Development and optimization of sectioning technique for the study of migration and differentiation potential of testicular stem cells in X. tropicalis tadpoles

The rapid development of regenerative medicine and the urgent need for cells which are able to modulate the immune system, or even differentiate into variable cell types, have led to the research of mesenchymal stem cells. The Sertoli cells, which are essential for the proper development of sperm in the testis, have a strikingly similar character. In the previous research, a cell culture expressing markers of mesenchymal stem cells and Sertoli cells from juvenile male testes of X. tropicalis was established. At the same time, the cells were modified by the introduction of gene for the red fluorescent protein (RFP) in their genome. The aim of this diploma thesis was to clarify their characteristics after microinjection to X. tropicalis tadpoles (allogeneic transplantation). For these experiments, it was necessary to develop a reliable technique for the preparation of sections, which won't be harmfull for the samples. Using the vibratome sectioning method along with immunohistochemical labeling, the cell culture has been found to contain precursors of Sertoli and peritubulare myoid cells.Rychlý rozvoj regenerativní medicíny a naléhavá potřeba buněk schopných ovládat imunitní systém nebo dokonce diferencovat do jiných buněčných typů vedla k výzkumu mesenchymálních kmenových buněk. Nápadně podobný charakter mají i Sertoliho buňky, které jsou nezbytné pro správný vývoj spermií ve varleti. V předcházejícím výzkumu byla založena buněčná kultura z varlat juvenilních samců X. tropicalis, která obsahovala buňky s markery mesenchymálních kmenových buněk tak i buněk Sertoliho. Zároveň byly buňky modifikovány a do genomu jim byl zaveden gen pro červený fluorescenční protein (RFP). Cílem této diplomové práce bylo tedy objasnit vlastnosti těchto buněk po jejich mikroinjikování do embryí jedinců stejného druhu (allogenní transplantace). Pro tyto experimenty bylo nutné vyvinout spolehlivou techniku přípravy řezových preparátů, při které by nedocházelo k ztrátě buněk či celých tkání. Metodou vibratomových řezů spolu s imunohistochemickým značením se podařilo zjistit, že buněčná kultura obsahuje buňky prekurzorů Sertoliho a peritubulárních myoidních buněk.Department of Cell BiologyKatedra buněčné biologieFaculty of SciencePřírodovědecká fakult

Analysis of gene expresion via next generation sequencing techniques

1 Abstrakt Cílem práce je vytvořit přehled dnes dostupných metod analýzy genové exprese, uvést výhody a nevýhody daných metod a porovnat je. Sekvenování je dnes jedna z nejpoužívanějších molekulárních metod. Sekvenovací metody se dají rozdělit na tři skupiny, sekvenování první generace, druhé generace a třetí generace. Nejvíce využívanými jsou metody sekvenování druhé generace. Avšak velký potenciál má i třetí generace, u které se ztrácí důležitost amplifikace vzorku pomocí PCR. Díky klesajícím nákladům a stoupající efektivitě se ale rapidně zvyšuje i množství dat. Rostou tak nároky na datová úložiště, výkon počítačů a důmyslnost softwarů, které velmi usnadňují zpracování získaných informací.1 Abstract The main task of this work is create review today's available methods for gene expression analysis, introduce advantages and disadvantages of this metods and compare them. Nowadays sequencing is one of the most usable molecular methods. Sequencing methods are divided to three groups, a first generation sequencing, a second generation sequencing and a third generation sequencing. The most useful are the second generation sequencing. However, the third generation sequencing have a big potencial too. It is not necessary amplificate samples using PCR thanks them. Amount of data raises rapidly, thanks decreasing costs and increasing efficiency. Demands on data starage, computers output are growing. And softwares for data analysis are much clever than ever before.Department of Cell BiologyKatedra buněčné biologiePřírodovědecká fakultaFaculty of Scienc

Development and optimalization of sectioning technique for the study of migration and differentiation potential of testicular stem cells in X. tropicalis tadpoles

