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Fanconi anemia and pancreatic cancer
Hucl, Tomáš ; Špičák, Julius (advisor) ; Nečas, Emanuel (referee) ; Vodička, Pavel (referee)
Inactivation of the Fanconi anemia (FA) pathway occurs in diverse human tumors including pancreatic cancer and renders those tumors hypersensitive to DNA interstrand-cross-linking agents (ICL). How to treat specificly pancreatic and other cancers harboring FA mutations has recently raised great interest, yet preclinical studies have been hampered by the lack of well-controlled human cancer models. We endogenously disrupted FANCC and FANCG in an adenokarcinoma cell line and observed a typical phenotype of FA pathway deficiency (abrogation of FANCD2 monoubiquitination; chromosomal instability, G2M arrest and decreased proliferation upon treatement with ICL, spontaneous chromosomal breakage). Homozygous deletion was achieved for FANCC and FANCG but not for FANCD2 and BRCA2/FANCD1 in RKO cells, suggesting a detrimental phenotype. It provided direct evidence for the paradoxical assumption that their inactivation could be predominantly selected against in cancer cells. Using high-throughput screening, we assessed the growth of our isogenic FANCC and FANCG cells upon treatment with 880 active drugs and 40 000 diverse compounds. The compound having the stronges effect, named 80136342, had a distinct mechanism of action from that of ICL agents. When applied in combination with ICL agents, 80136342 had at least...
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The influence of gene polymorphisms on the progression of renal insufficiency
Maixnerová, Dita ; Merta, Miroslav (advisor) ; Kohoutová, Milada (referee) ; Nečas, Emanuel (referee)
I worked on the referred to dissertation thesis in the Department of Biology and Human Genetics in cooperation with the Department of Nephrology of General Teaching Hospital and the First School of Medicine at Charles University. I concentrated on the possible influence of gene polymorphisms on the progression of renal insufficiency of IgAN and ADPKD to ESRD. I investigated the gene polymorphisms of Endothelin and Megsin myself and I participated in examinations of other gene polymorphisms. In our study we were concerned with the gene polymorphisms of G198T, T-1370G a 3A/4A ET-1 and we did not find any differences by comparing genotype frequencies among the IgAN groups with normal renal function and ESRD. The haplotype analysis demonstrated the negative influence of GG4A haplotype (defined as G-198, G-1370 and 4 A allele). The association of GG4A haplotype with the progression of chronic glomerulonephritides, especially IgAN, might be explained by shared interaction of all ET-1 polymorphisms. Then we dealt with the research of C2093T, C2180T Megsin gene polymorphisms on the progression of IgAN in Czech patients. No obvious effect of these polymorphisms was found in single-gene or in haplotype analysis. Nevertheless, the Megsin haplotype reconstruction revealed that the TT haplotype (defined as T-2093,...
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The role of stem and progenitor cells in regeneration of hematopoietic tissue
Faltusová, Kateřina ; Nečas, Emanuel (advisor) ; Hofer, Michal (referee) ; Filipp, Dominik (referee)
Tissue regeneration is a complex and highly orchestrated process dependent on cells with the potential to restore structures and functions and on controlling factors from the tissue microenvironment. Hematopoietic tissue has a high ability to regenerate, which is attributed to the presence of stem cells, but the regeneration of severely damaged adult tissue is still only partially understood. Hematopoietic tissue provides a unique opportunity to study tissue regeneration due to its well-established steady- state structure and function, easy accessibility, advanced research methods, and well-defined embryonic, fetal, and adult stages of development. Embryonic/fetal liver hematopoiesis and adult hematopoiesis recovering from damage share the needto expand populations of progenitors and stem cells in parallel with increasing production of mature blood cells. We analyzed adult hematopoiesis in mice subjected to a submyeloablative dose (6 Gy) of gamma radiation, in which only a few cells with reconstituting capacity survived. We targeted the period of regeneration characterized by the renewed massive production of mature blood cells and the ongoing expansion of immature hematopoietic cells. Cells from the top of the hematopoietic hierarchy, hematopoietic stem cells, and multipotent progenitors are almost missing...
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Bleeding after cardiac surgery. Importance of fibrinolysis in off-pump versus on-pump coronary artery bypass surgery
Jareš, Martin ; Vaněk, Tomáš (advisor) ; Dominik, Jan (referee) ; Málek, Jiří (referee) ; Nečas, Emanuel (referee)
The aim of our two prospective randomized studies was to evaluate and compare haemostatic effects of fibrinolytic inhibitors in patients undergoing off-pump coronary artery bypass surgery (OPCAB). The first study evaluated efficacy of tranexamic acid vs. placebo, the second one compared effects of tranexamic acid vs. aprotinin vs. placebo. In addition, the risk of per operative myocardial ischemia and trombotic events was assessed. In the first study postoperative bleeding was significantly lower in the tranexamic acid group compared with the control group. We observed two postoperative myocardial infarctions (one in each group) and any cerebrovacular event or pulmonary embolism. In the second study we conclude that both tranexamic acid and aprotinin significantly reduce the postoperative blood loss in OPCAB patients, and the efficacy of tranexamic acid and aprotinin, respectively, seems to be quite similar. We did not observe any statistically significant difference in the need for allogenic transfusion, although the total number of re-transfused patients was the highest in placebo group. Treated groups did not demonstrate postoperative increase in mean levels of myocardial enzymes, compared with placebo group. Significantly higher mean values of D-dimer were found in placebo group 24 h...
