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  • About
  • The Global ETD Search service is a free service for researchers to find electronic theses and dissertations. This service is provided by the Networked Digital Library of Theses and Dissertations.
    Our metadata is collected from universities around the world. If you manage a university/consortium/country archive and want to be added, details can be found on the NDLTD website.
51

METHODS DEVELOPMENT AND APPLICATION OF TWO DIMENSIONAL CHROMATOGRAPHY AND TANDEM MASS SPECTROMETRY IN PROTEOMICS

Wenner, Brett Romain 01 January 2004 (has links)
Although molybdenum blue solutions have been known for more than twocenturies, an understanding of their chemical nature is only beginning to emerge.This dissertation aimed at elucidating the structural nature of the polydisperse,nanoscopic components in the solution phases and the solid states of partiallyreduced polyoxomolybdate (Mo-POM). The study offered at least fourcontributions to the area: (1) a rational protocol for the molecular recognition ofMo-POM with de novo organic hosts. (2) demonstration of kinetic precipitation ofa dynamic mixture of polyoxomolybdates and application of the technique to thestudy of the dynamic mixture by TEM (3) characterization of the Mo-POMnanostructures by an unusual combination of complementary analyticaltechniques. (4) a general approach for the synthesis of crown-ethers-containingtripodal molecules.The molecular recognition of Mo-POM with designer tripodal hexaminetris-crown ethers opened a window to the solution phase structures of Mo-POMnanoscopic components. Studies with a series of structurally analogous hostsprobed the relationship between the structure of the molecular host and theformation of nanostructures.An unusual combination of complementary analytical protocols: flow fieldflowfractionation, electron microscopy (transmission and scanning), andinductively coupled plasma – emission spectroscopy, was used to monitor thesolution-phase evolution of Mo-POM nanostructures. The crystallization – drivenformation of keplerate Mo-POM and solution-phase evolution of structurallyrelated nanoscopic species were apparent in the self-assembling process ofpartially reduced Mo-POM.
52

Comparison of Two Methods for Detecting Intrathecal Synthesis of Borrelia Specific Antibodies

Holmqvist, Stephanie January 2010 (has links)
<p>In Europe, Lyme disease<em> </em>is caused by the species <em>Borrelia (B.) burgdorferi</em> sensu stricto,<em> B. garinii </em>and <em>B. afzelii.</em> The disease is the most common vector-borne infection in Europe and the United States,<em> </em>and the resulting manifestation can involve the skin, nervous system, heart and joints. The symptoms that arise are associated with the <em>Borrelia </em>species causing the infection. The species most associated with neuroborreliosis is <em>B. garinii</em> whilst <em>B. burgdorferi </em>sensu stricto is associated with arthritis and <em>B. afzelii </em>is associated with dermatological symptoms. Lyme disease normally has three phases in untreated patients. The first phase is characterised by erythema migrans, a reddening of the skin around the area of the tick bite. If the disease develops to the second phase the patient will suffer from neuroborreliosis which is characterised by neurological symptoms such as headache and peripheral facial paralysis. Cerebrospinal fluid (CSF) analysis is used to diagnose neuroborreliosis. The diagnosis is complicated by variations between the different <em>Borrelia</em> species and that many healthy individuals have antibodies directed against <em>Borrelia</em>. Antibodies in CSF can be found in different diseases. The antibodies can be produced in the central nervous system or come across the blood-brain barrier and thus derive originally from the blood. By measuring the concentration of total albumin in serum and in CSF it can be determined if the antibodies present in the CSF have been produced in the central nervous system or if they originate from the blood. The typical manifestation in the last phase of Lyme disease is severe arthritis. The aim of this examination project was to compare two ELISAs for detection of antibodies directed to <em>Borrelia</em>. Indirect ELISAs from DAKO and Euroimmun were compared for the diagnosis of neuroborreliosis in 100 individuals. <em>Borrelia </em>specific antibodies of class IgM or IgG were found in 16 of 100 patients by DAKO’s ELISA and in 20 of the same 100 patients by Euroimmun’s ELISA. The reason that Euroimmun’s method detected more cases of neuroborreliosis is probably that this method detects antibodies directed to all three pathological species of <em>Borrelia </em>while DAKO’s method only detects antibodies directed to <em>B. burgdorferi</em>. In conclusion, this study indicates that Euroimmun’s method to detect antibodies of class IgM and IgG directed to <em>Borrelia </em>is superior to DAKO’s method. The obtained results were confirmed by Western blot analysis which gave results in accordance with those of Euroimmun’s ELISA.</p>
53

