Trabalhamos com 8 jabutis, 4 fêmeas e 4 machos da espécie Geochelone carbonaria, para descrição da morfologia macroscópica e microscópica de luz e eletrônica de varredura do rim. Observamos que os rins são órgãos compactos, de forma piramidal, coloração vermelha escuro, e de aspecto cerebróide. Histologicamente o néfron apresenta glomérulo de Malpighi, túbulo contornado proximal, túbulo contornado distal, alça de Henle, túbulo coletor e ureter. Observamos também que todas estas estruturas secretam grânulos de muco neutro. / Eight tortoises have been used in this study i.e., 4 females and 4 males belonging to the Geochelone carbonaria species, in order to describe the macroscopic morphology, light microscopic morphology and the scanning electron microscopic morphology of the kidney. It has been observed that the kidney are compact, pyramid-shaped organs with a dark reddish coloring and circunvolutioned appearance. From a histological point of view, the nephron presents Malpighian corpuscle, proximal contoured tubule, loop of Henle, collector tubule and ureter. It has been also noticed that every renal structure secrete neutral-mucus granules.

Foram utilizados 10 eqüinos clinicamente sadios, machos, adultos, de diferentes raças, pesando entre 350 e 450 Kg, escolhidos aleatoriamente e divididos em dois grupos: em cinco animais foi realizada biópsia renal guiada por laparoscopia com agulha descartável do tipo tru-cut, e nos outros cinco através de pinça de biópsia laparoscópica de 5mm. Os procedimentos foram efetuados com os animais em posição quadrupedal, mediante sedação e analgesia, e bloqueio anestésico local em 2 pontos no flanco esquerdo, sendo um para introdução do trocarte e óptica, e outro para introdução da agulha ou pinça. Parâmetros clínicos e laboratoriais foram avaliados dois dias antes do procedimento e acompanhados nas três semanas subseqüentes, permitindo determinar as alterações decorrentes. As técnicas foram comparadas quanto à segurança, qualidade, tempo de execução e significância da amostra colhida. Durante o acompanhamento pós-operatório dos animais, não houve variação clínica e laboratorial significativa em ambos os grupos, porém, a análise histológica dos fragmentos demonstrou uma maior porcentagem de biópsias adequadas no grupo em que foi utilizada pinça laparoscópica. / Ten healthy adult male horses, of different breeds, weighing between 350 and 450kg were randomly chosen and divided into two groups of five animais each: in one group, laparoscopic guided renal biopsy was pertormed using tru-cut needles and in the other group using 5mm forceps. Procedures were carried out with animais in quadrupedal position, under sedation and analgesia and local anesthetic nerve block was made in two points on the left flank: one for the introduction of the trocar and optical system and one for the introduction of the needle or forceps. Laboratory and clinical parameters were evaluated two days prior to the proceeding and followed up in the subsequent three weeks in order to verify the resulting alterations. The techniques were compared as regards safety, quality, time for execution and significance of the sample obtained. During the post-surgery follow-up period, no significant clinical and laboratory alterations were observed in either group, although the histological analysis of the fragments a higher percentage of adequate biopsies in the group where Laparoscopic forceps were used.

