The subject of this thesis is five-storey concrete basement multifunctional object. The building plot is located in Brno. The Multifunctional object is designed with the view of the location. The Object will be used for trade and services and for living of a families. The plan view of the Object is letter L. The main entrance and the driveway to the Object is at 1 st ground floor level. The vertical supporting structures are formed by concrete pillars and walls that are insulated on the above floors. The ceiling structure is made of concrete monolithic panels. The multifunctional object covers a flat single-skin roof. The project was prepared by a computer program called Autocad.

The diploma thesis deals with a project documentation of the building kindergartens in Brno, on the estate Lesná. The design and architectural design of the building is based on a combination of mutually intersecting blocks, while in the spatial division of the building is put great emphasis on simple rectangular shapes. Building facade is completed unusual modifications that are in perfect harmony, and also contrast with the overall shape design construction. Business can complete single-storey building comprising two independent departments divided into four functional zones: preschool education, social, administrative and economic. Specializations in appropriate field are a part of the project.

The diploma thesis deals with a project documentation of single-storey building kindergarten in Hluk. The purpose is to make functional layout solution for everyday activity. New building contains from two parts for forty - eight children. Place emphasis on general visual impression and construction solution. The building is designed from constructional system KM BETA. The part of this project is specialization in specific field.

Data streams being transmitted over a network channel with capacity less than the data rate of the data streams is very common when using network channels such as dial-up, low bandwidth wireless links. Not only does this lower capacity creates delays but also causes sequential network problems such as packet losses, network congestion, errors in data packets giving rise to other problems and creating a cycle of problems hard to break out from. In this thesis, we present a new approach for shedding the less informative attribute data from a data stream with a fixed schema to maintain a data rate lesser than the network channels capacity. A scheme for shedding attributes, instead of tuples, becomes imperative in stream data where the data for one of the attributes remains relatively constant or changes less frequently compared to the data for the other attributes. In such a data stream management system, shedding a complete tuple would lead to shedding of some informative-attribute data along with the less informative-attribute data in the tuple, whereas shedding of the less informative-attribute data would cause only the less informative data to be dropped. In this thesis, we deal with two major problems in load shedding: the intra-stream load shedding and the inter-stream load shedding problems. The intra-stream load shedding problem deals with shedding of the less informative attributes when a single data stream with the data rate greater than the channel capacity has to be transmitted to the destination over the channel. The inter-stream load shedding problem refers to shedding of attributes among different streams when more than one stream has to be transferred to the destination over a channel with the channel capacity less than the combined data rate of all the streams to be transmitted. As a solution to the inter-stream or intra-stream load shedding problem, we apply our load shedding schema approach to determine a ranking amongst the attributes on a singe data stream or multiple data streams with the least informative attribute(s) being ranked the highest. The amount of data to be shed to maintain the data rate below the capacity is calculated dynamically, which means that the amount of data to be shed changes with any change in the channel capacity or any change in the data rate. Using these two pieces of information, a load shedding schema describing the attributes to be shed is generated. The load shedding schema is generated dynamically, which means that the load shedding schema is updated with any change in (i) the rankings of attributes that capture the rate of change on the values of each attribute, (ii) channel capacity, and (iii) data rate even after load shedding has been invoked. The load shedding schema is updated using our load shedding schema re-evaluation algorithm, which adapts to the data stream characteristics and follows the attribute data variation curve of the data stream. Since data dropped at the source may be of interest to the user at the destination, we also propose a recovery module which can be invoked to recover attribute data already shed. The recovery module maintains the minimal amount of information about data already shed for recovery purpose. Preliminary experimental results have shown that recovery accuracy ranges from 90% to 99%, which requires only 5% to 33% and 4.88% to 50% of the dropped data to be stored for weather reports and stock exchanges, respectively. Storing of recovery information imposes storage and processing burden on the source site, and our recovery method aims at satisfactory recovery accuracy while imposing minimal burden on the source site. Our load shedding approach, which achieves a high performance in reducing the data stream load, (i) handles wide range of data streams in different application domains (such as weather, stocks, and network performance, etc.), (ii) is dynamic in nature, which means that the load shedding scheme adjusts the amount of data to be shed and which attribute data to be shed according to the current load and network capacity, and (iii) provides a data recovery mechanism that is capable to recover any shedded attribute data with recovery accuracy up to 90% with very low burden on the source site and 99% with a higher burden on some stream data. To the best of our knowledge, the dynamic load shedding scheme we propose is the first one in the literature to shed attributes, instead of tuples, along with providing a recovery mechanism in a data stream management system. Our load shedding approach is unique since it is not a static load shedding schema, which is less appealing in an ever-changing (sensor) network environment, and is not based on queries, but works on the general characteristics of the data stream under consideration instead.

