The cellular prion protein (PrPc) is essential for pathogenesis of fatal neurodegenerative prion diseases. Recently reported four cases of vCJD transmission by blood transfusion raise concerns about the safety of blood products. Proper understanding of PrPc in blood is necessary for development of currently unavailable blood screening tests for prion diseases. Flow cytometry is an attractive method for prion detection, however, the reports on the quantity of PrPc on human blood cells are contradictory. We showed that the majority of PrPc in resting platelets is present in the intracellular pool and is localized in α-granules. We demostrated that both, human platelets and red blood cells (RBC) express significant amount of PrPc and thus may play an important role in the transmission of prions by blood transfusion. Our results suggest a unique modification of PrPc on human RBC. Such modification of pathological prion protein could distort the results of blood screening tests for prions. Further we showed that the storage of blood prior to analysis and the choice of anti-prion antibody greatly affect the detection of PrPc by flow cytometry and we identified platelet satellitism as a factor contributing to the heterogeneity of PrPc detection in blood cells. Moreover, we demonstrated existence of...

Nanotechnology is a rapidly growing field focused on the manipulation and control of materials with dimensions under 100 nm. The novel electronic, optical and mechanical properties observed at the nanoscale have resulted in a number of applications in catalysis, light emitting devices, solar power, self-cleaning surfaces and medicine. Medical applications of nanotechnology (“nanomedicine”) are particularly promising for rapid clinical diagnosis and targeted treatments. Understanding the interactions of nanoparticles with living matter is of fundamental importance for all application areas: manufacture, use and disposal of the growing number of nanoproducts will result in increased environmental exposure in addition to direct exposure through nanomedical applications. However, there is a lack of standard methodologies for assessing these interactions. In this work the stability of silver-based nanoparticles was established by UV- Visible (UV-Vis) spectroscopy, atomic force microscopy (AFM) and transmission electron microscopy (TEM). The presence of a higher valence metal or polymer on the nanoparticle surface was demonstrated to improve stability. A standard methodology was developed to study nanoparticle-cell interactions: an “atlas” of the effects of known drugs on a cell is created, and compared with the effects of a nanoparticle. Escherichia coli was selected as a model organism and the effects of a range of antibiotics were characterised through a combination of microbiological assays and AFM. Susceptibility, population cell growth and individual heights, widths, lengths and volumes of bacteria were obtained on a 2% agarose substrate in air. The methodology was applied and adjusted for silver nanoparticles due to the interactions of silver with the bacterial growth medium. 10 and 30 nm silver nanoparticles and ions were found to kill E. coli through an internal mechanism of action, with a size-specific effect on the height of bacteria. Finally, a novel AFM characterisation method is described to examine the mechanical properties of live bacterial and human cells in liquid.

A transfusão de sangue continua a ser uma importante ferramenta terapêutica, porém como qualquer outra, apresenta riscos. O sangue após coletado é processado e origina os hemocomponentes, dentre eles o Concentrado de Hemácias (CH). O CH é o hemocomponente mais utilizado na prática clínica. É indicado para o tratamento de anemias e em casos de hemorragia aguda ocorridas em cirurgia ou em decorrência de doenças ou traumas. Dentre os principais riscos da transfusão de CH estão a aloimunização e as reações transfusionais, imediatas ou tardias. Deve-se avaliar a real necessidade de se transfundir um paciente, principalmente, no período intra operatório. A transfusão de CH no intra operatório está associada a um pior prognóstico, maior tempo de internação, sobrevida menor e aumento do risco de infecção. Este estudo tem por objetivo analisar as solicitações de reserva de CH, e a partir desta análise, elaborar um protocolo de reservas