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Honey bee (Apis mellifera) virome
Kadlečková, Dominika ; Tachezy, Ruth (advisor) ; Rothová, Olga (referee)
Microbiome is composed of all microorganism (fungi, bacteria, archea, protozoa, virus) sharing the same space at the same time. Virome which consists of viruses either of the host or viruses of other microorganisms and of genetic elements derived from viruses, is part of the microbiome. Microbiome plays a role in a number of processes which are vital for development and survival of the host. Virome can affect host either directly or indirectly by interaction with other parts of microbiome. This thesis is focused on virome of Apis mellifera, globally managed pollinator. Viral diseases are one of the factors responsible for high colony losses of this pollinator and therefore this work is mostly dedicated to those viruses which were confirmed as pathogens of Apis mellifera. Also, their interaction with other pathogens of Apis mellifera, which leads to great losses of these pollinators will be discussed. Key words: virome, honey bee, Apis mellifera, honey bee viruses, colony losses
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Glutamate Receptors and Endoplasmic Reticulum Quality Control
Tachezy, Ruth ; Horák, Martin (advisor) ; Adámek, Pavel (referee)
Quality control (QC) is a collection of processes taking part in the biogenesis and architecture of proteins. The objective of this thesis is to describe these processes in detail. QC takes place on many different levels in various compartments of the cell. The focus is on the endoplasmic reticulum (ER) QC interconnected with cytosolic QC. There are multiple steps involved in ERQC: several types of protein translocation to the ER lumen, glycosylation, disulfide bond formation via protein disulfide isomerase, chaperones that assist to achieve a correct conformation, and ER- associated degradation pathway for retranslocation of misfolded proteins back to the cytoplasm, where they are degraded. Cytosolic QC is interconnected with the ERQC through various ways of translocation of proteins to the ER membrane or lumen. Proteins that are retranslocated from ER to the cytosol are ubiquitinated and subsequently degraded in the proteasome. Ubiquitination is a process of targeting a protein for degradation. Cytosolic chaperones and other cellular structures, such as aggresomes, juxtanuclear compartments, and insoluble protein deposits, take part in the ubiquitination. Calcium dysregulation that is linked to QC and correct protein folding in ER is also described. Some of the possible consequences of protein...
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Molecular analysis of head and neck carcinomas associated with HPV infection
Glendová, Kristýna ; Staněk, Libor (advisor) ; Tachezy, Ruth (referee)
Head and neck cancers (HNSCC) are highly heterogeneous disease, results from two major carcinogens - tobacco and/or alcohol, or HR HPV infection. This thesis was based on 60 biopsies of head and neck tumours embedded into paraffin after histological verification. HPV infection, including particular types was monitored in different HNSCC regions by multiplex qPCR. Subsequent IHC demonstrated expression of p16INK4A and p53 as a possible diagnostic biomarker. Based on the information, patients with HNSCC can benefit from antiEGFR therapy by Cetuximab, but so far without defined predictors, the analysis of point mutations of Ras gene family (Kras, Nras) and Braf gene was performed. These mutations were monitored as potential predictive biomarkers, in correlation with gender, age and other risk factors. For all statistical processing the Chi-x2 test was used. Key words: Head and neck cancers, biopsy, HPV types, PCR, p16INK4A, p53, molecular predictors, Kras, Nras, Braf
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Optimization of proces for detection of free tumor DNA in plasma and its clinical utility for colorectal cancer, lung cancer and pancreatic cancer patients
Belšánová, Barbora ; Benešová, Lucie (advisor) ; Tachezy, Ruth (referee)
In current days, examination of circulating tumor DNA (ctDNA) finds new use across different cancers. It is directed at tumor-derived short fragments of DNA present in peripheral blood of patiens (mainly in advanced stages). Due to its minimal invasivity, almost 100 % specificity and relatively high sensitivity in stage IV patients, this approch found its main potential clinical utility especially in early detection of disease relapse or progression after tumor resection (i.e. post-operative follow-up), prediction and monitoring of therapy response and estimation of prognosis. As a result of minute levels of ctDNA on a high background of other non-tumor DNA fragments present in plasma, a suitable method exhibiting highest sensitivity is the key for proper detection of this marker. The approach is predominantly based on initial identification of a mutation in tumor tissue and its subsequent detection in plasma. The present work is aimed at optimization of ctDNA isolation and method of its detection based on PCR amplification followed by heteroduplex analysis by denaturing capillary electrophoresis (DCE) to achieve highest sensitivity for detection of mutated fraction in plasma sample. I have applied the optimized protocol to examine ctDNA in three types of cancers, namely colorectal cancer (122...
