Дисертації з теми "Tc cells"

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Dissertacao (Mestrado)
IPEN/D
Instituto de Pesquisas Energeticas e Nucleares - IPEN-CNEN/SP

Thesis (MTech (Radiography (Nuclear Medicine)))--Cape Peninsula University of Technology, 2010
Radioactively labelled red blood cells (RBC) are used in various nuclear medicine studies. In order to obtain accurate results when performing these studies, it is of paramount importance that a good binding of the radioactivity (Tc-99m) with the red blood cells is ensured. The literature indicates that certain drugs can influence red cell membrane properties and biochemistry. These drugs can potentially influence the binding of radionuclides to cells. Antibiotics may possibly alter the labelling efficiency of Tc-99m RBC. Due to the high incidence of tuberculosis (TB) in South Africa, many patients receive anti-TB medication, and therefore the influence of these drugs on the labelling efficiencies of Tc-99m RBC was studied.

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Tese (Doutoramento)
IPEN/T
Instituto de Pesquisas Energeticas e Nucleares - IPEN-CNEN/SP

Thesis (MMed)--Stellenbosch University, 2014.
ENGLISH ABSTRACT: Background Occult blood loss from the gastrointestinal tract (GIT), causing iron deficiency often with anaemia, can be diagnostically and therapeutically challenging. This is because the endoscopic and radiologic tests may be negative due to the slow, chronic and intermittent nature of the gastrointestinal bleeding, making timing key in detection and localisation of the bleed. These limitations can be approached using two different radioactive isotopes. Firstly, we tested the sensitivity of extending Cr-51 RBC for 21 days relative to 5 days to detect GIT bleeding and its use to optimise timing of a Tc-99m RBC study for GIT blood loss localisation. Finally, we tested if the information provided by the Tc-99m RBC study aided gastroenterologic intervention for anatomical localisation of a lesion. Method In this retrospective review, after obtaining institutional and ethics committee approval, records of patients referred for evaluation of possible GIT blood loss were reviewed. In each; daily appearance of radiochromium in stool was measured in the whole body counter. In those cases exceeding 50 ml/day, a technetium-99m (Tc-99m) localization study was performed. These studies were correlated with clinical findings. Results A total of 59 Cr-51 RBC studies were carried out in 36 females and 21 males (n = 57). In 32 (54%) the radiochromium results were positive with 75% of the bleeding incidences occurring after 5 days of stool collection. Of 17 cases in whom Tc-99m RBC imaging studies were performed, 14 (82%) were positive with specific anatomical sites successfully defined in twelve. In all patients with blood loss of >100 ml/24h, Tc-99m RBC were positive and localised. Ten of the 17 Tc-99m RBC studies were further investigated and half diagnosed with small-bowel angiodysplasia. Conclusion This sequential twin isotope method is practical in revealing otherwise silent intestinal haemorrhage. Although it has good patient acceptability and clinical as well as diagnostic utility in management, further studies are required to clearly establish a cut-off level of blood loss for performing imaging studies and the impact of the findings on the overall patient management.
AFRIKAANSE OPSOMMING: Agtergrond Die evaluasie van okkulte bloedverlies uit die gastro-intestinale kanaal (GIT), met gevolglike ystertekort anemie, kan diagnosties en terapeuties uitdagend wees. Dit is omdat endoskopiese en radiologiese ondersoeke negatief mag wees as gevolg van die stadige, chroniese en intermitterende aard van die gastro-intestinale bloeding, wat die presiese tydstip van opsporing en lokalisering van die bloeding krities belangrik maak. Hierdie beperkings kan aangespreek word deur twee verskillende radioaktiewe isotope te gebruik. Eerstens is die sensitiwiteit van die verlenging van die Cr-51 RBS studie tot 21 dae in plaas van 5 dae om die GIT bloeding op te spoor, getoets, asook die gebruik daarvan om die optimale tyd vir ‘n Tc-99m RBS studie om die GIT bloedverlies te lokaliseer, vas te stel. Laastens is getoets of die inligting van die Tc-99m RBS studie wel bygedra het tot die gastroenterologiese ingreep om die letsel anatomies te lokaliseer. Metode Na institusionele en etiese komitee toestemming is inligting van pasiënte wat vir die evaluering van ‘n moontlike GI bloedverlies verwys is, in hierdie retrospektiewe oorsig nagegaan. Die daaglikse voorkoms van radioaktiewe chroom in stoelgangmonsters is in ‘n heelliggaamteller gemeet. In gevalle waar dit 50 ml/dag oorskry het, is ‘n tegnesium 99m (Tc 99m) studie gedoen. Hierdie studies is met die kliniese bevindinge gekorreleer. Resultate ‘n Totaal van 59 Cr-51 RBS studies is in 36 vroue en 21 mans (n = 57) gedoen. Die gemerkte chroomstudies was positief in 32 (54%), met 75% van die bloedings wat meer as 5 dae na versameling van die stoelgang plaasgevind het. In veertien (82%) van die 17 gevalle waar Tc-99m RBS studies gedoen is, was die studies positief. Spesifieke anatomiese gebiede van bloeding kon in 12 hiervan suksesvol bevestig word. Tc-99m RBS studies was positief in al die pasiënte met ‘n bloedverlies van >100 ml/24h, en kon gelokaliseer word. Tien van die 17 Tc-99m RBS studies is verder ondersoek en die helfte daarvan gediagnoseer met dunderm angiodisplasie. Gevolgtrekking Die opeenvolgende twee isotoopmetode om andersins asimptomatiese dermbloeding op te spoor, is prakties uitvoerbaar. Alhoewel die studies goed deur pasiënte aanvaar is, en ook van kliniese en diagnostiese waarde in die hantering van die pasiënte is, is verdere studies nodig om die afsnypunt vir die hoeveelheid bloedverlies om beeldingstudies uit te voer, sonder twyfel vas te stel, asook om die impak van die bevindings op ‘n groter pasiëntpopulasie vas te stel.

