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Úloha vrozené a získané imunity v imunoterapii melanomu a pankreatického adenokarcinomu
NEDBALOVÁ, Pavla
This study examines the role of innate and adaptive immunity in the immunotherapy based on the combination of the ligands stimulating phagocytosis anchored in the tumour cells membrane and the mixture of TLR agonists. This immunotherapy is primarily focused on the innate immunity activation and induces strong inflammatory infiltration, which neutrophils and NK cells are part of. Therefore, the next aim of this study was to evaluate the anti-tumour activity of neutrophils and NK cells. For examination malignant melanoma and pancreatic adenocarcinoma mouse tumour models were used.
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Preparation of recombinant forms of the extracellular part of mouse leukocyte receptors from NKR-P1 family.
Adámek, David
Mouse NK cell receptors belonging to NKR-P1 family plays role in activation, inhibition and cytokine secretion by these cells. Aim of this thesis is preparation of extracellular parts of C57BL/6 mouse strain activating receptors mNKR-P1A and mNKR-P1C. Production vectors with coding sequences of both proteins were prepared. Next, optimization of production in E. coli was done and appropriate in vitro refolding and purification protocol were developed. Purified proteins were characterized by mass spectrometry and labeled by a fluorescent dye. Primary screening for potential ligand was performed. Further work will involve structural characterization of the receptors and identification of their ligands. These data may help to clarify the function of NK cells.
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Preparation of recombinant forms of the extracellular part of mouse leukocyte receptors from NKR-P1 family.
Adámek, David ; Man, Petr (advisor) ; Brdička, Tomáš (referee)
Mouse NK cell receptors belonging to NKR-P1 family plays role in activation, inhibition and cytokine secretion by these cells. Aim of this thesis is preparation of extracellular parts of C57BL/6 mouse strain activating receptors mNKR-P1A and mNKR-P1C. Production vectors with coding sequences of both proteins were prepared. Next, optimization of production in E. coli was done and appropriate in vitro refolding and purification protocol were developed. Purified proteins were characterized by mass spectrometry and labeled by a fluorescent dye. Primary screening for potential ligand was performed. Further work will involve structural characterization of the receptors and identification of their ligands. These data may help to clarify the function of NK cells.
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Potentiation of the biological activity of IL-2 and IL-15 in vivo
Votavová, Petra ; Kovář, Marek (advisor) ; Reiniš, Milan (referee)
5 Abstract Interleukin-2 possesses strong stimulatory activity for activated T and NK cells and thus it is an attractive molecule for immunotherapy. However, its unfavourable pharmacological properties, extremely short half-life and severe toxicities associated with high-dose IL-2 are the most serious and limiting drawbacks. Moreover, IL-2 has been also implicated in the homeostasis of T regulatory cells where it plays a decisive role as an essential growth factor of these cells. Several different approaches to improve the therapeutic potential of IL-2 have been studied. Recently described IL-2/anti-IL-2 mAb immunocomplexes which show much higher and selective biological activity in contrast with free IL-2 in vivo are probably the most promising of them. In this study, we compared the biological activity of free IL-2 with IL-2/anti-IL-2 mAb immunocomplexes in order to demonstrate their benefits over free IL-2. We also demonstrated that IL-2/anti-IL-2 mAb immunocomplexes possess noticeable antitumor activity in two syngeneic mouse tumor models, namely EL4 T lymphoma and B16F10 melanoma, if administered early in tumor progression. Therefore, we justified potential use of IL-2/anti-IL-2 mAb immunocomplexes in tumor immunotherapy. We covalently conjugated IL-2 to synthetic semitelechelic polymeric carrier based...
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Expression of the recombinant extracellular parts of leukocyte receptors AICL a NKR-P1CBALB
Čonka, Martin ; Novák, Petr (advisor) ; Cebecauer, Marek (referee)
v anglickém jazyce NK cells represent a population of lymphocytes which are able to kill certain tumor cells or virally infected cells. The subject of the diploma thesis is a mouse NK cell receptor mNKR- P1CBALB and a human leukocyte receptor hAICL. The mNKR-P1CBALB belongs to the activating receptors and is able to activate the cytotoxic functions of NK cells. The hAICL receptor is a ligand to the NKp80 which is an activating receptor of NK cells. Interaction between these two proteins leads to the activation of effector functions of NK cells as well. The aim of this work was the preparation of the recombinant extracellular parts of receptors mNKR-P1CBALB and hAICL, the optimalization of their in vitro refolding and the characterization of proteins using mass spectrometry. The proteins samples will be used for further structural study of the extracellular parts of these leukocytes receptors.
