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The cross-talk between opioid and TLR4 receptor signaling systems
Hrdličková, Daniela ; Drastichová, Zdeňka (advisor) ; Březina, Jiří (referee)
Opioids are commonly used in clinical practice to treat acute and chronic pain. They stereoselectively bind to opioid receptors and trigger a signal cascade through inhibitory Gi/o- proteins, resulting in analgetic effects. However, their use can lead to various adverse effects, such as the development of addiction, analgetic tolerance, allodynia, hyperalgesia, constipation, and peripheral immunosuppression. Recent studies have found that opioids also bind non- stereoselectively to Toll-like receptor which belongs to innate immune system receptors recognizing lipopolysacharide. Findings suggest that some of these adverse effects may be a result of this interaction. It induces the expression of proinflammatory mediators in the central nervous system, but under certain conditions, it may also suppress inflammation. Communication between these different receptor systems occurs at multiple levels. Understanding these mechanisms could greatly benefit the development of new therapeutic approaches in opioid analgetic treatment, aiming to minimize their adverse effects, it could also lead to the therapeutic use of the antiinflammatory potential of opioids. Keywords: Opioids, Toll-like receptor 4, cellular signaling, inflammation, lipopolysaccharide, cytokine
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Impact of nanomaterials on mesenchymal stem cells and tissue regeneration
Echalar, Barbora ; Holáň, Vladimír (advisor) ; Hubálek Kalbáčová, Marie (referee) ; Buchtová, Marcela (referee)
Nanomaterials (NMs) are widely used in medicine for their antimicrobial properties. They are part of antibacterial coatings, creams, pharmaceutical vehicles or additives in drugs and other medical products. However, the impact of NMs on human organism is still not completely established. Nanoparticles (NPs) penetrate the cell membrane and enter to intracellular compartments including the nuclei. Different types of NPs could have various side effects on cell functions. These side effects include the damage of stem cells (SCs) or immune cells lead to slower regeneration and impaired wound healing. Therefore, the simultaneous application of NPs during SC therapy could decrease the therapeutic abilities of SCs. One type of SCs tested in clinical therapies nowadays are mesenchymal stem cells (MSCs). Therefore, we studied the impact of metal NPs (i.e. silver, copper oxide, zinc oxide and titanium dioxide) on characteristics and functional properties of mouse MSCs. Additionally, the effect of NPs on the expression of phenotypic markers, metabolic activity, differentiation potential, expression of genes for immunoregulatory molecules and on production of cytokines and growth factors was analyzed. We found that all types of tested NPs had a negative impact on the activity of MSCs and thus could alter tissue...
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Specificita interakcí protein-protein a jejich modulace
Pham, Phuong Ngoc ; Schneider, Bohdan (advisor) ; Damborský, Jiří (referee) ; Vaněk, Ondřej (referee)
(EN) Protein-protein interactions (PPI) have essential roles in life processes, and abnormal PPI are associated with many human diseases. Given their importance, PPI have received increasing attention and became drug targets. However, the design of specific PPI and their modulation is challenging. Cytokine-receptor interactions are especially important in the regulation of the immune system. Interleukin-10 (IL-10) over-production results in excessive immunosuppressive effects, tumor growth and infection. The interaction between interferon gamma receptor 2 (IFN- γR2) and interferon gamma (IFN-γ) leads to activation of downstream signaling pathways but the mechanism of such interaction is elusive. Interleukin-24 (IL-24) is another cytokine that signals through receptors sharing the interleukin-20 receptor two (IL-20R2) subunit and has important roles in autoimmunity and cancer. The aims of this Ph.D. thesis are to study PPI from several aspects emphasizing their specificity. The first goal is to develop a novel protein scaffold and subsequently evolve it into a high-affinity binder specific for human IL-10. The second goal is to understand the structural basis for receptor specificity of human IFN-γ. The third goal is to modulate the binding affinity between human IL- 24 and its receptor IL-20R2 by...
