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The ion channel TRPV1 and its role in dendritic cells and macrophages
Trnková, Nella ; Krulová, Magdaléna (advisor) ; Vlachová, Viktorie (referee)
This bachelor's thesis deals with the TRPV1 channel and its role in cells of the innate immune system, such as dendritic cells and macrophages. TRPV1 is a membrane ion channel that is activated by physical stimuli such as pH and temperature as well as by substances from the vanilloid group. The most significant of these is capsaicin. Activation of the TRPV1 channel has several effects on different types of cells. Based on the literature review, the bachelor's thesis concludes that the TRPV1 channel tends to lead dendritic cells to have an anti- inflammatory effect. However, there is evidence for both an anti-inflammatory and a pro-inflammatory response in macrophages and the role of TRPV1 is unclear in this cell type. Thus, further research is needed to understand the exact mechanisms of modulation of immune responses and to clarify the role of the TRPV1 channel. Key words: ion channels, TRPV1, capsaicin, dendritic cells, macrophages
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Early phase of anti-Leishmania immunity in the host skin
Máčalíková, Bára ; Leštinová, Tereza (advisor) ; Kolářová, Iva (referee)
Leishmania parasites are parasitic protozoans that cause disease called leishmaniasis, which primarily affects mammals. Throughout evolution, Leishmania has adapted to the host's immune system, using it to its advantage. This bachelor's thesis describes the relationship between Leishmania and early immune components in the host's skin, as well as the parasite's ability to inhibit the microbicidal activities of cells. The infection begins with the inoculation of infectious promastigotes into the skin, and before reaching their target cells, Leishmania primarily interacts primarily with the complement system, keratinocytes, fibroblats, eosinophils, neutrophils, mast cells and dendritic cells. Understanding the mutual interaction between the host and the parasite is essential for vaccine development and the treatment of leishmaniasis. KEYWORDS: leishmania, skin, early imunity, complement system, keratinocytes, fibroblasts, eosinophils, neutrophils, mast cells, dendritic cells
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Effect of early postnatal supplementation by probiotic bacteria Escherichia coli O83:K24:H31 on proportional and fucntional characteristics of selected cellular population
Věcek, Jan ; Hrdý, Jiří (advisor) ; Funda, David (referee)
The hygiene hypothesis proposes that exposure to microorganisms during the postnatal period is crucial for proper immune system development and may help to prevent development of autoimmune diseases and allergies. Probiotics, live microorganisms with beneficial health effects, could be a safe way to promote the appropriate maturation of the immune system. Early postnatal administration of a specific probiotic strain, Escherichia coli O83:K24:H31 (EcO83), reduces the incidence of allergies later in life. To understand the immunomodulatory features of EcO83, we conducted a bioinformatic analysis of its genome and compared it to two other strains, E. coli Nissle and E. coli K12. Our analysis identified unique genes in EcO83 related to propionate and galactose metabolism, as well as genes that may enhance its ability to thrive in the gastrointestinal tract. Moreover, we transformed EcO83 with luciferase enzymes and observed that it effectively colonizes the gastrointestinal tract of newborn mice but not adult mice. Further analysis of mice treated with EcO83 revealed that the probiotic promotes the expression of genes involved in tight junction formation and increases costimulatory molecules on dendritic cells in the mesenteric lymph nodes (MLN). Induced RORγt+ Tregs in MLN displayed increased...
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Mechanisms of antigen presentation in the etiopathogenesis of celiac disease
Hudec, Michael ; Černá, Marie (advisor) ; Hrdý, Jiří (referee) ; Slavčev, Antonij (referee)
1 ABSTRACT Celiac disease (CeD) is a chronic autoimmune disease that develops as a response of the immune system to the presence of gluten in the small intestine. CeD is manifested not only by classic intestinal symptoms: abdominal pain, constipation or diarrhea, as well as complex less common symptoms: anemia, osteoporosis, psychiatric disorders or menstrual cycle disorders. HLA risk alleles predisposing to origin of celiac disease are HLA-DQ2 (DQA1*05:01 / DQB1*02:01) and HLA-DQ8 (DQA1*03:01 / DQB1*03:02). There are other celiac disease-associated polymorphisms outside of HLA locus (6p21.3) that are located in 5q32 and 19p13 regions with unclear connection to CeD development. HLA class II glycoproteins are expressed on antigen presenting cells (APC) that include dendritic cells, macrophages and B cells. Monocytes are one of several possible dendritic cell precursors that circulate in the bloodstream. Deviations in the frequency of intermediate monocytes are directly associated with autoimmune disorders such as Crohn's disease or rheumatoid arthritis. It is known that the monocytes of CeD patients show pro-inflammatory reaction in the presence of gluten. It means that, in the context of CeD, the response to gluten arises earlier than the activation of gluten-specific T cells. The conventional way of direct...