The cerebral surfactant system and its alteration in hydrocephalic conditions

Schob, Stefan, Lobsien, Donald, Friedrich, Benjamin, Bernhard, Matthias K., Gebauer, Corinna, Dieckow, Julia, Gawlitza, Matthias, Pirlich, Mandy, Saur, Dorothee, Bräuer, Lars, Bechmann, Ingo, Hoffmann, Karl-Titus, Mahr, Cynthia V., Nestler, Ulf, Preuß, Matthias 22 November 2016 (has links) (PDF)
Introduction: Pulmonary Surfactant reduces surface tension in the terminal airways thus facilitating breathing and contributes to host's innate immunity. Surfactant Proteins (SP) A, B, C and D were recently identified as inherent proteins of the CNS. Aim of the study was to investigate cerebrospinal fluid (CSF) SP levels in hydrocephalus patients compared to normal subjects. Patients and methods: CSF SP A-D levels were quantified using commercially available ELISA kits in 126 patients (0±84 years, mean 39 years). 60 patients without CNS pathologies served as a control group. Hydrocephalus patients were separated in aqueductal stenosis (AQS, n = 24), acute hydrocephalus without aqueductal stenosis (acute HC w/o AQS, n = 16) and idiopathic normal pressure hydrocephalus (NPH, n = 20). Furthermore, six patients with pseudotumor cerebri were investigated. Results: SP AÐD are present under physiological conditions in human CSF. SP-A is elevated in diseases accompanied by ventricular enlargement (AQS, acute HC w/o AQS) in a significant manner (0.67, 1.21 vs 0.38 ng/ml in control, p<0.001). SP-C is also elevated in hydrocephalic conditions (AQS, acute HC w/o AQS; 0.87, 1.71 vs. 0.48 ng/ml in controls, p<0.001) and in Pseudotumor cerebri (1.26 vs. 0.48 ng/ml in controls, p<0.01). SP-B and SP-D did not show significant alterations. Conclusion: The present study confirms the presence of SPs in human CSF. There are significant changes of SP-A and SP-C levels in diseases affecting brain water circulation and elevation of intracranial pressure. Cause of the alterations, underlying regulatory mechanisms, as well as diagnostic and therapeutic consequences of cerebral SP's requires further thorough investigations.
54

Complex Skull Base Reconstructions in Kadish D Esthesioneuroblastoma: Case Report

Palejwala, Sheri, Sharma, Saurabh, Le, Christopher, Chang, Eugene, Erman, Audrey, Lemole, G. 04 May 2017 (has links)
Introduction Advanced Kadish stage esthesioneuroblastoma requires more extensive resections and aggressive adjuvant therapy to obtain adequate disease-free control, which can lead to higher complication rates. We describe the case of a patient with Kadish D esthesioneuroblastoma who underwent multiple surgeries for infectious, neurologic, and wound complications, highlighting potential preventative and salvage techniques. Case Presentation A 61-year-old man who presented with a large left-sided esthesioneuroblastoma, extending into the orbit, frontal lobe, and parapharyngeal nodes. He underwent margin-free endoscopic-assisted craniofacial resection with adjuvant craniofacial and cervical radiotherapy and concomitant chemotherapy. He then returned with breakdown of his skull base reconstruction and subsequent frontal infections and ultimately received 10 surgical procedures with surgeries for infection-related issues including craniectomy and abscess evacuation. He also had surgeries for skull base reconstruction and CSF leak, repaired with vascularized and free autologous grafts and flaps, synthetic tissues, and CSF diversion. Discussion Extensive, high Kadish stage tumors necessitate radical surgical resection, radiation, and chemotherapy, which can lead to complications. Ultimately, there are several options available to surgeons, and although precautions should be taken whenever possible, risk of wound breakdown, leak, or infection should not preclude radical surgical resection and aggressive adjuvant therapies in the treatment of esthesioneuroblastoma.
55

Vyšetření počtu buněčných elementů v mozkomíšním moku na analyzátoru Sysmex XE-5000 metodou "Body Fluid". / Examination of Cellular Elements in the Cerebrospinal Fluid on the Analyser Sysmex XE-5000 by Method of "Body Fluid".

