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Adipose tissue functional changes during postnatal development and impact of high-fat diet feeding on adipose and liver tissue
Hájková, Simona ; Bardová, Kristina (advisor) ; Hlaváčková, Markéta (referee)
Caloric intake increased over a long period of time may induce the development of obesity, causing so-called low-grade inflammation. The organism responses to the inflammation by the activation and production of cellular components of the immune system, such as macrophages or proinflammatory cytokines. The adipose tissue itself is involved in the production of bioactive molecules, including leptin and adiponectin. Increased concentration of proinflammatory cytokines can lead to a dysfunction of important metabolic pathways and impair organ's function. For the purpose of closer knowledge of the etiology of obesity and its metabolic complications, inbred strains of mice with different genetic backround are most commonly used. We aimed to define the impact of high-fat diet (HFD) on adipose and liver tissue of C57BL/6J and A/J murine strains with a different susceptibility to diet-induced obesity. We focused on description of morphological and functional changes of adipose tissue and on the evaluation of plasma leptin and adiponectin levels of mice in the early postnatal development. Next, we measured the expression of leptin mRNA in four tissues. In this study we described how the increased caloric intake leads to increased triacylglycerides (TAG) storage in the liver and to a higher inflammatory...
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The role and function of stromal enzymes in keratoconus pathogenesis
Ďuďáková, Ľubica ; Jirsová, Kateřina (advisor) ; Svozílková, Petra (referee) ; Ardan, Taras (referee)
Lubica Dudakova Doctoral Thesis ABSTRACT Keratoconus (KC) is a non-inflammatory disease of the cornea, in which ectasia and thinning occur probably due to defects in the collagen fibers binding. It is one of the most common indications for corneal transplantation. KC is a complex disorder with the involvement of both genetic and environmental factors; however the exact pathogenic mechanisms leading to the disease development have not been elucidated. The main aim of our work was to compare the presence and enzyme activity of cross- linking enzymes lysyl oxidases (LOX and LOX-like enzymes), in control human cornea samples and explanted cornea gained from patients with KC. We also focused on diseases previously described to be associated with KC with the aim to identify common signs among them. Furthermore, we replicated association of single nucleotide polymorphisms (SNPs) in LOX and hepatocyte growth factor (HGF) with KC risk. We attempted to link all pathophysiological disturbances observed in KC into one common pathway. We have used a wide spectrum of methods (cell culturing, immunohisto- and immunocytochemistry, microscopy, fluorimetric enzyme activity measurement, genotyping and direct sequencing, statistical analysis). We demonstrated the presence of entire family of LOX enzymes in control and in KC...
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Investigation of proteolytic enzymes expression in different tissues at the transgenic animal model of Huntington disease by means of biochemical and immunohistochemical methods
Kocurová, Gabriela ; Dršata, Jaroslav (advisor) ; Novotná, Eva (referee)
Charles University in Prague Faculty of Pharmacy in Hradec Králové Department of Biochemical Sciences Candidate: Bc. Gabriela Kocurová Supervisor: Prof. MUDr. Jaroslav Dršata, CSc. Title of diploma thesis: Investigation of proteolytic enzymes expression in different tissues at the transgenic animal model of Huntington's disease by means of biochemical and immunohistochemical methods Background: Huntington's disease (HD) is a neurodegenerative disorder that is caused by an expansion of a polyglutamine (polyQ) domain in the huntingtin (Htt) protein. Because it is known that mutant Htt and especially its small proteolytic fragments are toxic to neurons (particularly those in the striatum and cortex), it has been suggested that proteolysis of mutant huntingtin (mHtt) might play an important role in HD pathogenesis. Therefore, the aim of the present study was to examine the expression of endogenous and mtHtt and possible participation of the proteolytic enzymes from the group of caspases, matrix metalloproteinases (MMPs), kallikreins (KLKs) and calpains in HD pathology of brain tissue. Methods: In this study we used WT and TgHD minipigs for N-terminal part of the human mtHtt (548aaHTT-145Q, both F2 generation, age 36 months; F3 generation, age 48 months in additional experiment), R6/2 mice were used as...