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Preparation and characterisation of mRNA/DNA transfection vectors
Horák, Tomáš ; Chmelíková, Larisa (referee) ; Skopalík, Josef (advisor)
This diploma thesis deals with genetic engineering, especially the transfection of DNA into MSCs (Mesenchymal stromal cells) and dendritic cells. Both lipoplexes and metal magnetic nanoparticles were tested to introduce the vector into cells. The research was focused on finding more efficient methods of transfection. According to analysis on MADLS and gel electrophoresis, aspects playing an important role in conjugation and subsequent transfection were found. Conjugation occurs after only 4 minutes, as evidenced by an increase in zeta potential, but to achieve full conjugation it is necessary to incubate the sample for 20 minutes. Incomplete conjugation to iron nanoparticles resulted in strong carrier-carrier interactions, which formed an unwanted conglomerates. Encapsulation into liposomes with cationic surface treatment was without complications. The success rate of GFP-labeled protein expression after transfection by these methods was calculated to be 95%, resp. 91%. This result is due to low cytotoxicity. However, commercial tested kits on dendritic cells had a success rate below 5% with high cytotoxicity.
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Preparation and characterisation of mRNA/DNA transfection vectors
Horák, Tomáš ; Chmelíková, Larisa (referee) ; Skopalík, Josef (advisor)
This diploma thesis deals with genetic engineering, especially the transfection of DNA into MSCs (Mesenchymal stromal cells) and dendritic cells. Both lipoplexes and metal magnetic nanoparticles were tested to introduce the vector into cells. The research was focused on finding more efficient methods of transfection. According to analysis on MADLS and gel electrophoresis, aspects playing an important role in conjugation and subsequent transfection were found. Conjugation occurs after only 4 minutes, as evidenced by an increase in zeta potential, but to achieve full conjugation it is necessary to incubate the sample for 20 minutes. Incomplete conjugation to iron nanoparticles resulted in strong carrier-carrier interactions, which formed an unwanted conglomerates. Encapsulation into liposomes with cationic surface treatment was without complications. The success rate of GFP-labeled protein expression after transfection by these methods was calculated to be 95%, resp. 91%. This result is due to low cytotoxicity. However, commercial tested kits on dendritic cells had a success rate below 5% with high cytotoxicity.
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Mechanisms underlying subversion of host immunity by Bordetella pertussis
Klímová, Nela ; Bumba, Ladislav (advisor) ; Černý, Jan (referee) ; Filipp, Dominik (referee)
Bordetella pertussis is a Gram-negative human-adapted pathogen of the respiratory tract and the causative agent of the whooping cough (pertussis) illness. The bacterium produces a number of virulence factors, of which adenylate cyclase toxin (ACT) and pertussis toxin (PT) play important roles in manipulation of host immune response and establishment of the early catarrhal stage of infection. Although the toxins exert their cytotoxic activity by elevation of intracellular cAMP levels, both are distinct from each other in terms of their structures, mechanisms of secretion and cell intoxication, as well as in their ability to modulate the adaptive immune response of the host. The aim of this thesis was to determine the structure- function relationship underlying the mechanism of the Type I secretion system (T1SS)- mediated secretion of ACT and to decipher the immunomodulatory properties of ACT and PT in the course of B. pertussis infection. Integrative structural biology approaches revealed that the RTX domain of ACT consists of a contiguous assembly of five Ca2+ -loaded β-roll blocks, whose co-secretional folding constitute an intramolecular Brownian ratchet that prevents backsliding of the translocating polypeptide in the T1SS conduit, thus accelerating the secretion of ACT from bacterial cells by a...
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In vitro modulation of immune cells for adoptive cellular cancer immunotherapy
Kalkušová, Kateřina ; Smrž, Daniel (advisor) ; Kverka, Miloslav (referee)
Cancer is one of the leading causes of death in developed countries. Its traditional treatment is based on surgical removal of the tumor and metastases, radiotherapy, and chemotherapy. Recently, many new therapy options, including immunotherapy, have been investigated. Cancer immunotherapy seems to be a very promising treatment option as it has experienced many successes in the last few decades. However, there is still a number of patients not responding to today's immunotherapy methods. Adoptive cellular immunotherapy is one of those immunotherapeutic methods. This immunotherapeutic modality uses ex vivo prepared immune cells that participate in anti-tumor responses. Nowadays, most research is focused on the use of T cells, although many other cell types are considered, including dendritic cells. This thesis is focused on the modulation of dendritic cells for adoptive cellular cancer immunotherapy. The aim of the practical part is to evaluate the influence of beta2-microglobulin on the maturation of monocyte-derived dendritic cells. Key words: Adoptive cellular immunotherapy, dendritic cells, beta2-microglobulin, cancer
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Computational models of dendritic cell development
Štráchalová, Sára ; Bílý, Tomáš (advisor) ; Hartman, David (referee)
Computational modelling is gradually establishing its place in biology and medicine as a tool for research of systems and prediction of their behaviour. In this thesis we propose and analyze a simple compartmental model of the immune system and its responses, focusing on dendritic cells. These cells are an important component of the immune system that is crucial for initiation of the specific part of immune response. The analysis focuses on the types of behaviour and stability of the model. We compare our results with already existing basic model of the immune system. 1
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