Davídková, Jana January 2014 (has links)
Cerebrospinal fluid is examined using a variety of methods, which also include determining the number and type of each cell. Now the method of the first choice to determine the cellular elements is a microscopic method. This determination, however, can also be done using the analyzer method, which is not yet so widespread. The aim of my thesis was to compare these two methods and determine whether the examination of cerebrospinal fluid on the analyzer Sysmex XE-5000 in the "Body Fluid" mode can replace commonly used microscopic methods. To this purpose, we gathered the laboratory data measured by using both these methods. These data was compared, evaluated and statistically processed. The resulting data suggest that the values measured on the analyzer Sysmex XE-5000 are more accurate than from microscopic determination, especially at highly cell samples of cerebrospinal fluid. To this end, we came evaluation of Bland-Altman graphs and comparison graphs with marked of limits of physiological oligocytosis. For checking of the accuracy of measurements, we verified the repeatability of the analyzer for the values of leukocytes and erythrocytes, coefficients of variation corresponding to the values specified by the manufacturer's documentation. We also investigated the stability of samples of...
56

Caractérisation des neurones bulbo-spinaux PKD2L1+ qui contactent le liquide céphalo-rachidien / Characterization of medullospinal cerebrospinal fluid contacting neurons PKD2L1+

Orts-Del'immagine, Adeline 20 May 2014 (has links)
Chez les vertébrés, les neurones qui contactent le LCR (NcLCR) sont présents autour des cavités ventriculaires et tout le long du canal central (cc). Par la combinaison d'enregistrements électrophysiologiques sur tranche de tronc cérébral et d'analyses immunohistochimiques, nous avons réalisé la première caractérisation de cette population neuronale chez la souris adulte. Nous montrons que les NcLCR sont présents autour du cc au niveau du complexe vagal dorsal (CVD), une structure bulbaire impliquée dans la régulation des fonctions autonomes, où ils sont principalement GABAergiques, reçoivent des afférences synaptiques GABA/Glycinergiques et expriment le canal PKD2L1 ("polycystin kidney disease 2-like 1"), un membre de la famille des canaux TRP ("transient receptor potential"). Nous montrons que l'activité de PKD2L1 est modulée par les variations de pH et d'osmolarité et que son augmentation module l'excitabilité des NcLCR. Finalement, nous démontrons que les NcLCR existent dans un état de maturité intermédiaire caractérisé par propriétés fonctionnelles de neurones matures combinées à la conservation de l'expression de marqueurs d'immaturités.Les NcLCR étant stratégiquement positionné entre le LCR et le parenchyme, ils pourraient détecter des signaux circulant grâce à l'activation de PKD2L1 puis distribuer le message collecté à leurs partenaires. Un tel rôle, apparaît particulièrement intéressant au niveau du CVD, un site de régulation majeur des fonctions autonomes et pourrait être démontré par l'identification du réseau neuronal où les NcLCR sont intégrés. / In vertebrates, cerebrospinal fluid contacting neurons (CSF-cN) are present around the ventricular cavities and along the central canal (cc). In this study, by the combination of whole cell patch-clamp recordings on brainstem slice and immunohistochemistry analysis, we realize the first characterization of this neuronal population in adult mice. We show that CSF-cN are present around the cc in the dorsal vagal complex (DVC), a major hindbrain structure regulating autonomic functions. There, CSF-cN are mostly GABAergics, receive GABA- and glycinergic synaptic entries and express functional polycystin kidney disease 2-like 1 (PKD2L1) channels. These channels are a subtype of the transient receptor potential (TRP) channels superfamily and this study represent the first analysis of PKD2L1 properties in a native system. We show that PKD2L1 channel activity is modulated by variations in extracellular pH and osmolarity and in turn, an enhanced activity of only few PKD2L1 channels participates in the modulation of CSF-cN excitability. Finally, we demonstrate that CSFcN exhibit another interesting property since they exist in an intermediate stage of maturity by displaying many mature functional properties combined to the conservation of the expression of immature markers.Because CSF-cN are strategically positioned between CSF and parenchyma, they could detect circulating signals through PKD2L1 activation and convey the collected messages to cellular partners. Such a role might be particularly relevant at the level of the DVC a major regulatory site for autonomic functions and should be demonstrated by identifying and characterizing the neuronal network CSFcN are involved in.
57

