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Physical factors influencing the development of corneal damage by UV rays
Čejka, Čestmír ; Rosina, Jozef (advisor) ; Štípek, Stanislav (referee) ; Amler, Evžen (referee) ; Salminen, Lotta (referee)
Purpose: The purpose of the Thesis "Physical factors influencing the development of corneal damage by UV rays" was a) to determine the light absorption properties in the rabbit cornea irradiated with UVB rays by measuring of physical values of absorbance A and transmittance T as functions of wavelength ; b) To investigate the absorption coefficient as a function of wavelength ; c) To evaluate the importance of this coefficient for corneal light absorption properties; d) To determine the light absorption properties in the rabbit cornea irradiated with UVA by measuring of physical values of absorbance A and transmittance T as functions of wavelength (to compare the effect of UVA with UVB rays). Material and Methods: To achieve individual aims, the new spectrohotometrical method was developed and following experimets were performed: a) Repeted irradiation of the rabbit cornea with UVB rays (daily dose 1.01 J/cm2 during 5 days); b) Repeated irradiation of the rabbit cornea with UVA rays (daily dose 1.01 J/cm2 or 2.02 J/cm2) during five days; c) Repeated irradiation of the rabbit cornea with the daily dose of 1.01 J/cm2 during 4 days and in individual time intervals investigation of corneal light absorption and hydration changes; d) The anti-UV efficacy of UV filter (actinoquinol combined with hyaluronic acid,...
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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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Control of creatine kinase activity due to structural changes of the enzyme molecule
Horníková, Daniela ; Mejsnar, Jiří (advisor) ; Amler, Evžen (referee) ; Kolář, František (referee)
74 5. ZÁVĚR Práce prokázala adekvátními chemickými a fyzikálními metodami následující vlastnosti myofibrilární kreatinkinázy (EC 2.7.3.2) (CK), vyčistěné z čisté myofibrilární frakce krysího a králičího svalu m. psoas. 1) Sekvenční 2D SDS elektroforéza odhalila tři izoelektrické body v úzkém rozsahu fyziologického pH 7.17, 7.28 a 7.47. Specifická enzymatická aktivita CK byla 82.35 IU/mg. 2) Rychlost výměny fluorescenčně značené CK-IAF mezi M-linií izolovaných myofibril a okolím (nepřímo úměrně vypovídající o síle vazby CK molekuly v M-linii), měřená v rovnovážném stavu metodou FLIP, se zpomaluje okyselováním okolního prostředí v rozmezí pH 6.5 - 7.2. 3) Rychlost výměny CK-IAF molekuly je ovlivněna přítomností substrátů (pH 7.10 - 7.17), když rychlosti CK se substráty (konformace "closed" a "intermediary") jsou průkazně sníženy oproti rychlosti CK bez substrátů (v konformaci "open"). Specifická enzymatická aktivita při těchto pokusech se snižovala podle vazby v myofibrilách až o dva řády k hodnotě 0.40 IU/mg. 4) Měření steady-state fluorescence vnitřních tryptofanových zbytků neznačené CK prokázala a) trojí pokles fluorescence v čase podle vazby substrátů, b) změnu střední doby života lifetime 2.72 ns volné CK, vlivem vazby ATP (2.38 ns) a vazbou ATP+Cr (2.42 ns). 5) Zhášení fluorescence vnitřních...
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Collagen structures from cell culture to intact tendon
Hadraba, Daniel ; Jelen, Karel (advisor) ; Amler, Evžen (referee) ; Plášek, Jaromír (referee)
CHARLES UNIVERSITY and HASSELT UNIVERSITY / tUL Doctoral dissertation Collagen structures from cell culture to intact tendon ABSTRACT Author: Daniel Hadraba Promoters: Assoc. Prof. Karel Jelen | Charles University Prof. Marcel Ameloot | Hasselt University Co-promoters: Dr. Frantisek Lopot | Charles University Prof. Virginie Bito | Hasselt University Annotation Author: Ing. Mgr. Daniel Hadraba Doctoral thesis title: Collagen structures from cell culture to intact tendon Year: 2010 - 2017 Doctoral program: Doctor of Biomechanics at Charles University Doctor of Biomedical Science at Hasselt University / transnational University Limburg Departments: Dept. Anatomy and Biomechanics | Faculty of Physical Education and Sport | Charles University Dept. Biophysics | Hasselt University Promoters: Assoc. Prof. Karel Jelen | Dept. Anatomy and Biomechanics | Faculty of Physical Education and Sport | Charles University Prof. Marcel Ameloot | Hasselt University / transnational University Limburg Co-promoters: Dr. Frantisek Lopot | Dept. Anatomy and Biomechanics | Faculty of Physical Education and Sport | Charles University Prof. Virginie Bito | Hasselt University / transnational University Limburg Bibliography details: Pages 102 Figures 30 Tables 2 Equations 17 Keywords: tendon, collagen, crimps, orientation, aging,...