Foram realizadas mensurações dos rins direito e esquerdo, bem como, de carcaças de bovinos Nelore, inteiros e castrados. As médias gerais dos grupos e os respectivos desvio padrão para as mensurações renais foram: comprimento do rim direito 16,85 &plusmn; 1,10 cm; comprimento do rim esquerdo 16,20 &plusmn; 1,15 cm. As larguras dos rins direito e esquerdo apresentaram médias e desvio padrão iguais a 9,20 &plusmn; 0,80 cm e 9,35 &plusmn; 0,90 cm, respectivamente. Para a variável peso dos rins direito e esquerdo, as médias e desvio padrão foram, respectivamente, de 347,90 Kg &plusmn; 32,55 Kg e 343,25 Kg &plusmn; 32,20 Kg. O número de lobos observados nos rins direito e esquerdo obtiveram médias e desvio padrão de 16,30 &plusmn; 4,50 e 15,90 &plusmn; 3,90, respectivamente. Para as variáveis de carcaça dos grupos envolvidos as médias e respectivos desvios padrão foram: para peso das hemicarcaças direita e esquerda 137,00 &plusmn; 8,00 e 141,10 &plusmn; 8,00 Kg; peso total da carcaça 278,10 &plusmn; 15,90 Kg; rendimento de carcaça 55,80 &plusmn; 1,30%; espessura de gordura 6,30 &plusmn; 2,6 cm; comprimento de carcaça 1,28 &plusmn; 0,03 m. Para comparação das diferentes mensurações, os animais foram divididos em dois grupos (inteiros e castrados). Diferenças significativas (P < 0,01) foram observadas para quase todas as variáveis de carcaça exceto para comprimento da mesma. Para as variáveis de rim, resultados significativos foram observados para largura de rim direito (P < 0,01), peso de rim direito, peso de rim esquerdo e peso total dos rins (P < 0,05). As estimativas de correlações entre todas as características foram de baixa magnitude, indicando ausência de associação entre as diferentes medidas. / Measurement of the right and left kidneys were accomplished, non castrated and castrated carcasses of bovine Nelore. The averages managed of the groups and the respective deviation pattern for the renal measurement were: length of the right kidney 16.85 ± 1.10 cm, respectively; length of the left kidney 16.20 ± 1.15 cm. The widths of the kidneys right and left presented averages and standard deviation to 9.20 ± 0.80 cm and 9.35 ± 0.90 cm, respectively. For the variable weight of the kidneys right and left, the averages and standard deviation were, respectively, of 347.90 Kg ± 32.55 Kg and 343.25 Kg ± 32.20 Kg. The number of lobes observed in the kidneys right and left obtained averages and standard deviation of 16.30 ± 4.50 and 15.90 ± 3.90, respectively. The averages and respective deviation pattern of variables of the involved groups for carcass were: for weight of right and left hemicarcass 137.00 ± 8.00 and 141.10 ± 8.00 Kg total weight of the carcass 278.10 ± 15.90 Kg; carcass exploitation 55.80 ± 1.30%; fat thickness 6.30 ± 2.6 cm, carcass length 1.28 ± 0.03 m. For comparison of the different measurement, the animals were divided in two groups (non castrated and castrated). Significant differences (P <0,01) were observed for almost all the carcass variables except for length. For kidney variables, significant results were observed for width of right kidney (P <0,01), weight of right kidney, weight of left kidney and total weight of kidneys (P <0,05). The estimates of correlations among all the characteristics were of low magnitude, indicating association absence among the different measures.

Nos mamíferos, o fígado e o rim são órgãos fundamentais para uma adequada homeostase. Nos cetáceos, são de especial importância frente aos desafios da vida no ambiente marinho. O presente estudo teve como objetivo investigar as principais lesões hepáticas e renais de cetáceos do Brasil, utilizando-se amostras mantidas junto ao Banco de Tecidos de Mamíferos Marinhos (BTMM), Laboratório de Patologia Comparada de Animais Selvagens. Para a caracterização das lesões foram utilizadas técnicas anatomopatológicas, imuno-histoquímicas e ultraestrutural. Foram estudados 197 cetáceos de 18 espécies, encontrados mortos em decorrência de captura incidental em apetrecho de pesca, encalhe ou após tentativas de reabilitação. A principal espécie amostrada foi toninha (Pontoporia blainvillei) com 65,9% (130/197) dos casos. Quanto à distribuição geográfica as amostras provieram principalmente do estado de São Paulo (41,6%, 82/197), seguido do Rio Grande do Sul (36,5%, 72/192) e Ceará (11,7%, 23/197). Entre as principais lesões hepáticas diagnosticadas, as inclusões hialinas citoplasmáticas (IHC) apresentaram