Zákostelská, Zuzana ; Tlaskalová - Hogenová, Helena (advisor) ; Prokešová, Ludmila (referee) ; Rada, Vojtěch (referee)
Dynamic molecular interactions between the microbiota and the intestinal mucosa play an important role in the establishment and maintenance of mucosal homeostasis. Aberrant host- microbiota interaction could lead to many diseases such as inflammatory bowel disease. The aim of our study was to evaluate the commensal and probiotic bacteria activities and their ability to induce pathological or exert beneficial effects. The most important trigger for immune system development is an exposure to microbial components. Here, we show that there is a time window at about three weeks of age, which enables the artificial colonization of germ free mice by a single oral dose of cecal content. The delayed colonization by either inoculation or co-housing causes permanent changes in immune system reactivity, which may downgrade the results of experiments performed on first generation of colonized animals. In this thesis we report that even non-living commensal bacteria such as Parabacteroides distasonis (mPd) or well known probiotics such as L. casei DN-114 001 (Lc) possess anti-inflammatory effects in experimental model of colitis. The mechanisms that this effect is achieved by the lysate of L. casei DN-114 001 comprise: a) improvement in the gut barrier function, b) correction of the dysbiosis, and c) modulation of the...
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Impact of Probiotic Bacteria on Allergic Sensitization in Type I Allergy Model
Schwarzer, Martin ; Kozáková, Hana (advisor) ; Prokešová, Ludmila (referee) ; Nevoral, Jiří (referee)
The main goal in reversing the allergy epidemic is the development of effective prophylactic strategies. Early life events, such as exposures to microbes, have a major influence on the development of balanced immune responses. Due to their ability to interact with host immune system and to modulate host immune responses probiotics, mainly bifidobacteria and lactobacilli have been used with some success in prevention of allergic disease. In order to be referred to as probiotic, bacterial strain has to undergo rigorous testing. We have selected three new Lactobacillus (L.) strains out of 24 human isolates according to their antagonistic activity against pathogenic bacteria, resistance to low pH and milieu of bile salts. Safety of these strains was proven upon intragastric administration to mice; moreover, we have shown their ability to shift cytokine Th1 - Th2 balance towards non-allergic Th1 response in isolated splenic cells. Allergen specific prophylaxis using probiotics as vehicles for mucosal delivery of recombinant allergen is an attractive concept for development of well-tolerated and effective allergy vaccines. We have shown that neonatal mono-colonization of germ-free mice with the L. plantarum NCIMB8826 strain producing the major birch pollen allergen Bet v 1 attenuates the development of...
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Food allergy to wheat flour proteins
Šotkovský, Petr ; Tučková, Ludmila (advisor) ; Panzner, Petr (referee) ; Prokešová, Ludmila (referee)
THESIS SUMMARY Food allergy is one of the frequent disorders and its incidence in paediatric as well as adult population is continuously rising, having doubled in the last two decades. Although wheat belongs to major food allergens and is a staple food in most diets, we have only little knowledge of wheat proteins causing IgE mediated hypersensitivity reaction. Diagnostic approaches of food allergy to wheat have a high sensitivity, but low specificity. Poor predictability and specificity may be associated with the insufficient purity of wheat extracts used in sIgE assays or with the lack of major allergens in these extracts. In the first step, we characterized 19 potential allergens recognised by IgE Abs of allergic patients, using proteomic techniques (1-DE, 2-DE, MALDI-TOF, QTOF and LCQDECA nLC-MS/MS ion trap technique). We identified these IgE-binding molecules such as: α-amylase inhibitors, β-amylase, profilin, serpin, β-D-glucan exohydrolase and 27K protein. To quantify sIgE in patient's sera we developed ELISA using the whole wheat extract and two commercially available α-amylase inhibitors. Second, we developed a procedure that allows isolation of wheat allergens from natural sources using Rotofor cell and HPLC. Twenty-seven potential wheat allergens have been successfully identified; of these, the...
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Dental materials and imunne reaction
Chrástová, Iveta ; Bártová, Jiřina (advisor) ; Prokešová, Ludmila (referee)
Adverse reactions to metals is relatively frequent in sensitive individuals, and may lead to the development of various diseases, such as lichen planus and lichenoid reaction and was also described effect of amalgam to the pathology of autoimmune diseases. In sensitive patients with known reactions to metals leads to atrophy of the oral mucosa and discomfort (burning, cutting, excessive salivation). Adverse effects of dental materials are described as an allergic reaction type IV delayed hypersensitivity. It disrupt the immune system, increased vascular permeability, tissue is often damaged by chronic inflammation and it is also activated antigen- nonspecific component of immunity. Monocytes are the key cells of the immune system, which act as antigen presenting cells, in particular after conversion in the tissues into macrophages are directly involved in the activation of immune responses and their stimulation is therefore crucial for the development of adverse reactions to dental materials. In the thesis was measured cytokine production by monocytes after stimulation with metal salts. To determine the extent of the patients' responses to the metals was used lymphocyte proliferation assay (MELISA® ), which was developed and patented in Sweden. We compared lymphocyte proliferation after stimulation...