Increases in herd size and enforcement of water quality regulations have created an effluent disposal problem for the New Zealand dairy industry. Spray disposal to land and lagooning are commonly used but mechanical failures, management requirements and pressure on land have limited their suitability in many situations. This project was established to consider an alternative system. Initial studies revealed that anaerobic treatment in unmixed, non-insulated tanks, followed by trickling filter aeration, might be suitable. Two laboratory scale and one field treatment plant (1/15 - 1/20 full scale) were constructed to investigate the system. A factorial experimental design allowed investigation into three anaerobic treatment levels with a 3 x 3 aerobic treatment interaction nested within each anaerobic treatment. Anaerobic residence times of 5, 7.5 and 10 days provided loading rates of 1.35 - 0.63 kg COD/m3-day and 1.36 - 0.67 kg T S/m3-day. Removals between inlet and outlet averaged 71% and were insensitive to loading rate. Total solids accumulation rates of 40-50% TS input rate suggests that anaerobic tank design should be based on solids accumulation rate and cleaning frequency. The stone media trickling filter was loaded at approximately 0.61 kg COD/m3-day. Aeration periods of 1, 2 and 3 days and hydraulic loads of 2.8, 10.1 and 18.2 m3/m2-day were studied to determine their influence on treatment efficiency. Multiple regression analysis indicated that the longer residence times and higher recycle rates improved treatment efficiency. Removals varied with the measured parameters but ranged from 42-66% for COD. Design alterations to allow the final discharge to be taken from the bottom of the filter, after settling, would increase aerobic treatment efficiency above 75% COD removal. Prediction of treatment efficiencies beyond the monitored operating conditions suggested that only marginal improvements could be made. The TS accumulation rate in the aerobic phase was approximately 13% of the TS input rate or 56% of the BOD removal rate. Overall plant treatment efficiencies of 80-89% were obtained. Removals in excess of 92% could be achieved with minor design alterations. Maintenance and operational requirements were minimal. The only problem with the system was an average 15 fold increase in NO3-N and 4 fold increase in DIP under conditions for optimum removal of the other parameters. Intermittent land disposal could reduce this problem. Treatment comparison between similar laboratory plants, and between laboratory and field plants which varied by a scale factor of 56, suggests that identically designed plants would give a similar performance and that there is little scale effect. Increasing the scale only improved treatment efficiencies under unstable aerobic conditions, i.e., high recycle rates and low residence times. Increasing scale gave some decrease in maintenance and operational problems. Design of a full scale plant, based on daily pollution loads from a 250 cow dairy shed, suggests that the system is a viable proposition.