maior frequência (46,3%, 88/190), seguidas pelas hepatites portais linfoplasmocíticas crônicas observadas em 36,5% (69/190), esteatose, em 14,2% (27/190), hepatite necrótica, em 4,7% (9/190), e colangiohepatite parasitária, em 2,6% (5/190) dos casos. A ocorrência de IHC foi mais frequente em animais capturados do que encalhados. Entre as principais lesões renais diagnosticadas, a glomerulonefrite membranosa apresentou maior frequência (14,5%, 28/192). Foram observadas também glomerulonefrine membranoproliferativa, em 10,4% (20/192), nefrite intersticial, em 10,9% (21/192), cistos simples, em 4,16% (8/192), doença glomerulocística primária, em 4,6% (9/192), doença glomerulocística secundária (DGCS), em 8,3% (16/192), e doença renal policística e adenoma tubular, com 0,5% (1/192) de ocorrência cada. A incidência de DGCS apresentou diferença entre as espécies, sendo menos frequente em toninhas do que nos demais cetáceos. Um boto-cinza (Sotalia guianensis) morto em decorrência de captura incidental na baia de Paranaguá, Paraná, foi diagnosticado com toxoplasmose e devido à sua importância, fragmentos de todos seus órgãos, disponíveis no BTMM, foram avaliados. O presente estudo reflete a relevância em manter o BTMM, o qual consiste em uma fonte de informação ímpar, que possibilita a realização de estudos retroativos em tecidos de cetáceos e outras espécies de mamíferos aquáticos. O presente trabalho traz contribuições sobre as enfermidades em cetáceos, e aborda de maneira sistemática as lesões hepáticas e renais nestas espécies. Futuros estudos são necessários para elucidar aspectos sobre o impacto das lesões renais e hepáticas e sua relação com as condições mórbidas dos cetáceos, bem como para avaliar o impacto da toxoplasmose, nos cetáceos e outros mamíferos marinhos brasileiros. / In mammals, the main organs for an adequate homeostase are the liver and the kidney. These organs in Cetaceans have especial importance because of the challenges of life in a marine environment. This study had as main objective find the principal hepatic and renal lesions in Cetaceans from Brazil. Samples from the Marine Mammal Tissue Bank (BTMM) of the Laboratory of Comparative Pathology of Wild Animals were used. Anatomopathological, immunohistochemical and ultrastructural studies were performed. A total of 197 cetaceans belonging to 18 species were studied. They were found dead because of incidental capture or after attempts of rehabilitation for the stranded ones. Franciscana (Pontoporia blainvillei) was the principal specie sampled with a 65,9% (130/197) of the cases. Related to geographic distribution, samples were more frequent in São Paulo state (41,6%, 82/197), then Rio Grande do Sul (36,5%, 72/192) and Ceará (11,7%, 23/197). The hepatic lesions found include: hyaline cytoplasmatic inclusions (IHC) (46,3%, 88/190), lymphoplasmacytic chronic portal hepatitis (36,5%, 69/190), steatosis (14,2%, 27/190), necrotic hepatitis (4,7%, 9/190) and parasitic colangiohepatitis (2,6% , 5/190). The occurrences of IHC were more frequent in captured animals than stranded. The main kidney lesion found was the membranous glomerulonephritis (14,5%, 28/192). Additionally, there were observed membranoproliferative glomerulonephritis (10,4%, 20/192), intersticial nephritis (10,9%, 21/192), simple cysts 4,16% (8/192), glomerulocystic primary disease (4,6%, 9/192), glomerulocystic secondary disease (DGCS) (8,3% ,16/192) and polycystic kidney disease and tubular adenome (0,5%, 1/192). The incidence of DGCS differ among species, in Fransiscanas it was less frequent than in other cetaceans. A Guiana Dolphin (Sotalia guianensis) dead by incidental capture in the bay of Paranaguá, Paraná, was diagnosed with toxoplasmosis and because of its importance, fragments of all its organs available on BTMM, were evaluated. This study reflects the relevance to maintain the BTMM as an important primary source of information, enabling the realization of future reprospective studies in tissues of whales and other species of aquatic mammals. Furthermore, this study presents contributions on cetacean diseases and addresses in a systematic way lesions in the liver and kidney in these species. Future studies are necessary to elucidate aspects of the impact of renal and hepatic lesions and their relation to the morbid conditions of cetaceans, as well as to evaluate the impact of toxoplasmosis in cetaceans and other marine mammals in Brazil.