cirúrgicas de CH com a finalidade de determinar o quantitativo de bolsas a serem reservadas ou não para a realização procedimentos cirúrgicos realizados no Hospital Regional de Ceilândia, no Distrito Federal. Para a elaboração do protocolo, foi realizado o levantamento de dados referentes às solicitações de reservas realizadas neste hospital entre março e agosto de 2017. Baseado nestes dados, calculou-se o Índice de Pacientes Transfundidos (IPT) que definiu o percentual das reservas que foram utilizadas. Verificou-se que a maioria das reservas solicitadas não foi utilizada. Do total de 216 pacientes que tiveram reservas preparadas, para as quais foram compatibilizadas 438 bolsas, apenas 48 destas bolsas foram transfundidas, ou seja, somente 10,95%. A partir do resultado do IPT, foi definida uma conduta hemoterápica cirúrgica para cada tipo de cirurgia. E, a partir destas, foi elaborado, um protocolo de reservas de CH para cirurgias, com o intuito de racionalizar a solicitação de reservas para ser implementado no hospital. Podendo assim ser um instrumento, para a equipe médica e para a agência transfusional, contribuindo para a economia de recursos financeiros e de sangue. / Blood transfusion continues to be an important therapeutic tool but it presents risks. The blood after collection is processed originating the blood components, among them the Red blood cells (RBC). RBC is the most widely used blood component in clinical practice. It is indicated for the treatment of anemias and in cases of acute hemorrhage occurring in surgery or due to pathologies or traumas. Among the main risks of RBC transfusion are alloimmunization and transfusion reactions. The actual need to transfuse a patient, especially during the intraoperative period, should be evaluated. Intraoperative RBC transfusion is associated with a worse prognosis, longer length of stay, shorter survival and increased risk of infection. The aim of this study was to analyze the RBC reserve requests and then to elaborate a protocol of RBC surgical reserves in order to determine the quantity of blood bags to be reserved or not for surgical procedures performed at the Regional Hospital de Ceilândia (HRC), in the Federal District. For the elaboration of this protocol, the data of the reservations requests performed in HRC March 2017 to August 2017 was carried out. The transfusion Index was calculated from the data, which defined the percentage of reservations that were used. It was found that most of the requested reserves were not used. Of the total of 216 patients who had prepared reservations, for which 438 bags were compatibilized, only 48 of these bags were transfused, that is, only 10.95%. From the result of the transfusion Index, so for each elective surgical procedures a guide surgical hemotherapy was defined. In addition, a reserve protocol of RBC for surgeries was elaborated, with the purpose of rationalizing the reservation request to be implemented in the hospital. This can be an instrument for the medical team and for the transfusion agency, contributing to the rational use of financial and blood resources.

O sangue total passa por processo de centrifugação a fim de que o paciente receba apenas o componente sanguíneo do qual necessita. O controle de qualidade, inicia com inspeção visual, onde são avaliados: coloração, lipemia, presença de coágulos e vazamentos. No Hemocentro Coordenador do Paraná, a principal causa de descarte de plasma fresco (PF) é a lipemia. Os PF turvos são descartados e os respectivos concentrados de hemácias (CH) permanecem em estoque. Durante o armazenamento, os CH passam por processo de alterações bioquímicas e morfológicas conhecido como lesão de armazenamento, cuja última etapa é a hemólise. É comprometida a função terapêutica e segurança transfusional. Alguns autores relatam a relação entre lipemia e hemólise. Lipemia é o aspecto turvo do plasma, devido à presença de lipoproteínas e tem relação principalmente com a dieta do doador. O objetivo deste estudo foi comparar os parâmetros de qualidade dos CH que tiveram os plasmas descartados por lipemia com os CH de plasmas límpidos. Os PF lipêmicos foram separados e foi realizado registro por fotografia e dosagem dos triglicerídeos. Conforme o grau de turbidez os plasmas foram classificados como: ligeiramente turvos, moderadamente turvos, intensamente