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Thesis (doctoral)--Lund University, 1991. / Added t.p. with thesis staement inserted.

Thesis (doctoral)--Lund University, 1991. / Added t.p. with thesis staement inserted.

Diss. (sammanfattning) Uppsala : Univ., 2003. / Härtill 5 uppsatser.

Diss. (sammanfattning) Stockholm : Karolinska institutet, 2007. / Härtill 4 uppsatser.

Lau Tung. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2004. / Includes bibliographical references (leaves 142-157). / Abstracts in English and Chinese. / Abstract --- p.i / 摘要 --- p.iii / Acknowledgements --- p.v / Table of Contents --- p.vi / List of Abreviations --- p.x / Chapter Chapter 1 --- Introduction / Chapter 1.1 --- Pancreas and its functions --- p.1 / Chapter 1.1.1 --- Structure of pancreas --- p.1 / Chapter 1.1.2 --- Exocrine function --- p.4 / Chapter 1.1.3 --- Endocrine function --- p.7 / Chapter 1.1.3.1 --- Pancreatic islet and islet cells --- p.7 / Chapter 1.1.3.2 --- Regulation of insulin secretion --- p.10 / Chapter 1.1.3.3 --- Mechanism for glucose-stimulated insulin release --- p.14 / Chapter 1.1.3.4 --- Bi-phase response of insulin secretion --- p.16 / Chapter 1.2 --- Pancreatic Renin-Angiotensin System --- p.19 / Chapter 1.2.1 --- Circulating RAS and local RAS --- p.19 / Chapter 1.2.2 --- RAS inhibitors --- p.25 / Chapter 1.2.2.1 --- Angiotensin converting enzyme inhibitor --- p.25 / Chapter 1.2.2.2 --- Non-specific Ang II receptor blocker --- p.28 / Chapter 1.2.2.3 --- Specific AT1 receptor antagonist --- p.29 / Chapter 1.2.2.4 --- Specific AT2 receptor antagonist --- p.30 / Chapter 1.2.3 --- RAS and Pancreas --- p.30 / Chapter 1.2.3.1 --- Expression and localization of pancreatic RAS --- p.30 / Chapter 1.2.3.2 --- Regulation of pancreatic RAS and its clinical relevance --- p.32 / Chapter 1.3 --- Islet Transplantation and RAS --- p.34 / Chapter 1.3.1 --- Whole pancreas and islet transplantation --- p.34 / Chapter 1.3.2 --- Problems encountered in islet transplantation --- p.36 / Chapter 1.3.3 --- Potential role of RAS in islet transplantation --- p.38 / Chapter 1.4 --- Diabetes Mellitus and RAS --- p.40 / Chapter 1.4.1 --- Diabetes Mellitus --- p.40 / Chapter 1.4.2 --- Type 1 diabetes and its animal model --- p.42 / Chapter 1.4.3 --- Type 2 diabetes and its animal model --- p.44 / Chapter 1.4.4 --- RAS blockade in diabetes patients --- p.46 / Chapter 1.4.5 --- Potential role of RAS in Diabetes Mellitus --- p.47 / Chapter 1.5 --- Aims of Study --- p.49 / Chapter Chapter 2 --- Materials and Methods / Chapter 2.1 --- Experimental animals and mouse models --- p.50 / Chapter 2.1.1 --- Experimental animals for islet isolation and transplantation --- p.50 / Chapter 2.1.2 --- Mouse model for type 2 diabetes --- p.51 / Chapter 2.2 --- Islet isolation and transplantation --- p.52 / Chapter 2.2.1 --- Enzymatic islet isolation --- p.52 / Chapter 2.2.2 --- Islet transplantation --- p.53 / Chapter 2.3 --- Biological assay on islet functions --- p.53 / Chapter 2.3.1 --- Measurement of islet insulin release --- p.53 / Chapter 2.3.2 --- Measurement of islet glucose