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior
As células dos seres vivos são constantemente ameaçadas por agentes químicos ou físicos que possam causar danos ao DNA. Um dos agentes deste estudo foi o cloreto estanoso (SnCl2), utilizado na medicina nuclear como redutor de um isótopo radioativo do tecnécio, o 99mTc. O SnCl2 é um agente cujo mecanismo de produção de lesões em estruturas celulares envolve a geração de espécies reativas de oxigênio (ERO), tais como o H2O2 e o radical OH. Essas ERO podem causar uma série de doenças, o envelhecimento e até mesmo a morte celular, por apoptose ou necrose. Sendo assim, torna-se importante a pesquisa sobre os efeitos biológicos, tanto desse sal, quanto das outras substâncias que compõem os radiofármacos utilizados em medicina nuclear. Desta forma, nosso objetivo geral foi avaliar a toxicidade do cloreto estanoso, associado, ou não, ao kit 99mTc-MDP, bem como dos demais componentes do kit, em diferentes sistemas biológicos. Eletroforese em gel alcalino de agarose em células de E. coli AB 1157 para a avaliação da genotoxicidade; Ensaio do Cometa em células de sangue total de ratos Wistar para estudar a genotoxicidade; Ensaio do Micronúcleo em células da medula óssea de ratos Wistar para verificar o potencial aneugênico e clastogênico; Ensaio Cometa em células de sangue total e em células mononucleares de sangue periférico humano para estudar a genotoxicidade; Ensaio de Viabilidade com Azul Trypan e citometria de fluxo para analisar a citotoxicidade em PBMC; Ensaio do Micronúcleo em linfócitos humanos para verificar o potencial aneugênico e clastogênico. Em cepas de E. coli AB1157, o SnCl2 e o MDP induziram quebras no DNA genômico, quando isolados; porém quando usados de forma associada, ocorreu uma atenuação do número de quebras. Em ratos Wistar, o 99mTc-MDP não foi genotóxico e também não induziu clastogênese ou aneugênese. Em Sangue total, in vitro, o SnCl2 apresentou efeito dose-resposta. Em PBMC, in vitro, o 99mTc-MDP causou redução da viabilidade celular e apresentou genotoxicidade, porém não induziu clastogênese e nem aneugênese.
Alive cells are constantly threated by chemical and physical agents that can generate DNA damage. Here, the studied agent was stannous chloride (SnCl2), a 99mTc reducing agent employed in nuclear medicine. This salt can produce lesions through generation of reactive oxygen species (ROS) as H2O2 and OH. These ROS can be the origin of several diseases and cell death by apoptosis or necrosis. In this way it is important the research about the biological effects of this salt and the other substances composing the radiopharmaceuticals used in nuclear medicine.The aim of this work was to evaluate, in different biological systems, the stannous chloride toxic potentiality, associated or not to 99mTc-MDP radiopharmaceutical as well as the other kit components. Alkaline electrophoresis agarosis gel of E. coli AB 1157 to evaluate the genotoxicity in prokariotic cells; Comet assay in Wistar rats total blood to evaluate the genotoxicity in eukaryotic cells; Micronucleous assay in Wistar rats bone marrow cells to verify the aneugenic and clastogenic effects; Comet assay in human in peripherical total blood and mononuclear cells to evaluate the genotoxicity; Trypan blue viability assay and flow cytometry to evaluate citotoxicity in PBMC; Micronucleous assay in human lymphocyte cells to verify the aneugenic and clastogenic effects; SnCl2 and MDP when isolated induced breaks in E. coli genomic DNA. But, when used in an associated way it was observed an atenuation of the breaks number. 99mTc-MDP was not genotoxic, clastogenic nor aneugenic in Wistar rats In ex vivo human total blood, SnCl2 presented a dose-response effect In ex vivo PBMC; 99mTc-MDP induced a cellular viability reduction and presented genotoxic but not clastogenic or aneugenic effects.

The 45kDa isoform of TC-PTP is a ubiquitous, predominantly nuclear protein. Reports demonstrate that it can exist outside the nucleus; however, immunofluorescence and cellular fractionation techniques point to an almost exclusive residence in the nucleus, and even within the nucleolus. Though the conditions of nucleolar enrichment appear to be serum-dependent, and may involve S52 phosphorylation, the exact circumstances necessary for this localization remain unknown, though TC-PTP C216S mutant, and not an S52A/C216S double mutant had the ability to trap a phosphorylated substrate of ∼105kDa, thus encouraging further research into the identity of this phosphoprotein. There is also data implicating TC-PTP in the p53-dependent DNA damage response. TC-PTP WT pMEF's are more susceptible to apoptosis than KO pMEF's, thus suggesting a possible role as a tumor suppressor. Finally, in order to explore the physiological significance of possible non-hematopoietic roles of TC-PTP, a Cre-Lox targeting vector was constructed for the eventual generation of a conditional knockout.