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Proteomic characterization of human NK cell membrane microdomains
Kádek, Alan ; Man, Petr (advisor) ; Martínková, Markéta (referee)
Proteomic characterization of human NK cell membrane microdomains. (in Czech) Bc. Alan Kádek (Department of Biochemistry, Faculty of Science, Charles University in Prague, Czech Republic) Natural killer (NK) cells are one of the important components of innate immune system. Their main function is to fight against tumors, virally infected or otherwise malformed cells. Plasma membranes of NK cells contain regions with specific lipid composition compared to the surrounding membrane (called membrane microdomains or rafts). Because of their lipid composition, microdomains preferentially accommodate some immunologically relevant proteins and play a role during cellular polarization and signalization. Characteristic feature of membrane microdomains is their partial resistance to solubilization by mild non-ionogenic detergents. In this thesis, microdomains were isolated in a detergent-resistant membrane fraction (DRM) from human NK-92MI cell line and from NK cells immunomagnetically enriched from peripheral blood of non-leukemic donors. For the isolation, Triton X-100 or Brij-98 detergent solubilization and ultracentrifugation in a sucrose density gradient was used. Protein composition of isolated DRMs was analyzed by mass spectrometry employing an LC-MALDI-TOF/TOF method. Protein lists generated in these...
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Immunologic profile of experimental autoimmune encephalomyelitis
Novosádová, Iva ; Fišerová, Anna (advisor) ; Zajícová, Alena (referee)
5 Anglický abstrakt Experimental autoimmune encephalomyelitis (EAE) is widely accepted as a murine model of human multiple sclerosis autoimmune disease. Murine EAE is usually actively induced by immunization with a suitable myelin antigen. Following immunization, CD4+ T helper lymphocytes Th1 and Th17 accumulate in the nervous tissue and via the production of cytokines, they mediate an inflammatory reaction and the subsequent destruction of myelin. The main goal of this study was the induction of EAE with clinically observable symptoms and the observation of changes in the counts and phenotypes of cells, mainly NK and T cells. NK cells express a wide range of inhibitory and activation receptors from the C-lectin-like receptor superfamily. The specific ligand of the activating NKR-P1C isoform is still unknown and thus this receptor's involvement in EAE was also observed. Another goal was the use of medication with regard to the disease progress improvement. For the purposes of this study, two inbred murine strains with distinct NKR-P1 surface expression were used - the SJL/J strain (expressing inhibitory NKR-P1B) and C57BL/6 (expression activating NKR-P1C). SJL mice elicited a relapse-remitting of EAE, while C57BL/6 had chronic EAE. Both mouse strains exerted changes in the counts of NK...
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Carbohydrate dimers in tumor immunotherapy
Krupová, Monika ; Bezouška, Karel (advisor) ; Ryšlavá, Helena (referee)
Carbohydrate dimers in tumor immunotherapy Monika Krupová (Department of Biochemistry, Faculty of Science, Charles University in Prague) One possible approach to tumor immunotherapy is an activation of killer lymphocytes through specific ligands for their surface receptors. CD69 is a molecule greatly widespread among various cells of haematopoietic origin. Since the physiological ligand for this receptor is still unknown, ligand mimetics are used for modulation of its activity. The mimetics tested in this work are based on monomeric or oligomeric carbohydrated attached through two different chemical groups to the central linker of varying length, giving rise to thiourea and triazole series. The ability to precipitate soluble NKR-P1 and CD69 receptors was evaluated in precipitation assays and the optimal length of the linker for NKR-P1 receptor was found to be decyl. On the other hand, cross-linking of CD69 is not so dependent on the length of the linker. The aim of this work was to describe in vitro effect of the tested compounds on cellular signalization, natural killing of leukemic cell lines and activation-induced apoptosis. Compounds of triazole series containing two disaccharides (GalNAc β1→4 GlcNAc) linked by a linker were found to have the strongest effect on the production of...
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The role of LIF gene mutations and the relative distribution of NK cells, NKT and T lymphocytes in follicular fluid and blood of women with different history of infertility
Křížan, Jiří ; Šíma, Petr (advisor) ; Jonáková, Věra (referee) ; Kročová, Zuzana (referee)
Charles University in Prague Faculty of Natural Sciences Summary of Ph.D. thesis The role of LIF gene mutations and the relative distribution of NK cells, NKT and T lymphocytes in follicular fluid and blood of women with different history of infertility Jiří Křížan Prague 2010 1 | P a g e Doctoral degree programs in biomedicine Charles University in Prague and the Academy of Sciences of the Czech Republic Programme: Biomedicine Chairman of the Subject Board: Doc. RNDr. Vladimír Holáň, Dr.Sc. Place of study: Institute of Microbiology, v.v.i., Academy of the Czech Sciences Vídeňská 1083, 142 20 Prague 4 phone: +420 296 442 318 Autor: Mgr. Jiří Křížan Supervisor: RNDr. Petr Šíma, CSc. The dissertation can be found at Dean's Office Faculty of Charles University in Prague 2 | P a g e CONTENTS Contents 2 Summary 3 1. Introduction 5 2. Hypotheses and aims 6 3. Material and methods 7 Material: 7 Methods: 8 4. Results 10 5. Discussion 12 6. Conclusion 15 7. References 16 Bibliography of Autor: 18 1. papers in extenso (thesis background) 18 2. papers in extenso (without regard to thesis) 19 3 | P a g e SUMMARY The aim of the dissertation thesis "The role of LIF gene mutations and the relative distribution of NK cells, NKT, and T lymphocytes in follicular fluid and blood of women with different history of infertility"...
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