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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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The role of T regulatory cells in kidney transplantation
Urbanová, Anna ; Stříž, Ilja (advisor) ; Zajícová, Alena (referee)
T regulatory lymphocytes (Treg) belong to the CD4+ cell group. They are an essential part of the immunity system. Treg cells prevent from excessive activation of effector T cells and they keep the tolerance to the tissues of the body. They have high expression of CD25 and the transcription factor Foxp3. We distinguish two basic populations of Treg cells: natural Treg cells (nTreg) created in the thym and representing 5-10 % of all CD4+ cells, and induced Treg cells (iTreg), created from naive CD4+ cells in the periphery.Their regulatory effect is well-known, therefore using of Treg cells could bring about a huge treatment potential for patients with a transplantated kidney. Healthy people and patients tolerant to the transplantated kidney show higher occurance of circulating Treg cells and the Treg cells present in the graft unlike patients with chronical rejection. The tolerance is cancelled with the damage of CD4+ CD25+ cells.For a graft acceptance it is necessary to treat the patient after the transplantation with immunosuppressive medicaments resulting in suppression of immunity reaction against the graft. Their disadvantages are side effects often resulting in the patient's death. Moreover they often have a negative impact on survival and expansion of Treg cells. The analysis of flow cytometry has...
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Cytokines in the effector function of regulatory T cells
Zadražil, Zdeněk ; Holáň, Vladimír (advisor) ; Stříž, Ilja (referee)
Regulatory T cells (Treg) are an important control mechanism within the Immune system (IS). Tregs prevent overactivation of effector T cells or autoreactive cells from invading organism-derived tissues. Treg are characterised by expression of surface molecules, CD4, CD25 and by an intracellular transcription factor forkhead box protein 3 (FoxP3). There are two basic populations of Treg, naturally occuring Treg (nTreg) developing in the thymus and induced Treg (iTreg) rising from CD4+ T cells in periphery, which are also precursors for T helper cells. In spite of an outgoing intensive research, there is still no clear clue which mechanisms are used by Treg to inhibit other effector cells. First in vitro experiments showed, that those mechanisms are of a contact dependent manner and do not use secreted molecules. But in vivo experiments showed the exact opposite. Those studies showed that secretory molecules, such as interleukin (IL)-10, IL-35 or transforming growth factor beta (TGF-β), are important in the effectory phase of Treg. Since the first experiments other distinct mechanisms of supression by Treg cells have been discovered. Those mechanisms seem to be important only in particular situations, particular cell assays or with using of specific experimental models. The reasons for this...
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Immunoprofiling in patients with HPV-associated and non-associated head and neck squamous cell carcinoma
Lukešová, Eva ; Klozar, Jan (advisor) ; Šlapák, Ivo (referee) ; Laco, Jan (referee)
Head and neck squamous cell carcinomas (HNSCC) remain a significant cause of morbidity worldwide, with approximately 550,000 new cases diagnosed each year. The main etiological factors include smoking and alcohol consumption. The incidence of non-oropharyngeal HNSCC is gradually decreasing while the incidence of squamous cell oropharyngeal carcinoma (OPSCC) is still on the rise. This increasing incidence can be most likely attributed to an increasing prevalence of human papillomavirus (HPV) infection. From the clinical point of view the most significant fact is that patients with HPV positive OPSCC have better prognosis. HNSCC is linked to an alteration in the immune system. Only a limited number of studies have correlated both the immunological parameters and HPV status with patient prognosis. Therefore, we focused on the research of the immunological profile of patients with HNSCC of viral and non-viral etiology. In our study, 110 patients with HNSCC were enrolled. They were divided into HPV-positive and HPV-negative groups based on the expression of HPV 16 E6 mRNA detected in the tumor tissue. Basic lymphocyte subpopulations (CD3+, CD4+ CD25+ Treg, CD4+ CD25+ FoxP3 Treg, CD4+, CD8+, CD19, and CD3- CD16+ CD56+ cells) were determined by flow cytometry in the peripheral blood (PB). We observed...
