Accessing the inner ear presents a significant challenge for the diagnosis and treatment of inner ear diseases. Many existing techniques to access the inner ear are invasive and can cause permanent damage to the cochlea. Recently, a novel microneedle has been fabricated to perforate the round window membrane (RWM) – a membrane sealing one of the two openings in the cochlea. These perforations enhance drug delivery into the inner ear, potentially improving the efficacy of therapeutics. Furthermore, they allow for the aspiration of perilymph samples, which is essential for diagnosing inner ear diseases. However, owing to limited knowledge about the mechanical properties of the RWM, certain technical aspects remain unexplored. Specifically, the interaction between the RWM and the microneedle during perforation is yet to be examined. This investigation is pivotal for the optimal design of microneedles — those robust enough to perforate RWMs yet delicate enough to minimize damage. In this thesis, we conduct a thorough examination of the guinea pig RWM, encompassing its geometry and its mechanical responses to pressures from the middle ear and inner ear. Additionally, we also formulate a comprehensive constitutive law for the guinea pig RWM. Our exploration begins with the creation of a U-Net model tailored to automatically segment the RWM. Despite the presence of other structures in the same image—such as bone, the basilar membrane, and ambient noise—the model proved invaluable for efficiently and automatically segmenting the RWM. To enhance accuracy, post-processing techniques like connected component analysis and majority voting were incorporated. Using this 3D model, we proceeded to study the RWM’s geometry. Recognizing the shrinkage observed in fixed RWMs, we integrated fresh RWM data to estimate the shrinkage ratio. Subsequently, we analyzed both the overall RWM thickness and that of the middle connective tissue layer—crucial metrics for future RWM modeling. Next, we proposed a method to evaluate the in-plane deformation of the RWM due to applied pressure. This involved using a bulge test system to pressurize and deform the RWM, combined with confocal microscopy to track stained nuclei or pre-introduced fluorescent beads on the RWM. We then utilized the coherent point drift (CPD) algorithm to measure the displacement of beads and nuclei. Results indicated that both markers could be successfully used to measure the RWM’s displacement. Further analysis revealed the in-plane Lagrangian strain of the RWM, with a significant observation being that the direction of maximum in-plane Lagrangian strain is perpendicular to the fiber direction. This underscores the crucial role of collagen fibers in determining the RWM’s mechanical properties. To conclude our study, we devised a constitutive law for the RWM, conceptualizing it as a combination of the ground substance and a family of dispersed fibers. This model was integrated into a FEBioStudio plugin, facilitating simulations of the RWM’s mechanical reactions to different pressures. Although our simulations closely aligned with experimental findings, some discrepancies were noted, likely stemming from an incomplete understanding of fiber dispersions. Nevertheless, our constitutive law reinforces the notion that fibers primarily govern the RWM’s mechanical characteristics.

The thesis deals with the issue of care of drug addicts in the in the Czech Republic, particularly the Harm Reduction. The theoretical part of the thesis describes the above-mentioned working procedures with clients, the legislative framework of the services and the current situation on the drug scene in the Czech Republic. The practical part uses a qualitative form of research. The information was collected by means of structured interviews with open questions. The respondents were eleven workers of contact centres in the Central Bohemian region which has five such facilities. I cooperated with four of them for the purposes of the thesis. The objective of the thesis was to demonstrate that the Harm Reduction services are important and beneficial from the viewpoint of workers of contact centres and to find out whether the contact workers have any suggestions about how to improve the Harm Reduction services in the Czech Republic.

Both testate amoebae and fungi are common inhabitants of coniferous litter. Their interactions in this environment were rarely studied, although they reach high biodiversity and can play a significant role in nutrient cycling in this environment. In this study, a cultivation of litter needles in the damp chambers was used to investigate interactions between fungi and testate amoebae. Observation of spruce litter needles in environmental scanning electron microscope was used to better characterize testate amoebae communities directly on the needles. Additionally, two experiments changing the biotic conditions in the microcosm were used to follow a principle of the interactions. Three species of testate amoebae from litter needles were able to colonize the filter paper on the bottom of the damp chambers. Occurrence of Phryganella acropodia and Assulina muscorum on the filter paper was significantly fuelled by the presence of mycelium. Assulina muscorum was associated with the fungal spores and Arcella discoides was attracted by sporulating colonies of Cladosporium spp. in the damp chambers. By contrast, no association of putatively mycophagous Phryganella acropodia with fungal spores was observed. Arcella discoides was attracted both by live and death mycelium in additional experiment. Interestingly, the...

