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Changes in mitochondrial DNA in colorectal cancer patients
Danešová, Natálie ; Vodenková, Soňa (advisor) ; Büchler, Tomáš (referee)
Mitochondrial DNA (mtDNA) changes occur more frequently in the cell than alterations in nuclear DNA (nDNA) due to factors such as proximity to reactive oxygen species (ROS) or fewer DNA repair pathways. While nDNA damage is known to play a role in colorectal cancer (CRC), there is a lack of studies concerning mtDNA damage. Changes in mtDNA copy number, a possible indirect marker of mtDNA damage known as mtDNA content, have been reported in CRC with conflicting results. Various changes in mtDNA have been observed in multiple cancer types and proposed as potential biomarkers, including CRC; however, its exact role in disease progression, patients' prognosis, or prediction of treatment is yet to be determined. The primary hypothesis of this study was that mitochondrial dysfunctions resulting from mtDNA changes play a role in colorectal carcinogenesis and could serve as potential CRC biomarkers. To test this hypothesis, we measured mtDNA damage, mtDNA content, and the expression of selected DNA repair genes in both tumor and adjacent non-malignant mucosa. Initially, we conducted a Pilot study involving 7 patients, utilizing RNA sequencing and qPCR. Subsequently, a Validation study was performed on a larger cohort of 50 patients using qPCR. Our findings revealed that adjacent mucosa exhibited higher...
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A study of chemoresistance in patients with colorectal cancer treated with 5-fluorouracil
Dostál, Petr ; Vymetálková, Veronika (advisor) ; Šeborová, Karolína (referee)
The choice of treatment strategy in patients with malignant disease depends on various clinical and molecular biological factors. Although several molecular predictive biomarkers have already been proposed, only a few of them are used in clinical practice and the number is still not enough for reliable personalized medicine. Given the lack of treatment success results in many colorectal cancer (CRC) patients, there is an urgent need for personalized medicine to identify new predictive biomarkers. One of the main clinical challenges in the treatment of advanced CRC is the development of chemoresistance to systematic chemotherapy. Early detection of resistant cancer cells clones could lead to changes in treatment regimens but it requires a long-term follow up of patients and monitoring of specific markers of chemoresistance. The aim of this master's thesis has been to determine the biomarkers associated with chemoresistance to 5-FU drug, one of the most used chemotherapeutics in the treatment of CRC patients. In the first step, the whole transcriptome of maternal (sensitive) and resistant DLD-1 (line resistant to 40 µM 5-FU and line resistant to 160 µM 5-FU) cell lines using the next generation sequencing (NGS) has been analyzed. Through bioinformatic analyses, potential candidate genes (HIST1H2BE,...
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Evaluation of perfusate solution after ex vivo lung perfusion
Dobrovolná, Terezie ; Enghuber, Florian (referee) ; Paštěka, Richard (advisor)
The aim of this master’s thesis is to introduce lung perfusion procedures and determine the parameters that are connected to the state of the lungs after perfusion and during preservation. In the perfusion process and for storing the lung, the phosphate-buffered saline (PBS) solution is used. The work focuses mainly on the parameters that can be measured and evaluated in the perfusate solution during the perfusion and preservation of the lungs such as pH, total dissolved solids, and proteins. These factors can be measured according to several procedures that are based on different principles. The mentioned process has been selected and implemented with regard to availability, feasibility, and appropriateness given. The pH and TDS parameters have been acquired by electrodes. Therefore, sensor calibration is an integral part of the measure ment. The concentration of the proteins was measured with the Bradford assay where it is important to follow the established protocol. Materials for that were present and provided in the Tissue engineering laboratory. Another intention is to assess the state of the lung tissue, meaning how the lungs gradually change their appearance, weight, and mechanical properties over time. The weight is obtained with the calibrated load cell sensor. For this sensor, a platform had to be assembled in order to function correctly and with great accuracy. From the field of mechanical properties of the lungs, the compliance and PV loops have been selected and presented. Five lungs were used for measurement. The appearance of the lungs changed over time and the necrosis was progressing. The lung viability was preserved with the PBS solution with balanced pH. The value of pH was maintained around pH 7. The concentration of the total dis solved solids (TDS) was determined and similar values were obtained in all lungs because the same solution was used. The TDS fluctuated slowly same as pH because they are correlated. The weight of the lungs decreased with passage of the time as expected. Protein concentration increased over time. That can indicate the inflammatory processes in the lung tissue or lung injury. The surface temperature was also obtained with a laser thermometer and decreasing values were noted with increasing storage time. Static compliance had unexpected charac teristics in some of the lungs. This was probably influenced by leakage due to an unsecured trachea. The last parameter was PV loops. The shape of the PV loops looked good until 48h in the majority of the loops. Then, the shape flatted and the shortage of curves appeared due to shortness of breath.
