Waran används sedan 60 år som blodförtunnande läkemedel för att förebygga eller förhindra progress av blodproppssjukdom. I Sverige behandlas årligen cirka 1 % av befolkningen med waran. I Östergötland uppskattas antal waranpatienter till cirka 3000. Waran hämmar enzymet VKORC1 som ansvarar för vitamin K omsättningen i kroppen. Vitamin K behövs som kofaktor för flera koagulationsfaktorer. Behandling med waran är förenad med en svår balansgång och kräver en noggrann dosering. Stora skillnader i dosbehov mellan olika individer, beroende på ärftliga och miljöfaktorer, gör waran till ett svårhanterligt läkemedel. För låg dos medför otillräcklig effekt och därmed risk för minskat skydd mot blodproppssjukdom. För hög dos leder till allvarliga blödningskomplikationer. Uppskattningsvis 1 – 3 % livshotande blödningsfall registreras årligen efter waranbehandling. Därför måste behandlingen kontrolleras noga med analys av protrombinkomplex (PK) och dosjusteringar göras med ledning av resultaten. Två olika metoder finns att använda för mätning av PK. I Norden och i Japan används Owrens metod (utvecklat i Norge under 40- och 50-talet av Paul Owren). I de flesta andra länder används Quickmetoden (utvecklat i USA under 30-talet av Armand Quick). Den senare metoden är förenad med stora variationer mellan olika analyslaboratorier. I Norden, däremot, där Owrens metod används finns det ofta bra överensstämmelse mellan olika laboratorier i PK-resultat. Beroende på vilken PK-metod som används, kan samma patient få olika warandoser vilket ökar risker för under- eller överbehandling. Vi har i samarbete med flera sjukhus och antikoagulationsmottagningar (AK-mottagningar) i sydöstra Sverige studerat dels mekanismerna bakom skillnader i warandos mellan olika patienter, och dels tittat varför de olika PK-metoder skiljer sig så mycket som de gör. I studien har vi identifierat genetiska varianter av enzymet VKORC1. Av de undersökta patienter som gick på waran under längre tid, har vi identifierat en grupp som markant skiljde sig från de övriga. Denna grupp hade warandoser som var betydligt lägre än de övriga. När vi kartlade deras arvsmassa, fann vi att lågdospatienterna hade genvarianten VKORC1*2. Dessutom hade patienter med denna variant svårigheter att få stabila PK-värden. De gjorde också fler besök på AK-mottagningar än andra patienter. Vi har därför konstaterat att en del av de problem som är förenade med waranbehnadlingen kan förklaras av VKORC1*2 varianten. Vetskap om denna variant skulle troligen underlätta behandlingen framför allt under inledningsfasen då patienter med VKORC1*2 riskerar blödningar på grund av överdosering. Vi har identifierat att provförspädning enligt Owrens metod är nödvändig för harmonisering av PK-resultatet mellan olika länder. Quickmetoden använder inte förspädning av patientprov till skillnad från Owrens metod. När vi modifierade en Quickmetod genom att förspäda prover enligt Owrens metod noterade vi en bra överensstämmelse mellan de två olika metoderna. Däremot var resultatet sämre utan provförspädning. Vi anser att Quickmetoder kan uppnå lika bra resultat som Owrens metod om prover förspäds som i Owrens metod. Det skulle gynna patienter som reser mellan olika regioner eller länder och leda till en bättre övervakning av waranbehandling internationellt. I studien har dessutom en metod för mätning av waran i