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Preparation, characterization and testing of blood derivatives for applications in regenerative medicine
Sovková, Věra ; Amler, Evžen (advisor) ; Kolářová, Hana (referee) ; Vránová, Jana (referee)
Platelet products can be used, thanks to the broad range of bioactive molecules, either as a supplement for cell cultering in vitro alone or for development of cell- or cell-free scaffolds in diverse fields in regenerative medicine. The aim of this study was to prepare several types of platelet products. The concentration of selected molecules were observed. Subsequently, these products were tested with cell cultures in vitro alone or in combination with nanofibres scaffolds prepared by electrospinning or centrifugal spinning. It was found out, that platelets products contains chemokine RANTES and growth factor PDGF in the highest concentrations. It was further discovered the content of pro and antiinflammatory in terleukins and other growth factors. Platelet lysat in concentration 7% is sufficient to replace FBS in keratinocytes and fibroblasts cultures. In the other experiments, platelets in different concentrations were adhered to the scaffolds prepared by electrospinning and centrifugal spinning. Thus prepared scaffolds promote the proliferation and viability of all tested cell types in dose-dependent manner. In the last experiment, the individual components of platelet concentrate were separated and characterized. Their effect to the cell culture were tested. It was examinated the synergic...
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Intelligent nanofibres functionalized with growth factors and blood derivatives for dermatology applications
Vocetková, Karolína ; Amler, Evžen (advisor) ; Rosina, Jozef (referee) ; Arenberger, Petr (referee)
Platelet derivatives are an attractive source of natural growth factors and they are widely used in various tissue engineering and regenerative medicine applications. The aim of this study was to optimize cell culture conditions using platelet lysate and to develop platelet-functionalized fibrous scaffolds as a controlled drug delivery system for native growth factors. Fibrous scaffolds were prepared by electrostatic and centrifugal spinning of PCL and they were functionalized by the platelets by surface adhesion or their encapsulation using emulsion spinning techniques. The cell culture study determined the 7% platelet lysate to be the optimum concentration as a medium supplement in keratinocyte and fibroblast culture. Additionally, following surface adhesion of the platelets to PCL electrospun nanofibres, the platelets were activated due to their contact with the nanofibre nanotopography, resulting in formation of fibrin network. Fibrin served as a reservoir of the growth factors, prolonging the half-time of EGF release to 1.7 days. Such platelet-functionalized samples fostered proliferation of keratinocytes, fibroblasts and melanocytes. Furthermore, adhesion of platelets to centrifugally spun nanofibrous scaffolds resulted in almost two-fold increase in the amount of immobilized platelet-derived...