turvos ou leitosos. Conforme a concentração de triglicerídeos foram classificados como normal (<175 mg/dl), limítrofe (175 a 199 mg/dl), elevado (200 a 499 mg/dl) ou muito elevado (>500 mg/dl). Os respectivos CH foram avaliados durante o período de validade. Os experimentos nos CH foram realizados entre 1º e 10º, entre 11º e 22º, entre 23º e 34º e entre 35º e 42º dias de armazenamento. Foram avaliados: esterilidade, índices hematimétricos, ERO, TBARS, metemoglobina. Nos sobrenadantes dos CH foram avaliados: Na+, K+, Cl-, lactato, glicose, pH e grau de hemólise. Os experimentos também foram realizados com controles, CH de plasmas límpidos do mesmo dia de doação do grupo teste. Houve aumento da hemólise mais expressivo nos CH teste, demonstrando que a lipemia impacta negativamente na qualidade do CH durante o armazenamento. A partir dos resultados do estudo foram elaboradas estratégias para processamento, controle de qualidade, modificação e distribuição dos CH de doações com lipemia, garantindo a distribuição de um hemocomponente seguro e eficaz, minimizando descarte por hemólise e efeitos adversos à transfusão. / The blood processing allows the patient receives only the blood component that he needs. The quality control starts with visual inspection and are evaluated: coloration, lipemia, clots and leaks. At the HEMEPAR, the main loss of fresh plasma (FP) is caused by lipemia. The lipemic F are discarded and the related red blood cell (RBC) remain in stock. During storage, RBCs go through the process of biochemical and morphological changes known as storage lesions, and the last one hemolysis. Therapeutic function and transfusion safety are compromised. Some authors report the relationship between lipemia and hemolysis. Lipemia is the turbid aspect of plasma, due to the presence of lipoproteins and is mainly related to the diet of the donor. The objective of this study was to compare the quality parameters of the RBCs from lipemic donations with RBC from clear donations. Lipemic FP were photographed and quantificated to triglycerides. According to the degree of turbidity the plasmas were classified as: slightly cloudy, moderately cloudy, intensely turbid or milky. Triglyceride concentrations were classified as normal (<175 mg / dl), borderline (175 to 199 mg / dl), high (200 to 499 mg / dl) or very high (> 500 mg / dl). The respective RBCs were evaluated during the period of validity. The CH experiments were performed between 1 and 10º, 11º and 22º, 23º and 34º and 35º and 42º days of storage. The following: sterility, hematimetric indexes, ROS, TBARS and methemoglobin were evaluated. In the supernatants Na +, K +, Cl-, lactate, glucose, pH and degree of hemolysis were evaluated. The experiments were also carried out with controls, RBC from clear donations of the same day of donation of the test group. There was an increase in hemolysis in the RBC test. It shows that lipemia negatively impacts the quality of RBC during storage. From the results of the study, were established strategies for processing, quality control, modification and distribution of RBC of donations with lipemic plasma to ensure the use of safe and effective blood product for transfusion.

Introdução. O transporte é um procedimento crítico na manutenção da cadeia do frio do sangue. Falhas neste procedimento podem comprometer a sua qualidade e a segurança transfusional. Objetivo. Avaliar as não conformidades (NC) do Hemocentro de Ribeirão Preto relacionadas ao transporte de concentrado de hemácias (CH). Analisar caixas térmicas e gelos recicláveis submetidos à temperatura de 20°C e 30°C e propor melhorias neste processo. Métodologia. Estudo retrospectivo de NC relacionadas ao transporte de CH no software do sistema da qualidade do Hemocentro de Ribeirão Preto e pesquisa experimental realizada com três caixas diferentes em combinação com três gelos recicláveis distintos, simulando uma situação de transporte, nas temperaturas externas de 20°C e 30°C. Realizou-se uma estatística descritiva com os dados. Utilizou-se a metodologia de análise de variância (ANOVA), com o auxílio do software SAS® 9, utilizando a PROC GLM. Para as comparações foram utilizados contrastes ortogonais baseados na distribuição t. Resultados. Os resultados mostraram diferenças significativas no desempenho entre as caixas e entre os gelos recicláveis. No