Estudo de bandas oligoclonais restritas ao líquido cefalorraquidiano em pacientes com esclerose múltipla na cidade de São Paulo / Study of oligoclonal bands restricted to the cerebrospinal fluid in multiple sclerosis patients in the city of São Paulo

Gama, Paulo Diniz da 02 October 2009 (has links)
Introdução: O diagnóstico da esclerose múltipla (EM) embora seja clínico, se completa com os resultados de imagem de ressonância magnética, somados ainda com a análise do líquido cefalorraquidiano (LCR), que se constituem em ferramentas indispensáveis. A presença das bandas oligoclonais (BOC) no LCR faz parte do estudo da EM, assim como auxilio no diagnóstico. Existem grandes variações quanto à frequência de BOC em pacientes com EM nas diferentes populações, desde 90% em países nórdicos europeus, até 30 a 60% no Japão, China, Índia e Líbano. O presente estudo tem o objetivo de estabelecer o valor da análise do LCR para o diagnóstico da EM em nossa população, com ênfase na pesquisa de BOC. O estudo também objetiva correlacionar os resultados destas análises com as características clínicas e demográficas da amostra selecionada. Casuística e Métodos: Foram estudados 145 pacientes selecionados do Centro de Referência de Doenças Desmielinizantes do Hospital das Clínicas da Faculdade de Medicina de Universidade de São Paulo, no período de agosto de 2005 a janeiro de 2008. Foram registrados para o estudo os dados demográficos, clínicos e da evolução da doença. O diagnóstico da EM foi estabelecido segundo o painel internacional de McDonald, revisado em 2005. A técnica utilizada para a pesquisa de BOC foi a focalização isoelétrica, seguida do immunoblotting, simultaneamente no LCR e no soro sanguíneo. Os resultados desta análise não foram utilizados para estabelecer o diagnóstico, ou para incluir pacientes na casuística da pesquisa. O grupo controle foi selecionado entre aqueles que se submeteram à anestesia raquidiana para cirurgias de pequeno porte. Resultados: Apresentaram BOC: 54,4% dos 90 pacientes com EM; 31,2% dos 16 pacientes com síndrome clínica isolada; 17,9% dos 39 pacientes com doenças neurológicas inflamatórias. No grupo controle, os 19 pacientes não apresentaram BOC. A sensibilidade das BOC foi calculada em 54,4%. A especificidade foi de 100% quando comparado ao grupo controle. Quando comparado com o grupo de pacientes com doenças neurológicas inflamatórias, a especificidade foi de 82,1%. Nos pacientes que se autodeclaram de cor parda ou preta, a frequência de BOC foi maior, com significância na borda do nível de significância de 5% (p=0,0518). Quanto aos aspectos clínicos evolutivos foi constatada maior presença de BOC nas formas progressivas recorrentes (100%); seguida da forma primariamente progressiva (87,5%); secundariamente progressiva (54,5%) e forma remitente recorrente (44,3%). A homogeneidade das BOC segundo as formas clínicas foi significante (p=0,0103), ao nível de significância de 5%. Conclusões: Este trabalho demonstra que a frequência de BOC em pacientes com EM foi de 54,4%, sendo menor que em outras séries mundiais. Estes resultados também não confirmam aqueles obtidos em estudos brasileiros prévios. A etnia e as formas de evolução clínica da doença influenciaram com significância na frequência de aparecimento das BOC. A baixa frequência das BOC no LCR de pacientes com EM pode ser decorrente de fatores associados à baixa e média prevalência da doença nesta região. / Introduction: The diagnosis of multiple sclerosis (MS) although clinical, is rounded out with the results of magnetic resonance imaging, in addition to an analysis of the cerebrospinal fluid (CSF), which form a set of indispensible tools. The presence of oligoclonal bands (OCB) in the CSF is used in the study and for the diagnosis of MS. The frequency of OCB in MS patients varies widely in different populations, ranging from 90% in Nordic countries, to 30 to 60% in Japan, China, India and Lebanon. The objective of the present study is to establish a reference value for the analysis of CSF for the diagnosis of MS in our population, with an emphasis on the study of OCB. A further objective of the study is to correlate the results of these analyses with the clinical and demographic characteristics of the sample selected. Methods: The sample was composed of 145 patients selected from the Demyelinating Diseases Reference Center of the Clinical Hospital of the University of São Paulo, from August 2005 to January 2008. Records containing demographic, clinical and disease progression were used for the study. The diagnosis of MS was established according to 2005 Revisions of the McDonald Criteria. To detect OCB, isoelectric focusing was employed, followed by immunoblotting, simultaneously in both the CSF and blood serum. The results of this analysis were not used to establish the diagnosis or to include patients in the study sample. The control group was selected from those who were submitted to spinal tap for anesthesia in minor surgery. Results: OCB were found in 54.4% of the 90 patients with MS, 31.2% of the 16 patients with isolated clinical syndrome, and 17.9% of the 39 patients with inflammatory neurological diseases. In the control group, 19 patients did not present OCB. The sensitivity of OCB was calculated to be 54.4%. The specificity was 100% when compared to the control group. When compared with the group of patients with inflammatory neurological diseases, the specificity was 82.1%. For the patients who classified themselves as colored or black, the frequency of OCB was higher, with borderline significance using a significance level of 5% (p=0.0518). With regard to clinical evolution of MS, the highest presence of OCB was found in the relapsing progressive types (100%); followed by the primary progressive type (87.5%); secondary progressive type (54.5%); and relapsing remitting type (44.3%). The homogeneity of the OCB in relation to the clinical types was significant (p=0.0103), at a significance level of 5%. Conclusions: This study shows that the frequency of OCB in patients with MS was 54.4%, lower than the findings of other worldwide series. Moreover, these results do not confirm those obtained in other Brazilian studies. Race and clinical progression of the disease influenced, in a statistically significant manner, the frequency of the presence of OCB. The low frequency of OCB in the CSF of patients with MS may be the result of factors associated with the low and medium prevalence of this disease in this region.
58