oxidation rate --- p.56 / Chapter 2.3.3 --- Measurement of islet (pro)insulin biosynthesis --- p.59 / Chapter 2.3.4 --- Measurement of islet total protein synthesis --- p.60 / Chapter 2.4 --- Chronic losartan treatment --- p.62 / Chapter 2.5 --- Perfusion experiment of transplanted islet graft --- p.62 / Chapter 2.6 --- Insulin content of the islet graft --- p.63 / Chapter 2.7 --- Islet graft (pro)insulin and total protein biosynthesis --- p.64 / Chapter 2.8 --- Real-time RT-PCR Analysis --- p.64 / Chapter 2.8.1 --- Design of primers and probes --- p.67 / Chapter 2.8.2 --- Use of internal control --- p.69 / Chapter 2.8.3 --- RT-PCR reaction --- p.69 / Chapter 2.8.4 --- Calculation using the comparative CT method --- p.70 / Chapter 2.9 --- Western Blot Analysis --- p.71 / Chapter 2.10 --- Immunocytochemistry --- p.72 / Chapter 2.11 --- Statistical data analysis --- p.73 / Chapter Chapter 3 --- Results / Chapter 3 .1 --- Effect of Angiotensin II and Losartan on islet insulin release --- p.74 / Chapter 3.1.1 --- Insulin release from normal islets --- p.74 / Chapter 3.2 --- "Effect of Angiotensin II and Losartan on islet glucose oxidation rate, (pro)insulin and total protein biosynthesis" --- p.77 / Chapter 3.2.1 --- Glucose oxidation rate of isolated normal islets --- p.77 / Chapter 3.2.2 --- (pro)insulin and total protein biosynthesis of isolated normal islets --- p.77 / Chapter 3.3 --- Regulation of RAS components in islet transplantation --- p.81 / Chapter 3.3.1 --- Expression of RAS components in endogenous islets and transplanted islets --- p.81 / Chapter 3.3.2 --- Localization of AT1-receptor in endogenous islets --- p.87 / Chapter 3.3.3 --- Expression of AT1-receptor protein in endogenous and transplanted islets --- p.89 / Chapter 3.3.4 --- Relative abundance of RAS components in kidney and liver --- p.91 / Chapter 3.3.5 --- Insulin release from perfused transplanted islet graft --- p.93 / Chapter 3.3.5 --- (pro)insulin and total protein biosynthesis of transplanted islet graft --- p.96 / Chapter 3.4 --- Effect of Angiotensin II and losartan on diabetic islets --- p.99 / Chapter 3.4.1 --- Expression of RAS components in diabetic pancreas --- p.99 / Chapter 3.4.2 --- Localization of AT1 receptors in diabetic pancreas --- p.105 / Chapter 3.4.3 --- Insulin release from islets of type 2 diabetic mice --- p.107 / Chapter 3.4.4 --- (pro)insulin and total protein biosynthesis of islets from type 2 diabetic mice --- p.112 / Chapter Chapter 4 --- Discussion / Chapter 4.1 --- Effect of angiotensin II and losartan on islet insulin release --- p.116 / Chapter 4.2 --- Existence of local RAS in pancreatic islets --- p.119 / Chapter 4.3 --- Regulation of islet RAS components in transplanted islets --- p.122 / Chapter 4.4 --- Clinical relevance of islet RAS in transplantation --- p.125 / Chapter 4.5 --- Regulation of islet RAS by type 2 diabetes --- p.126 / Chapter 4.6 --- Clinical relevance of islet RAS in type 2 diabetes --- p.134 / Chapter 4.7 --- Conclusion --- p.140 / Chapter 4.8 --- Further studies --- p.141 / Chapter Chapter 5 --- Bibliography --- p.142