O objetivo desta pesquisa foi estabelecer uma metodologia para avaliar carcinomas espinocelulares (CEC) de cabeça e pescoço, identificando e distinguindo áreas de maior atividade metabólica dentro da neoplasia, associando dados simultaneamente adquiridos, obtidos por diferentes modalidades de aquisição de imagens, combinando informações metabólicas e anatômicas num único exame. A população estudada consistiu de 17 pacientes com carcinoma espinocelular (CEC) de cabeça e pescoço pertencentes aos arquivos do Departamento de Imagem do Hospital do Câncer, São Paulo. As imagens de TC (tomografia computadorizada) e do 18 F-FDG-PET (tomografia por emissão de pósitrons) foram simultaneamente adquiridas utilizando um aparelho não dedicado. Os dados originais foram transferidos para uma estação de trabalho independente com programa de computação gráfica para o processamento dos grupos individuais e fusão em um único grupo, contendo os dados fisiológicos e metabólicos. Os achados foram definidos como positivos na presença de focos com aumento da concentração do radiofármaco em áreas não relacionadas à distribuição normal do mesmo. Em 77% dos casos (n=13), a hipercaptação foi detectada ao centro da lesão e em 23% (n=4) dos casos houve comportamento diferente, com hipercaptação excêntrica. A fusão de imagens simultaneamente adquiridas num único exame ( 18 F- FDG PET e TC) possibilitou o mapeamento topográfico metabólico das lesões estudadas e foi possível localizar áreas de maior atividade metabólica dentro do próprio 13 tumor, verificando recidivas ou metástases, possibilitando aumentar as opções quanto ao planejamento radioterápico ou cirúrgico a serem seguidos.
The aim of this study is to propose a methodological approach to evaluate head and neck squamous cell carcinoma (SCCA) in order to identify and to distinguish areas of higher metabolic activity inside the lesion combining the functional metabolic and morphological data simultaneously acquired in a non dedicated PET-CT device. The study population consisted of 17 patients, with SCCA of the head and neck carcinoma. These patients were submitted to a non-dedicated 18 F- FDG- PET imaging using a system with low dose CT and Positron emission coincidence acquisition capabilities. The image acquisition was then transferred to an ENTEGRA 2 NT workstation to generate groups of individual images (metabolic and anatomical data) and image fusion (CT + PET). In those patients with anomalous concentrations of 18 F-FDG, the lesion was depicted on three planes (axial, coronal and sagittal) in CT, PET, and the image fusion at the computer screen. The findings were defined as positive in the presence of well-defined focal area of increased uptake in regions unrelated to the normal biodistribution of the tracer on visual inspection. Two examiners interpreted the images in different sessions, in order to get an agreement. Subsequently, the sites of higher metabolic activity inside the tumor were identified and classified in centric or eccentric, according to their relative location. Observing the images, we found 77.00% of the patients with the site of higher activity at the center of lesion. In 23.00% of the patients a different 15 behavior, with the tracer increased eccentrically to the lesion. This technique gave a realistic view of the functional metabolism, locating the anatomical tumor area and helping in future treatment planning.