O presente estudo utilizou o modelo fatia-dental/matriz-polimérica para avaliar a influência do tratamento dentinário e das BMPs dentinárias na diferenciação das células-tronco da polpa de dentes decíduos (SHED). Secções transversais (1mm) foram preparadas a partir de terceiros molares humanos extraídos. Matrizes poliméricas a base de ácido poli-L-lático (PLLA) foram criadas no interior da cavidade pulpar das secções dentinárias, tratadas com solução de EDTA a 10%; NaOCl a 5.25%; ou permanecendo sem tratamento. Matrizes poliméricas confeccionadas sem as fatias dentais foram utilizadas como controle. As células (5x104) foram semeadas nas matrizes e, após 7, 14, 21 e 28 dias de cultura in vitro, a expressão de marcadores de diferenciação odontoblástica (DSPP, DMP1 e MEPE) e a proliferação celular (WST-1) foram avaliadas. Células (5x105) semeadas nas matrizes foram transplantadas em camundongos imunodeficientes e cultivadas in vivo por um período de 14 e 28 dias. Para avaliar a atividade das BMPs dentinárias, 5x104 células foram semeadas em matrizes poliméricas com fatia dental e cultivadas na presença de anticorpos anti-BMP- 2, -4, ou -7 (2 μg/ml) durante 14 dias. Adicionalmente, 5x105 células foram tratadas com rhBMP-2, -4, ou -7 (100ng/mL) por 24hs. As células cultivadas in vitro e in vivo alteraram sua expressão genética durante o curso do tempo. DSPP, DMP-1 e MEPE foram expressos por células cultivadas in vitro após 14 dias (tratamento com EDTA e dentina sem tratamento) e in vivo após 28 dias (EDTA), não sendo detectados nos grupos NaOCl e nas células cultivadas nas matrizes sem fatia dental. A proliferação foi reduzida com a diferenciação celular (p<0.05). A utilização de BMP-2/4Ab no meio de cultura exerceu um efeito inibitório na expressão dos marcadores de diferenciação celular, não ocorrendo quando do cultivo das SHED na presença de BMP-7Ab. DSPP, DMP-1 e MEPE foram expressos por células tratadas com rhBMP-2, e DSPP e DMP-1 por células tratadas com rhBMP-4 e -7. Células sem tratamento não expressaram os marcadores. O modelo fatia-dental/matriz-polimérica demonstrou ser adequado para o estudo da diferenciação de células-tronco pulpares, sugerindo que a dentina possa fornecer um microambiente favorável para a diferenciação de celular. As proteínas ósseas morfogenéticas dentinárias BMP-2 e BMP-4 parecem exercer um papel relevante nesse processo. / The effect of dentin pre-treatments and dentin-derived BMPs on SHED differentiation was tested using the Tooth-Slice Scaffold model (TSS). Dentin slices (1mm thickness) were prepared from extracted human third molars. Biodegradable PLLA scaffolds were prepared inside the pulp chamber of the tooth-slices, treated alternatively with a 5.25% NaOCl or 10% EDTA solution, or remaining untreated (WO-T). PLLA sponge scaffolds with no tooth-slice (PSS) were used as control. SHED (5x104) were seeded in TSS and PSS and after 7, 14, 21 and 28 days in culture, RT-PCR (DSPP, DMP1 and MEPE) and WST-1 proliferation assay were performed. Additionally, cells (5x105) were seeded in TSS and PSS and transplanted into SCID mice (14 and 28 days). To verify the dentinderived BMPs bioactivity, SHED (5x104) were cultured in TSS in the presence of antihuman BMP-2, -4, and -7 antibodies for 14 days. Besides, cells in culture were treated with rhBMP-2; -4; or -7 for 24 hours. After in vitro and in vivo time course, SHED altered their genetic expression. The cells cultured in vitro in the TSS (EDTA or WO-T) expressed the differentiation markers after 14 days and maintained expression thereafter. Cell proliferation rate was reduced following the differentiation (p<0.05). Cells transplanted in vivo expressed DSPP, DMP-1 and MEPE after 28 days (EDTA). No transcripts were found in tooth-slices treated with NaOCl or in PSS groups. BMP-2/4Ab prevented the differentiation process and no inhibitory effect was detected for BMP-7Ab. After 24 hours, expression of DSPP, DMP-1 and MEPE was found for rhBMP-2, and DSPP and DMP-1 for rhBMP-4 and rhBMP-7 treated SHED, but not for untreated cells. The tooth slice scaffold model suggests that dentin can provide the environment for SHED differentiation and dentin-derived morphogenic signals BMP-2 and BMP-4 play an important role in this process.