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Lipid emulsions in lonf-term parenteral nutition
Ševela, Stanislav ; Novák, František (advisor) ; Cahová, Monika (referee) ; Lahoda Brodská, Helena (referee)
The combination of intestinal failure and parenteral nutrition is a condition associated with mild chronic inflammation and organ injury, the most significant of which is hepatic injury. This crossover-controlled trial investigated the effects of different intravenous lipid emulsions (IVLE) enriched with increased doses of omega-3 polyunsaturated fatty acids (n-3 PUFA) from fish oil (FO) in patients on home parenteral nutrition (HPNP) for chronic intestinal failure. Twelve patients were consecutively given three different IVLEs, with an FO emulsion added after each IVLE. The study also included twelve paired healthy controls for comparison. The aim of this study was to investigate how long-term administration of these IVLEs affected primarily the erythrocyte and plasma fatty acid (FA) spectrum, as well as the inflammatory response, antioxidant status, markers of liver function and bile acid spectrum in HPNP, and how these changes differed from healthy subjects. We demonstrated successful incorporation of n-3 PUFA into erythrocyte phospholipids and corresponding changes in the plasma FA spectrum, characterized in particular by significant increases in n-3 versus n-6 PUFA, with a decrease in the n-6/n-3 PUFA ratio. We also found a more prolonged effect on these changes with high dose FO...
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Influence of the citrulination of histon proteins on the expression of selected genes in myeloid cells
Tučková, Kristýna ; Dobeš,, Pavel (referee) ; Číž,, Milan (advisor)
Neutrophils are major cell type of innate immunity, that can eliminate pathogens by different mechanisms. One of these mechanisms is called NETosis, which leads to release of decondensed chromatin and citrullinated histone proteins. Citrullination is post-translational modification catalysed by peptidylarginine deiminase (PAD) and causing transformation of possitively charged arginin to neutral citrullin and can change expression of cytokine genes. Concetrations of pro-inflammatory cytokines (IL-8, TNF, IL-1) were measured after activation of PAD4 and induction of citrullination. Calcium ionophore was used to induce citrullinaton, Cl-amidine and TDFA were used as inhibitors. Production of cytokines was assessed by ELISA on protein level and by qPCR on mRNA level. It was found that induction of citrullination led to increased concentrations of IL-8 and IL-1. Elevated gene expression of IL-8 was confirmed on mRNA level. Both inhibitors were able to decrease level of histone H3 citrullination and IL-8 and IL-1 concentrations. Expression of TNF was not detected on protein and mRNA level.
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Monitoring of sublingual allergen immunotherapy using serum cytokine levels
Brzáková, Adéla ; Řezáčová, Vladimíra (advisor) ; Andrýs, Ctirad (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Biological and Medical Sciences Author: Bc. Adéla Brzáková Supervisor: Mgr. Vladimíra Řezáčová, Ph.D. Title of thesis: Monitoring of sublingual allergen immunotherapy using serum cytokine levels Keywords: allergen immunotherapy, cytokine, ELISA, grass pollen allergy Background: The aim of this thesis was to find out whether there are detectable changes in the levels of IFNγ, IL-4 and IL-5 during allergen immunotherapy and whether these cytokines could thus serve as biomarkers of the efficiency of this treatment. Methods: Selected cytokines were determined using Quantikine® HS ELISA Human IFN-γ Immunoassay, Quantikine® HS ELISA Human IL-4 Immunoassay and Quantikine® ELISA Human IL-5 Immunoassay. The monitored population consisted of 32 polinotics, 19 of which were administered allergen immunotherapy and 13 of which constituted the control group. As part of the follow-up, both groups also continuously recorded the severity of their symptoms using a visual analog scale. Results: In the group of treated patients, a decrease in IL-4, IL-5 and IFNγ concentrations was observed during treatment, but no significant differences in the values of these cytokines were found between patients treated with Oralair and control individuals. Using...
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Produkce cytokinů v průběhu mastitid skotu
Tylčová, Natálie
The aim of this work was to detect the pro-inflammatory cytokine TNF-α and the anti-inflammatory IL-10. The experiment included 20 dairy cows at different stages of mastitis, which came from the Nová Ves agricultural cooperative. 80 samples were totaly taken, from all quarters, from each dairy cow, which were divided into three groups. The first group represented healthy dairy cows, without clinical signs of mastitis, with a somatic cell count of up to 100 000, from 5 dairy cows, that is 20 milk samples. The second group consisted of dairy cows with clinical signs of mastitis and somatic cell counts up to 400 000, from 10 cows, that is 40 milk samples. The third group consisted of dairy cows with somatic cell counts of more than 1 000 000, from 5 cows, that is 20 milk samples. These samples were processed in the laboratory of the Department of Animal Morphology, Physiology and Genetics at Mendel University in Brno. The concentration of TNF-α and IL-10 was detected by ELISA. The average concentration of TNF-α in the first group was 138.70 pg/ml, in the second group 133.60 pg/ml, in the third group 95.05 pg/ml. The average concentration of IL-10 in the first group was 9.52 pg/ml, in the second group 11.03 pg/ml, in the third group 8.66 pg/ml.
