京都大学 / 新制・論文博士 / 博士(医学) / 乙第13419号 / 論医博第2227号 / 新制||医||1052(附属図書館) / 京都大学大学院医学研究科脳統御医科学系専攻 / (主査)教授 高橋 淳, 教授 中本 裕士, 教授 大森 孝一 / 学位規則第4条第2項該当 / Doctor of Medical Science / Kyoto University / DFAM

Neuropeptide W-23 (NPW23) and neuropeptide W-30 (NPW30) are 23- and 30-amino acid peptides recently isolated from the porcine hypothalamus. Immunohistochemical studies using a rabbit polyclonal antiserum against the rat NPW23 peptide revealed a limited distribution in the rat brain. NPW23-immunoreactive (irNPW) cells were detected in the paraventricular nucleus (PVH), mainly in the parvocellular division, supraoptic nucleus (SO), accessory neurosecretory nuclei, dorsal and lateral hypothalamic areas, perifornical nucleus, arcuate nucleus, and anterior and posterior pituitary; whereas, irNPW fibers were noted in the PVH and SO, retrochiasmatic nucleus, dorsal and lateral hypothalamic areas, median eminence, amygdala, and posterior pituitary. The pattern of distribution of irNPW in the hypothalamus corroborates a possible role of NPW on prolactin release and feeding behavior reported by others.

No description available.

Pineal-mediated depressions in prolactin cell activity after light deprivation were studied in the male and female Golden Syrian hamster. Prolactin cell activity was determined by measuring radioimmunoassayable prolactin, newly synthesized prolactin and prolactin mRNA levels in the pituitary. Serum prolactin was also measured by radioimmunoassay. Use of the recombinant DNA plasmid, pPRL-1, which contains the rat prolactin complimentary DNA sequence, was validated in this dissertation for measuring prolactin mRNA in the hamster. Male hamsters blinded for 11, 21, or 42 days showed significant and progressively greater declines in prolactin mRNA levels which were completely prevented by pinealectomy. The decline seen after 11 days is the earliest depression in prolactin cell activity reported after light deprivation in the hamster. Female hamsters blinded for 28 days, however, showed no such decreases in prolactin cell activity if they continued to display estrous cyclicity. This supports the hypothesis that, unlike the male, there is not a gradual decline in prolactin cell activity after blinding in the female hamster and that loss of estrous cyclicity may precede or possibly accompany declines in prolactin cell activity. After 12 weeks of blinding, females were acyclic and had dramatically depressed levels of prolactin cell activity. However, pinealectomy did not completely prevent this decline due to blinding unless the females continue to display estrous cyclicity. Thus, when pinealectomy was ineffective in preventing the loss of estrous cyclicity due to blinding, it was also ineffective in preventing declines in prolactin cell activity. In ovariectomized females, blinding caused a decline in prolactin cell activity. Pinealectomy was not consistently effective in preventing this decline after 12 weeks of treatment, although, in females blinded for 4 weeks (at which time all animals were cycling) and then ovariectomized for an additional 4 weeks, pinealectomy completely prevented this decline in prolactin cell activity. In a separate study, significant changes in prolactin cell activity during the estrous cycle were seen in untreated normally cycling female hamsters. These changes in prolactin mRNA, prolactin synthesis, and radioimmunoassayable prolactin in the pituitary were measured in the morning, when, consistent with other reports, no differences in serum prolactin were observed.

The C57 BL/KsJ db/db mouse is obese, hyperglycemic, hyperinsulinemic and serves as a model for noninsulin dependent diabetes mellitus (NIDDM). This study reports a dysfunction in the pituitary-thyroid axis and apparent peripheral resistence to thyroid hormones due to a reduction in T3 receptor binding. Diabetic mice have subnormal serum T4 concentrations and supranormal T3 concentrations which are most pronounced between 8 and 10 weeks of age. Thyroid glands of diabetic animals appear hypoactive histologically. Serum TSH concentrations approximate those found in normal mice. In vitro studies show that thryroid glands from diabetic animals are responsive to TSH. Pituitary glands from the same animals hypersecrete TSH and are responsive to TRH. Ultrastructural analysis of pituitary thyrotropes from diabetic mice indicate that these cells are hypersecretory and may be under chronic stimulation by TRH. Diet restriction maintains diabetic mice at a normal total body weight but these animals still possess abnormally large fat deposits. The thyroid hormone profile of these mice appears normal as does the histological appearance of the thyroid gland. Similarly, the blockade of peripheral deiodination by daily injection of iopanoic acid returns the thyroid hormone profile to normal.