blodet utvecklats. Metoden som är den enda i sitt slag i Norden ger möjlighet att studera hur läkemedlet beter sig i kroppen. Vi har med denna metod kunnat upptäcka patienter som har onormala nedbrytningar av waran. / Warfarin was introduced more than 60 years ago and is used worldwide for the prophylaxis of arterial and venous thromboembolism in primary and secondary prevention. The drug is orally administered as a racemic mixture of (R)- and (S)-enantiomers. The (S)-form is mainly responsible for the anticoagulant effect and is metabolised by CYP2C9 enzyme in the liver microsomes. Warfarin exerts its pharmacological action by inhibiting the key enzyme (VKORC1) that regenerates vitamin K from an oxidised state to a reduced form. The latter is a cofactor for the post-translational modification of a number of proteins including coagulation factors II, VII, IX and X. The vitamin K-dependent modification provides these factors with the calcium-binding ability they require for the interaction with cell membranes of their target cells such as platelets. Warfarin is monitored by measuring prothrombin time (PT) expressed as INR. Two main methods exist for PT analysis. The Owren method is used mainly in the Nordic and Baltic countries, in Japan, whereas the Quick method is employed in most other countries. Warfarin management is associated with some complications. Unlike many other drugs the dose for a given patient cannot be estimated beforehand, dose-response relationship is not predictable, and the prevention of thrombosis must be balanced against the risk of bleeding. Furthermore, the different PT methods used to monitor the drug are sometimes not in agreement and show significant discrepancies in results. In an attempt to clarify the mechanisms influencing the inter-individual variations in warfarin therapy and to detect the factors that contribute to differences between PT methods, studies were conducted in collaboration with hospitals and anticoagulation clinics in the south-eastern region of Sweden. First, a stereo-specific HPLC method for measurement of warfarin enantiomers was developed and validated. With this method, the levels of plasma warfarin following its oral administration can be studied and evaluated. Abnormal clearance in some patients can be detected, and patient compliance can be verified. Furthermore, differing ratios of (S)- and (R)-isomers can be identified. The impact of common VKORC1 polymorphisms on warfarin therapy was investigated. This study has shown that the VKORC1*2 haplotype is an important genetic determinant for warfarin dosage and is associated with difficulties in attaining and retaining therapeutic PT-INR. Further, significant differences in warfarin S/R-ratio was detected between patients with VKORC1*2 and VKORC1*3 or VKORC1*4 variants. This difference was not coupled with CYP2C9 genotype. The effects of predilution of patient plasma samples, sources of thromboplastin and deficient plasma on between PT methods agreement were studied. This study has revealed that sample predilution according to the Owren method is to be preferred for the harmonisation of PT results. Undiluted samples, in contrast, according to the Quick method have shown reduced correlation between two different thromboplastin reagents. Sources of thromboplastin and deficient plasma were only of minor importance.