Relação entre os padrões de secreção central e periférica de ocitocina: implicações sobre produção de leite em ovelhas / Relation between central and peripheral oxytocin realese: implications on milk production in sheep

Bochini, João Carlos 07 August 2008 (has links)
Esse projeto de pesquisa teve como objetivo estudar simultaneamente as concentrações da ocitocina no líquido cefalorraquidiano (LCR) e no plasma de ovelhas multíparas durante a ordenha, estabelecendo suas possíveis correlações entre os dois compartimentos corporais, bem como em relação à produção e ejeção do leite. Para tanto, foram utilizadas 10 ovelhas multíparas (Ovis aries) da raça Santa Inês apresentado peso médio de 40 Kg. Os animais foram divididos em quatro grupos, sendo eles: ordenha mecânica exclusiva (OME; cordeiros apartados 3 dias após o parto), manejo misto com ordenha mecânica (MMom; cordeiros permanecem com as mães durante o período diurno, após a ordenha mecânica, sendo apartados durante a noite), manejo misto com ordenha manual (MMoma) e amamentação exclusiva (AE; cordeiros apartados durante o período noturno). Para a obtenção de LCR foram realizadas a punção do espaço subaracnóideo e implantação de um cateter epidural, enquanto que o plasma foi colhido com o auxílio de um scalp. As amostras de LCR e sangue foram coletadas simultaneamente antes (-0,5 min), durante (0,5; 1 e 4 min) e após (10; 15 min) a amamentação ou ordenha para posterior quantificação das concentrações de ocitocina por ensaio imunoenzimático. Foi obtido um total de 503 amostras, sendo 241 de LCR e 262 de plasma. As estimativas de média, desvio padrão, coeficiente de variação, mínimo e máximo para as concentrações de ocitocina no LCR e plasmáticas foram 257,880 ± 265,90 pg/ml; 103,11%; 11,70 pg/ml e 1000,00 pg/ml, respectivamente. A análise estatística não revelou correlação significante entre LCR e plasma para os quatros grupos experimentais avaliados. O coeficiente de correlação para os grupos OME, AE, MMom e MMoma foram, respectivamente: -0,26, -0,19, 0,05 e 0,04. Com relação ao LCR, não ocorreu diferença significante entre os 4 grupos experimentais em relação às concentrações de ocitocina. Já para o plasma, os animais do grupo MMom (679,80 ± 25,63) e MMoma (591,82 ± 30,56) apresentaram maiores concentrações plasmáticas médias de ocitocina em relação a OME e AE. Assim também, o grupo OME (381,04 ± 22,09) apresentou maior concentração média de ocitocina em relação ao grupo AE (218,82 ± 27,04). Conclui-se que, não existe correlação positiva entre as concentrações de ocitocina central e na circulação periférica durante a ordenha ou amamentação. Os padrões de liberação de ocitocina plasmática diferem de acordo com o tipo de manejo ao qual o animal é submetido, o que pode ter conseqüências para a ejeção do leite e conseqüentemente, para a produção. Finalmente, as concentrações de ocitocina presentes no LCR não sofrem influência do tipo de manejo de ordenha ao qual o animal foi submetido, ao contrário daquilo que foi observado para o plasma. / The aim of the present work was to study a possible relationship between central and peripheral oxytocin release and its consequences to milk production during milking in experimental ewes. Ten multiparous Santa Ines ewes (Ovis aries) were divided in 4 groups according to milk ejection stimuli: exclusive machine milking (OME), mixedmanagement of milking and suckling (MMom: lambs separated during night and reunited to their mother after morning milking; MMoma: mixed-management with manual milking) and exclusive suckling (AE: lambs separated also during night). Cerebrospinal fluid (CSF) was collected through a implanted sub-arachnoid catheter and plasma was collected from the jugular vein. imultaneous sampling was performed at -0.5, 0.5, 1, 4, 10 and 15 min (0 min was teat attachment to either machine or manual milking system or lamb suckling). A total of 241 samples of CSF and 262 plasma samples were processed and oxytocin concentrations were quantified by immunoenzimatic assays. Estimated means, standard deviation, variation coefficient and minimum and maximun values of CSF and plasma oxytocin concentrations were, respectively: 257.880 ± 265.90 pg/ml; 103.11%; 11.70 pg/ml e 1000.00 pg/ml. No statistical strong positive correlations (OME= -0.26, AE= -0.19, MMom= 0.05 and MMoma= 0.04) were found between CSF and plasma samples. Also, CSF was not influenced by milk ejection stimuli, although plasmatic oxytocin was higher in MMom (679.80 ± 25.63) and MMoma (591.82 ± 30.56) compared to OME and AE. In addition, OME (381.04 ± 22.09) plasmatic oxytocin concentration was higher when compared to AE (218.82 ± 27.04). In conclusion, no positive correlations between central and peripheral oxytocin concentrations were observed during milking or suckling. Plasma oxytocin oncentrations differ as a function of management and have consequences to milk ejection as well as to milk production Also, plasma, but not CSF oxytocin, was influenced by different milk ejection stimuli.
59

Avaliação de um ensaio utilizando-se MULTIPLEX-PCR para a detecção de meningites por diferentes agentes bacterianos / A Test Evaluation using a MULTIPLEX-PCR for the meningitis detection by different bacterial agents