A isquemia e reperfusão (IR) é a principal causa de insuficiência renal aguda. Evidências mostram a participação de linfócitos T e células dendríticas (DC) na lesão por IR. Entretanto, os mecanismos envolvidos não estão claros. A enzima heme oxigenase (HO)-1 está relacionada à diminuição de respostas inflamatórias. Neste trabalho, investigamos a participação de linfócitos T CD4+ e DC na proteção da lesão por IR induzida pela HO-1. Injetamos em camundongos um indutor de HO-1 (Hemin), e realizamos a IR. Avaliamos a lesão pela uréia e creatinina no soro, a expressão de citocinas por RT-PCR, nível de proteínas por bioplex e perfil celular por FACS. Observamos que a HO-1 protegeu da lesão por IR. A diminuição de INFg com a HO-1 sugeriu uma menor ativação de linfócitos T. No entanto, a transferência de células CD4+ tratadas com Hemin não apresentou diferença. Vimos que a HO-1 alterou o fenótipo das DC após IR e suprimiu a produção de TNFa pelas mesmas. Concluímos que a proteção pela HO-1 é capaz de modular a resposta inflamatória de DC, diminuindo o TNFa. / The ischemia and reperfusion (IR) is the main acute kidney injury. Evidences have shown the role of CD4+ T cells and dendritic cells (DC) in the IR injury. However, the mechanisms involved in the participation of these cells are not clear. The heme oxygenase (HO)-1 enzyme is associated to decrease in inflammatory responses. In this work, we investigated the role of CD4+ T cells and DC in renal injury protection induced by HO-1. We injected in mice a HO-1 inducer (Hemin), and we did the IR. Renal injury was evaluated by serum levels of urea and creatinine, cytokines expression by RT-PCR, proteins level by bioplex and cell profile by FACS. We observed that HO-1 lead to IR injury protection. The IFNg decrease with HO-1 suggested less T cells activation. However, transfer of hemin treated CD4+ cells did not differ from the other group. We observed that HO-1 altered DC phenotype after IR and suppressed TNFa production by these cells. We concluded that the protection by HO-1is able to modulate DC inflammatory response, diminishing TNFa.

A leptospirose é uma zoonose de importância mundial, causada por leptospiras patogênicas. Aproximadamente 5 a 10% das infecções humanas cursam com a forma grave. A Doença de Weil é a forma mais comum de doença grave e pode apresentarse com duas formas de evolução, aguda progressiva monofásica ou de curso prolongado. A doença grave se caracteriza por uma combinação de hemorragia, mais comumente pulmonar, icterícia e insuficiência renal, com letalidade de 5 a 15%. O rim é um órgão muito acometido na Leptospirose. Clinicamente o envolvimento renal ocorre de 16 a 40%, com manifestações peculiares como poliúria, hipocalemia, e perda de sódio. A disfunção tubular renal, é característica da leptospirose forma grave, com envolvimento dos transportadores renais de sódio ao longo do néfron, levando às manifestações observadas. A terapêutica antimicrobiana é recomendada na Leptospirose, porém com controvérsias à sua indicação após o quarto dia de doença. Quando da instalação da lesão não haveria benefícios com a utilização de antibióticos. O tratamento pode diminuir a morbidade e letalidade, assim como interferir no envolvimento renal e na expressão dos transportadores renais de sódio. A patogênese pode estar relacionada a efeitos diretos da leptospira ou a resposta inflamatória assim como o estresse oxidativo. A utilização de antioxidantes, pode ser considerada como terapia adjuvante. Nós avaliamos a expressão no túbulo proximal do trocador Na+-H+ (NHE3) e na porção espessa