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Immune response and tumor microenvironment in the treatment with polymer cytotoxic drugs
Malátová, Iva ; Šírová, Milada (advisor) ; Stollinová Šromová, Lucie (referee)
Chemotherapy is still one of the most widely used anticancer therapies. It is mostly about inhibiting the proliferation of rapidly dividing cells, so it is not selective for tumor cells. As a result, many undesirable side effects are associated with chemotherapy. The disadvantageous properties of chemotherapeutics can be largely eliminated by using conjugates of polymers with low molecular weight drugs. An example of such a conjugate is a doxorubicin-linked HPMA polymer. In addition to the properties obtained by polymer binding, such as achieving solubility in aqueous solutions, reducing systemic toxicity, increasing the maximum tolerated dose, or passive targeting by the EPR effect, the fact that doxorubicin induces immunogenic cell death is used in therapy with this drug. It has already been shown that after complete cure of the experimental mice with polymeric conjugates of HPMA with doxorubicin, some mice develop long-term resistance to re-inoculation with a lethal dose of tumor cells. Resistance is specific to the particular line of tumor cells from which the mouse was cured, and CD8+ cytotoxic T cells and IFNγ play an important role. In this work, we monitored changes in the proportion of immune populations and their activation markers after treatment with HPMA-based polymers with doxorubicin...
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Immunoregulatory characteristics of immune cells of children of allergic and non-allergic mothers and the possibility of their modulation with probiotic E. coli strain O83:K24:H31
Černý, Viktor ; Hrdý, Jiří (advisor) ; Tlaskalová - Hogenová, Helena (referee) ; Černá, Marie (referee)
Due to high incidence, medical and socioeconomic burden and impact on individual quality of life and productivity, allergic disorders are a crucial issue for 21st century immunology. Much still remains to be elucidated, particularly regarding the very early processes in allergy development. In order to introduce timely, effective preventive measures, novel, more reliable predictive factors of allergy risk also need to be established. Dysregulation of proper balance between the branches of immune response, particularly unwarranted dominance of Th2, is the underlying cause of allergy. After birth, new immune balance needs to be established to prepare the neonate for adequate reactivity towards newly encountered environmental stimuli. Regulatory T cells (Treg) play a central role in finely setting this balance and inducing tolerance towards harmless environmental antigens, including allergens. Interactions with external factors, most importantly microbiota, modulate this process during the early postnatal "window of opportunity." Analysis of cord blood Treg of children of allergic mothers uncovered decreased presence of function-associated surface markers and lower production of IL-10. Furthermore, decreased proportion of Helios- induced Treg was observed in children with higher risk of allergy....
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Different capacity of in vitro generated monocyte-derived dendritic cells of newborns of healthy and allergic mothers to prime immune responses
Súkeníková, Lenka ; Hrdý, Jiří (advisor) ; Javorková, Eliška (referee)
(EN) Reduced microbial stimulation of an immature neonatal immune system can lead to a poor balance adjustment of immune responses, thus contributing to the development of allergic diseases, whose incidence continues to rise. One of the promising precautionary measures seems to be an early preventive administration of probiotic bacteria to pregnant or nursing mothers, or to newborns. Previous works have described a beneficial effect of Escherichia coli O83:K24:H31 (E. coli O83) in the prevention of allergic diseases. In order to contribute to the clarification of E. coli O83 effects on the neonatal immune system, its immune- modulating properties were tested in vitro on umbilical cord blood cells. The ability of E. coli O83 to support the maturation of in vitro-derived dendritic cells from cord blood precursors (moDCs) of the children of healthy (children with a relatively low risk of allergy) and allergic (children at a relatively high risk of developing allergies) mothers was tracked by flow cytometry, qPCR and ELISA. Probiotic bacteria-stimulated moDCs were subsequently cultured with autologous naive CD4+ T lymphocytes and immune response polarization was also characterised by flow cytometry, qPCR, and ELISA. It was evident from the results that E. coli O83 promoted moDCs maturation. The presence of...