L’arteritis de cèl·lules gegants és la vasculitis més freqüent en persones majors de 50 anys. Es classifica segons els criteris de Hunder (1990), essent la cefalea el símptoma més freqüent. L’afectació vascular s’ha estudiat de forma extensa amb diferents proves d’imatge i amb la biòpsia de l’artèria temporal, havent-se suggerit que existeixen dos subtipus de la malaltia. Es coneix que una proporció elevada de pacients presenta aortitis i que aquesta s’associa a una major dosi acumulada de corticoides i a un major risc d’aneurisma d’aorta toràcica. Per altra banda, l’afectació ocular constitueix la complicació més temuda en l’actualitat, ja que un cop instaurada no és reversible. L’objectiu d’aquest treball va ser identificar quins són els patrons de captació per PET/TC en el moment del diagnòstic de l’ACG d’acord amb la presència o absència de clínica isquèmica, incloent l’afectació ocular. A més, s’estudià quins eren els patrons clínics i histològics en aquesta cohort de pacients d’acord a la presència o absència d’aortitis en el PET/TC. En el primer estudi, realitzat amb una cohort de pacients amb ACG de nou diagnòstic, s’inclogueren 30 pacients i s’objectivà que els pacients amb clínica isquèmica presentaven una major proporció d’afectació d’artèries vertebrals (OR 5.0, IC 95%: 0.99 – 24.86, p=0.051). Aquests pacients, en canvi, presentaven una menor proporció d’afectació d’artèries de gran cabal, essent l’aortitis un factor protector tant per la clínica isquèmica, en global (OR 19.0, IC 95%: 2.79 – 127.97, p=0.001), com per la pèrdua de visió permanent (OR 10.67, IC 95%: 1.12 – 101.34, p=0.04). En aquesta cohort, la presència de símptomes isquèmics s’associà a patrons de PET/TC diferenciats (p=0.001). En el segon estudi es demostrà que els pacients amb aortitis eren més joves (69.9 anys vs. 83.7 anys, p=0.04) i presentaven menys freqüentment clínica isquèmica, incloent clínica ocular (25.0% en els pacients amb aortitis vs. 84.2% en els pacients sense aortitis, p=0.006). La presència de cèl·lules gegants multinucleades en la biòpsia de l’artèria temporal s’associà de forma independent a la presència d’aortitis en el PET/TC (OR 12.23, p=0.046). En el grup de pacients amb aortitis no s’objectivà halo en el Doppler d’artèria temporal en cap cas. Així, podem concloure que el PET/TC és una eina útil en el diagnòstic de l’ACG, que permet identificar dos patrons de captació vascular diferents d’acord a la presència o absència de clínica isquèmica al diagnòstic de l’ACG. Així mateix, la presència d’aortitis en el PET/TC s’associa amb la presencia de cèl·lules gegants multinucleades en la biòpsia.
Giant cell arteritis is the most frequently diagnose vasculitis among patients older than 50 years. Classification is done by Hunder criteria (1990). Headache is the most frequent symptom. Vascular involvement has been extensively studied by imaging techniques, including temporal artery biopsy, suggesting the existence of two different clinical subsets of the disease. Evidence suggests that a high proportion of patients present with aortitis and that this situation is associated to a higher corticosteroid dose requirement and a higher risk of developing aortic aneurisms. Moreover, ocular involvement is the most feared complication nowadays as it is usually irreversible. The objective of this study was to identify different patterns of vascular involvement on PET/CT at GCA diagnosis according to the presence or absence of ischaemic manifestations at disease onset, including ocular involvement. Furthermore, we sought to describe clinical and histological patterns on TAB according to the presence or absence of aortitis in PET/CT. In the first study, performed with a cohort that comprised 30 patients with a newly diagnosed GCA, patients with ischaemic symptoms showed a higher proportion of vertebral involvement when compared to those patients without these symptoms (OR 5.0, CI 95%: 0.99 – 24.86, p=0.051). Patients with ischaemic manifestations showed a lower proportion of large vessel vasculitis, being aortitis a protector factor against ischaemic manifestations (OR 19.0, CI 95%: 2.79 – 127.97, p=0.001) and against permanent visual loss (OR 10.67, CI 95%: 1.12 – 101.34, p=0.04). In this cohort, the presence or absence of ischaemic symptoms was associated to different vascular patterns of PET/TC (p=0.001). In the second study we showed that patients with aortitis were younger (69.9 years vs. 83.7 years, p=0.04) and had less frequently ischaemic manifestations, including ocular involvement (25.0% of patients with aortitis vs. 84.2% of patients without aortitis, p=0.006). The presence of giant multinucleated cells on TAB was an independent risk factor for the presence of aortitis on PET/CT (OR 12.23, p=0.046). The halo sign was absent in patients with aortitis. We can conclude that PET/CT is a useful technique at GCA diagnosis that allows identifying two patterns of vascular involvement according to the presence or absence of ischaemic symptoms. When we compared PET/CT and TAB findings, we identified that aortitis was related to the presence of giant multinucleated cells.

A mathematical model is developed for investigating time dependent surface deformations of a hydrostatic water volume, when it is subjected to a sudden partial collapse or rise of the sea bottom. The model solves two-dimensional Navier-Stokes Equations on a vertical plane numerically by using Marker and Cell Method (MAC) for viscous and compressible fluid including all the nonlinear effects in the solution. For demonstration, a vertical motion was given to a section in a hypothetical reservoir bed within a short time period and the resulting velocity and pressure fields and the surface profile of the water body are obtained. Computational and physical aspects are discussed.

Background: Asthma during pregnancy has been associated with poor pregnancy outcomes such as pre-eclampsia, small for gestational age babies and preterm birth. Depression and anxiety are associated with reduced asthma control in non-pregnant individuals. This study investigated whether depression/anxiety in combination with pregnancies complicated by asthma has a negative effect on asthma control. Potential immune mechanisms that may drive worsening asthma were also investigated. Methods: One hundred and eighty-nine asthmatic women with and without depression/anxiety were followed throughout their pregnancies. Incidences of uncontrolled asthma and exacerbations were measured throughout gestation. At 18 and 30 weeks of gestation, monocyte inflammatory profile was examined using flow cytometric analysis of cell surface molecules (FACS) and peripheral blood mononuclear cell (PBMC) chemotaxis was also examined. Results: The incidence of uncontrolled asthma increased in women with depression/anxiety compared to women without depression/anxiety during pregnancy (unadjusted incidence rate ratio (IRR) 1.739, adjusted IRR 1.633, CI 1.092-2.442, p=0.017). Relative risk of experiencing uncontrolled asthma during pregnancy was also increased with depression/anxiety (unadjusted RR 1.619; adjusted RR 1.538, CI 1.114-2.122, p=0.009). There was no increase in the incidence rate ratio (unadjusted IRR 0.770; adjusted IRR 0.755, CI 0.412-1.382, p=0.362) or relative risk (unadjusted RR 0.867; adjusted RR 0.859, CI 0.496-1.489, p=0.589) of asthma exacerbations during pregnancies complicated by depression/anxiety. Asthma without depression/anxiety was associated with an increase in peripheral blood total monocyte percentage at 18 but not 30 weeks gestation when compared to asthmatic women with depression/anxiety (p=0.027). There were no changes in PBMC chemotaxis at 18 or 30 weeks gestation in pregnant women regardless of the presence of asthma or depression/anxiety. Conclusion: The presence of asthma and depression/anxiety during pregnancy is associated with an increase in uncontrolled asthma, but not a change in exacerbation risk. This increase in uncontrolled asthma in women with depression/anxiety was not a result of alterations in monocyte inflammatory profile or PBMC chemotaxis.
Thesis (M.Phil.) -- University of Adelaide, School of Paediatrics and Reproductive Health, 2015.