Thanks to their ability to differentiate into variable cell types and migrate to the site of an injury mesenchymal stem cells (MSC) are broadly used in regenerative medicine. Their relative easy availability together with the property to control the immune system determines them as a cure of autoimmune diseases or a recovery of wounded tissues. Similar features posses Sertoli cells which take place in the seminiferous tubule of testis. Cell culture of testicular stem cells from juvenile male testes of X. tropicalis (XtTSC) was established in supervisor's laboratory. This cell culture showing both MSC's and SeC's properties was transformed to carry red fluorescent protein RFP. The aim of this diploma thesis was to investigate an behavior of transformed XtTSC in living organism, therefore cells were transplanted into the X. tropicalis tadpoles in stage 41. Subsequently, their migration potential was explored. To study of XtTSC's differentiation potential it was necessary to introduce a reliable sectioning techniques for the subsequent immunohistochemical analysis. Based on our experiments, we found that the XtTSC's cell culture contains precursors of SeC and peri-tubular myoid cells, however in vivo these cells turned into the dedifferentiated MSC-like state allowing a strong migration through the...Mesenchymální kmenové buňky (MSC) díky svým vlastnostem diferencovat v jiné buněčné typy a migrovat do místa poranění našly široké využití v regenerativní medicíně. Jejich snadná dostupnost spolu se schopností modulovat imunitní systém je předurčuje k léčbě autoimunitních onemocnění nebo obnově poraněných tkání. Stejně jako MSC i Sertoliho buňky (SeC) nacházející se ve varleti mají velmi podobné vlastnosti. V laboratoři školitele byla založena buněčná kultura testikulárních kmenových buněk z varlat juvenilních samců drápatky tropické (Xenopus tropicalis) (XtTSC). Kvůli jejich vizualizaci in vivo byla tato kultura buněk vykazující vlastnosti MSC a SeC transformována plasmidovým vektorem kódujícím červený fluorescenční protein RFP. Cílem této diplomové práce bylo zkoumat chování transformovaných XtTSC v živém organismu. Z tohoto důvodu byly buňky transplantovány do pulců X. tropicalis ve stádiu 41. Následně byl vyhodnocen jejich migrační potenciál. Aby bylo možné studovat také diferenciační potenciál těchto buněk, bylo nezbytné zavést spolehlivou techniku přípravy řezových preparátů. Na základě provedených experimentů se podařilo zjistit, že buněčná kultura XtTSC obsahuje prekurzory SeC a peritubulárních myoidních buněk, avšak in vivo buňky zřejmě přechází do dediferencovaného stavu podobného MSC,...Katedra buněčné biologieDepartment of Cell BiologyFaculty of SciencePřírodovědecká fakult

Repository of drought event impacts across the danube catchment countries between 1981 and 2016 using publicly available sources

Drought directly and indirectly affects human society in a number of ways. In many regions of the world climate change will exasperate the effects of droughts, affect national economies more intensely. The main aim of this article was to catalogue and analyze the drought impacts in the 11 Central and South Eastern European states located in the Danube river basin. The identification of dry episodes was based on information from publicly available sources, namely, newspaper and journal articles that reported drought impacts. Information on drought impact occurrences was classified into one of five defined categories in which the drought impact report was most clearly manifested (i.e., agriculture, forestry, soil systems, wildfires and hydrology). In terms of the spatial distribution of drought impacts, individual recorded events were analyzed at the level of EU NUTS regions (or their equivalent in non-EU countries). The analysis highlights country-specific vulnerability to drought. Furthermore, gradual increases in drought events and the number of reported impacts were identified, which was particularly evident in the agricultural sector

Validity and reliability of drought reporters in estimating soil water content and drought impacts in central Europe

Increasing drought is considered one of the major threats associated with climate change in central Europe. To provide an objective, quantitative tool that represents current drought conditions, the Czech Drought Monitor System (CzechDM) was established in 2012. Like other drought monitoring systems worldwide, the CzechDM uses several approaches to provide drought data. However, the CzechDM is unique internationally due to its utilization of a network of voluntary reporters (farmers) who complete a weekly online questionnaire to provide information about soil water content and the impacts of drought on crop yield. In this study, the results from the questionnaires from individual farms were aggregated by district. Reporters’ data were compared and validated with the outputs of the SoilClim model (a core tool of the CzechDM) and with other drought monitoring tools, such as the water balance model, the soil water index and the evaporative stress index. The soil water content estimated by the reporters was significantly correlated (on average r = 0.8) with the outputs of the SoilClim model. Conversely, the correlation between the drought impacts on yield estimated by the reporters and the SoilClim outputs was lower (on average r = 0.4), suggesting that in situ observations by farmers provide additional insights into the occurrence of drought impacts. Importantly, it was found that farmers reported significant drought impacts on yield earlier in the season than any other methods (models or remote sensing). The main findings of this study are that the drought monitoring provided by reporters is a useful and reliable component of the CzechDM. We conclude that weekly reports by farmers represent a significant enhancement to drought monitoring and have potential for use in developing automated approaches that combine in situ, modeling and remote sensing data within a data fusion or machine learning framework

The 2022 drought needs to be a turning point for European drought risk management