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Use of biomarkers for assessing environmental pollution by platinum group metals
Nedvědická, Lucie ; Řezáčová, Veronika (referee) ; Komendová, Renata (advisor)
This diploma thesis in the theoretical part analyses the characteristic feature of individual platinum group elements, their toxicity, anthropogenic sources and the presence of platinum group elements in the environment of the Czech Republic. In addition, there are specified types of bioindicators focusing mainly on lichens, mosses and needles. There are described individual bioindication techniques and pre-treatment of samples for platinum metals determination in bioindicators. The subject of the experimental part is to verify the sorption efficiency and ability of platinum and palladium using the SPE method and to determine the actual platinum metal contamination in selected localities in the city of Brno. There was also the additionally subject to verify the sorption potential of selected bioindicators and determination of the environmental pollution by platinum metals using the bag technique. In the experimental part, the bioindicators of moss Pleurozium schreberi, lichen Xanthoria parietina, lichen Flavoparmelia caperata and needles Pinus nigra were used.
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Importance of biosensors and biomarkers for environment contamination assessment
Mišo, Milan ; Čáslavský, Josef (referee) ; Zlámalová Gargošová, Helena (advisor)
This bachelor thesis is purely theoretical. It aims to explain the principle and nature of operation of biosensors and biomarkers, their properties, distribution and use in the assessment of environmental contamination. It describes their practical use. In the first part of the thesis are described biosensors. Biosensors combine biorecognition molecules and a suitable converter, this combination allows you to convert (bio)chemical information into electrical signals that can be easily measured and therefore are becoming more used for determination of different substances. The second section describes the early warning signs called biomarkers. Biomarkers provide information on changes in the ecosystem before it is irreversible damaged, and we can intervene early.
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Proteomic analysis of posttranslation modifications in breast cancer cell line profiles
Predná, Nikola ; Laštovičková,, Markéta (referee) ; Strouhalová,, Dana (advisor)
Estrogenové a progesteronové receptory, stejně jako HER2 protein, jsou v současnosti klinicky nejužitečnějšími metabolickými markery u karcinomu prsu. Tyto markery umožňují určit typ nádoru a nejlepší možnosti jeho léčby. Jeden z nejagresivnějších typů tohoto onemocnění, triple-negative breast cancer (TNBC), však tyto klinicky stanovené biomarkery postrádá. To znamená, že hormonální terapie nebo cílené léky nepřicházejí v úvahu, takže je na výběr méně možností léčby. Aby bylo možné vyvinout nové léky na míru, je zásadní pochopení molekulárního základu onemocnění. V poslední době se mnoho studií zaměřuje na hledání biomarkerů na úrovni proteinů pomocí proteomiky. Proteiny, zejména jejich post-translační modifikace (PTM), jsou jádrem mnoha buněčných událostí a jejich odhalení může pomoci při pochopení mechanismů rakoviny prsu. Pro objevení molekulárních rysů TNBC, je cílem této studie porovnat proteomická data neléčených rakovinných buněčných linií s buňkami, které podstoupily retinoidní terapii. Důraz bude kladen na PTM, zejména glykosylaci a fosforylaci, Vimentinu a CD44, které byly navrženy jako potenciální biomarkery TNBC v předchozích studiích. Proteinová separace bude provedena pomocí 1D a 2D gelové elektroforézy nebo pomocí SEC-HPLC. Vzorky budou také podrobeny enzymatickému štěpení před identifikací pomocí MALDI-TOF hmotnostní spektrometrie. V případě fosfoproteinového selektivního záchytu bude obohacení provedeno afinitní chromatografií s použitím hrotů pro obohacení fosfopeptidu TiO2 (TopTip). Glykosylované proteiny budou obohaceny pomocí WGA lektinové afinitní chromatografie. Proteiny s významnými rozdíly v PTM mezi ošetřenými a neošetřenými buňkami budou blíže hodnoceny pomocí proteinových databází (MASCOT, STRING a další). Data získaná ze studie budou případně použita k navržení potenciálních biomarkerů pro TNBC.
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Proteomic approach for the study of cancer cell line profiles.