Diss. (sammanfattning) Stockholm : Karol. inst., 2005. / Härtill 3 uppsatser.

Diss. (sammanfattning) Stockholm : Karolinska institutet, 2007. / Härtill 4 uppsatser.

Thesis (Ph.D. in Immunology) -- University of Colorado, 2005. / Typescript. Includes bibliographical references (leaves 133-151). Free to UCDHSC affiliates. Online version available via ProQuest Digital Dissertations;

O transplante (Tx) de ilhotas pancreáticas (IP) é uma atraente alternativa para o tratamento do diabetes melito tipo 1. No entanto, para evitar a rejeição há necessidade de imunossupressão. Uma nova idéia de tolerância surge a partir do paradoxo imunológico, onde a mãe, imunologicamente competente, não rejeita o embrião durante a gravidez. Uma das hipóteses é que células da placenta expressam a molécula IDO, a qual protege o embrião do ataque imunológico materno. O objetivo do estudo foi analisar o efeito da indução da expressão da IDO em IP em transplante experimental de IP. Para tanto, as seguintes etapas de padronização foram necessárias. Etapa 1: Padronização da perfusão e digestão do tecido pancreático de rato e determinação do método para a purificação das IP, comparando-se diferentes gradientes de densidade: descontínuo de Ficoll, contínuo de Ficoll e contínuo de iodixanol. Foi demonstrado que o gradiente contínuo de iodixanol fornece maior pureza e maior número de IP íntegras e funcionais. Etapa 2: Padronização do Tx experimental de IP sob a cápsula renal para avaliação do número mínimo de IP transplantadas para reverter o diabetes induzido por estreptozotocina, definido como glicemia >300mg/Kg. Foram transplantadas entre 200 a 3.000 IP por experimento. A rejeição das IP foi analisada pela sobrevida das IP (permanência da glicemia <300mg/dL), tanto em Tx isogênico (Lewis-Lewis) como em alogênico (Sprague-Dawley-Lewis). Para reverter o diabetes foram necessárias no mínimo 2.500 IP. No transplante entre ratos isogênicos (n=6) não houve rejeição das IP. Já no transplante entre animais alogênicos (n=12), as IP apresentaram uma curta sobrevida pós-Tx (11±1 dias; p<0,01 vs. Tx isogênico). Dez dias pós-Tx, houve um grande infiltrado de macrófagos e linfócitos T no enxerto alogênico e uma diminuição significativa da expressão de insulina (p<0,001 vs. Tx isogênico). Etapa 3: Construção do vetor de expressão para IDO. A partir de RNA extraído de placenta de rata no 10º dia de gestação, foi amplificada a seqüência completa do cDNA para IDO, utilizando-se RT-PCR. Em seguida, o cDNA para IDO foi inserido em vetor de expressão (vetor-IDO). Etapa 4: Transfecção do vetor-IDO nas IP. O vetor-IDO foi introduzido nas IP através de lipofecção (Lipofectamina 2000), testando-se diferentes concentrações do vetor-IDO (0, 0,5, 1 e 10 ng/uL) e diferentes períodos de incubação (1h, 15h e 24h). A expressão de IDO nas IP foi confirmada por RT-PCR e imuno-histoquímica. A incubação com 10 ng/uL de vetor-IDO durante 24h foi eficaz para induzir a expressão de IDO nas IP, confirmada a nível de RNAm (RT-PCR) e de proteína (imuno-histoquímica). A eficiência da transfecção em nível funcional foi confirmada pela degradação de triptofano em cultura (dosagem de triptofano por HPLC). Etapa 5: Onze transplantes alogênicos (Sprague-Dawley-Lewis) com IP transfectadas com vetor-IDO foram realizados para analisar o efeito da IDO. Três animais foram sacrificados para análise de imuno-histoquímica e 8 animais foram acompanhados por 45 dias. A sobrevida das IP transfectadas com vetor-IDO foi significativamente maior comparada com a sobrevida de IP não-transfectadas (p<0,01). O estudo conclui que a expressão da IDO protege as IP aumentando a sobrevida das IP. / Transplantation (Tx) of pancreatic islets (PI) is an attractive alternative of treatment for type 1 diabetes mellitus. However, continuous immunossupression is necessary in order to avoid allograft rejection. A new idea of tolerance is based on the immunological paradox, during pregnancy, in that the mother, immunologically competent, does not reject the semi-allogeneic fetus. The hypothesis is that the placenta produces IDO molecules, which protect the embryos against the maternal immunologic attack. The aim of this study was to analyze the effect of the induction of the IDO expression into PI in an experimental model of PI transplantation. The following steps for standardization were necessary. Step 1: Besides the standardization of the rat pancreas perfusion and digestion, the best method for purification of the PI was determined, comparing several density gradients: Ficoll discontinuous, Ficoll continuous and iodixanol continuous. The iodixanol continuous gradient was able to provide high purity and a high number of intact and functional PI. Step 2: The transplantation of the PI between rats was established determining the minimal number of PI to reverse the diabetes (glycemia > 300mg/dL) induced by streptozotocin. In addition, the rejection was analyzed by PI survival (time with glycemia <300mg/dL) in syngeneic (Lewis-Lewis) and allogeneic (Sprague-Dawley-Lewis) transplantation. To reverse the diabetes at least 2,500 PI were necessary. Transplantation between syngenic rats (n=6) disclosed no rejection of the PI. In the allogeneic transplantation (n=12), the PI had a short survival (11±1 days). Ten days post-Tx, a higher number of macrophages and T lymphocytes were observed in the grafts, accompanied by very low insulin expression. Step 3: The expression vector for IDO was constructed from RNA extracted from rat placenta. RT-PCR was carried out to amplify the IDO cDNA, which was inserted into expression vector (IDO vector). Step 4: The IDO vector was introduced into PI through lipofection (Lipofectamine 2000) analyzing several concentrations of the IDO vector (0, 0.5, 1.0 and 10 ng/uL) and several periods of incubation (1h, 15h e 24h). The IDO expression in PI was confirmed by RT-PCR and immunohistochemistry. The incubation with 10 ng/uL of IDO vector during 24h was efficient to induce IDO expression in PI. The function of the IDO was confirmed by tryptofan degradation in culture (measurement of tryptofan by HPLC). Step 5: Eleven allogenic transplants (Sprague-Dawley to Lewis) of PI expressing IDO were performed to analyze the effect of the IDO in the rejection. Eight animals were accompanied for 45 days, whereas three were sacrificed after 10 days for immunohistochemistry analysis. Finally, the survival of the PI expressing IDO was significantly higher than nontransfected PI. The study concludes that the induction of the IDO into PI protects the PI increasing the PI survival.