Thesis (Ph. D.)--Lehigh University, 2003. / Includes vita. Includes bibliographical references (leaves 150-165).

Glucose uptake occurs in skeletal muscle under basal conditions, and increases in response to stimuli such as insulin and exercise. Exercise is known to increase blood flow, and it appears that insulin has similar hemodynamic effects, including increased blood flow and capillary recruitment, which can modify the amount of glucose uptake occurring under each condition. Here we study factors affecting both basal and stimulated myocyte glucose uptake, with a particular focus on vasoactive agents. Insulin stimulates the release of endothelin-1 (ET-1), a potent vasoconstrictor, from endothelial cells in culture. As yet it is unknown whether ET-1 is a type A (causing nutritive perfusion) or a type B (non-nutritive) vasoconstrictor, so here we use the pump-perfused rat hindlimb to characterize the distribution effects of ET-1. We show that ET-1 causes a type A vasoconstriction, stimulating basal metabolism at low doses, while at high doses the distribution of flow changes to become non-nutritive, inhibitory to metabolism. As a general vasodilator prevents both metabolic and hemodynamic effects, the effects on metabolism are due to the redistribution of flow. These redistribution effects are confirmed by the ability of high dose ET-1 to decrease aerobic tension development in the contracting hindlimb, and by the ability of low dose ET-1 to increase the interstitial glucose concentration. Given this understanding of the effects of ET-1 alone, we can investigate the interactions between ET-1 and insulin. In the perfused rat hindlimb, insulin has not been observed to have any vasodilatory effect, whereas here for the first time insulin appears to have vasodilator-like actions against ET-1 mediated vasoconstriction. Also, the redistribution of flow by ET-1 does not appear to alter the metabolic effect of insulin to cause glucose uptake at either dose of ET-1 used. Nitric Oxide (NO) is thought to be the mechanism by which insulin causes vasodilation in muscle. A previous study has shown that methacholine (MC), by increasing NO, was able to augment insulin-mediated glucose uptake and capillary recruitment, while other NO donors were unable to do so. Here we show that, at the dose used to increase glucose uptake in the previous study, MC has only a vasodilatory effect, and no direct effect on glucose uptake, in the perfused rat hindlimb. At higher doses, an effect on glucose uptake can be observed. This means that the increase in capillary recruitment by MC was responsible for the elevated insulin-mediated glucose uptake, and there was no direct effect of MC on glucose uptake. A recent publication suggested that the Na+-D-glucose cotransporter (SGLT1) was essential for insulin-mediated glucose uptake, although not required for basal glucose uptake. The implications of this detract from our proposed role of blood flow redistribution in insulin action. In attempting to reproduce these results in the perfused rat hindlimb we found that SGLT1 is not required for insulin-mediated glucose uptake, and confirmed this using a low sodium buffer, which would also inhibit the transporter. We conclude that SGLT1 is not required for insulin-mediated glucose uptake. Our results therefore suggest that complex interactions are involved in insulin action, some of which involve hemodynamic actions that are capable of altering insulin-mediated glucose uptake, and others in which insulin itself can limit the action of other vasomodulators, such as ET-1. It is apparent, however that SGLT1 in the endothelium may not be necessary for the metabolic effects of insulin, and that blood flow distribution, or capillary recruitment is therefore of great importance in delivering glucose to myocytes.

<p>Analysis of M-components is an important part of the diagnosis of monoclonal gammopathies and for the evaluation of disease response during treatment. In this project, two widely used electrophoresis methods and their corresponding immunotyping method were compared to evaluate the sensitivity of each method for the detection of small M-components. The project included 30 plasma samples from patients with identified M-components; 10 samples containing each IgG, IgA and IgM, respectively. All samples were diluted with normal EDTA plasma to achieve M-components of 5,00g/L. The samples were then serially diluted to achieve M-component concentrations of; 5,00, 2,50, 1,25, 0,63, 0,31 and 0,16g/L. All 180 samples were analysed with agarose gel electrophoresis and capillary electrophoresis. The dilutions above and below the detection level of each method were then analysed with immunofixation and immunosubtraction. The results showed good agreement between agarose gel electrophoresis and capillary electrophoresis in the highest concentrations of IgG and IgM. With agarose gel electrophoresis, IgA was detected in the same location as transferrin and the lowest concentration detected were therefore 1,25g/L. Besides the samples containing IgG, immunofixation was the most sensitive method.</p>

<p>Haematology instruments using optical and fluorescence techniques have improved the platelet count in domestic animals. There are still some difficulties present, especially when counting cat thrombocytes due to their ability to aggregate and the occurrence of large platelets.</p><p>The objective of this study was to evaluate and compare the platelet count, mean platelet volume and platelet crit in dogs, horses and cats on Advia 2120 and Sysmex XT-2000iV.</p><p>Fresh blood samples from 64 dogs, 40 horses and 39 cats with various medical conditions were analysed on both instruments. Manual blood smears of all feline samples were scrutiniously analysed to evaluate the aggregation warning flag from Advia.</p><p>There was good agreement between the instruments for the optical platelet count in dogs and cats. Slightly higher values were reported from Advia. Samples from horses presented poor correlations for all studied parameters. Platelet clumps appeared in 70% of the 37 scrutinized feline blood smears, while 46% of the samples generated aggregation warning flags from the Advia instrument.</p><p>Advia and Sysmex showed good agreement for platelet counts in blood from dogs and cats. Mean platelet volume and platelet crit need further evaluation before conclusions can be made concerning their clinical relevance. The sensitivity of the platelet aggregation warning flag from the Advia instument needs further elevation.</p>