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Immunologic Characteristics of Cord Blood in Children with Increased Risk of Allergy Development Preventive Use of Probiotics
Hrdý, Jiří ; Prokešová, Ludmila (advisor) ; Tlaskalová - Hogenová, Helena (referee) ; Ulčová-Gallová, Zdeňka (referee)
Allergy is one of the most common diseases. Identification of early prognostic markers pointing to an increased risk of allergy development is therefore of increasing importance. Cord blood represents an easily attainable clinical material for searching for prognostic markers signalizing future allergy development. Proportions of Th1 cytokines, Th2 cytokines and regulatory cytokines were tested in cord blood of children of allergic mothers (children in relatively high risk of allergy development) in comparison with cord blood of children of healthy mothers (low risk children). Also the activities of lymphocytes, dendritic cells (DC) and regulatory cells (Tregs) were compared in children of healthy and allergic mothers. The generally increased activity of both in vitro stimulated and non-stimulated mononuclear cord blood leukocytes was proved in children of allergic mothers in comparison with low risk children. The increased activity of DC of high risk children was detectable only after polyclonal stimulation. Significantly less pronounced functional properties of cord blood Tregs were found in children of allergic mothers when compared with children of healthy mothers. The increased reactivity of lymphocytes and DC together with the decreased activity of Tregs can support an easier...
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Function of gut mucosal barrier: methodology and modulation.
Hornová, Michaela ; Tlaskalová - Hogenová, Helena (advisor) ; Prokešová, Ludmila (referee)
ABSTACT Function of gut mucosal barrier: methodology and modulation Right function of the intestinal barrier is essential for maintaining the homeostasis of the inner environment and it serves as a mechanical barrier against penetration of macromolecular compounds and microbes from outer environment. Impaired gut mucosa cannot effectively protect from invasion of infectious microorganisms, antigens from food, bacteria, viruses and xenobiotics into the subepithelial gut layer. Excessive interaction among the immune system and antigens from intestinal lumen can lead to immune mediated damage of the organism. Damaged intestinal barrier plays an important role in the pathogenesis of inflammatory and autoimmune diseases, like inflammatory bowel disease, diabetes, celiac disease and rheumatoid arthritis. The aim of this study was to define methods for measuring intestinal permeability, which would reflect the gut barrier damage. We were trying to detect the gut damage caused by indomethacin (non-steroidal anti-inflammatory drug), alcohol and bacterial LPS by measuring the intestinal permeability for 4,4 kDa FITC-dextran. We were also evaluating changes in gene expression of tight junction proteins, IL-6 and iNOS in mice after intraperitoneal administration of LPS, which induces septic shock. Another goal was to...
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The Role of Gut Microbiota and Lipopolysachaide Content of the Diet in the Development, Maturation and Function of the Immune System
Hrnčíř, Tomáš ; Tlaskalová - Hogenová, Helena (advisor) ; Prokešová, Ludmila (referee) ; Macela, Aleš (referee)
Mammals are essentially born germ-free but the epithelial surfaces are promptly colonized by astounding numbers of bacteria soon after birth. The most extensive microbial community is harboured by the distal intestine. The gut microbiota outnumbers ~10 times the total number of our somatic and germ cells. The hostmicrobiota relationship has evolved to become mutually beneficial. Studies in germfree mice have shown that gut microbiota is essential for the proper development of the immune system. The pivotal role of the innate immune system in the complex and dynamic host-microbiota interactions has become increasingly evident. The principal aims of the present study were: firstly, to determine whether LPS-rich sterile diet can promote maturation of the immune system in germ-free mice, secondly, to elucidate whether gut microbiota and LPS-rich sterile diet influence the LPS susceptibility, and finally, to investigate a role of the adaptive immunity in endotoxin shock. Our data clearly show that both live gut microbiota and LPS-rich sterile diet increase susceptibility to endotoxin shock. Further, we demonstrate that immunodeficient SCID mice, which lack mature B and T cells, are more sensitive to endotoxin shock than immunocompetent Balb/c mice. In addition, we show that not only live gut microbiota but also...
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