da medula ascendente o cotransportador Na+-K+-2Cl- (NKCC2), no modelo de hamster com as duas formas de evolução de doença grave, mimetizando a doença de humanos realizados em dois experimentos. Os experimentos envolveram animais infectados não tratados e tratados com ampicilina associado ou não ao antioxidante, N-acetilcisteína. A presença de antígenos de Leptospira e a expressão dos transportadores foram avaliadas por imunohistoquímica, e o ácido tiobarbitúrico, marcador de estresse oxidativo, (TBARS) foi quantificado.Hamsters infectados, apresentaram altas quantidades de antígenos nos tecidos-alvo, enquanto que a expressão de ambos os transportadores apresentou-se diminuída. O tratamento com ampicilina esteve associado com mínima detecção ou ausência de antígenos, restabelecimento da expressão dos transportadores nos respectivos locais e redução dos níveis de TBARS. O tratamento precoce e tardio com ampicilina restabeleceu os defeitos tubulares na leptospirose forma grave em ambos experimentos, sem benefícios com a utilização da N-acetilcisteína. / Leptospirosis is a zoonosis of worldwide distribution. About 5-10% of all human infections presents with severe forms. Weil\'s syndrome, the most common presentation of severe forms of leptospirosis, may courses either as a single monophasic disease or as a disease with prolonged course, characterized by a combination of hemorrhage, particularly in the lung, renal failure, and jaundice, with fatality rates ranging from 5 to 15%. The kidney is an important target organ in leptospiral infection. Clinically, renal involvement in leptospirosis occurs in 16% to 40% of cases and is unique because of the atypical presentation of polyuria, hypokalemia, and sodium wasting, suggestive of a special form of tubular dysfunction related to the major renal sodium transporters expressed along the nephron. A wide range of antimicrobial therapy for leptospirosis was described and benefits have been disputed for cases with more than four days of clinical disease, because after a threshold of leptospiremia, the delayed use of antibiotics is unlikely to reduce fatality. Antimicrobial therapy is thought to interfere on fatality, renal involvement, and renal sodium transporters expression during severe disease. The pathogenesis may be related to direct effects of leptospiral compounds or inflammatory response due to oxidative stress. Antioxidant could be considered for adjunctive therapy.We evaluated the expression of proximal tubule type 3 Na+/H+ exchanger (NHE3) and thick ascending limb Na+-K+-2Cl- cotransporter (NKCC2) in infected non treated and treated hamsters reproducing the two forms of clinical human presentations of Weil\'s syndrome divided in two experiments. Animals were treated or not with ampicillin and/or N-acetyl-cysteine (NAC). Leptospiral antigen/s and expression of renal transporters were evaluated by immunohistochemistry, and serum thiobarbituric acid (TBARS) was quantified. Infected hamsters had high amounts of detectable leptospiral antigen/s in target tissues while renal expression of NHE3 and NKCC2 decreased. Ampicillin treatment was associated with minimal or no detection of leptospiral antigens, normal expression of NHE3 and NKCC2 transporters, and reduced levels of TBARS. Early and late ampicillin treatment rescued tubular defects in leptospirosis severe disease in both experiments, and there was no evidence of benefit from antioxidant therapy.