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Risk factors of HPV infection and head and neck tumours
Sekavová, Alžběta ; Tachezy, Ruth (advisor) ; Bobák, Martin (referee)
Epidemiology of head and neck cancers is currently intensively studied topic. Recent shift in the age of incidence towards younger population is generally attributed to growing proportion of head and neck cancers caused by human papillomavirus - sexually transmitted virus which causes asymptomatic, but sometimes persisting infection that can lead to malignant transformation of the infected tissue. Significance of the topic lies mainly in the prognostic ad- vantage of patients with virally induced head and cancers and preventabil- ity of infections with human papillomavirus. First aim of this thesis is to demonstrate epidemiological trends of head and neck cancers in the Czech Republic, with the focus on change of age-specific incidence and mortality in the last three decades. Second aim of this thesis is to identify risk factors of oral infections with human papillomaviruses and head and neck cancer in a case-control and cross-sectional study of a hospital-based cohort. 1
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Immune cells infiltrating premalignant lesions and invasive carcinomas associated with HPV infection
Dalewská, Natálie ; Tachezy, Ruth (advisor) ; Španielová, Hana (referee)
The immune system participates in many defence mechanisms of the body which in- volve the action of different types of immune cells. Immune cells play an important role in tumour development and progression. They infiltrate tumours and contribute to the tumour microenvironment. They are also implicated in diseases associated with human papillomavi- rus infection. Human papillomaviruses are viruses with oncogenic potential which cause in- fection in a number of anatomical locations of the human body. Such infection can lead to benign lesions, pre-invasive lesions, and invasive tumours. Pre-invasive lesions often re- gress spontaneously, but at advanced stages of the disease, progression to malignancy can occur. The tendency to regress or progress varies depending on the HPV genotype involved in the lesion and the quantity, type, and location of infiltrating immune cells. The synergy of these factors influences the prognosis and treatment of the disease, and, therefore, deter- mining the immunological score as an important diagnostic parameter becomes of utmost relevance. The knowledge of the effect of immune cells on tumour cells is useful in immunotherapy, which is based on the activation of the patient's own anti-tumour immune mechanisms. Immunotherapy is used in combination with systemic therapy...
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The role of biomarkers in erosive osteoarthritis of the hands
Lennerová, Tereza ; Šenolt, Ladislav (advisor) ; Tachezy, Ruth (referee)
Hand osteoarthritis (OA) is a degenerative joint disease that causes pain, functional limitation and negatively affects the patients' quality of life. The most severe subtype of this disease is erosive OA. Erosive hand OA is characterized by an abrupt onset, inflammation and is linked to worse outcomes than non-erosive hand OA. Current methods do not allow early diagnosis or to distinguish between patients with different forms at disease onset. This could be changed by the utilization of biomarkers in clinical practice. Biomarkers are molecules released into circulation that reflect biological processes. The main goal of this study was to analyze the levels of circulating biomarkers with the aim to differentiate patients from healthy subjects and patients with erosive OA from patients with non-erosive disease. Serum concentrations of seven biomarkers and the expression of plasma microRNAs were determined. Patients with hand OA showed altered cartilage metabolism, increased levels of adiponectin, decreased levels of clusterin and a dysregulated expression of several microRNAs in comparison to the healthy population. Patients with erosive OA had lower levels of clusterin and decreased expression of miR-151-3p than those with the non-erosive form of the disease. These findings suggest the potential...
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Methylation of the regulatory sequences of integrated and extrachromosomal form of HPV 16 in tonsillar tumors
Pokrývková, Barbora ; Tachezy, Ruth (advisor) ; Hubáček, Petr (referee)
Human papillomavirus is connected with induction of cervical carcinoma as well as for some other anogenital carcinomas and subset of carcinomas of head and neck. Presence of viral E6 and E7 oncoproteins may induce cell transformation, higher load of oncoproteins is caused by the regulatory E2 protein inactivation. Aims of recent study are mechanisms of E2 protein inactivation. One option is integration of viral DNA into the host genome, which is located into the E2 gene region. Some carcinomas, where virus with extrachromosomal form was presented, were found. It appears that epigenetic changes can play the role in the development of this type of tumors, especially DNA methylation or mutation in the regulatory region of the virus. The methylation degree analysis on samples of tonsillar carcinomas with extrachromosomal and integrated form of the virus was conduced, as well as viral load of both groups was compared and the expression of E6 and E7 gene was confirmed. The results of methylation analysis showed increased methylation of the virus with integrated DNA. Mutations in the E2 protein binding sites are not revealed. The expression of the viral oncogenes were confirmed in all tumors regardless of the form of the viral genome. The mechanism of tumors induction, especially for virus with...
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MicroRNAs in Human Cancers Associated with Viral Infections
Dvořáková, Lucie ; Tachezy, Ruth (advisor) ; Drda Morávková, Alena (referee)
MicroRNA (miRNA) are short single-stranded RNAs that do not encode proteins. Their main function is the regulation of the gene expression on the level of translation. This regulation is mediated by the binding of miRNA to the partially complementary segments of mRNA, both cellular or viral. It is estimated that miRNAs affect expression of at least one third of human genes and thereby influence regulation of cellular growth, differentation and apoptosis of cells. Recently the miRNAs encoded mainly by DNA viruses were discovered. These miRNAs enhance the persistence of viral infection in the host and can contribute to malignant transformation. However, the oncogenesis is also significantly affected by the regulation of cellular miRNAs expression by viral proteins. The miRNA research is topical. MiRNAs are considered as potential biomarkers and their utilization as a cancer therapy is being intensely explored. In this thesis, I'm describing the biogenesis and regulatory functions of miRNAs. I'm also presenting an overview of viral miRNAs focusing on human oncogenic viruses which do not only code their own miRNAs but also influence the expression of the host miRNAs. Finally, I am focusing on current clinical applications of miRNA. Key words: viral miRNA, cellular miRNA, oncogenesis, viral infections,...
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