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Regulatory B lymphocytes and mechanisms of their action
Boháčová, Pavla ; Holáň, Vladimír (advisor) ; Krulová, Magdaléna (referee)
Regulatory B lymphocytes (Bregs) represent a small population of B cells which participate in immunomodulations and in suppression of immune responses. These cells can regulate the immune system by different mechanisms, but the main mechanism of their action is a production of anti-inflammatory cytokine interleukin 10. The regulatory effects of Bregs were described in various models of inflammation, autoimmune diseases, transplantation reactions and in anti-tumor immunity. During autoimmune diseases Bregs function as important regulatory elements which can support remission or repression of the disease. Bregs have also important therapeutic potential in transplantation immunity where they can suppress rejection reactions. However, Bregs can disturb immune surveillance due to their immunosuppressive influence and thus they can attenuate anti-tumor immunity. Therefore, the aim of this thesis is to summarize the recent knowledge about Bregs. The thesis is focused on the mechanisms of Breg action in regulation of immune responses. The imunoregulatory effects of Bregs are described in various models of autoimmune diseases, transplantation immunity and anti-tumor immunity. The recognition of mechanisms of Breg action may have a great impact for their potential use in a clinical setting.
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Role cytokinů ve vývoji a diferenciaci regulačních T buněk
Procházková, Jana ; Holáň, Vladimír (advisor) ; Kovář, Marek (referee) ; Stříž, Ilja (referee)
The development and function of T helper (Th) cells and regulatory T cells (Tregs) are plastic processes that are regulated by cytokines. In our project we first analyzed the effect of different cytokines on the development of induced (i) Tregs. It has been demonstrated that iTregs arise from CD4+ CD25- T cells upon stimulation with alloantigen in the presence of transforming growth factor β (TGF-β). The development of these Tregs and their proliferation were inhibited by interleukin (IL)-4 and IL-12. The aquired results also demonstrated distinct responses of naturally occuring (n) Tregs and iTregs to the regulatory action of IL-4 and an opposite role of IL-4 in maintenance of nTregs and iTregs phenotype. An important role in the induction of T cell subsets may play also mesenchymal stem cells (MSCs) which can, under specific conditions, produce TGF-β and IL-6. Depending on the current production of TGF-β or IL-6, MSCs can qualitatively regulate the ration between Tregs and Th17 cells. Anti-inflammatory Tregs and pro-inflammatory Th17 cells are induced upon stimulation in the presence of TGF-β and TGF-β and IL-6, respectively. In addition to our previous work we studied the role of IL-12 in the development of Tregs and Th17 cells. It was shown that Treg and also Th17 cell differentiation was...
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Pre-clinical model of acute promyelocytic leukemia:/study of the anti-leukemic effect induced by ATRA and DNA vaccination
Pokorná, Kateřina ; Holáň, Vladimír (advisor) ; Stöckbauer, Petr (referee) ; Degos, Laurent (referee)
DOCTORAL THESIS 2012 POKORNA Abstract We have used a well characterized transplantable transgenic mouse model which mimics human acute promyelocytic leukemia (APL), both in its biological characteristics and its response to conventional therapeutic drugs. The aim of our study was to better characterize the efficacy of the combined treatment and to determine molecular markers of clinical outcome. We established a minimal residual disease monitoring based on the high sensitivity of detection of PML-RAR transcripts by polymerase chain reaction (PCR) technology in APL mice. We showed that oncogene-specific PCR-based assays allow, like in patients, the diagnosis, follow-up and prediction of disease evolution. Furthermore, PCR assay was used to assess various tissues and organs for the presence of PML-RAR-positive cells in minimal residual disease free long-term survivors. As expected, majority of mice had no measurable tissue level of PML-RAR demonstrating the efficacy of immunotherapy. However, tracking the oncogene-positive cells reveals for the first time that extramedullary PML-RAR-positive cell reservoirs such as the brain may persist and be involved in the leukemia relapse. We aimed at investigating the immune responses involved in the anti-leukemic effect of the combined immutherapy. To evaluate the...
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Clinical significance of cytokine gene polymorphism
Kolesár, Libor ; Stříž, Ilja (advisor) ; Holáň, Vladimír (referee) ; Mrázek, František (referee)
Univerzita Karlova v Praze Přírodovědecká fakulta Studijní program: Doktorský studijní program v biomedicíně Studijní obor: Imunologie Mgr. Libor Kolesár Klinický význam polymorfismu cytokinových genů Clinical significance of cytokine gene polymorphism Disertační práce Vedoucí závěrečné práce/Školitel: Prof. MUDr. Ilja Stříž, CSc Praha 2012 Abstract The human genome is full of different sequence variants. They are different mainly in size but also in their influence on phenotype. The smallest unit of genetic polymorphism is single nucleotide polymorphism (SNP). SNPs represent a single nucleotide change between two alleles and might affect the gene expression. We have studied SNPs in three distinct fields as: (1) marker of risky patients after the organ transplantation, (2) diagnostic marker of patients with interstitial lung diseases (ILD) or (3) with uterine fibroid (UF). We have come to the following results. Ethnicity or even nationality plays a role in the distribution of genetic polymorphism. This must be absolutely taken into account when one would like to transfer findings of a clinical study from a certain nation or ethnic and applied them to his studied group for the comparative purposes. Our first clinical gene-association study has found that even gene polymorphism of the IL-18 gene may...