碩士
長庚大學
生物醫學研究所
99
Cervical cancer is the second largest cause of cancer-related death in women Worldwide, which occurs following persistent infection with specific high-risk human papillomaviruses (HPV). Even though the prophylactic vaccines were approved and became available in the market in recent years. There also need therapeutic vaccines to combat high-risk HPV-associated cancers of those had been infected. In order to increase tumor antigen presentation of therapeutic vaccines, we previously modified wild-type GM-CSF gene (wtGM) into codon-optimized GM-CSF (cGM), and then used lentivirus as a vector to deliver GM-CSF gene into a HPV-16 E6/E7 transformed cell line, TC-1. It was verified that TC-1/cGM cells significantly increased steady-state mRNA levels of GM-CSF and further enhanced GM-CSF protein expression. Granulocyte-macrophage colony-stimulating factor (GM-CSF) is an important hematopoietic growth factor and has profound effects on the functional activities of various leukocytes including T-cells and dendritic cells. In this study, the mice were inoculated with irradiated TC-1/cGM cell-based vaccine, and evaluated its effects on antigen-specific T-cell and dendritic cells (DCs) proliferation and activation by flow cytometric analysis. The antitumor immunity in vivo was monitored using a non-invasive animal position-emission tomography (microPET) imaging. It showed that mice vaccinated with TC-1 or TC-1/wtGM increased a lower level of CD8+ T-cells and DCs immune response. However, administration of TC-1/cGM vaccine revealed more potent immunity to elicit anti-tumor efficacy.