entanto, as maiores diferenças ocorreram entre as temperaturas externas de 20°C e 30°C, indicando a interferência da temperatura externa na temperatura do conteúdo interno. Outra diferença encontrada foi entre pontos de aferição das temperaturas, dentro da mesma caixa, indicando que a distribuição do ar, no interior da caixa não é homogênea. Conclusão. São vários os interferentes existentes durante o transporte de CH, que podem comprometer a qualidade do sangue. O transporte deve ser validado, padronizando o maior número de variáveis possíveis, para minimizar as adversidades que ocorrem durante o procedimento / Introduction. The transport is a critical procedure in the maintenance of the blood cold chain. Failures in this procedure can compromise the blood quality and transfusion safety. Purpose: Evaluate the non-conformances (NC) of Hemocentro de Ribeirão Preto related to the Red Blood Cell (RBC) transport. Verify coolers, recycle ice at a temperature of 20°C and 30°C and propose improvements in this process. Methodology: Retrospective study of NC related to RBC transport in Hemocentro de Ribeirão Preto quality system software. Experimental research performed with three different insulated boxes combined with three different ice packs, simulating a situation of transport in the external temperatures of 20°C and 30°C. It was made a descriptive statistic of the data. The methodology of analysis of variance was used, through the software SAS® 9, using a PROC GLM. To do the comparisons orthogonal contrasts based on t distribution were used. Results: The results shown significant differences between insulated box and ice packs. However, the larger differences occurred between the temperatures of 20°C and 30°C, indicating that the external temperature interfere in the temperature inside the box. Another difference found was between points of temperature, inside the same insulated box, indicating that the air distribution inside the insulated box is not homogeneous. Conclusion. During RBC transport, many interfering exist that can compromise the blood quality. The transport must be validated, standardizing the largest number of details that is possible, to minimize adversity that occur during the procedure

Células T reguladoras (Treg) são cruciais na tolerância periférica e no controle da inflamação. Nós usamos dois modelos bem estabelecidos de tolerância de mucosas para a asma alérgica e a tolerância inalatória local induzida pela exposição crônica a OVA para estudar o aparecimento e função das Treg. Nós mostramos que a tolerância nasal distinguiu da tolerância oral pela produção sistêmica de IgG1 e desenvolvimento da inflamação alérgica na cavidade peritoneal ou pela indução da inflamação das vias aéreas de camundongos RAG-/- reconstituídos com células T CD4+ após desafios com OVA. Observamos também que Treg Foxp3+ migraram para o pulmão alérgico e expressaram fenótipo de ativação e memória que distinguiu essas células das Treg presentes nos linfonodos drenantes. Células T CD4+CD25+ do pulmão dos animais alérgicos suprimiram a proliferação das células T CD4+CD25-, mas não a produção de citocinas Th2. Finalmente, a exposição crônica a OVA levou ao aumento da apoptose de eosinófilos que infiltraram o pulmão resultando na resolução da inflamação alérgica pulmonar. / Regulatory T cells (Treg) are critical for peripheral tolerance and control of inflammation. We used two well established models of mucosal tolerance to allergic airway disease and the local inhalational tolerance induced by chronic OVA exposure to study the appearance and function of Treg cells. We found that nasal tolerance distinguished from oral tolerance by systemic IgG1 antibody production and development of allergic inflammation in the peritoneal cavity or by induction of airway inflammation in RAG-/- mice reconstituted with CD4+ T cells after OVA challenge. We also found that Foxp3+ T cells migrated to allergic lung and expressed an effector/memory phenotype that distinguished them from Treg cells present in lung draining lymph nodes. Lung infiltrating CD4+CD25+ T cells from allergic mice suppressed CD4+CD25- T cell proliferation but not Th2 cytokines production by these cells. Finally, chronic OVA exposure leaded to increased apoptosis of infiltrating lung eosinophils resulting in the resolution of allergic lung inflammation.