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Effect of gene expression in hyperproliterative skin diseases
Lysá, Barbora ; Arenberger, Petr (advisor) ; Kohoutová, Milada (referee) ; Brdička, Radim (referee) ; Nečas, Emanuel (referee)
The introductory part of this thesis gives insight into selected aspects of the issue deals with hyperproliferative outlining the molecular basis of skin diseases, which are included in addition to psoriasis as well as the characteristic representative of non-melanoma skin cancers. Further analyzes partly exogenous influence of UV radiation on their formation. The actual research work is focused on a group of genes whose expression was studied in actinic keratosis disease in relation to its treatment of topical immunomodulators - imiquimod. Examined genes can be divided into groups of genes modulating the immune response, genes involved in apoptotic processes and the genes playing a potential role in tumor formation.
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Factors influencing the metabolism of homocysteine in selected diseases with complex etiopathogenesis
Veselá, Kamila ; Anděl, Michal (advisor) ; Nečas, Emanuel (referee) ; Rušavý, Zdeněk (referee) ; Poledne, Rudolf (referee)
At present, the great medicine to develop molecular genetic techniques, which significantly help to explain the causes of illnesses. In some diseases the genetic basis is well known. These are primarily contingent on monogenic diseases. Unfortunately, many diseases remain the genetic cause is still unknown. The largest group is the so-called complex diseases, where the origin and development of disease involving both genetic and nongenetic factors. These diseases include atherosclerosis, diabetes mellitus, orofacial clefts, preeclampsia, neural tube defects, and many others. Complex diseases are serious health and social problem in developed countries. Clasification of risk factors is the subject of much attention, because knowledge of these factors offer opportunities for effective prevention and treatment. The aim of my work was to obtain new knowledge of factors affecting homocysteine metabolism with regard to the formation of some complex diseases in the Czech population.
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A sudden cardiac arrest and use of capnometry in cardiopulmonary resuscitation
Pokorná, Milana ; Nečas, Emanuel (advisor) ; Málek, Jiří (referee) ; Bělohlávek, Jan (referee)
The study is focused on advanced life support (ALS) performed by rescue team physicians in an out-of-hospital setting. The first part of the study analyzes diagnostic possibilities and correctness of assumed aetiology of the sudden cardiac arrest during cardiopulmonary resuscitation (CPR) in the field. It introduces an original method of "Crosscheck Tables" and applies this method to 211 cases of CPR provided by physicians of the Emergency Care Service. The study demonstrates that significantly misleading conclusions can result from a global analysis of a set of cases. It stresses importance of diagnostic analyses applied to individual cases. The second part of the study concentrates on changes in PETCO2 level as related to the return of spontaneous circulation (ROSC - Return of Spontaneous Circulation) as opposed to the circulation fully dependent on chest compressions. The study demonstrates that in constantly ventilated patients undergoing CPR in an out-of-hospital setting, PETCO2 is significantly higher (about 10 mmHg) after ROSC than before ROSC. It demonstrates that a sudden increase in PETCO2 exceeding 10 mmHg is likely to indicate the moment of ROSC. The study also support a view that steadily low levels of PETCO2 values (<10 mmHg) indicate a low chance for a successful resuscitation outcome. Key...
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Histomorphological Changes in Normal and Pathological Cartilage Tissues and after their Experimental and Clinical Transplantation
Kaňa, Radim ; Nečas, Emanuel (advisor) ; Slípka, Jaroslav (referee) ; Ludvíková, Marie (referee)
1 Abstract Introduction Autologous transplants of the cartilage tissue from the pinna is commonly used in reconstructive surgery of the nasal skeleton. The present study used animal models to elucidate responses of the auricular cartilage to its damage or transplantation to ectopic sites. Histomorphological analysis of changes observed in auricular cartilage including immunohistochemical study of different isoforms of actin and S-100 proteins was performed. Human articular cartilage prepared by in vitro cultivation using artificial scaffolds was also studied after its transplantation. Aims of the study The aim was to study histological changes and expression of chondrocytic markers (α- SMA and S-100 proteins) in intact, artificially traumatised, or in a human auricular cartilage cultivated in culture medium. An attempt to grow human auricular cartilage chondrocytes implanted in vitro into various types of three dimensional scaffolds aimed at testing chondrocyte survival and phenotype both in the culture and after transplantation to immunodeficient mice. A human auricular cartilage transplanted into the nasal skeleton of patients during a reconstruction surgery should be submitted to a histomorphological examination. Research assumed also comparison of the auricular cartilage responses to a damage,...
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The utility of Toll-like receptor 2 in defining the progenitors of definitive embryonic hematopoiesis
Šplíchalová, Iva ; Filipp, Dominik (advisor) ; Nečas, Emanuel (referee) ; Kalina, Tomáš (referee)
Hematopoiesis is a vital process in which red blood cells and cells of the immune system are formed. It is initiated during early embryonic development when we find hematopoietic progenitors in separate anatomical sites. Embryonic hematopoiesis comprises three successive and partly overlapping waves of progenitors with a different hematopoietic potential. The primary anatomical place where hematopoiesis takes place shortly before the birth is the bone marrow (BM). Since at this time point of development BM is already populated by hematopoietic stem cell (HSCs) progenitors, it becomes also the site of hematopoiesis in adulthood. However, the bone marrow is not the only place where hematopoietic progenitors emerge and develop. The Yolk sac (YS) and the Aorta-Gonad-Mesonephros (AGM) region are the initial sites of the appearance of the three waves of progenitors in the early embryogenesis. These progenitors and their descendants play an indispensable role during the development of an individual. Because there are no specific markers that would unambiguously characterize progenitors of these individual waves, their physical separation and hence also functional characterization is still incomplete. Recent studies have shown that Toll-like receptors (TLRs) are expressed on adult HSCs. The stimulation of...
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