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Nanofibrous scaffolds in controlled delivery of autologous growth factors
Buzgo, Matej ; Amler, Evžen (advisor) ; Gášková, Dana (referee)
Platelet preparations are a source of various autologous growth factors and have numerous applications in tissues engineering. The aim of this work was to development electrospun nanofiber scaffolds with platelet preparations. Scaffolds based on the adhesion of platelets on nanofiber meshes were developed. The scaffolds were able to enhance chondrocyte proliferation in vitro. The main disadvantage of this system is the burst release of growth factors immediately after adhesion. To overcome this, we developed coaxially electrospun scaffolds with incorporated alpha granules. Alpha granules are novel platelet preparations with high amounts of growth factors. This system was able to stimulate chondrocyte proliferation and maintain TGF- 1 concentrations for 7 days. Additionally, a novel drug delivery system with coaxially incorporated liposomes was developed. Liposomes incorporated into nanofibers remain intact and can be used for the delivery of various molecules. The ability to maintain HRP activity was compared to systems based on coaxial electrospinning with liposomes, coaxial electrospinning without liposomes and blend electrospinning. When compared to other systems, coaxial electrospinning with liposomes preserves enzyme activity twice as long. These results clearly indicate the potential of...
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Competition of cells within the population of yeast colony
Očková, Veronika ; Váchová, Libuše (advisor) ; Gášková, Dana (referee)
Competition is a very important natural phenomenon, which causes the rivalry of organisms, in cases such as space limitation or lack of nutrients. It occurs mainly in situations where organisms, including microorganisms live in large populations. Multicellular yeast colonies represent an example of such a population. After the population of yeast cells spends nutrients from the environment, the cells in colonies are able to respond to these changes by production of ammonia functioning as a signaling molecule. Subsequently, the cells are able to change their morphology and metabolism and, dependently on their location within the colony, to create a subpopulation of cells with specific characteristics and functions. It is likely that in the case of mixed colonies formed by the two different strains, a competition between the cells of these two strains could exist. Such rivalry can result in changes in the ratio of cells of the two strains within the colony population, so that the cells of one strain outweigh the other. In this diploma thesis, I compared the growth and development of giant colonies and competition between the cells of selected pairs of strains forming mixed colonies. I focused on the parental strain Saccharomyces cerevisiae BY and its variants labeled with fluorescent proteins. For...
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Study of membrane transport processes in yeast using potentiometric fluorescent porbe diS-C3(3)
Bartl, Tomáš ; Gášková, Dana (advisor) ; Krůšek, Jan (referee)
1 Title: Study of membrane transport processes in yeast using potentiometric fluorescent probe diS-C3(3) Author: Tomáš Bartl Department: Institute of Physics of Charles University Supervisor: doc. RNDr. Dana Gášková, CSc., Institute of Physics of Charles University Abstract: Yeast membranes contain a number of transporters. Some are responsible for flow of nutrients to the inside of the cell, others for disposing of waste and foreign substances and some for transport of small ions or protons across the membrane. The focus of this work is on the activity of specific transport membrane proteins, so-called MDR pumps, which are responsible for transport of foreign substances or drugs, out of the cell. Using the series of mutant strains of the yeast Saccharomyces cerevisiae (AD1-3, AD1-8 and AD12) differentiated in the presence of specific MDR pumps in their membrane, an influence of various chemical substances on the intracellular concentration of the potentiometric fluorescent probe diS-C3(3), which is actively being transported out of the cell by some of the MDR pumps, was observed. By the examination of the effect of 2-deoxyglucose we proved the active contribution of not only the main MDR pump, Pdr5p, but also of some other pumps, in lowering the intracellular probe concentration. It was observed that...
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Functionalized nanofiber system for accelerated regeneration of internal tissues
Kráľovič, Martin ; Amler, Evžen (advisor) ; Gášková, Dana (referee) ; Chvojka, Jiří (referee)
Functionalized nanofiber system for accelerated regeneration of internal tissues Abstract Functionalization can adjust the properties of nanofibers prepared by electrospinning to best replace the missing extracellular matrix in the healing wound. In the rabbit intestinal anastomoses, functionalization of nanofibers by cryogenic fractionation has proven to be crucial. While the use of functionalized PVA and chitosan nanofiber membranes led to intestinal strictures and severe inflammation, further functionalization of PVA and chitosan nanofibers by their cryogenic fractionation led to the possibility of reducing the polymer dose, which significantly reduced the inflammatory response and significantly increased the biocompatibility of the material used. The application of fractionated nanofibers increased the fraction of microvessels, fibroblasts and collagen, which resulted in an increase in the biomechanical strength of healed anastomoses. The functionalization of the polycaprolactone nanofibers by special adhesion to the polypropylene mesh made it possible to use them as a support for the healing of the fascia in the abdominal wall of pigs. Functionalization of PVA nanofibers with glyoxal led to their increased stability in the aqueous environment. By applying a membrane of PVA functionalized nanofibers to...
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Analysis of the arrangement of the binding pocket of the MDR pump Cdr1p of the pathogenic yeast Candida albicans - a major contributor to clinical drug resistance.
Bartl, Tomáš ; Gášková, Dana (advisor) ; Krůšek, Jan (referee)
Title: Analysis of the arrangement of the binding pocket of the MDR pump Cdr1p of the pathogenic yeast Candida albicans - a major contributor to clinical drug resistance. Author: Tomáš Bartl Department: Institute of Physics of Charles University Supervisor: doc. RNDr. Dana Gášková, CSc., Institute of Physics of Charles University Abstract: Candida infections are becoming an increasing cause of death in hospitalized patients. The main reason for drug resistance in the most common yeast pathogen Candida albicans is an increased production of transport proteins, which are removing the drug from the cell cytosol and thus producing the phenomenon called multidrug resistance - MDR. The goal of this thesis was to verify the suitability of the yeast strain Saccharomyces cerevisiae with heterologously expressed MDR pumps from the pathogenic yeast Candida albicans and comparison to the results from C. albicans with homologous expression. The results that the azole drugs miconazole, bifonazole, and ketoconazole, together with potentiometric fluorescent probe diS-C3(3), are substrates of the CaCdr1p and CaCdr2p pumps, but not, or minimally, of the pump CaMdr1p, are consistent with the previously published work. The binding pocket of CaCdr1p was explored using the disc diffusion assay and the diS-C3(3) fluorescent probe...
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