Albuquerque, Renata Chaves 06 February 2009 (has links)
No presente trabalho, foi realizada uma MULTIPLEX-PCR para a detecção do DNA bacteriano de S. agalactiae, S.pneumoniae, N. meningitidis, H. influenzae e outros possíveis agentes etiológicos bacterianos das meningites. Este ensaio combina cinco diferentes iniciadores que detectam simultaneamente o gene crtA de N. meningitidis, o gene p6 de H. influenzae, o gene fbsA de S. agalactiae, o gene lytA de S. pneumoniae e o gene universal 16S rDNA para identificar a presença de agente bacteriano. Foram analisadas 447 amostras de LCR, o ensaio detectou 27 amostras positivas para cultura bacteriana e 13 amostras com resultado de cultura negativa. Estas amostras com cultura negativa apresentavam alterações bioquímicas, hematológicas, imunológicas ou microbiológicas (bacterioscopia) sugestivas de meningite, estes dados auxiliaram na análise dos resultados do MULTIPLEX-PCR. Este ensaio não apresentou reações inespecíficas com fungos, vírus e com outros agentes bacterianos testados (amplificando somente o gene 16S rDNA). A MULTIPLEX-PCR é um ensaio rápido, confiável, de fácil execução e facilmente implementável para a confirmação de meningite bacteriana. E este método pode auxiliar no diagnóstico de meningite com cultura de LCR negativa, particularmente para pacientes que previamente iniciaram antibioticoterapia e no diagnostico diferencial de meningite bacteriana ou viral. / In this work, a MULTIPLEX-PCR has conducted for the bacterial DNA detection of S. agalactiae, S.pneumoniae, N. meningitidis, H. influenza and other possible etiologic bacterial meningitis agents. This test combines five different primers that detect simultaneously the crtA gene of N. meningitides, the p6 gene of H. influenza, the fbsA gene of S. agalactiae, the lytA gene of S. pneumoniae and the 16S rDNA universal gene to identify the presence of bacterial agent. From the 447 samples of CSF that were analyzed, the test detected 27 positive samples for bacterial culture and 13 samples with the result of negative culture. These negative culture samples presented biochemical changes, hematological, immunological or microbiological (bacterioscopy) suggestive of meningitis, these data helped in the analysis of the MULTIPLEX-PCR results. This test showed no nonspecific reactions with fungi, viruses and other bacterial agents tested (only amplifying the gene 16S rDNA). The MULTIPLEX-PCR test is a fast, reliable, easy to implement and easily implementable for of bacterial meningitis confirmation. And this method can aid in the meningitis diagnosis with negative culture of CSF, particularly for patients who previously started antibiotic therapy and in the differential diagnosis of bacterial or viral meningitis.
60

Estudo das concentrações de proteína C-reativa sérica e liquórica em cães com epilepsia idiopática / Study of C-reactive protein concentrations in serum and cerebrospinal fluid in dogs with idiopathic epilepsy