The chemical elicitor methyl jasmonate (MeJA) could replace insecticides in Europe and Asia for protection of young conifers against the pine weevil (Hylobius abietis). This thesis mainly focuses on the effects of MeJA treatment on chemical defenses of conifers from seedling batches with documented field resistance. Tissues of three pine species and one spruce species, with various treatments, were here extracted in hexane, whereafter volatile contents of tissues were separated and analyzed by 2D GC-MS. Induced responses of seedlings of Maritime pine (Pinus pinaster) and Monterey pine (Pinus radiata) to the folivore pine processionary (Thaumetopoea pityocampa), and the phloem-feeder H. abietis, have been studied. Amounts of mono- and sesqui-terpenes (and also non-volatile resin) in conifer tissues (needles and phloem) were less induced by T. pityocampa than by H. abietis. The MeJA-treated seedlings of Scots pine (Pinus sylvestris) changed their composition of phloem monoterpenes (induction of (-)-β-pinene), and were better protected in field than the seedlings of Norway spruce (Picea abies), which increased their total amounts of monoterpenes. Orientation bioassays with H. abietis showed deterrent effects of (-)-β-pinene, (+)-3-carene, (-)-bornyl acetate and 1,8-cineole. Conversely, (-)-α-pinene (induced in P. abies but not in P. sylvestris) was non-deterrent. MeJA-treated seedlings fed on by H. abietis contained higher amounts of the H. abietis antifeedant 2-phenylethanol, in tissues and emissions. Phloem of control seedlings instead induced the (+)-α-pinene, which is one of the enantiomers of the H. abietis attractant α-pinene. In response to MeJA treatment, the relative amounts of the (+)-α-pinene increased in the phloem of  P. radiata, while it decreased in P. pinaster phloem. The preference of H. abietis in the field for P. radiata before P. pinaster may be explained by these changes in enantiomers of the H. abietis attractant α-pinene. / Barrträd är viktiga för skogsindustrin. I Sverige består 57% av ytan av produktiv skogsmark, och av dessa träd utgörs 80% av gran och tall. Som försvar mot insekter och sjukdomar, producerar barrträden kådämnen. För unga barrträd i Europa och Asien är snytbaggen (Hylobius abietis) en allvarlig skadegörare, som orsakar skogsnäringen stora kostnader. För att skydda små gran- och tallplantor före utplanteringen, skulle växthormonet metyljasmonat (MeJA) kunna användas. I denna avhandling har effekter av MeJA-behandling och insektsangrepp undersökts på de två viktigaste barrträden i Sverige, tall och gran (Pinus sylvestris och Picea abies) och på två arter i medelhavsområdet (Pinus radiata och Pinus pinaster). De kemiska försvarsämnena i barrträd har analyserats för att utvärdera effekterna av MeJA-behandling. Vävnader från barrträd med olika behandling har extraherats i organiska lösningsmedel och dofterna från plantorna har insamlats genom fastfas-mikroextraktion. De flyktiga ämnena har separerats och identifierats med hjälp av gaskromatografi och masspektrometri (GC-MS). Dessutom har optiskt aktiva doftämnen separerats med hjälp av en tvådimensionell GC-MS. Plantornas olika försvarsreaktioner har studerats vid angrepp av snytbaggen, som är en floemätare, och av tallfjärilslarv (Thaumetopoea pityocampa), som huvudsakligen äter barr. De lättflyktiga ämnena (mono- och seskviterpener) och de icke-flyktiga ämnena (kådsyror) ökade mer i stam och barr från angrepp av H. abietis än av T. pityocampa. Sammansättning av monoterpener ändrades i de MeJA-behandlade tallplantorna i Sverige, specifikt ökade (-)-β-pinene i stamfloemet. Tallplantorna var bättre skyddade i fält än granplantorna, där den totala mängden monoterpener ökade. (-)-β-Pinene, (+)-3-carene, (-)-bornyl acetate och 1,8-cineole visade sig ha en negativ effekt på snytbaggens orientering mot talldoft i orienteringstester. En av monoterpenerna, (-)-α-pinene, som av MeJA-behandlingen inducerats i granen, men inte i tallen, hade ingen negativ effekt på snytbaggen. MeJA-behandlade tallplantor som angripits av snytbaggen, innehöll större mängder av snytbagge-äthämmaren 2-fenyletanol, både i vävnader och i emissioner. Stam från kontrollplantor inducerade istället (+)-α-pinene, vilken är en av de optiska isomererna av snytbagge-attrahenten α-pinene. För de spanska arterna, som svar på MeJA-behandling, ökade de lättflyktigaste terpenerna i barren hos P. radiata men inte hos P. pinaster, vilken redan innehöll en hög terpenhalt. De relativa mängderna av (+)-α-pinene ökade också i stammen hos P. radiata, medan de minskade i P. pinaster. I fält väljer snytbaggen att gå till P. radiata- framför P. pinaster-plantor, vilket kan vara kopplat till de skillnader i α-pinene-isomerer, vilka här har observerats. / <p>QC 20160304</p>

KUBÍNOVÁ, Zuzana. The effect of elevated CO2 concentration and irradiation on quantitative parameters of mesophyll cells of Norway spruce. Prague, 2010. 74 p. Master's degree thesis. Faculty of Science, Charles University in Prague. Abstract The aim of the present thesis was to choose and adjust a suitable methodology for counting particles in 3D space, which would be suitable for unbiased estimation of chloroplast number in needle mesophyll cells. The disector method was used for the first time to determine the number of chloroplasts. This method enables unbiased estimation of chloroplast number in needle volume from optical sections captured from fresh free-hand sections by confocal microscope. The sections did not need any pre-processing. Another aim was to compare selected photosynthetic and anatomical characteristics of sun and shade Norway spruce needles, which were grown under different CO2 concentration. The trees were grown for eight years in ambient (during the experiment increasing from 357 up to 370 µmol CO2 ∙ mol-1 ) CO2 concentration or elevated (700 µmol ∙ mol-1 ) CO2 concentration in special glass domes on an experimental research site of the Institute of Systems Biology and Ecology, Academy of Sciences of the Czech Republic at Bílý Kříž in Moravskoslezské Beskydy mountains. The sun needles...