Introdução: Células tronco mesenquimais (CTM) são uma população residente nos tecidos adultos de origem mesodérmica, com funções regenerativas de manutenção da integridade tecidual, com destaque no desempenho imunomodulador. Esse aspecto levou as CTM a tornarem-se ferramentas terapêuticas valiosas da pesquisa à assistência ao paciente em doenças autoimunes e de cunho inflamatório. Além disso, CTM podem ser isoladas de materiais tidos como descarte de procedimentos, como dentes decíduos, filtros de transplante de medula óssea e gordura. Nesse panorama, torna-se necessário estabelecer o efeito que a origem tecidual tem na eficiência imunoreguladora e na possível aplicabilidade clínica destas células. Objetivo: Comparar o potencial imunomodulador de células mesenquimais isoladas a partir de filtros descartados após a infusão de medula óssea, de lipoaspirado e de polpa de dentes decíduos. Métodos: Foi realizada a comparação da capacidade proliferativa de CTMs, cultivadas na presença de lisado plaquetário, das diversas fontes através do cálculo de population doubling das CTM em co-cultura com linfócitos T isolados em coluna magnética e com células mononucleares de sangue periférico, estimuladas com fitohemaglutinina; e determinado por citometria de fluxo o efeito das CTM das diversas fontes sobre as subpopulações linfocitárias. Resultados: CTM das três fontes foram capazes de inibir a proliferação de linfócitos e CTM de tecido adiposo foram mais eficientes em induzir o fenótipo de células T reguladoras e na diminuição de células T citotóxicas. Conclusão: comparadas à CTM isoladas de medula óssea e de polpa dentária, as CTM originadas de tecido adiposo exibem efeito imunomodulador mais acentuado. / Background: Mesenchymla stromal cells (MSC) reside in most adult tissue of mesenchymal origen, with a broad functions envolving cell repopulation and maintenence of tissue homeostasis, trough immunemmodulatory action. MSC are valuable terapêutic instruments applied from research to autoimune and inflamatory diseases. MSC can be isolated from diverse discarted biological matherials, like lipoaspirate, exfoliated deciduous teeeth and boné marrow ransplant filters. There so it´s necessary to stablish how source can impact MSC efficiency and possible clinical aplications. Objective: Compare immunomodulatory potential of adipose MSC and dental pulp MSC to boné marrow MSC. Methods: MSC from three selected sources were cocultured with phytohemaglutinin stimulated and magnetically isolated T cells and peripheral blood mononuclear cells; immunephenotype of cocultivated lymphocytes were also conducted. Results: MSC from all analyzed sources were capable to inhibit lymphocyte proliferation. Adipose MSC were capable to induce Treg phenotype and decrease T CD8+ limphocytes. Conclusion: Cell culture and therapy with MSC present many paradigms and we address to some of those to elucidate the possible most efficient source.

O presente estudo utilizou o modelo fatia-dental/matriz-polimérica para avaliar a influência do tratamento dentinário e das BMPs dentinárias na diferenciação das células-tronco da polpa de dentes decíduos (SHED). Secções transversais (1mm) foram preparadas a partir de terceiros molares humanos extraídos. Matrizes poliméricas a base de ácido poli-L-lático (PLLA) foram criadas no interior da cavidade pulpar das secções dentinárias, tratadas com solução de EDTA a 10%; NaOCl a 5.25%; ou permanecendo sem tratamento. Matrizes poliméricas confeccionadas sem as fatias dentais foram utilizadas como controle. As células (5x104) foram semeadas nas matrizes e, após 7, 14, 21 e 28 dias de cultura in vitro, a expressão de marcadores de diferenciação odontoblástica (DSPP, DMP1 e MEPE) e a proliferação celular (WST-1) foram avaliadas. Células (5x105) semeadas nas matrizes foram transplantadas em camundongos imunodeficientes e cultivadas in vivo por um período de 14 e 28 dias. Para avaliar a atividade das BMPs dentinárias, 5x104 células foram semeadas em matrizes poliméricas com fatia dental e cultivadas na presença de anticorpos anti-BMP- 2, -4, ou -7 (2 μg/ml) durante 14 dias. Adicionalmente, 5x105 células foram tratadas com rhBMP-2, -4, ou -7 (100ng/mL) por 24hs. As células cultivadas in vitro e in vivo alteraram sua expressão genética durante o curso do tempo. DSPP, DMP-1 e MEPE foram expressos por células cultivadas in vitro após 14 dias (tratamento com EDTA e dentina sem tratamento) e in vivo após 28 dias (EDTA), não sendo detectados nos grupos NaOCl e nas células cultivadas nas matrizes sem fatia dental. A proliferação foi reduzida com a diferenciação celular (p<0.05). A utilização de BMP-2/4Ab no meio de cultura exerceu um efeito inibitório na expressão dos marcadores de diferenciação celular, não ocorrendo quando do cultivo das SHED na presença de BMP-7Ab. DSPP, DMP-1 e MEPE foram expressos por células tratadas com rhBMP-2, e DSPP e DMP-1 por células tratadas com rhBMP-4 e -7. Células sem tratamento não expressaram os marcadores. O modelo fatia-dental/matriz-polimérica demonstrou ser adequado para o estudo