Calvo, Daniel Bernardes 31 July 2012 (has links)
A epilepsia compreende um grupo de alterações neurológicas frequentes em humanos e animais, caracterizada pela ocorrência periódica de crises convulsivas. A causa do processo pode ter várias origens sendo necessária a realização de exames complementares para o diagnóstico definitivo. A análise de biomarcadores, em especial as proteínas de fase aguda, como a proteína C-reativa (PCR), auxilia na identificação de doenças neurológicas inflamatórias e infecciosas, já que após serem produzidas pelo fígado conseguem atingir o tecido danificado e ter suas concentrações elevadas rapidamente na circulação. A concentração de PCR está diretamente relacionada à resposta de fase aguda, independentemente da origem ou natureza do estímulo, podendo ser um processo inflamatório, infeccioso ou até mesmo de origem neoplásica. Embora a PCR tenha sido estudada e monitorada em pacientes com as mais variadas doenças, até o momento não foi determinada a presença de PCR no soro e líquor de cães com epilepsia idiopática. O objetivo deste estudo foi avaliar as concentrações de PCR no líquor e soro de cães apresentando epilepsia idiopática e verificar se essa protéina pode ser utilizada como biomarcador para auxiliar no diagnóstico da doença. Para tanto foram compostos três grupos. O primeiro, denominado A, com 23 animais clinicamente normais; o segundo denomindo B, composto por 17 cães manifestando convulsão em até 24 horas anteriores ao momento da coleta de mateiral; e o terceiro denominado C, com 16 cães apresentando convulsões de 24 horas até 120 horas antecedendo o momento da coleta do líquor e do soro. Foram mensuradas as proteínas totais séricas e realizadas as eletroforeses, além da análise do líquor e tomografia computadorizada. Animais com alterações estruturais detectadas na tomografia foram excluidos do estudo. As concentrações de PCR séricas foram avaliadas por meio da técnica ELISA utilizando-se kit comercial Tridelta Development Ltd, espécie específico. Além desta avaliação, os grupos B e C foram alivados quanto à concentração de PCR no liquor. Os resultados foram analisados pelo teste de Kruskal Wallis, seguido pelo teste de Dunn, enquanto para eletroforese e análise de PCR no líquor utilizou-se teste T não pareado. Não houve diferença significante em relação à eletroforese de proteínas séricas nos três grupos, assim como não se observaram alterações na análise do líquor nos grupos B e C. As concentrações séricas de PCR em cães normais variaram níveis não detectáveis a 6,36 &micro;g/mL, com média de 0,98 &micro;g/mL. As concentrações séricas nos animais do grupo B variaram de 1,04 &micro;g/mL a 5,03 &micro;g/mL, com média 2,14 &micro;g/mL, enquanto no grupo C as concentrações foram de níveis não detectáveis a 1,9 &micro;g/mL com média 0,51 &micro;g/mL. A análise estatística demonstrou diferença significante entre os grupos sendo a média do grupo B superior aos demais (p= 0,0002). As concentrações liquóricas de PCR foram muito baixas quando comparadas àquelas observadas em cães com afecções inflamatórias e infecciosas e não foram em sua maioria detectáveis no líquor quando o período entre a convulsão e a coleta foi superior ao período de 24 horas. Concluiu-se que as convulsões associadas à epilepsia idiopática promovem uma resposta de fase aguda caracterizada pelo aumento de PCR sérica e liquórica nas primeiras 24 horas e que essas concentrações decaem após esse período, podendo estar associadas à liberação de mediadores inflamatórios