O sangue total passa por processo de centrifugação a fim de que o paciente receba apenas o componente sanguíneo do qual necessita. O controle de qualidade, inicia com inspeção visual, onde são avaliados: coloração, lipemia, presença de coágulos e vazamentos. No Hemocentro Coordenador do Paraná, a principal causa de descarte de plasma fresco (PF) é a lipemia. Os PF turvos são descartados e os respectivos concentrados de hemácias (CH) permanecem em estoque. Durante o armazenamento, os CH passam por processo de alterações bioquímicas e morfológicas conhecido como lesão de armazenamento, cuja última etapa é a hemólise. É comprometida a função terapêutica e segurança transfusional. Alguns autores relatam a relação entre lipemia e hemólise. Lipemia é o aspecto turvo do plasma, devido à presença de lipoproteínas e tem relação principalmente com a dieta do doador. O objetivo deste estudo foi comparar os parâmetros de qualidade dos CH que tiveram os plasmas descartados por lipemia com os CH de plasmas límpidos. Os PF lipêmicos foram separados e foi realizado registro por fotografia e dosagem dos triglicerídeos. Conforme o grau de turbidez os plasmas foram classificados como: ligeiramente turvos, moderadamente turvos, intensamente turvos ou leitosos. Conforme a concentração de triglicerídeos foram classificados como normal (<175 mg/dl), limítrofe (175 a 199 mg/dl), elevado (200 a 499 mg/dl) ou muito elevado (>500 mg/dl). Os respectivos CH foram avaliados durante o período de validade. Os experimentos nos CH foram realizados entre 1º e 10º, entre 11º e 22º, entre 23º e 34º e entre 35º e 42º dias de armazenamento. Foram avaliados: esterilidade, índices hematimétricos, ERO, TBARS, metemoglobina. Nos sobrenadantes dos CH foram avaliados: Na+, K+, Cl-, lactato, glicose, pH e grau de hemólise. Os experimentos também foram realizados com controles, CH de plasmas límpidos do mesmo dia de doação do grupo teste. Houve aumento da hemólise mais expressivo nos CH teste, demonstrando que a lipemia impacta negativamente na qualidade do CH durante o armazenamento. A partir dos resultados do estudo foram elaboradas estratégias para processamento, controle de qualidade, modificação e distribuição dos CH de doações com lipemia, garantindo a distribuição de um hemocomponente seguro e eficaz, minimizando descarte por hemólise e efeitos adversos à transfusão. / The blood processing allows the patient receives only the blood component that he needs. The quality control starts with visual inspection and are evaluated: coloration, lipemia, clots and leaks. At the HEMEPAR, the main loss of fresh plasma (FP) is caused by lipemia. The lipemic F are discarded and the related red blood cell (RBC) remain in stock. During storage, RBCs go through the process of biochemical and morphological changes known as storage lesions, and the last one hemolysis. Therapeutic function and transfusion safety are compromised. Some authors report the relationship between lipemia and hemolysis. Lipemia is the turbid aspect of plasma, due to the presence of lipoproteins and is mainly related to the diet of the donor. The objective of this study was to compare the quality parameters of the RBCs from lipemic donations with RBC from clear donations. Lipemic FP were photographed and quantificated to triglycerides. According to the degree of turbidity the plasmas were classified as: slightly cloudy, moderately cloudy, intensely turbid or milky. Triglyceride concentrations were classified as normal (<175 mg / dl), borderline (175 to 199 mg / dl), high (200 to 499 mg / dl) or very high (> 500 mg / dl). The respective RBCs were evaluated during the period of validity. The CH experiments were performed between 1 and 10º, 11º and 22º, 23º and 34º and 35º and 42º days of storage. The following: sterility, hematimetric indexes, ROS, TBARS and methemoglobin were evaluated. In the supernatants Na +, K +, Cl-, lactate, glucose, pH and degree of hemolysis were evaluated. The experiments were also carried out with controls, RBC from clear donations of the same day of donation of the test group. There was an increase in hemolysis in the RBC test. It shows that lipemia negatively impacts the quality of RBC during storage. From the results of the study, were established strategies for processing, quality control, modification and distribution of RBC of donations with lipemic plasma to ensure the use of safe and effective blood product for transfusion.