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Pathophysiological and genetic factors affecting serum uric acid level.
Hasíková, Lenka ; Závada, Jakub (advisor) ; Hrnčíř, Zbyněk (referee) ; Horák, Pavel (referee)
Introduction: Serum uric acid level (SUA) depends on the balance between its production and excretion. SUA is associated with several transmembrane proteins responsible for reabsorption (mainly URAT1 and GLUT9) and secretion (ABCG2) on the apical and basolateral membranes of the proximal tubules in the kidney, and in the case of ABCG2, it also correlates with its significant excretion through the gastrointestinal tract. Gout is a metabolic disease caused by the deposition of urate crystals in the joints and tissues. Chronic hyperuricemia is a primary risk factor for the development of gout; however, gout patients usually have a lower SUA during an acute gout attack than in the intercritical periods. The exact mechanism of this phenomenon is unknown. It has been speculated that the systemic inflammatory response can explain this discrepancy. The aim of the study is to determine whether treatment with specific inhibitors of the proinflammatory cytokine TNF (TNFi) affects SUA in patients with systemic rheumatic disease (SRD), and whether changes in SUA correlate with changes in selected proinflammatory cytokines or with the biomarker of oxidative stress, allantoin. Another aim is to determine the frequency and effect of allelic variants in the ABCG2 urate transporter gene in patients with primary...
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Circadian system as a modulator of neuroinflammation
Kotková, Eliška ; Spišská, Veronika (advisor) ; Dočkal, Tereza (referee)
The circadian system is involved in the regulation of biological rhythms in physiological, behavioural and immune processes. These rhythms can be found in the central nervous system, including the blood-brain barrier, astrocytes, microglia, and the pineal gland, which produces the hormone melatonin. Neuroinflammation is a complex response of the central nervous system to inflammatory stimuli by rhythmic expression of pro-inflammatory and anti-inflammatory mediators or by rhythmic regulation of immune system cells. Studies have examined the influence of genes and proteins of the circadian system, suprachiasmatic nuclei, melatonin, and glial cell rhythms on neuroinflammation. Lipopolysaccharide was used to induce neuroinflammation in these studies. Based on these studies, the effect of melatonin on mikroglia and endothelial cells, and the responses of suprachiasmatic nuclei was evaluted as the most important circadian modulator of neuroinflammation. This thesis describes the basic principles of the circadian system and neuroinflammation, with the last section presenting the modulation of neuroinflammation by the circadian system. Keywords: astrocytes, blood-brain barrier, circadian system, cytokines, immune system, melatonin, microglia, neuroinflammation, suprachiasmatic nuclei
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Cytokine networks and their impact on the immune profile of acute myeloid leukemia (AML) patients
Ptáček, Antonín ; Musil, Jan (advisor) ; Fišer, Karel (referee)
Acute myeloid leukemia (AML) is a malignant hematological disorder characterized by aberrant expansion of blasts in the bone marrow and peripheral blood. The immune system protects the body from leukemia by eliminating transformed cells. However, in AML, the abilities of immune cells are affected both by direct contact between leukemic cells and effector cells, as well as by cytokines, metabolites and other soluble proteins that, together with the cells, form the specific AML microenvironment. The effects of cytokines and other soluble molecules in the AML microenvironment are not sufficiently described yet. This thesis aimed to optimize and implement a multiparametric flow cytometry panel for the measurement of cell populations and to implement multiparametric assays for the analysis of cytokine levels, chemokines and other soluble proteins in plasma. The following goal was to use these methods to characterize the frequency and functional phenotype of cell populations and the levels of the soluble proteins and to describe their influence on disease severity and overall survival of the patients. We also tried to find novel biomarkers of the immune escape of leukemic cells. In patients, we observed a suppressive microenvironment with aberrant levels of soluble receptors and other proteins. This...
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