da diferenciação de células-tronco pulpares, sugerindo que a dentina possa fornecer um microambiente favorável para a diferenciação de celular. As proteínas ósseas morfogenéticas dentinárias BMP-2 e BMP-4 parecem exercer um papel relevante nesse processo. / The effect of dentin pre-treatments and dentin-derived BMPs on SHED differentiation was tested using the Tooth-Slice Scaffold model (TSS). Dentin slices (1mm thickness) were prepared from extracted human third molars. Biodegradable PLLA scaffolds were prepared inside the pulp chamber of the tooth-slices, treated alternatively with a 5.25% NaOCl or 10% EDTA solution, or remaining untreated (WO-T). PLLA sponge scaffolds with no tooth-slice (PSS) were used as control. SHED (5x104) were seeded in TSS and PSS and after 7, 14, 21 and 28 days in culture, RT-PCR (DSPP, DMP1 and MEPE) and WST-1 proliferation assay were performed. Additionally, cells (5x105) were seeded in TSS and PSS and transplanted into SCID mice (14 and 28 days). To verify the dentinderived BMPs bioactivity, SHED (5x104) were cultured in TSS in the presence of antihuman BMP-2, -4, and -7 antibodies for 14 days. Besides, cells in culture were treated with rhBMP-2; -4; or -7 for 24 hours. After in vitro and in vivo time course, SHED altered their genetic expression. The cells cultured in vitro in the TSS (EDTA or WO-T) expressed the differentiation markers after 14 days and maintained expression thereafter. Cell proliferation rate was reduced following the differentiation (p<0.05). Cells transplanted in vivo expressed DSPP, DMP-1 and MEPE after 28 days (EDTA). No transcripts were found in tooth-slices treated with NaOCl or in PSS groups. BMP-2/4Ab prevented the differentiation process and no inhibitory effect was detected for BMP-7Ab. After 24 hours, expression of DSPP, DMP-1 and MEPE was found for rhBMP-2, and DSPP and DMP-1 for rhBMP-4 and rhBMP-7 treated SHED, but not for untreated cells. The tooth slice scaffold model suggests that dentin can provide the environment for SHED differentiation and dentin-derived morphogenic signals BMP-2 and BMP-4 play an important role in this process.

Introdução: Células tronco mesenquimais (CTM) são uma população residente nos tecidos adultos de origem mesodérmica, com funções regenerativas de manutenção da integridade tecidual, com destaque no desempenho imunomodulador. Esse aspecto levou as CTM a tornarem-se ferramentas terapêuticas valiosas da pesquisa à assistência ao paciente em doenças autoimunes e de cunho inflamatório. Além disso, CTM podem ser isoladas de materiais tidos como descarte de procedimentos, como dentes decíduos, filtros de transplante de medula óssea e gordura. Nesse panorama, torna-se necessário estabelecer o efeito que a origem tecidual tem na eficiência imunoreguladora e na possível aplicabilidade clínica destas células. Objetivo: Comparar o potencial imunomodulador de células mesenquimais isoladas a partir de filtros descartados após a infusão de medula óssea, de lipoaspirado e de polpa de dentes decíduos. Métodos: Foi realizada a comparação da capacidade proliferativa de CTMs, cultivadas na presença de lisado plaquetário, das diversas fontes através do cálculo de population doubling das CTM em co-cultura com linfócitos T isolados em coluna magnética e com células mononucleares de sangue periférico, estimuladas com fitohemaglutinina; e determinado por citometria de fluxo o efeito das CTM das diversas fontes sobre as subpopulações linfocitárias. Resultados: CTM das três fontes foram capazes de inibir a proliferação de linfócitos e CTM de tecido adiposo foram mais eficientes em induzir o fenótipo de células T reguladoras e na diminuição de células T citotóxicas. Conclusão: comparadas à CTM isoladas de medula óssea e de polpa dentária, as CTM originadas de tecido adiposo exibem efeito imunomodulador mais acentuado. / Background: Mesenchymla stromal cells (MSC) reside in most adult tissue of mesenchymal origen, with a broad functions envolving cell repopulation and maintenence of tissue homeostasis, trough immunemmodulatory action. MSC are valuable terapêutic instruments applied from research to autoimune and inflamatory diseases. MSC can be isolated from diverse discarted biological matherials, like lipoaspirate, exfoliated deciduous teeeth and boné marrow ransplant filters. There so it´s necessary to stablish how source can impact MSC efficiency and possible clinical aplications. Objective: Compare immunomodulatory potential of adipose MSC and dental pulp MSC to boné marrow MSC. Methods: MSC from three selected sources were cocultured with phytohemaglutinin stimulated and magnetically isolated T cells and peripheral blood mononuclear cells; immunephenotype of cocultivated lymphocytes were also conducted. Results: MSC from all analyzed sources were capable to inhibit lymphocyte proliferation. Adipose MSC were capable to induce Treg phenotype and decrease T CD8+ limphocytes. Conclusion: Cell culture and therapy with MSC present many paradigms and we address to some of those to elucidate the possible most efficient source.