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Nanodiamonds as an innovative system for intracellular delivery of mirna-34a inprostatic cancer therapy
Bitti, G. ; Abate, M. ; Neuhoferová, Eva ; Kindermann, Marek ; Petráková, V. ; Boccellino, M. ; Quagliuolo, L. ; Filová, Eva ; Benson, Veronika ; Caraglia, M. ; Amler, Evžen
The microRNA(miRNA)-34a is an important regulator of tumor suppression. It controls the expression of several target proteins involved in cell cycle, differentiation and apoptosis, and antagonizes processes that are necessary for basic cancer cell viability as well as cancer stemness, metastasis, and chemoresistance. It is downregulated in numerous cancer types, including prostatic cancer, and inhibits malignant growth by repressing genes involved in various oncogenic signaling pathways. Given the anti-oncogenic activity of miR-34a, here we proved the substantial benefits of a new therapeutic concept based on nanotechnology delivery of miRNA mimics. In order to monitor the miRNA-34a replacement, we used a fluorescent nanodiamond particles (FND) system with linked miRNA-34a mimic, which was delivered to PC3 and DU145 prostatic cancer cell lines. We used functionalized nanodiamonds coated with polyethylenimine to transfer miRNA-34a into PC3 and DU145 prostatic cancer cell lines and we measured the zeta-potential of these complexes before using them for in vitro experiments. A replacement of miRNA-34 was observed by monitoring levels of miRNA-34 via real-time PCR. Moreover, our in vitro experiments demonstrated that miRNA-34a replacement, using this FND delivery system, decreased viability and induced apoptosis in prostatic cancer cell lines. Our findings suggest the replacement of oncosuppressor miRNA-34a provides an effective strategy for cancer therapy and the FND-based delivery systems seems to be an excellent strategy for a safe and effective targeting of the tumor.
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Study of transport systems of microorganisms
Jančíková, Iva ; Gášková, Dana (advisor) ; Amler, Evžen (referee) ; Krůšek, Jan (referee)
Title: Study of transport systems of microorganisms Author: Mgr. Iva Jančíková Department: Institute of Physics of Charles University Supervisor: Assoc. Prof. RNDr. Dana Gášková, CSc., Institute of Physics Abstract: Overexpression of drug efflux pumps is responsible for a multidrug resistance (MDR). We used the potentiometric fluorescent probe diS-C3(3), which is a substrate of major MDR pumps in three yeast species, Saccharomyces cerevisiae (ScPdr5p, ScSnq2p), Kluyveromyces lactis (KlPdr5p) and Candida albicans (CaCdr1p, CaCdr2p), to monitor inhibition of selected membrane transporters caused by various chemical stressors (diS-C3(3) assay). The extent of inhibition of probe transport points to a tighter arrangement of the K1Pdr5p binding pocket compared to that of ScPdr5p. Furthermore, we discovered that while deletion of the KlPDR16 gene does not affect K1Pdr5p activity, it only caused cell hyperpolarization, deletion of the KlERG6 gene results in both change in membrane potential and in a suppression of the pump's activity. We developed an effective method to search for inhibitors of MDR proteins of. C. albicans, which is based on pre-screening their potential to block the probe efflux from S. cerevisiae cells. Using this method we identified the substance H, derivative of 1,4-dihydropyridine, which...
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Collagen structures from cell culture to intact tendon
Hadraba, Daniel ; Jelen, Karel (advisor) ; Amler, Evžen (referee) ; Plášek, Jaromír (referee)
CHARLES UNIVERSITY and HASSELT UNIVERSITY / tUL Doctoral dissertation Collagen structures from cell culture to intact tendon ABSTRACT Author: Daniel Hadraba Promoters: Assoc. Prof. Karel Jelen | Charles University Prof. Marcel Ameloot | Hasselt University Co-promoters: Dr. Frantisek Lopot | Charles University Prof. Virginie Bito | Hasselt University Annotation Author: Ing. Mgr. Daniel Hadraba Doctoral thesis title: Collagen structures from cell culture to intact tendon Year: 2010 - 2017 Doctoral program: Doctor of Biomechanics at Charles University Doctor of Biomedical Science at Hasselt University / transnational University Limburg Departments: Dept. Anatomy and Biomechanics | Faculty of Physical Education and Sport | Charles University Dept. Biophysics | Hasselt University Promoters: Assoc. Prof. Karel Jelen | Dept. Anatomy and Biomechanics | Faculty of Physical Education and Sport | Charles University Prof. Marcel Ameloot | Hasselt University / transnational University Limburg Co-promoters: Dr. Frantisek Lopot | Dept. Anatomy and Biomechanics | Faculty of Physical Education and Sport | Charles University Prof. Virginie Bito | Hasselt University / transnational University Limburg Bibliography details: Pages 102 Figures 30 Tables 2 Equations 17 Keywords: tendon, collagen, crimps, orientation, aging,...
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