Law, Pak Tsun = 研究氧化鋅納米粒子應用在生物醫學的可能性 / 羅百浚. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2011. / Includes bibliographical references (p. 62-63). / Abstracts in English and Chinese. / Law, Pak Tsun = Yan jiu yang hua xin na mi li zi ying yong zai sheng wu yi xue de ke neng xing / Luo Baijun. / Chapter 1 --- Introduction --- p.1 / Chapter 2 --- Background --- p.2 / Chapter 2.1 --- Introduction of Zinc Oxide Nanostructures --- p.2 / Chapter 2.1.1 --- Biomedical Application of ZnO nanostructure --- p.3 / Chapter 2.2 --- Current problems of Zinc Oxide Nanostructures on Biological Application --- p.5 / Chapter 2.2.1 --- Behaviours of ZnO Nanostructure in Cell Culture --- p.6 / Chapter 2.2.2 --- Cytotoxicity of ZnO Nanostructures --- p.8 / Chapter 2.2.3 --- Other Synthetic Problems --- p.9 / Chapter 2.3 --- Summary --- p.9 / Chapter 3 --- Instrumentation and Experimental Methodology --- p.11 / Chapter 3.1 --- Electron Microscopy --- p.11 / Chapter 3.1.1 --- SEM --- p.12 / Chapter 3.1.2 --- TEM --- p.13 / Chapter 3.1.3 --- Elemental Analysis --- p.14 / Chapter 3.2 --- X-ray Photoelectron Spectroscopy --- p.16 / Chapter 3.3 --- X-ray Diffraction --- p.17 / Chapter 3.4 --- Dynamic Light Scattering --- p.18 / Chapter 3.5 --- Photoluminescence Spectroscopy --- p.19 / Chapter 3.6 --- MTT Assay --- p.20 / Chapter 3.7 --- Cells Incubation --- p.22 / Chapter 4 --- Synthesis and Characterization of Zinc Oxide Nanostructure --- p.23 / Chapter 4.1 --- Hydrophobic Zinc Oxide Nanoparticle --- p.23 / Chapter 4.1.1 --- Synthetic Method --- p.23 / Chapter 4.1.2 --- Characterizations and Property measurements of the ZnO nanoparticles --- p.24 / Chapter 4.2 --- Hydrophilic Zinc Oxide Nanoparticle --- p.29 / Chapter 4.2.1 --- Synthetic Method --- p.29 / Chapter 4.2.2 --- Characterizations and property measurements of the ZnO nanoparticles --- p.30 / Chapter 5 --- Interaction of Zinc Oxide with Human Cells --- p.35 / Chapter 5.1 --- Can Zinc Oxide Nanoparticles Enter the cells? --- p.35 / Chapter 5.1.1 --- Exp erimental --- p.36 / Chapter 5.1.2 --- General Description --- p.36 / Chapter 5.2 --- What Affect the Cellular Uptake of ZnO Nanoparticles? --- p.37 / Chapter 5.2.1 --- Concentration of ZnO Nanoparticles --- p.37 / Chapter 5.2.2 --- Agglomeration of ZnO nanoparticles --- p.40 / Chapter 5.2.3 --- Reaction with Buffer Solution and Culture Medium --- p.41 / Chapter 5.2.4 --- Summary --- p.45 / Chapter 5.3 --- Cell Viability after Introducing ZnO Nanoparticle to the Cells ´Ø --- p.56 / Chapter 5.3.1 --- Experimental --- p.56 / Chapter 5.3.2 --- Results and Discussion --- p.57 / Chapter 6 --- Conclusion --- p.59 / Bibliography --- p.62 / Chapter A --- TEM sample preparation procedure for animal cell --- p.64

<p>Mantle cell lymphoma (MCL) is a non-Hodgkin B-cell lymphoma with very bad prognosis. The genetic hallmark of MCL, is the translocation t(11;14)(q13;q32) which leads to overexpression of cyclin D1, a D-type cyclin that is not usually expressed at high levels in normal B lymphocytes.</p><p> </p><p>Previous studies indicate that cannabinoid receptors are expressed in lymphoma and have shown that lymphoma cell death is induced as a result of exposure to cannabinoids (ligands).</p><p> </p><p>The aim of this diploma work was to combined cytostatics with the cannabinoid receptor ligand R (+)-Methanandmide (R-MA). Our data suggest that combination treatment with cytostatics and R-MA induces synergistic effects in most cases.</p>