by Leung Cheuk Wa. / Thesis (M.Sc.)--Chinese University of Hong Kong, 1994. / Includes bibliographical references (leaves 97-106). / ACKNOWLEDGEMENTS --- p.i / LIST OF FIGURES --- p.v / LIST OF TABLES --- p.vi / SUMMARY --- p.1 / Chapter 1. --- INTRODUCTION --- p.3 / Chapter 2. --- BASIC ASPECTS OF CARNITINE / Chapter 2.1 --- BIOSYNTHESIS OF CARNITINE --- p.5 / Chapter 2.2 --- CARNITINE TRANSPORT --- p.8 / Chapter 2.3 --- THE ROLE OF CARNITINE IN INTRACELLULAR METABOLISM --- p.10 / Chapter 2.4 --- THE ROLE OF KIDNEY IN CARNITINE METABOLISM --- p.16 / Chapter 3. --- CARNITINE DEFICIENCY --- p.18 / Chapter 3.1 --- PRIMARY CARNITINE DEFICIENCY --- p.20 / Chapter 3.1.1 --- MYOPATHIC CARNITINE DEFICIENCY --- p.21 / Chapter 3.1.2 --- SYSTEMIC CARNITINE DEFICIENCY --- p.21 / Chapter 3.2 --- SECONDARY CARNITINE DEFICIENCY --- p.22 / Chapter 4. --- CARNITINE METABOLISM IN SELECTED DISEASES / Chapter 4.1 --- CARDIOMYOPATHY --- p.23 / Chapter 4.2 --- ORGANIC ACIDURIAS --- p.24 / Chapter 4.3 --- VALPROIC ACID THERAPY --- p.26 / Chapter 4.4 --- RENAL DIALYSIS ANDTRANSPLANTATION --- p.28 / Chapter 5. --- ANALYTICAL METHODS FOR CARNITINE ASSAYS --- p.30 / Chapter 6. --- DETERMINATION OF TOTAL AND FREE CARNITINE / Chapter 6.1 --- PRINCIPLE OF THE ASSAYS --- p.32 / Chapter 6.1.1 --- FREE CARNITINE DETERMINATION --- p.32 / Chapter 6.1.2 --- TOTAL CARNITINE DETERMINATION --- p.33 / Chapter 6.2 --- INSTRUMENTATION --- p.34 / Chapter 6.3 --- PREPARATION OF REAGENTS AND STANDARDS --- p.36 / Chapter 6.4 --- SPECIMEN COLLECTION --- p.42 / Chapter 6.5 --- SAMPLE PREPARATION --- p.43 / Chapter 6.6 --- ASSAY PROTOCOL FOR FREE CARNITINE --- p.44 / Chapter 6.7 --- ASSAY PROTOCOL FOR TOTAL CARNITINE --- p.46 / Chapter 6.8 --- FACTORS AFFECTING THE PERFORMANCE OF ASSAYS --- p.48 / Chapter 6.9 --- EVALUATION OF FREE AND TOTAL CARNITINE ASSAYS --- p.50 / Chapter 7. --- RESULTS OF EVALUATION OF TOTAL AND FREE CARNITINE ASSAYS / Chapter 7.1 --- CALIBRATION --- p.52 / Chapter 7.2 --- PRECISION --- p.55 / Chapter 7.3 --- LINEARITY RANGE --- p.56 / Chapter 7.4 --- RECOVERY --- p.58 / Chapter 7.5 --- INTERFERENCE OF ACETYLCARNITINE ON FREE CARNITINE ASSAY --- p.59 / Chapter 7.6 --- DISCUSSION --- p.59 / Chapter 8. --- STUDY IN NORMAL SUBJECTS / Chapter 8.1 --- SUBJECTS --- p.61 / Chapter 8.2 --- RESULTS OF THE NORMAL SUBJECTS --- p.61 / Chapter 8.3 --- DISCUSSION --- p.63 / Chapter 9. --- PATIENTS STUDY / Chapter 9.1 --- PATIENTS WITH CARDIOMYOPATHY / Chapter 9.1.1 --- SUBJECTS --- p.66 / Chapter 9.1.2 --- RESULTS OF THE STUDY --- p.66 / Chapter 9.1.3 --- DISCUSSION --- p.69 / Chapter 9.2 --- PATIENTS WITH METABOLIC DISEASES / Chapter 9.2.1 --- SUBJECTS --- p.71 / Chapter 9.2.2 --- RESULTS OF THE STUDY --- p.71 / Chapter 9.2.3 --- DISCUSSION --- p.74 / Chapter 9.3 --- PATIENTS ON VALPROIC ACID THERAPY / Chapter 9.3.1 --- SUBJECTS --- p.75 / Chapter 9.3.2 --- RESULTS OF THE STUDY --- p.75 / Chapter 9.3.3 --- DISCUSSION --- p.77 / Chapter 9.4 --- PATIENTS ON RENAL DIALYSIS AND AFTER TRANSPLANTATION / Chapter 9.4.1 --- SUBJECTS --- p.79 / Chapter 9.4.2 --- RESULTS OF THE STUDY --- p.79 / Chapter 9.4.3 --- DISCUSSION --- p.81 / Chapter 10. --- GENERAL DISCUSSION --- p.84 / Chapter 11. --- REFERENCES --- p.97