Orientador: Maria Cristina Thomaz / Banca: Vera Maria Barbosa de Moraes / Banca: Fábio Enrique Lemos Budiño / Banca: Danísio Prado Munari / Banca: Jacinta Diva Ferrugem Gomes / Resumo: Com o objetivo de estudar o uso de probiótico em dietas de suínos foram realizados cinco ensaios. No ensaio 1 foi avaliado a adição de diferentes concentrações de probiótico (0, 200 e 300 mg/kg de ração) em rações de leitões recém desmamados e em diferentes idades, sobre as características do sistema digestório e desempenho zootécnico, nos ensaios 2 e 3, os objetivos foram determinar os parâmetros sangüíneos em diferentes idades dos leitões e a digestibilidade de rações para suínos, alimentados com rações contendo ou não probiótico, nos ensaios 4 e 5, foram comparadas a composição das fezes de suínos obtidas por dois métodos de colheita e foi determinado o balanço de minerais em rações contendo ou não probiótico para suínos em fase de crescimento. As diferentes concentrações de probiótico nas dietas de suínos não alteraram a maioria das variáveis estudadas, afetadas apenas pela idade dos animais. O ganho de peso e a conversão alimentar dos animais suplementados com 200 mg de probiótico foi melhor que o grupo com 300 mg de probiótico. A adição do probiótico nas rações diminuiu as quantidades de leucócitos, globulinas e beta + gama globulinas e melhorou a utilização da matéria mineral da dieta pelos suínos. Colheitas de fezes do reto ou da caixa coletora da gaiola, em estudos de balanço de minerais podem ser realizadas sem alterar a composição das fezes e a adição de probiótico nas rações de suínos melhorou a utilização da matéria mineral. / Abstract: With the objective of studying the probiotic use in diets of swine five assays were accomplished. In the assay 1 the addition of different probiotic concentrations was evaluated (0, 200 and 300 mg/kg of ration) in rations of pigs recently weaned and in different ages, on the characteristics of the digesting system and performance, in the assays 2 and 3, the aims were to determine the blood parameters in age different piglets and the ration digestibility for swine fed with rations containing or no probiotic, in the assays 4 and 5, were compared the composition of the feces of swine obtained by two collection methods and was determiner the minerals balance in rations containing or no probiotic for growth swine phase. The different probiotic concentrations in the diets of swine did not alter most of the studied variables, just affected by the age of the animals. The gain weight and feed:gain ratio of the animals supplemented with 200 mg of probiotic was better than the group with 300 mg of probiotic. The addition of the probiotic in the rations reduced the amounts of leukocytes, globulins and beta + gama globulins and it improved the use of the mineral matter of the diet for the swine. Collect of feces of the rectum or of the collector cage box, in studies of minerals balance can be accomplished without altering the feces composition and the probiotic addition in the rations of swine improved the use of the mineral matter. / Doutor

Koblasa,Vladimír - Comparison of control measurements on various types of haematology analyzers used by ÚKBLD CHLTC at University Hospital in Prague First Faculty of Medicine, Charles University in Prague, Praha 2, Kateřinská 32 Head of the work: prof. MUDr. Jan Kvasnička, DrSc. Supervisor -consultant: Mgr. Ivana Malíková Blood cell count is essential testing method in hematology, where it is necessary to ensure properquality control. Aim of this study was to compare the results of measurement control materials with defined parameters and the same samples at different haematological analyzers to obtain evidence for the expression of measurement uncertainties. There are used more types of blood analyzers in ÚKBLD CHLTC at University Hospital in Prague, which operate on different principles. For comparsion were selected analyzers using the impedance working principle, where individual blood cell passes between two electrodes controlled by low voltage. Variation of this voltage is recorded and accurately defined for each type of blood cells. It was also chosen analyzer that works with optical detection. Analyzer illuminates the individual blood cell by light beam. A cell that enters into the path of light rays, reduce its optical density incident on the photocell. The change of the light density...

Major challenges in the management of blood supply chain are related to the shortage and wastage of the blood products. Given the perishability characteristics of blood which can be stored up to a limited number of days, if hospitals and blood centers keep an excessive number of blood units on inventory, wastages may occur. On the other hand, if sufficient number of blood units are not stored on inventory, shortages of this resource may cause the cancellations of important activities and increase the fatality rates at hospitals. Three mathematical models have been developed with the goal to improve the efficiency of blood related activities at blood centers and hospitals. The first model uses an integer programming (IP) approach to identify the optimal order levels that minimizes the total cost, shortage and wastage levels of blood products at a hospital within a specified planning horizon. The IP model explicitly considers the age of blood inventory, uncertain demand, the demand for two types of patients and crossmatch-to-transfusion ratio. The second model formulates the different shortage and inventory distribution strategies of a blood center supplying blood products to multiple hospitals. The third model develops a vehicle routing problem for blood centers to minimize the daily distance travelled by bloodmobiles during the blood collection process. Optimal routing for each bloodmobiles is identified using CPLEX solver, branch \& bound and column generation algorithms and their solution times are compared.