no SNC e às contrações musculares. Assim sendo, a PCR sérica pode ser utilizada como um biomarcador para diferenciar a epilepsia idiopática de outras causas de convulsão. A técnica ELISA para análise de PCR no líquor, pode se somar às outras análises liquóricas, necessitando ainda de validação. / Epilepsy is a group of neurological disorders of humans and animals characterized by recurrent seizures. Epilepsy can have a number of causes and some complementary tests can help with a precise diagnosis. Biomarkers analysis, in special acute protein phase such as C reactive protein (PCR) can help identify inflammatory and infection neurological disease. Acute phase proteins are produced by liver and reach damaged tissue increasing blood concentration. Today studies show that increases in the PCR blood concentration is related to acute inflammatory response independent of mint, whether it is inflammatory, neoplastic or infection. Although PCR has been studied in many diseases, in special neurological disorder followed or not by seizures, until now PCR has not been founded in blood or liquor of dogs with idiopathic epilepsy. The purpose of this study is to evaluate PCR concentration in blood and liquor of patients with idiopathic epilepsy and verify if the protein can be considered a biomarker to help its diagnose. The study has 3 groups. The first named control group A, with 23 healthy animals, the second named B with 17 dogs that have had seizures within 24h, and the third named C with 16 dog that have had seizures after 24 to 120 hours from blood or liquor collection. The investigation is based on analyzing total protein and electrophoretic protein profile, liquor analysis and tomography. Patients with structural brain damages detected by tomography were excluded from the study. In the control group PCR concentration were analyzed by ELISA method and kit Tridelta Development Ltd, species specific. In groups B and C were also procedure PCR analyses in liquor sample. The results were analyzed by the Kruskal Wallis test and the Dun test, while electrophorese and PCR of liquor where analyzed by the T test not parried. There was no significant difference in electrophorese in the three groups and there were not found alterations in the liquor analyzes of the groups B and C. PCR blood concentration in healthy dogs vary between not detectable values to 6,36mcg/ml, with an average of 0,98mcg/ml. Blood concentrations from animal of group B vary from 1,04 mcg/ml to 5,03, with and average of 2,14mcg/ml. Meanwhile in group C blood concentration values were from not detectable to 1,9 mcg/dl, with an average 0,50 mcg/ml. Statistic analyses show significant difference between groups. Group B average was higher (p=0,0002). PCR liquor concentration was lower to those found on dogs with inflammatory infection diseases and the majority were not detectable in the liquor when the sample has been collected after 24 hours from the seizures. It is able to conclude that seizures associated with idiopathic epilepsy promote an acute phase response characterized by an increase of blood and liquor PCR concentrations within 24 hours, and after this period PCR concentrations declined due to the liberation of inflammatory mediators by the CNS and muscle contractions. Therefore blood can be used as a biomarker to differentiate idiopathic epilepsy from other seizures causes. The ELISA technique for PCR liquor analysis still needs to be validated.

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