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Significance of HNF-1B in different types of carcinomas and non-neoplastic lesions of the female genital system at the level of protein expression, epigenetic and genetic changes
Němejcová, Kristýna ; Dundr, Pavel (advisor) ; Zámečník, Josef (referee) ; Nenutil, Rudolf (referee)
Introduction HNF-1β is a transcription factor that plays a crucial role in the ontogenesis, regulates expression of multiple genes involved in cell cycle modulation and seems to be involved in cancerogenesis of various tumors. HNF-1β protein is coded by the HNF1B gene. Genetic and epigenetic changes of HNF1B play role in tumorigenesis and these changes can be accompanied by loss of expression or increased protein expression as detected by immunohistochemistry. In gynecopathology, expression of HNF-1β was considered as specific marker of clear cell carcinomas of the ovary and endometrium. However, more recent studies described HNF-1β expression also in tumors of other histogenesis. Aims: Our study focused on the immunohistochemical and molecular analysis of HNF1B in the normal tissue, various types of tumors and non-neoplastic lesions of the female genital tract. The goals of our study were: 1. Analysis of HNF-1β expression in cervical carcinomas. 2 Analysis of HNF-1β expression in endometrial carcinomas and non-neoplastic tissues of the female genital tract. 3. Analysis of epigenetic and genetic changes of HNF1B in endometrioid carcinomas and ovarian clear cell carcinomas. 4. Comprehensive analysis of atypical polypoid adenomyomas. Material and methods: A total of 574 samples including 399...
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Methods of Molecular Biology in Predictive Diagnostics of Her/neu in Stomach Cancer
Důra, Miroslav ; Staněk, Libor (advisor) ; Gemperle, Jakub (referee)
Stomach cancer is a malignant neoplastic disease, which is caused by malignant transformation of the gastric mucosal epithelium. Elderly patients are more frequently affected, the majority of the patients are males. Stomach cancer has some typical features like minor symptomatology and early metastasis founding. Nowadays, a mild decrease of incidence is registered. Treatment of stomach cancer depends on its location and stage. Because of its relative chemo- and radioresistance, surgical resection is the only one potentially curable method, although there is high number of recurrences. An amplification and overexpression of HER2 receptor is detected in approximately 10 - 20 % patients with stomach cancer. This overexpression correlates with worse prognosis of disease. Treatment with a monoclonal antibody named trastuzumab, Herceptin® could significantly elongate their life. Trastuzumab is now widely and successfully used for the treatment of female patients with breast carcinoma. Detection of HER2 amplification is there needed. Treatment with trastuzumab can be useful only when HER2 positivity of tumor is determined. This determination should be made by a reliable test with sufficient sensitivity and specificity. The aim of this work is to summarize the methods of molecular biology that are used in...
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Relation of tumor genotype and phenotype to diagnosis, prognosis and prediction of colorectal cancer
Pitule, Pavel ; Králíčková, Milena (advisor) ; Vožeh, František (referee) ; Slabý, Ondřej (referee)
Colorectal cancer is one of the most common type of malignity. Despite of the existence of numerous studies focused on this carcinoma, there are still many unknown features regarding its diagnosis, treatment or prognosis. In the thesis we focused on the identification of novel prognostics markers that could be useful for the stratification of patients based on the disease outcomes. In the first study we immunohistochemically assessed expression of two proteins associated with cancer stem cells in the samples of primary colorectal cancer and matched liver metastasis. Goal of the study was to evaluate relation among expression of CD44 and CD133 and overall survival and disease free interval in our set of patients. We observed that increased ratio of CD133 positive compared to CD133 negative tumor glands resulted in longer disease free interval, finding which is opposite to the general view on the CD133 role in the cancer development. Our hypothesis is that we analyzed confined group of patients and followed a bit different goal, where we measured ratio between positive and negative glands in the view-field and not the intensity of staining as the previous studies did. Our second study was focused on the transcriptional analysis of the selected set of twelve genes using frozen samples from colorectal...