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The dynamics of T regulatory lymphocytes generation during pregnancy
Kopřivová, Helena ; Krulová, Magdaléna (advisor) ; Frič, Jan (referee)
Regulatory T cells (Tregs) are CD 4+ T lymphocytes with regulatory and reparative properties. Tregs are functionally and differentially controlled by Foxp3 transcription factor, and they are considered a key factor inducing feto-maternal tolerance. Disruption of this unique type of time-limited immune tolerance leads to pregnancy complications including miscarriages, intrauterine growth retardation of the fetus, and premature delivery. Tregs' production and functional properties are influenced by the cytokine environment, especially by TGF-β1, IL-2, and retinoic acid. These pleiotropic factors are involved in the differentiation of tolerogenic populations of decidual cells. In the present work, a detailed immunophenotyping of Tregs was performed, and a determination of serum concentrations of TGF-β1 during physiological pregnancy was realized. The examined subpopulations of Tregs have shown different signs or markers of their functional and maturity properties. In some subpopulations, apparent changes in Treg counts were observed during pregnancy. Moreover, typical development was observed for TGF-β1 serum levels. The main goal of the current work was an examination of differences in Treg subpopulation counts in women with premature deliveries compared to females with full term births....
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Immunoprofiling in patients with HPV-associated and non-associated head and neck squamous cell carcinoma
Lukešová, Eva ; Klozar, Jan (advisor) ; Šlapák, Ivo (referee) ; Laco, Jan (referee)
Head and neck squamous cell carcinomas (HNSCC) remain a significant cause of morbidity worldwide, with approximately 550,000 new cases diagnosed each year. The main etiological factors include smoking and alcohol consumption. The incidence of non-oropharyngeal HNSCC is gradually decreasing while the incidence of squamous cell oropharyngeal carcinoma (OPSCC) is still on the rise. This increasing incidence can be most likely attributed to an increasing prevalence of human papillomavirus (HPV) infection. From the clinical point of view the most significant fact is that patients with HPV positive OPSCC have better prognosis. HNSCC is linked to an alteration in the immune system. Only a limited number of studies have correlated both the immunological parameters and HPV status with patient prognosis. Therefore, we focused on the research of the immunological profile of patients with HNSCC of viral and non-viral etiology. In our study, 110 patients with HNSCC were enrolled. They were divided into HPV-positive and HPV-negative groups based on the expression of HPV 16 E6 mRNA detected in the tumor tissue. Basic lymphocyte subpopulations (CD3+, CD4+ CD25+ Treg, CD4+ CD25+ FoxP3 Treg, CD4+, CD8+, CD19, and CD3- CD16+ CD56+ cells) were determined by flow cytometry in the peripheral blood (PB). We observed...
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Cytokines in the effector function of regulatory T cells
Zadražil, Zdeněk ; Holáň, Vladimír (advisor) ; Stříž, Ilja (referee)
Regulatory T cells (Treg) are an important control mechanism within the Immune system (IS). Tregs prevent overactivation of effector T cells or autoreactive cells from invading organism-derived tissues. Treg are characterised by expression of surface molecules, CD4, CD25 and by an intracellular transcription factor forkhead box protein 3 (FoxP3). There are two basic populations of Treg, naturally occuring Treg (nTreg) developing in the thymus and induced Treg (iTreg) rising from CD4+ T cells in periphery, which are also precursors for T helper cells. In spite of an outgoing intensive research, there is still no clear clue which mechanisms are used by Treg to inhibit other effector cells. First in vitro experiments showed, that those mechanisms are of a contact dependent manner and do not use secreted molecules. But in vivo experiments showed the exact opposite. Those studies showed that secretory molecules, such as interleukin (IL)-10, IL-35 or transforming growth factor beta (TGF-β), are important in the effectory phase of Treg. Since the first experiments other distinct mechanisms of supression by Treg cells have been discovered. Those mechanisms seem to be important only in particular situations, particular cell assays or with using of specific experimental models. The reasons for this...
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