Tsang Kam Sze, Kent. / Thesis (M.Phil.)--Chinese University of Hong Kong, 1992. / Includes bibliographical references (leaves 155-174). / Abstract --- p.i / Acknowledgements --- p.v / List of Abbreviations --- p.viii / Table of Contents --- p.x / List of Figures --- p.xvi / List of Tables --- p.ixx / Chapter Chapter 1. --- Introduction --- p.1 / Chapter 1.1. --- Idiotype Network --- p.2 / Chapter 1.2. --- Anti-idiotype Classification --- p.8 / Chapter 1.3. --- Blood Transfusion Effect --- p.11 / Chapter 1.4. --- Transfusion Protocol --- p.12 / Chapter 1.5. --- Mechanism of Beneficial Transfusion Effect --- p.15 / Chapter 1.5.1. --- Donor Selection --- p.15 / Chapter 1.5.2. --- Clonal Deletion --- p.16 / Chapter 1.5.3. --- Suppressor Cells Induction --- p.18 / Chapter 1.5.4. --- Prostaglandins Mediation --- p.19 / Chapter 1.5.5. --- Mixed Chimerism Motivation --- p.20 / Chapter 1.5.6. --- Fc-receptor Blocking Antibodies Stimulation --- p.22 / Chapter 1.5.7. --- Anti-idiotypic Antibodies Instigation --- p.23 / Chapter 1.6. --- Study Aims --- p.25 / Chapter 1.7. --- Technical Strategy --- p.26 / Chapter Chapter 2. --- Materials and Methods --- p.30 / Chapter 2.1. --- Materials --- p.31 / Chapter 2.1.1. --- Patient Population --- p.31 / Chapter 2.1.2. --- Normal Control Group --- p.31 / Chapter 2.1.3. --- Serum Samples --- p.32 / Chapter 2.1.4. --- Additional Specimens --- p.32 / Chapter 2.1.5. --- Chemicals --- p.32 / Chapter 2.1.6. --- Antisera --- p.34 / Chapter 2.1.7. --- Buffers --- p.35 / Chapter 2.1.8. --- Consumables --- p.38 / Chapter 2.1.9. --- Apparatus and Equipment --- p.39 / Chapter 2.2. --- Methods --- p.40 / Chapter 2.2.1. --- Purification of Human Polyclonal Anti-HLA Antisera --- p.40 / Chapter 2.2.1.1. --- Affinity Chromatography --- p.41 / Chapter 2.2.1.2. --- Dialysis --- p.41 / Chapter 2.2.1.3. --- Concentration --- p.42 / Chapter 2.2.1.4. --- Quantitation --- p.42 / Chapter 2.2.2. --- Generation of F(ab')2 fragments from the Purified Human Anti-HLA Antibodies --- p.42 / Chapter 2.2.2.1. --- Buffer Exchange --- p.43 / Chapter 2.2.2.2. --- Pepsin Digestion --- p.43 / Chapter 2.2.2.3. --- Purification of (ab')2、 --- p.43 / Chapter 2.2.3. --- Enzyme-Linked Immunosorbent Assay for anti-Idiotypes against anti-HLA antibodies --- p.44 / Chapter 2.2.3.1. --- Optimization --- p.44 / Chapter 2.2.3.2. --- Quality Control --- p.45 / Chapter 2.2.3.2.1. --- F(ab')2 Specificity --- p.45 / Chapter 2.2.3.2.2. --- Fc Contamination --- p.46 / Chapter 2.2.3.2.3. --- Precision Test --- p.47 / Chapter 2.2.4. --- Anti-Casein Interference --- p.47 / Chapter 2.2.5. --- Test Protocol --- p.48 / Chapter 2.3. --- Statistical Analysis --- p.48 / Chapter Chapter 3. --- Purification of Anti-HLA IgG and F(ab')2 --- p.50 / Chapter 3.1. --- Immunoglobulin Concentration --- p.51 / Chapter 3.2. --- F(ab')2 Specificity --- p.51 / Chapter 3.3. --- Fc-fragments Contamination --- p.53 / Chapter 3.4. --- Discussion --- p.56 / Chapter Chapter 4. --- ELISA Optimization --- p.57 / Chapter 4.1. --- Coating F(ab')2 Quantitation --- p.58 / Chapter 4.2. --- Blocking and Diluting Agent Concentration --- p.61 / Chapter 4.3. --- Serum Analyte Dilution --- p.61 / Chapter 4.4. --- Conjugated Detector Antibody Titration --- p.64 / Chapter 4.5. --- Discussion --- p.66 / Chapter Chapter 5. --- Quality Control --- p.70 / Chapter 5.1. --- Avoidance of Prozone Phenomenon --- p.71 / Chapter 5.2. --- Inter-assay and Intra-assay Precision --- p.71 / Chapter 5.3. --- Discussion --- p.74 / Chapter Chapter 6. --- Adjustment of Anti-casein Interference --- p.77 / Chapter 6.1. --- Casein Allergy --- p.78 / Chapter 6.2. --- Prevalence of Anti-casein --- p.80 / Chapter 6.3. --- Discussion --- p.81 / Chapter Chapter 7. --- Prevalence of Anti-idiotypic Antibodies --- p.86 / Chapter 7.1. --- Formation Kinetics --- p.87 / Chapter 7.2. --- Occurrence in Transplant Patients --- p.87 / Chapter 7.3. --- Transfusion Effect --- p.101 / Chapter 7.3.1. --- Comparison between Transfused Transplant Patients and Normal Controls --- p.103 / Chapter 7.3.2. --- Comparison between Transfused Transplant Patients and Non-transfused Transplant Patients --- p.116 / Chapter 7.3.3. --- Association with Graft Survival --- p.117 / Chapter 7.4. --- Discussion --- p.128 / Chapter Chapter 8. --- Correlation of Transfusion with the Outcome of Transplant --- p.137 / Chapter 8.1. --- Rejection Episode --- p.138 / Chapter 8.2. --- Graft Survival --- p.139 / Chapter 8.3. --- Discussion --- p.142 / Chapter Chapter 9. --- General Conclusions --- p.149 / References --- p.153