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Contribution to radiodiagnostics and to treatment of chosen pathological lesions of femur in childhood and in adults
Horák, Martin ; Poučková, Pavla (advisor) ; Marek, Josef (referee) ; Vítek, František (referee)
7 Abstract (AJ) Introduction Radiology examination using specialized modern imaging methods, including CT and MRI, is essential in the diagnosis of congenital and acquired diseases of the musculoskeletal system. The first part of the dissertation deals with certain congenital defects of the short femur, known in the literature as proximal femoral focal deficiency (PFFD). This part summarizes our experience with the radiological findings in the preoperative and postoperative period, with the main attention to the vascular supply to the affected area. The second part of the presentation deals with some aspects of autologous chondrocyte transplantation fixed at two different carriers implanted into post-traumatic articular cartilage defects of the distal femur. Radiological findings are evaluated in the relation to the histopathological findings. Objectives The first part of the study after the distribution of patients with PFFD by current commonly used radiographic classification sets the objective in the extent of scans of the hip joints to specify diagnosis PFFD in each patient and to evaluate in detail changes in the area of disability, especially a course of blood vessels. The evaluation of the radiation burden of repeated X-ray measurements was done with respect to the age of the patients. Tissue samples...
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Bilirubin influence on the progression of inflammatory bowel disease.
Patková, Anna ; Nachtigal, Petr (advisor) ; Hronek, Miloslav (referee)
Charles University in Prague Faculty of Pharmacy in Hradec Králové Department of Biological and Medical Sciences Bilirubin influence on the progression of inflammatory bowel disease Diploma thesis Anna Patková Supervisor: doc. PharmDr. Petr Nachtigal, Ph.D. Background: Inflammatory bowel diseases, including Crohn's disease and ulcerative colitis, are chronic inflammatory disorders of the gut caused by an interaction of genetic and environmental factors. It is thought that tissue damage is also partly caused by an oxidative stress. Heme oxygenase-1 and bilirubin are strong antioxidants and both of them provide an anti-inflammatory effect in various tissues. The aim of this diploma thesis was to detect changes of expression of HO-I in the large intestine of normobilirubinemic and hyperbilirubinemic rats after the induction of acute or chronic experimental colitis. Methods: We used Gunn rats with hereditary defect of UDP-glucuronyltransferase, which causes hyperbilirubinemia. The control group of animals was made up of heterozygous littermates of the Gunn rats, which have normal serum bilirubin levels. All animals were treated by dextran sulfate sodium in order to induce an experimental colitis. Rats were divided into two groups. Each of them contained hyperbilirubinemic and normobilirubinemic...
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Analysis of embryotoxic effect of hydrocortisone using chick embryotoxicity screening test (CHEST).
Janíková, Michaela ; Peterka, Miroslav (advisor) ; Hovořáková, Mária (referee)
Cleft lip is one of the most common human birth deffects. Its etiopathogenesis is multifactorial and many aspects of its occurrence remain unknown in the fields of both genetics and teratology. One of the set of known negative external factors causing cleft lip is chemical hydrocortisone. Its effect on cell proliferation is highly heterogeneous and depends on attributes of a specific cell population. In this work we studied the cleft beak origin after the hydrocortisone treatment on the basis of Chick Embryotoxicity Screening Test (CHEST). Our main aim was to detect cell cycle changes in the chick frontonasal process after hydrocortisone injection via flow cytometry analysis. Hydrocortisone caused S phase arrest within a minor subpopulation of highly granular cells with specific cell cycle. This sensitive subpopulation was localized in the areas of previously defined proliferative centers within the frontonasal process using immunohistochemistry of frozen sections. Quantitative analysis of cells in these areas revealed significant decrease of M phase portion in the hydrocortisone treated samples in comparison with the control samples. The TUNEL staining of histological sections was used to determine the apoptotic rate in the frontonasal process. The comparison between the control and the...
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Localization of individual opioid receptor subtypes in CNS
Lišháková, Michaela ; Hejnová, Lucie (advisor) ; Bendová, Zdeňka (referee)
In last decades, the research focused on opioid receptors has been intensively conducted in order to determine their role in various homeostatic functions, control of movement, neurotransmission and drug addiction. An important factor determining the pharmacologic role of opioid receptors is their distribution within the brain regions as well as at the cellular level. Over the past 40 years, a great deal of information concerning their distribution in the central nervous system has been collected, allowing us to determine their localization and mechanisms of their action. In general, results obtained from previous studies are in agreement. However, there are some inconsistencies that impede the accurate determination of receptor distribution and need to be clarified. This paper aims to summarize results of the previously published studies observing the localization of opioid receptors in the rat CNS. Emphasis is placed on the comparison of results obtained by all available methods.
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