<p>The insulin-like 3 peptide is autocrine/paracrine insulin-related hormone with a size of approximately 6kDa [1]. It mediates through a leucine richG-coupled receptor named LGR8. INSL3 is mainly expressed in human Leydig cells and is directly responsible for migration of the testis during the pre-natal period in maledevelopment. [2]</p><p>INSL3 mRNA has recently been verified in human platelets whereas no mRNA has been detected for LGR8 (by Sanofi-Aventis GmbH in Frankfurt,Germany), indicating that INSL3 might be released through paracrine functions at sites of platelet adhesion and aggregation upon a vascular injury.Furthermore, has activated platelets been shown to translate essential proteins upon activation, in a term called “signal-dependent protein synthesis”.The B-Cell lymphoma-3 protein (BCL-3) is an example of such a protein [3], and there is a possibility that INSL3 might be also.</p><p>In this thesis we wanted to detect the relaxin- like peptide 3 hormone (INSL3). (Its function, location and the timeframe of its release, when/if it issecreated in stimulated platelets).The source of platelet-derived INSL3 can be found with Western blotting and Enzyme immunoassay.</p><p>Detection of the insulin-like 3 peptide in human platelets turned out to be a difficult challenge due to the small amount of INSL3 secretion uponplatelet activation; hence the total amount of INSL3 produced might be below detection limit.</p>

<p><strong>Objective</strong><em>:</em> Estimation of glomerular filtration rate (GFR) is an important tool in the diagnosis and management of chronic kidney disease. Today creatinine is the most frequently used marker for kidney function though several studies indicate that cystatin C is a superior marker. The purpose of this study was to validate Abbott Diagnostics turbidimetric cystatin C assay and enzymatic creatinine assay.</p><p><strong>Methods</strong><em>:</em> The validation was performed by studies of CV for the two methods and correlations between the two and other available methods for assessing GFR. The stability of cystatin C at room temperature was also evaluated.</p><p><strong>Results</strong><em>: </em>Both methods showed good precision. The Abbott cystatin C assay generally gave lower values and thereby higher estimated GFRs than the correlated Gentian method. The Abbott enzymatic creatinine assay gave higher values than the correlated Jaffe method. Those results are generally unexpected, but in this study the cause is an automatically applied negative intercept used together with the Jaffe method. Cystatin C showed high stability when stored at room temperature.</p><p><strong>Conclusions</strong><em>:</em> Estimated GFRs tend to differ depending on the choice of method for analyzing cystatin C or creatinine and this study gives an overview of the range of variation. The study also enlightens the need for an international calibrator for the cystatin C methods presented by different manufacturers.</p>

<p> </p><p>Genotyping can be used to link genetic variation among individuals to certain diseases or conditions. Some known disorders and states that are dependent on single nucleotide polymorphism (SNPs) are lactose intolerance, venous thrombosis, hereditary hemochromatosis and the difference in sensibility among people to metabolise drugs.</p><p>In this project a new kit, TaqManÒ Sample-to-SNP KitÔ <strong>for extraction of DNA and preparation of the extract for genotyping with real-time PCR and allelic discrimination, was evaluated. QIAamp® DNA Blood Biorobot® MDx Kit was used as the reference method.</strong></p><p>The purpose of the comparison was to find a method that makes DNA extraction from blood samples cheaper and faster, but with the same reliability as the reference procedure.</p><p>The results of the evaluation showed a complete agreement of the genotype results between the methods tested, which means that the new method was as reliable as the reference method. The costs of reagents and material would be reduced with 52% if the new method is adopted, that alone would result in a cost reduction of 144 000SEK a year with a sample volume of 650 samples/month. The time for DNA extraction would also be reduced with the new procedure.</p><p> </p>