Wong Wai Lun.
Thesis (M.Phil.)--Chinese University of Hong Kong, 1998.
Includes bibliographical references (leaves 119-125).
Abstract also in Chinese.
Acknowledgments --- p.i
Legend for Figures --- p.ii
Legend for Tables --- p.iv
Abstract --- p.v
Abstract in Chinese --- p.ix
Chapter Chapter I --- Introduction --- p.1
Objective --- p.5
Chapter Chapter II --- Literature Review
Chapter II.1. --- Anatomy of the urinary system --- p.6
Chapter II.2. --- Physiology of the urinary system --- p.10
Chapter II.3. --- Methods for investigating the urinary system --- p.12
Chapter II.3.1. --- Plain film radiography --- p.12
Chapter II.3.2. --- Excretory Urogram --- p.12
Chapter II.3.3. --- Ultrasound --- p.13
Chapter II.3.4. --- Computed Tomography --- p.15
Chapter II.3.5. --- Renal Angiography --- p.16
Chapter II.3.6. --- Magnetic Resonance Imaging (MRI) --- p.16
Chapter II.3.7. --- Radionuclide Imaging --- p.17
Chapter II.4. --- Radiopharmaceuticals for renal parenchyma imaging --- p.17
Chapter II.4.1. --- Tc-99m GHA --- p.18
Chapter II.4.1.1. --- Chemistry of Tc-99m GHA --- p.18
Chapter II.4.1.2. --- Preparation --- p.18
Chapter II.4.1.3. --- Doses --- p.18
Chapter II.4.1.4. --- Biological behavior --- p.19
Chapter II.4.2. --- Tc-99m DMSA
Chapter II.4.2.1. --- Chemistry of Technetium-99m Dimercaptosuccinic Acid (Tc-99m DMSA) --- p.20
Chapter II.4.2.2. --- Chemical property of Tc-99m DMSA --- p.21
Chapter II.4.2.3. --- Preparation --- p.22
Chapter II.4.2.4. --- Radiochemical purity measurement --- p.22
Chapter II.4.2.5. --- Doses --- p.23
Chapter II.4.2.6. --- Pharmacokinetic of Tc-99m DMSA --- p.23
Chapter II.4.2.7. --- Renal handling of injected Tc-99m DMSA --- p.25
Chapter II.5. --- General consideration for quantitative uptake measurement in organs --- p.26
Chapter II.5.1. --- Clinical significance of renal Tc-99m DMSA uptake --- p.28
Chapter II.5.2. --- Special consideration and problems for quantitative renal Tc-99m uptake measurement --- p.29
Chapter II.5.3. --- Suggestions and solutions for quantitative renal Tc-99m uptake measurement --- p.29
Chapter II.5.3.1. --- Planar images Vs SPECT images for quantification --- p.29
Chapter II.5.3.2. --- Background subtraction --- p.31
Chapter II.5.3.3. --- Choice of location for background ROI --- p.32
Chapter II.5.3.4. --- Attenuation --- p.35
Chapter II.5.3.5. --- Principle of the conjugate view method --- p.36
Chapter II.5.3.6. --- Body thickness and kidney depth measurement --- p.37
Chapter II.6. --- Glomerular Filtration
Chapter II.6.1. --- Introduction --- p.39
Chapter II.6.2. --- Gold standard for GFR measurement --- p.40
Chapter II.6.3. --- Laboratory studies for the measurement of glomerular filtration : Serum Creatinine and Blood Urea Nitrogen (BUN) levels --- p.41
Chapter II.6.3.1. --- Calculation of Creatinine Clearance Rate --- p.43
Chapter II.6.3.2. --- Critique for using creatinine clearance as a measurement of renal function --- p.44
Chapter II.6.3.3. --- Limitation of the serum creatinine concentration used alone as a measurement of renal function --- p.46
Chapter II.6.4. --- Radionuclide technique for the assessment of the glomerular function --- p.48
Chapter II.6.4.1. --- Diethylene Triamine Penta Acetic acid (DTPA) --- p.49
Chapter II.6.4.2. --- Methods
Chapter II.6.4.2.1. --- Measurement of Glomerular Filtration Rate using Tc-99m DTPA with single injection techniques --- p.51
Chapter II.6.4.2.2. --- Compartment model --- p.52
Chapter II.6.4.2.2a. --- Two-compartment model --- p.52
Chapter II.6.4.2.2b. --- Single-compartment model --- p.54
Chapter II.6.4.2.3. --- Single blood sample technique: a modification of Tauxe's OIH method in which counts in a single plasma sample correlated with a GFR nomogram --- p.56
Chapter II.6.4.2.4. --- Gamma camera based method --- p.58
Chapter II.6.4.2.4a. --- Gates-modification of Schlegel's OIH technique --- p.58
Chapter II.6.4.2.4b. --- Critique for the Gamma camera technique for measuring GFR --- p.62
Chapter II.7. --- The relationship between the Tc-99m DMSA uptake and GFR --- p.67
Chapter Chapter III --- Material and Methods --- p.69
Chapter III.1. --- Subjects and Sampling Methods --- p.69
Chapter III.2. --- Quantitation of Absolute DMSA uptake --- p.70
Chapter III.2.1. --- Parameters for Tc-99m DMSA uptake study --- p.70
Chapter III.2.1.1. --- Materials and methods --- p.70
Chapter III.2.1.1.1. --- Instrumentation --- p.70
Chapter III.2.1.1.2. --- Dosage --- p.70
Chapter III.2.1.1.3. --- Optimum acquisition start time --- p.70
Chapter III.2.1.1.4. --- Length of acquisition time --- p.71
Chapter III.2.1.1.5. --- Acquisition parameter --- p.71
Chapter III.3. --- Calculation of absolute renal DMSA uptake --- p.72
Chapter III.3.1. --- Attenuation Coefficient factor(μ) --- p.73
Chapter III.3.2. --- Table attenuation --- p.75
Chapter III.3.3. --- Body thickness measurement --- p.77
Chapter III.3.4. --- Decay correction --- p.78
Chapter III.3.5. --- Calculation of DMSA uptake --- p.78
Chapter III.3.6. --- Counting dose injected --- p.80
Chapter III.3.7. --- Calculation of absolute quantitation of Tc-99m DMSA uptake --- p.80
Chapter III.3.8. --- Dose infiltration --- p.81
Chapter III.4. --- GFR measurement --- p.82
Chapter III.4.1. --- Instrumentation --- p.82
Chapter III.4.2. --- Methods --- p.82
Chapter III.5. --- Statistical and analytical methods --- p.84
Chapter Chapter IV --- Results --- p.87
Chapter IV. 1. --- Characteristics of experimental subjects and their serum creatinine profile --- p.88
Chapter IV.2. --- Absolute Tc-99m DMSA uptake
Chapter IV.2.1. --- The change of absolute Tc-99m uptake with time --- p.89
Chapter IV.2.2. --- Absolute Tc-99m DMSA uptake measurement at 6 and 24 hours --- p.90
Chapter IV.2.3. --- Gender difference in absolute Tc-99m uptake measurement at 6 hour --- p.92
Chapter IV.3. --- GFR measurement --- p.93
Chapter IV.3.1. --- GFR measurement by single (3hr) and double (1&3 hrs) plasma sampling --- p.93
Chapter IV.3.2. --- Gender difference in GFR measurement using single plasma sampling --- p.96
Chapter IV.4. --- Univariate Correlation --- p.97
Chapter IV.4.1. --- Correlation between GFR using single plasma sampling and absolute Tc-99m uptake --- p.97
Chapter IV.4.2. --- Correlation between GFR using single plasma sampling and plasma creatinine levels --- p.98
Chapter IV.4.3. --- Correlation between anthropometric variables on GFR(3 hr) --- p.99
Chapter IV.4.4. --- Correlation between anthropometric variables and serum creatinine plasma level on absolute Tc-99m DMSA uptake measurement at 6 hour --- p.101
Chapter IV.4.5. --- Multiple linear stepwise regression --- p.103
Chapter Chapter V. --- Discussion
Chapter V. 1 --- . Review of the study --- p.104
Chapter V.1.1. --- Experimental subjects and their absolute Tc-99m DMSA uptake (%) at 6 hr --- p.104
Chapter V.1.2. --- Experimental subjects and their GFR(3 hr) --- p.105
Chapter V.2. --- Discussion on subject --- p.105
Chapter V.2.1. --- Subject preparation --- p.106
Chapter V.3. --- Discussion of method --- p.106
Chapter V.3.1. --- Equipment --- p.106
Chapter (a) --- Dose calibrator --- p.106
Chapter (b) --- The sensitivity of the head 1 and 2 of the gamma camera --- p.106
Chapter (c) --- Validation of quantification of injected activity by gamma camera method--------constancy of performance for gamma camera --- p.110
Chapter (d) --- LEHR Collimator --- p.112
Chapter (f) --- Dead time loss --- p.112
Chapter V.4. --- Discussion on measurement --- p.113
Chapter (a) --- Length of acquisition time --- p.113
Chapter (b) --- Attenuation Coefficient factor (\x) --- p.113
Chapter (c) --- "Body thickness, L, measurement" --- p.113
Chapter (d) --- Optimum acquisition time for data collection --- p.115
Chapter v.5. --- Discussion on overall error estimation --- p.115
Chapter (a) --- Tc-99m DMSA uptake measurement at 6 hr --- p.115
Chapter (b) --- GFR measurement by single (3 hr) sample --- p.116
Chapter Chapter VI --- Conclusion --- p.117
Reference --- p.119
Appendix I --- p.126
Appendix II --- p.128
Appendix III --- p.134