Ce travail porte sur le rôle des microparticules procoagulantes (MPs) générées par les cellules vasculaires en réponse à un état septique. Après une introduction rappelant la structure et les propriétés des microparticules et la réponse del’hôte à un agent pathogène, en particulier en terme d’activation de la coagulation, nous rapportons nos travauxexpérimentaux et cliniques. Le premier travail a été réalisé sur un modèle cellulaire de vésiculation induite par le LPS. Il nous a permis de caractériser le transfert du complexe CD14/TLR4 à différents types cellulaires in-vitro dépourvus du récepteur au LPS. Ainsi, les MPs monocytaires pourraient avoir un rôle d’amplification de la réponse inflammatoire mais aussi dans la réponse anti-inflammatoire secondaire en participant à l’apoptose lymphocytaire. Le second travail aété réalisé chez l’animal. Après induction d’un choc septique, nous avons observé une amélioration hémodynamique enréponse à la perfusion de protéine C activée associée à une modulation du phénotype des MPs. Réinjectées à des ratsnaïfs, les MPs issues des rats septiques traités par protéine C activée développaient une moindre vasoplégie. Enfin, nous avons réalisé une étude prospective sur 100 patients en choc septique. Nous avons ainsi pu caractériser la présence d’une concentration élevée de microparticules procoagulantes, avec une variation phénotypique en présence de coagulation intravasculaire disséminée (CIVD) : réduction du contingent plaquettaire au profit des MPs d’origine leucocytaires qui deviennent prépondérantes et témoignent d’une activation leucocytaire accrue, et surtout une activation des cellules endothéliales avec génération de MPs porteuses d’endogline (CD105). En analyse multivariée,CD105+-MPs étaient fortement associée à la CIVD et pourraient constituer un marqueur précoce de l’atteinte endothéliale au cours du choc septique. / This work focused on procoagulant microparticles shed after vascular cells stress during sepsis. The first part gives an overview on MPs and host response during pathogen challenge. The first lab experimental work confirms direct and functional transfer of CD14/TLR4 LPS sensor by MPs shed to target cells after monocytic THP-1 challenge by LPS.CD14-MPs amplify LPS-induced apoptosis in monocytes but also prompted lymphocyte apoptosis and could play a role in secondary anti-inflammatory response. Then, septic shock was induced in rats after caecal ligature and puncture.Activated protein C (APC) infusion improved haemodynamic parameters and alter septic microparticular content. Infused in naïve rats, APC-treated MPs were associated with reduced hypotension and inflammatory response, confirming cytoprotective effect of both APC and APC-induced MPs. Finally, we performed a clinical prospective study in 3 medical ICU in France. Patients referred for septic shock had an increased level of circulating procoagulant MPs regardless disseminated intravascular coagulopathy (DIC) diagnosis. Nevertheless, DIC patients evidenced a specific pattern with lower platelet-MPs, increased leucocyte-MPs and specific endothelial cells activation with endoglin (CD105) shedding. In multiple logistic regression analysis, CD105-MPs were strongly associated with DIC and were evidenced before DIC diagnosis according to routine laboratory assays.