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Chemokines of epithelial cells in bronchial asthma
Volfová, Dominika ; Stříž, Ilja (advisor) ; Holáň, Vladimír (referee)
Asthma is an allergic disease caused by adverse reactions to harmless antigens (allergens) and is characterized by the recruitment of eosinophils, Th2 lymphocytes, mast cells and neutrophils into the tissue site of inflammation, to the lungs. This accumulation of leukocytes is mediated by the generation of chemotactic cytokines (chemokines). Chemokines are low molecular weight proteins, functioning by binding to specific receptors on the cell surface. Binding of chemokines and their receptors is highly promiscuous and subsequent activation of effector cells is very heterogeneous, which can often complicate research in this area. However, chemokines and their receptors are important potential therapeutic targets in allergic diseases including asthma, mainly because of their central role in cell activation and inflammation. Chemokines are secreted by cells of the immune system and cells of various tissues of the body. Recently, attention turns to the role of epithelial cells in the pathogenesis of asthma. Bronchial epithelial cells stimulated by antigens produce cytokines and defense molecules used for the amplification of inflammatory processes and regulate the aktivity of effector cells. Impaired cytokine regulation may lead to the development of various lung diseases including asthma. This work...
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Effect of diabetogenic autoantigens on the cytokine production of peripheral blood mononuclear cells from type 1 diabetic patients
Labiková, Jana ; Štechová, Kateřina (advisor) ; Holáň, Vladimír (referee)
5 Abstract Type 1 diabetes (T1D) is a serious organ-specific autoimmune disease characterised by irreversible destruction of pancreatic β-cells by immune system. This process results in an absolute insulin deficiency. Both genetical predisposition and environmental factors influence the development of the disease. β-cell destruction is mediated by cellular components of an immune system. Proinflammatory Th1 response is considered as most pathological. Autoimmune destruction of β-cells can be identified by the detection of specific serum autoantibodies a long time before the T1D clinical onset. Currently, there is no efficient cure available to prevent or at least to delay the destructive insulitis. This diploma thesis describes the influence of synthetic diabetogenic autoantigens GAD65 and IA2 on the cytokine response of peripheral blood mononuclear cells (PBMC) obtained from T1D patients with regards to their antibody profile. The study has been carried out on patients with confirmed T1D diagnosis who tested positive for anti-GAD65 and/or anti-IA2 autoantibodies. By using flow cytometry we measured the cell type ratio in PBMC samples. The cells have been stimulated by three different concentrations of antigens and their IFNγ and IL-17 production has been detected by ELISPOT assay. In the case of both...
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Mesenchymal stem cells and the possibility of their transdifferentiation into insulin producing cells.
Dostálová, Veronika ; Holáň, Vladimír (advisor) ; Čečrdlová, Eva (referee)
Mesenchymal stem cells (MSCs) have been demonstrated in almost all tissues of the body. Their main source is bone marrow and adipose tissue. These cells are multipotent, e.g. they are capable of differentiating into a variety of cell types. They are able to migrate into damaged tissues. Their other relevant property is a specific suppression of imunity. In the body they serve as precursors for specialized cell types and they also participate in formation of specific tissue microenvironment. Their properties represent a great potential in a wide range of clinical therapies. Besides other possible applications they could be used in the therapy of diabetes mellitus type one. During this disease insulin producing -cells are destroyed. MSCs have been used in experimental in vitro and in vivo studies to differentiate into insulin producing cells. However these cells are not able to produce sufficient amounts of insulin to exclude the supportive administration of exogenous insulin. Therefore there is a need for further research in this field of possible therapy.
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Interaction between NKT and myeloid derived suppressor cells and antitumour immunity
Straňavová, Lucia ; Reiniš, Milan (advisor) ; Holáň, Vladimír (referee)
Myeloid- derived suppressor cells (MDSCs) are a heterogeneous population of cells, which plays an important role in the suppression of anti-tumor immune responses. NKT cells represent an additional heterogeneous cell population that plays a crucial role in the regulation of immune responses. It shows that MDSCs and NKT cells may be similar to other populations imunoregulatory cells interact with each other and influence their functions. These interactions are important regulatory factor that may contribute to activation and to suppress anti-tumor immunity. Through interactions with type I NKT cells could differentiate these immunosuppressive MDSCs to immunogenic APC, which could form the basis for immunotherapeutic vaccine. All interactions between the NKT and MDSCs but have a positive effect of imunoregulatory. Interaction between MDSCs and CD4 + NKT cells II. type are immunosuppressive and may subsequently suppress the activity of cytotoxic T-lymphocyte (CTL). In some tumor models it was found that the immunosuppressive nature may also be interactions between MDSCs and type I NKT cells He had, however, alleviate the use of all-trans-retinol acid (ATRA), which induces differentiation of MDSCs.
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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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Targeted differenciation of limbal and mesenchymal stem cells and their therapeutic application
Kuthanová, Hana ; Holáň, Vladimír (advisor) ; Indrová, Marie (referee)
The research of stem cells slowly transfers from the experimental to the preclinical and clinical level. They are in the centre of interest thanks to their potential to treat many of severe injuries and genetically determined diseases. However, the clinical application of these cells has to be based on a basic research of their characteristics and differential potential. Adult stem cells are in organism in minor populations in unique niches. In comparison with embryonic and induced pluripotent stem cells, the adult stem cells have lower differential potential but they also tend less to making teratomas. The therapeutic use of differential and transdifferential potential of limbal and mesenchymal stem cells is described here in more detail with focus on their use in damaged ocular surface treatment. Limbal stem cells are the only source of stem cells for corneal epithelium regeneration in most organisms. Deficiency of these stem cells leads to severe eye disorders even to blindness. Nowadays, a transplantation of allogeneic limbal stem cells or allogeneic limbus is the only chance for patients with total limbal stem cell deficiency. In clinical trials with patients with particular limbal stem cell deficiency, autologous limbal stem cells were successfully transplanted. Mesenchymal stem cells derived...
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