The MHC class I status of tumour cells during immunotherapy is often underestimated. It represents one of important tumour escape mechanisms and thus can contribute to the failure of most of the cancer clinical trials that are usually based on the induction of cytotoxic T cell responses. Epigenetic changes in the promoters of genes involved in the MHC class I Ag presentation can result in decreased expression of the cell surface MHC molecules on tumour cells. Thus, epigenetic modifiers can restore an expression of the MHC class I molecules and make tumours visible to the CD8+ effector cells. Besides the epigenetic changes on the tumour cells, epigenetic modulators affect cells of the immune system such as dendritic cells (DC). Tumour cells can escape from the immune response not only by changes in the cancer cells, but also by influencing, expanding and/or activating immunoregulatory cell populations, such as regulatory T cells (Treg). This thesis focuses on the potential of the DC-based vaccines against HPV-16-associated tumours with a different MHC class I expression, on the combination of cancer immunotherapy with the treatment using epigenetic modifiers, with special attention paid to their effects on DC, and, finally, on the impacts of the anti-CD25 antibody (used for Treg elimination) on Treg and NKT...

Introduction: Multidrug resistance (MDR) is a condition defined by the cross-resistance to several non-structurally related drugs, representing one of the major setbacks to the success of chemotherapy. One of the best studied MDR mechanisms is the overexpression of efflux pumps, such as P-glycoprotein (Pgp), multiple resistance-related protein 1 (MRP-1) and major vault protein (LRP). These proteins confer resistance to a large spectrum of similar substrates, despite their different extrusion mechanisms. Pharmacologic inhibition of MDR transporters is the major strategy to overcome this phenotype. Verapamil is an L-type calcium-channel blocker and a modulator for Pgp and MRP-1. L-buthionine-sulfoximine (BSO) is a γ-glutamylcysteine synthase inhibitor and can be used to functionally decrease MRP-1 activity. Aim: In this study we aim to compare transport kinetics for human colorectal adenocarcinoma cell lines, one sensitive and another resistant to chemotherapy, in the presence and absence of MDR reversers, using 99mTc-Sestamibi. Methods: MDR proteins expression was evaluated in sensitive (WiDr) and resistant (LS1034) human colorectal adenocarcinoma cell lines. Intracellular and plasma membrane Pgp and MRP1, and LRP expression was analyzed by flow-cytometry; Pgp expression was also analyzed by western blot. Cellular transport kinetics was analyzed in the presence and absence of MDR modulators, verapamil and BSO, using 99mTc-Sestamibi. Uptake and retention studies were performed using sensitive and resistant cells. MDR modulation was evaluated by performing retention studies in resistant cells after incubation with the referred drugs, for different time intervals (10 and 60 minutes) and concentrations (10, 25, 50 and 100 µM). Results: Pgp and MRP-1 expression was significantly higher (p<0.05) in resistant cells when comparing with the sensitive ones, although LRP was also expressed. Western blot studies confirmed flow-cytometry results. 99mTc-Sestamibi uptake and retention percentage were significantly higher (p<0.05) in the sensitive cell line, comparing with the resistant one for all time-points considered. In resistant cells incubated with MDR modulators there were no statistically significant differences (p>0.05) when considering the curves as a whole. However, for the first minutes after incubation with 99mTc-Sestamibi, there were differences among the MDR modulators used. Conclusions: In vitro kinetic studies using 99mTc-Sestamibi could be an indicator of MDR phenotype in colorectal adenocarcinoma cells. As the modulators used showed a reversion of the retention profile only for the first minutes, their administration should occur immediately before the administration of cytotoxic drugs.
Introdução: A multirresistência a fármacos (MDR) é definida pela resistência cruzada a diversos fármacos não relacionados estruturalmente, representando um dos principais factores de insucesso da quimioterapia. Um dos mecanismos mais estudados de MDR é a sobre-expressão de proteínas de efluxo, como a glicoproteína P (Pgp), a multiple resistance-related protein-1 (MRP-1) e a major vault protein (LRP). A inibição farmacológica dos transportadores associados a MDR é a principal estratégia investigada para superar este fenótipo. O verapamil é um bloqueador dos canais de cálcio do tipo L e um modulador da Pgp e da MRP-1. A L-butionina-sulfoximina (BSO) é um inibidor da γ-glutamilcisteina sintetase e pode ser utilizada para diminuir a actividade funcional da MRP-1. Objectivo: Neste estudo pretendemos comparar a cinética de transporte para linhas celulares de adenocarcinoma colorrectal, uma sensível (WiDr) e outra resistente (LS1034), na presença e ausência de reversores de MDR, utilizando 99mTc-Sestamibi. Métodos: A expressão de proteínas MDR foi avaliada nas duas linhas celulares referidas. A expressão de Pgp e MRP-1 intracelular e membranar, e a expressão de LRP foram analisadas por citometria de fluxo; a expressão de Pgp foi também analisada por western blot. A cinética de transporte foi analisada na presença e ausência dos moduladores de MDR verapamil e BSO, usando 99mTc-Sestamibi. Foram efectuados estudos de captação e retenção utilizando células sensíveis e resistentes. A modulação de MDR foi avaliada pela realização de estudos de retenção em células resistentes após a incubação com os fármacos referidos, durante diferentes intervalos de tempo (10 e 60 minutos) e concentrações (10, 25, 50 e 100 µM). Resultados: A expressão de Pgp e MRP-1 foi significativamente superior (p<0,05) nas células resistentes, embora a LRP também estivesse expressa. Os estudos de western blot confirmaram os resultados da citometria de fluxo. A captação e retenção de 99mTc-Sestamibi foram significativamente superiores (p<0,05) na linha celular sensível para todos os tempos de amostra considerados. Nas células resistentes incubadas com moduladores de MDR não houve diferenças estatisticamente significativas (p>0,05) quando se consideraram as curvas como um todo. No entanto, para os primeiros minutos após a incubação com 99mTc-Sestamibi, houve diferenças entre as células incubadas com os moduladores. Conclusões: Estudos cinéticos in vitro com 99mTc-sestamibi podem ser um indicador de fenótipo MDR em células de adenocarcinoma colorrectal. Como os moduladores usados apenas mostraram reversão do perfil de retenção para os primeiros minutos, a sua administração deverá ser feita imediatamente antes da administração de citotóxicos.