by Wing-cheung Pang. / Thesis (M.Sc.)--Chinese University of Hong Kong, 1988. / Bibliography: leaves 45-56.

Mycophenolate mofetil (MMF) is widely used to minimise acute rejection following solid organ transplantation as it inhibits inosine monophosphate dehydrogenase (IMPDH) and thereby reduces lymphocyte activation. The effects of MMF and azathioprine on renal allograft outcome were examined by analysis of the national transplant database held at National Health Service (NHS) Blood and Transplant, Stoke Gifford, Bristol, UK. In a paired kidney analysis, MMF treated patients had a 3 year death censored graft survival of 91% (n=217) contrasts to 97% (n=231) in azathioprine treated patients (p=0.07) with an increased acute rejection rate in the first year after transplantation (44 v 31%, n=105 v 74, p<0.01). In a further study, 13% (n=71) of patients were found to be taking less than 1 g of MMF which was associated with a 3-fold increased risk of graft failure and inferior graft function up to 36 months. One strategy to improve graft outcome would entail targeting MMF dose according to pre-transplant IMPDH activity, which is known to display wide variability between patients, in order to maximize efficacy and minimize toxicity. Therefore, it was decided to measure pre-transplant IMPDH activity and to investigate associations with renal allograft outcome and MMF dose tolerated after transplantation. IMPDH activity was measured by detection of generated XMP by a validated HPLC method in the peripheral mononuclear cells of 55 patients waiting for renal transplantation and was found to exhibit a 4-fold variation of IMPDH activity. Black males had significantly increased IMPDH activity contrasts to Black females (p=0.01). Within the first year of transplantation, 71% (n=12) patients required a reduction in MMF dose. There was no association between pre-transplant IMPDH activity and MMF dose achieved at 1 year or MMF associated side effects or eGFR up to 36 months. It was proposed that the inter-individual variability of IMPDH activity may be associated with genetic polymorphisms and therefore sequencing of the exons of IMPDH I and II was undertaken. Two novel single nucleotide polymorphisms (SNPs), Leu244Leu and Ala285Thr, were identified in the IMPDH I gene. Patients with these variants did not exhibit differential IMPDH activity. Genotyping for established intronic SNPs was undertaken in our patient cohort as well as a random sample of 1040 recipients from the Collaborative Transplant Study DNA bank based at the University of Heidelberg, Germany. The presence of these SNPs did not increase the risk of rejection or affect graft function or MMF dose tolerated at 1 year after transplantation and there was no association between pre-transplant IMPDH activity, 5 year graft and patient survival and genotype. In our study, MMF treatment did not result in improved renal allograft outcomes in comparison to azathioprine therapy. Furthermore, we suggest that measurement of pre-transplant IMPDH activity or genotyping of the IMPDH enzymes is unlikely to assist in optimizing MMF dose and renal allograft outcome.