Predná, Nikola ; Langová, Denisa (referee) ; Strouhalová,, Dana (advisor)
Triple-negativní karcinom prsu (TNBC), velice agresivní podtyp rakoviny prsu, je známý svou nepříznivou prognózou a omezenými možnostmi léčby. V tuto chvíli je chemoterapie považována za hlavní způsob léčby. Za účelem vyvinutí nových účinných léčiv je snaha pochopit molekulární základ této nemoci. V důsledku toho bylo již několik potenciálně aktivních látek pro tento konkrétní typ rakoviny prsu podrobeno výzkumu. V poslední době se mnoho studií zabývající touto záležitostí provádí za použití proteomiky jakožto prostředku ke studiu proteomů rakovinných buněk. Rakovinové buňky obsahují klíčové rozdíly v proteinech, které regulují mechanismy buňky. Mapování těchto mechanismů může nakonec umožnit diagnostikovat stav organismu. Tato práce se zaměřuje na proteomické studium buněk TNBC a porovnává neošetřené buňky s buňkami, které byly podrobeny léčbě retinoidy. Separace proteinů a peptidů byla úspěšně provedena elektroforézou na 1D a 2D gelu. Kromě toho byly vzorky podrobeny enzymatickému štěpení vybraných proteinů, které byly poté identifikovány pomocí hmotnostní spektrometrie MALDI-TOF (MS). Proteiny, které se podílejí na procesu epiteliálně-mezenchymálního přechodu (EMT), byly poté kvantifikovány a porovnány mezi vzorky.
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The diagnostic and prognostic ability of selected serum and urinary markers of prostate cancer
Do Carmo Silva, Joana Isabel ; Veselý, Štěpán (advisor) ; Čapoun, Otakar (referee) ; Fedorko, Michal (referee)
The diagnostic and prognostic ability of selected serum and urinary markers of prostate cancer Abstract Serum prostate specific antigen (PSA) is the only widely approved marker in prostate cancer (PC) diagnosis and follow up after treatment. Its role has remained controversial due to lack of specificity and the risk of overdiagnosis of insignificant PC. The aim of this work was to explore promising markers of PC and to improve current patient stratification to adjuvant treatment. Three main studies were performed using different media (urine and serum). The first study included the evaluation of Engrailed-2 (EN2) - a urinary marker of interest - in 90 patients with localized PC, 30 healthy controls, and 40 patients indicated for prostate biopsy. The second study evaluated 205 men with high-risk PC-features who underwent radical prostatectomy (RP) and were subject to a strict follow-up protocol of ultrasensitive PSA (UPSA) at close time intervals. The ability of particular measurements to predict biochemical recurrence (BCR) and thus the need for adjuvant therapy was assessed using the area under the curve (AUC) and a stratification model was created. The third study involved 128 patients who underwent RP. PSA and its serum isoforms normally used in the diagnostic context were evaluated both preoperatively...
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Influence of autism on facial morphology
Cenková, Klára ; Velemínská, Jana (advisor) ; Harnádková, Katarína (referee)
Autism (ASD) is a severe pervasive neurodevelopmental disorder with an as yet unclear etiology but increasing incidence. Early diagnosis and early initiation of therapy are crucial to the success of its treatment and to improving the quality of life of individuals with PAS. The identification of biomarkers in the form of specific facial features of individuals with autism can contribute significantly to facilitating diagnosis. These facial dysmorphologies have their basis in the prenatal period as a consequence of the common and interacting embryogenesis of the face and brain and reflect the neurodevelopmental basis of autism. Using special anthropometric methods, specific facial phenotypes can be determined, which can also be used to classify individuals with PAS into groups according to the severity of the disorder. Key words: Autism, facial dysmorphology, ASD, biomarkers, anthropometry
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Carotid Artery Stenosis
Svoboda, Norbert ; Beneš, Vladimír (advisor) ; Peisker, Tomáš (referee) ; Herzig, Roman (referee)
The basis of the doctoral thesis consists of three studies focused on the diagnosis of carotid stenosis. The first study examines the histological structure of carotid plaques and their relationship to clinical manifestations. The study included 269 patients with 280 analyzed plaques. Multiple logistic regression demonstrated a significant association between symptomatic stenosis and foam cells (OR=1.9, p=0.03) as well as plaque thrombosis (OR=3.5, p=0.02). A significant correlation was also found between symptomatic plaque and unstable plaque according to the comprehensive AHA plaque classification (OR=1.8, p=0.03).The second study addresses the accuracy of carotid stenosis diagnosis using digital subtraction angiography (DSA). Carotid stenosis was measured on DSA and histologically processed plaques obtained during carotid endarterectomy in 644 patients. The study did not establish a correlation between the findings on the histological plaque and DSA (correlation coefficient of 0.22 for ECST and 0.20 for NASCET). The last study focuses on the analysis of the biomarker of oxidative stress (malondialdehyde, MDA) as a predictor of unstable carotid plaques. The study involved 35 patients. Patients with asymptomatic carotid stenosis and those who experienced amaurosis fugax had significantly lower...
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