碩士
長庚大學
生物醫學研究所
103
The antigens of tumor cells are highly similar to those in the normal host cells, suggesting that they are not easily recognized by the host immune system. Early research found that immunization with tumor cell based vaccines may effectively induce tumor antigen-specific T cell activation and proliferation. Granulocyte-macrophage colony-stimulating factor (GM-CSF) has been broadly used as adjuvant with DNA vaccines and tumor vaccines. Immunizations with codon-optimized GM-CSF plasmid DNA-vaccine result in an effective cytotoxic T lymphocyte response. Previous studies have shown that a highly-productive GM-CSF -based tumor cell vaccine may induce immune-suppressive effects that are dependent on the number of immunization. However, in this study, we found that different numbers of vaccinations resulted in different tumor rejection effects. Thus, we hypothesized that the GM-CSF-secreting tumor cell vaccine induces tumor rejection and improves prognosis after tumor cell challenge, which may be associated with interferon-producing killer dendritic cell (IKDC) activation. In this study, we used the high GM-CSF-producing TC-1 cells as a tumor cell vaccine that, simultaneously expresses the HPV E6 and E7 oncoproteins. The mice were subcutaneously injected either one, three or five times. The mice were then sacrificed, and the frequencies of Treg cell and IKDC in splenocytes were analyzed via flow cytometry. We found that the percentage of Treg cells in splenocytes is associated with the number of immunizations; however, this association was not only due to tumor rejection and survival. The IKDC percentage in the splenocytes from the group of 3 times vaccinations was significantly higher than the other groups, which showed a similar trend with survival rate. These data suggest that the efficacy of tumor rejection was not only associated with Treg cell effects but also the activation of IKDC proliferation.

Human thyroid cancer is the most commonly occurring cancer of the endocrine gland having good survival rate, but some patients show recurrence with an invasive phenotype and treatment failures. The mechanisms behind this invasive phenotype are not well understood in TC. Previously our group has identified a pro-migratory role of relaxin-like peptides in thyroid cancer that is mediated by S100A4. We have observed in human TC cells that extracellular S100A4 induces migration and activates ERK1/2, JNK/SAPK and NFkB signaling pathways. Employing immunohistochemistry and immunofluorescence we have identified the expression of RAGE in human TC primary cells, cell lines, and in tumor tissues but not in normal thyroid tissues. We showed that S100A4 binds to RAGE in TC cells and that RAGE and its cytoplasmic partner Dia-1 mediate the S100A4-induced migration of TC cells. This study identified a crucial role of RAGE in TC cell migration induced by S100A4.

碩士
亞洲大學
生物科技學系碩士班
93
Human papillomaviruses (HPV) are a family of small DNA viruses that have been linked to a variety of human diseases. The high-risk types of HPV, including HPV-16 and HPV-18, have been implicated in the causation and malignant progression of human cervical cancer. The consistent expression of two viral genes, E6 and E7, are required for the establishment and maintenance of a fully transformed phenotype. Therefore, the E6 and E7 genes can be considered relevant targets for anti-cancer therapy. Recently, a powerful reverse genetic tool termed small interfering RNA (siRNA) has been developed which can induce sequence-specific gene silencing through RNA-mediated interference (RNAi) in mammalian cells. this thesis, we demonstrated that synthetic siRNA targeted to various region of HPV-16 E6 and E7 mRNAs can specifically block the expression and function of respective viral oncogenes in HPV-positive TC-1 tumor cell lines. It is shown that siRNAs to E6 and E7 messages were useful in inhibiting cell growth and in reversing the transformed phenomena like anchorage-independency. These observations augurs well for the therapeutic potential of anti-E6 and anti-E7 siRNA in HPV-related cancers.