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Proliferace a dynamika tkání během vývoje zubů a jim pribuzných patrových lišt
Rothová, Michaela ; Peterková, Renata (advisor) ; Matalová, Eva (referee) ; Černý, Robert (referee)
My PhD thesis has addressed specific questions regarding cell proliferation and tissue dynamics in three key areas of craniofacial development: during suppression and revitalization of tooth buds that develop in the mouse embryonic dentition as rudiments of lost premolars; during dental papilla and follicle formation of the first mouse molar; and during origin of palatal rugae on the mouse hard palate. By evaluation of cell proliferation, we recorded a change in the proliferation pattern along the cheek region of the mandible between less and more advanced embryos at embryonic day 13.5. Thus during the time period, when the development of the large mouse rudimental premolar primordium (R2) is stopped, we showed that the arrest of the rudiment R2 is caused by exhibiting low rate of cell proliferation and high rate of apoptosis. When Sprouty gene signalling is disrupted, the premolar primordium shows rates of proliferation and apoptosis similar to the growing first molar. The R2 subsequently revitalizes and develops into a supernumerary tooth in front of the first molar. Furthermore, we discovered that the dental mesenchyme is very dynamic tissue during bud and cap stages of tooth development and that the dental papilla of the first molar originates only from a restricted region of the dental...
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Sample preparation for single cell analysis
Adamová, Eva ; Basova, E. Y. ; Potáčová, Anna ; Foret, František ; Matalová, Eva ; Klepárník, Karel
Development of sensitive, miniaturized and fast methods aims to reliable analyze even small amounts of samples. Micromanipulation systems such as laser capture microdissection enable to collect just single cells from cell suspensions or tissue samples. One option for further analysis is the bioluminescence reaction based on luciferin/luciferase conversion. This approach was used for detection of active caspases, caspase-3/7. Droplets based microfluidic systems provide benefits in biological, pharmaceutical and chemical research.
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Analysis of tissue-bound cells: Novel approaches using laser capture microdissection
Matalová, Eva ; Adamová, Eva ; Klepárník, Karel
Recent biomedical research prefers analysis of homogenous samples obtained from primary cells to cell lines. Moreover, attention is paid to cells of low number but high biological impact, such as signalling centres in embryonic development, stem cells or tumour cells. To localize cells of interest within an organ/tissue, histological sections are widely used. However, the methods of choice for further analysis of such samples are mostly limited to histochemistry, immunohistochemistry and in situ hybridization.
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Senzitive method of Caspase-3 detection in single stem cell
Adamová, Eva ; Klepárník, Karel ; Matalová, Eva
Caspases are involved in physiological process (e.g. cellular differentiation) but also can cause serious disorders. It is getting necessary to develop sensitive, miniaturized and fast methods amenable to analyze small amounts of samples. Luciferin/luciferase chemiluminescence reaction is one of the methods of caspase-3 detection. We have developed a miniaturized device enabling detection of caspase-3 and quantitation just in femtogram level (10–15 g) in single apoptotic human stem cells (neural crest derived). The technology is based on the specific cleavage of modified luciferin by caspase-3, emissions of photons and their detection by photomultiplier tube working in the photon counting regime.
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Modulace intracelulární kaspázové mašinérie s využitím explantátových kultur
Matalová, Eva ; Fleischmannová, Jana ; Norek, Adam ; Míšek, Ivan
Organ explant cultures eliminate animal suffering during experiments but simultaneously allow investigations of intact organ, tissue and cell systems with preserved interactions corresponding to the situation in vivo. Moreover, the explants can be manipulated in several ways using modern micromanipulation techniques (e.g. electroporation, inhibition). We show possible usage of explant culture approaches in embryonal apoptosis research in three models – mouse limb digitalization, tooth germ morphogenesis, separation of middle ear ossicles. Data from specific pharmaceutical inhibitions focused on key apoptotic molecules – caspases - are demonstrated.
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Molekulární aspekty hypodoncie
Fleischmannová, Jana ; Krejčí, P. ; Matalová, Eva ; Míšek, Ivan
Numeric dental anomalies are the most common craniofacial congenital malformations in humans. More than 20 % of human population miss one or more third molars, approximately 5 % of population display agenesis of another tooth. In this contribution molecular events underlying tooth formation (more than 200 genes have been identified so far) and defects in tooth germ formation are correlated. Both syndromic forms of hypodonia (e.g. Rieger syndrome, anhidrotic ectodermal dysplasia) and non-syndromic forms (related to pax9, msx1, axin2) were investigated.
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