The 2022 European drought has underscored critical deficiencies in European water management. This paper explores these shortcomings and suggests a way forward for European drought risk management. Data for this study was gathered through a continent-wide survey of water managers involved in this event. The survey collected 481 responses from 30 European countries and is comprised of 19 questions concerning sectorial impact in the 55 regions of the responders and drought risk management practices of their organizations. Information from the survey is enriched with climate-related information to offer a comprehensive overview of drought risk management in Europe. Our research focuses on four key aspects: the increasing risk of drought, its spatial and temporal impacts, current drought risk management approaches, and the evolution of drought risk management across the continent. Our findings reveal a consensus on the growing risk of drought, which is confounded by the rising frequency and intensity of droughts. While the 2022 event affected most of the continent, our findings show significant regional disparities in drought risk management capacity among the various countries. Our analysis indicates that current drought risk management measures often rely on short-term operational concerns, particularly in agriculture-dominated economies, leading to potentially maladaptive practices. An overall positive trend in drought risk management, with organizations showing increased awareness and preparedness, indicates how this crisis can be the ideal moment to mainstream European-wide drought risk management. Consequently, we advocate for a European Drought Directive, to harmonize and enforce drought risk management policies across the continent. This directive should promote a systemic, integrated, and long-term risk management perspective. The directive should also set clear guidelines for drought risk management at the national level and for cross-boundary drought collaboration. This study and its companion paper "The 2022 Drought Shows the Importance of Preparedness in European Drought Risk Management " are the result of a study carried out by the Drought in the Anthropocene (DitA) network

Experiences of kindergarten teachers with shared custody and their attitude to this custody.

This Bachelor's thesis inquires shared child custody after divorce. In a theoretical part it is mentioned, when the shared custody, as a form of a child custody after divorce, was implemented. It also refers to its low frequency. The problem of shared custody is clarified here and a presentation of a research all over the world comes after. A difficult situation after divorce is mentioned afterward, an explanation haw difficult this period is, not only for parents, but also for children, who are overlooked usually, themselves. Further forms of child custody are mentioned in the end. Methods, used in the research, are presented in a methodological part. Information got from nursery school teachers are analyzed in a practical part. It is about their point of view of shared custody and of their experience with this form of custody

Development and optimization of sectioning technique for the study of migration and differentiation potential of testicular stem cells in X. tropicalis tadpoles

The rapid development of regenerative medicine and the urgent need for cells which are able to modulate the immune system, or even differentiate into variable cell types, have led to the research of mesenchymal stem cells. The Sertoli cells, which are essential for the proper development of sperm in the testis, have a strikingly similar character. In the previous research, a cell culture expressing markers of mesenchymal stem cells and Sertoli cells from juvenile male testes of X. tropicalis was established. At the same time, the cells were modified by the introduction of gene for the red fluorescent protein (RFP) in their genome. The aim of this diploma thesis was to clarify their characteristics after microinjection to X. tropicalis tadpoles (allogeneic transplantation). For these experiments, it was necessary to develop a reliable technique for the preparation of sections, which won't be harmfull for the samples. Using the vibratome sectioning method along with immunohistochemical labeling, the cell culture has been found to contain precursors of Sertoli and peritubulare myoid cells

Elementary introduction to the theory of Groebner bases

V první části práce je připomenuta historie, je ukázáno, jak řešit příklady "lidským" způsobem. Druhá část je zaměřena na Gröbnerovy báze. Nejdříve jsou zavedeny pojmy S-polynom, uspořádání termů a normálnní tvar polynomů. Po definování pojmu Gröbnerovy báze je řešeno několik příkladů. Na konci práce je ukázáno, jak řešit soustavy algebraických rovnic pomocí Gröbnerových bází s využítím počítače.Katedra matematiky, fyziky a technické výchovyObhájenoIn the first part is remind the history, it is shown how to solve examples of "human" way. The second part focuses on Groebner bases. First, the concepts are introduced S-polynomial ordering of terms and normal form polynomials. After defining the concept of Groebner bases is dealt with several examples. At the end of the thesis, it is shown how to solve systems of algebraic equations using Groebner bases using computers

Analysis of gene expresion via next generation sequencing techniques

1 Abstract The main task of this work is create review today's available methods for gene expression analysis, introduce advantages and disadvantages of this metods and compare them. Nowadays sequencing is one of the most usable molecular methods. Sequencing methods are divided to three groups, a first generation sequencing, a second generation sequencing and a third generation sequencing. The most useful are the second generation sequencing. However, the third generation sequencing have a big potencial too. It is not necessary amplificate samples using PCR thanks them. Amount of data raises rapidly, thanks decreasing costs and increasing efficiency. Demands on data starage, computers output are growing. And softwares for data analysis are much clever than ever before