Magister Pharmaceuticae - MPharm / Cardiovascular diseases (CVDs) such as ischaemic heart diseases, heart failure and stroke remain a major cause of death globally. Various deep-rooted factors influence CVD development; these include but are not limited to elevated blood lipids, high blood pressure, obesity and diabetes. A considerable number of proteins are involved directly and indirectly in the transport, maintenance and elimination of plasma lipids, including high and low-density lipoprotein cholesterol (HDL-C and LDL-C). There are several mechanisms involved in the removal of LDL particles from systemic circulation. One such mechanism is associated with the gene that encodes proprotein convertase subtilisin/kexin type 9 (PCSK9), which has become an exciting therapeutic target for the reduction of residual risk of CVDs. Currently, statins are the mainstay treatment to reduce LDL-C, and a need exists to further develop more effective LDL-C-lowering drugs that might supplement statins. This study was aimed at contributing to the generation of knowledge regarding the effect of niacin in reducing LDL levels through PCSK9 interaction.

This thesis studies the problematic of the new polymorphism APOA2 gene and that association with fatty acids contain in a group of Czech Large White pigs. APOA2 gene (ID: 100153243) is a candidate gene for porcine meat quality. The aim of thesis was to analyze the influence of selected polymorphisms on fatty acids and intramuscular fat contain. Among fatty acids was observed a contain of: tetradecenoic acid, palmitic acid, palmitoleic acid, stearic acid, oleic acid, linoleic acid, linolenic acid, arachidonic acid, arachidic acid a eicosapentaenoic acid (EPA). Have been used molecular-genetic methods such as primers design in the OLIGO software, PCR, gel electrophoresis a sequencing by Sanger method. The results were processed by form of genotype frequency and followed by associative analysis with a mixed linear model. The values of the relative alleles frequency of polymorphism APOA2 T>A rs80803879 were as follows: A = 0,086, T = 0,914 and relative alleles frequency of APOA2 G>A rs331415849: A = 0,068 a G = 0,948. Polymorphism associations were found for fatty acids: myristoleic acid, acid palmitoleic acid, oleic acid, arachidonic acid, and arachidic acid.

京都大学 / 0048 / 新制・課程博士 / 博士(医学) / 甲第18145号 / 医博第3865号 / 新制||医||1002(附属図書館) / 31003 / 京都大学大学院医学研究科医学専攻 / (主査)教授 萩原 正敏, 教授 柳田 素子, 教授 稲垣 暢也 / 学位規則第4条第1項該当 / Doctor of Medical Science / Kyoto University / DFAM

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<p> Gene-targeted studies in mice have shown that the murine scavenger receptor class B type I (mSR-BI) is atheroprotective and plays a key role in the clearance of high density lipoprotein (HDL) cholesterol by the liver. We focused on the analysis of human SR-BI (hSR-BI) and the role of its C-terminal cytoplasmic tail on its localization, lipid uptake activity, and regulation in hepatocytes both in vitro and in vivo. Full length hSRBI and hSR-BI lacking its C-terminal cytoplasmic tail (hSR-BI-DM) localized to vesiclelike structures in the cytoplasm, to juxtanuclear regions and to the cell surface in HepG2 cells. Similar cytoplasmic punctate distribution was observed in transfected human and mouse aortic endothelial cells. </p> <p> In HepG2 cells both hSR-BI and hSR-BI-DM mediated HDL-lipid uptake; however, the truncation mutant displayed only half ofthe activity, suggesting that removal ofthe C-terminal cytoplasmic tail reduced but did not eliminate SR-BI's activity. In HepG2 cells treated with the PKC inhibitor, calphostin C, hSR-BI or hSR-BI-DM mediated HDL-lipid uptake was decreased by 40 and 50%, respectively, indicating that this activity is regulated by PKC. </p> <p> In order to determine the effects of hSR-BI and hSR-BI-DM in vivo, we set out to generate transgenic mice with hepatic overexpression ofeach protein using a bipartite expression system requiring driver and responder transgenes. Mice expressing the responder transgenes, PTREhSR-BI and PTREhSR-BI-DM, as well as a reporter transgene (PTRdacZ), driven by the same bi-directional promoter, were generated and mated to mice with a liver-specific driver trans gene, PMuptTA. The mice were analyzed and showed the presence of a reporter protein, ~-galactosidase, in their livers, but not in other tissues tested. Total and HDL cholesterol levels were not altered in PMuPtTA I PrREhSRBI or PMuptTA I PrREhSR-BI-DM transgenic mice. Further characterization ofthe double transgenic mice revealed that hSR-BI m.RNA transcripts were detected in the livers of PMuPtTA I PrREhSR-BI mice, but not in those ofPMuPtTA I PrREhSR-BI-DM mice. However, neither PMuptTA I PrREhSR-BI nor PMuptTA I PrREhSR-BI-DM mice showed increased expression of SR-BI in their livers. </p> / Thesis / Master of Science (MSc)

La apoA-I cumple un rol muy importante en el transporte reverso del colesterol (TRC), es el componente mayoritario de las lipoproteínas de alta densidad (HDL) que desempeñan diversas funciones en las distintas etapas del TRC. Resultados previos de este laboratorio permiten postular la hipótesis de que la región central de la apoA-I, formada por el par de hélices tipo Y, estaría involucrada en la interacción con la membrana celular, que sería importante para el eflujo de lípidos y la movilización de depósitos intracelulares de colesterol (como el disponible a ser esterificado por ACAT) hacia la membrana plasmática. Como la conformación del dominio central es influenciada por el tamaño y composición lipídica (contenido de colesterol) de las HDL, también se postula que esto podría modular la capacidad de interacción con la membrana celular y el consecuente eflujo lipídico. El objetivo general de este trabajo fue someter a prueba esta hipótesis y aportar información relevante para entender los mecanismos implicados en las etapas iniciales del TRC, como en la interacción de las HDL con membranas celulares y el eflujo celular de lípidos. Como objetivos específicos, nos propusimos: 1) Reconstituir partículas discoidales HDL similares a las pre-β-HDL del plasma, de diferente composición y tamaño, mediante la técnica de diálisis con el detergente colato. Estas fueron comparadas en cuanto a su capacidad de unirse a la membrana celular, y de promover el eflujo de colesterol y fosfolípidos de dos líneas celulares diferentes: CHO-K1 (células de ovario de hámster chino) y RAW 264.7 (macrófagos murinos). 2) Estudiar en comparación con apoA-I salvaje, la funcionalidad y las respuestas celulares a dos mutantes de deleción de un residuo de lisina en las regiones de hélices tipo Y: una con la deleción en la región central de la hélice 4 (ΔK107) y la segunda con la deleción en la posición homóloga de la hélice 10 (ΔK226). La primera de estas mutantes es una variante natural cuyos portadores presentan un metabolismo alterado de las HDL e incrementado riesgo aterogénico, por lo que los resultados de estos estudios también podrían ayudar a la comprensión de los síntomas presentados por estos pacientes. Es de esperar que estas mutaciones desplacen en ~100º la orientación relativa entre las caras hidrofílica e hidrofóbica de la hélice anfipática a ambos lados de la mutación, lo que puede afectar tanto la interacción con lípidos como con los receptores celulares.