Call number: LD2668 .T4 1950 S666 / Master of Science

De nombreux modèles ont été proposés pour expliquer les mécanismes de développement et de progression tumorale, néanmoins certains aspects comme la nature et la hiérarchie des altérations primaires et secondaires sont encore discutés. Pour répondre à ces questions, nous nous sommes intéressés à la progression tumorale des tumeurs hypophysaires à prolactine humaines. Ces tumeurs d'origine monoclonale sont souvent bénignes mais certaines présentent un phénotype agressif voire malin. Afin d'identifier les mécanismes impliqués dans la progression vers le phénotype agressif nous avons utilisé des techniques de génomique intégrative (puces à ADN, séquençage haut débit) couplant l'étude du transcriptome, du génome (variation du nombre de copie chromosomique, polymorphismes) et du miRNome à partir des mêmes échantillons tumoraux. Par cette stratégie nous avons identifié et hierarchisé des altérations spécifiques des tumeurs agressives et / ou malignes dans un modèle expliquant la progression tumorale des tumeurs hypophysaires à prolactine humaines. Nous avons montré que la sous-expression des microARNs miR-183, miR-744 et miR-98 stimule la prolifération via la surexpression de leurs cibles KIAA0101, TGFB1 et E2F2 spécifiquement dans les tumeurs agressives. Ceci entraine l'acquisition d'altérations chromosomiques (perte du chromosome 11 et le gain du chromosome 1q) permettant l'activation de la dissémination métastatique. Enfin, ce travail montre que l'approche de génomique intégrative multidimensionnelle permet d'apporter de nouveaux éléments pour la caractérisation des phénotypes tumoraux, le diagnostic des tumeurs agressives et la prédiction du comportement tumoral / Numerous models have been proposed to explain the mechanisms of tumor development and progression. Nevertheless some aspects such as the nature and hierarchy of primary and secondary alterations are still debated. To answer these questions, we decided to focus on the tumoral progression of human prolactin pituitary tumors. These monoclonal tumors are usually benign but some present an aggressive or malignant phenotype. To identify the molecular events involved in tumoral progression of human PRL towards aggressive phenotype we used an integrative genomics approach (microarrays, high-throughput sequencing) coupling analysis of transcriptome, genome (variation in the number of chromosomal copy polymorphisms) and miRNome from the same human tumor. Using this strategy we identified and prioritized specific alterations of aggressive and / or malignant tumors in a model explaining the tumor progression of human prolactin pituitary tumors. We have shown that under-expression of micro-RNA miR-183, miR-744 and miR-98 stimulates proliferation through overexpression of their targets KIAA0101, TGFB1 and E2F2 specifically in aggressive tumors. This leads to the acquisition of chromosomal damage (loss of chromosome 11 and gain of chromosome 1q) which allowed the activation of the metastatic processes. Finally, this work shows that the integrative genomic multi-dimensional approach can provide new clues for the characterization of tumor phenotypes, diagnosis of aggressive tumors and prediction of tumor behavior

Orientador: Cezinande de Meira / Banca: Sony Dimas Bicudo / Banca: Cássia Maria Barroso Orlandi / Resumo: O presente estudo visou avaliar a resposta ovariana e a taxa de recuperação embrionária em éguas tratadas com EPE nas doses de 6, 8 e 12,5 mg sendo o tratamento iniciado após a aplicação de prostaglandina F2 no oitavo ou sexto dia após a ovulação com a finalidade de reduzir o tempo e custo do tratamento. Foram realizados dois experimentos, para o experimento 1, 40 ciclos estrais de éguas Mangalarga Marchador, entre cinco e 24 anos, e para o segundo experimento 30 ciclos estrais de éguas mestiças entre quatro e 12 anos de idade, foram estudados. Foi aplicada 7,5 mg i.m.de prostaglandina F2 (Dinoprost Trometamina) no oitavo (experimento 1) ou sexto (experimento 2) dia após a ovulação. Nesse momento as éguas foram distribuídas aleatoriamente em cinco grupos para o experimento 1: (n=8 ciclos estrais/grupo): Grupo F20-23mm 6mg e F20-23mm 8mg receberam 6 e 8mg, respectivamente, de EPE por via i.m. a cada 12 horas, a partir da detecção de folículo(s) entre 20-23mm de diâmetro. Nos Grupos D8 - 6mg e D8 - 8mg foi aplicado 6 ou 8 mg de EPE, respectivamente, a cada 12 horas por via i.m. a partir do D8 (concomitante a PGF2 ). No Grupo F20-23mm Salina (Controle): as éguas receberam solução salina respeitando os mesmos intervalos que os grupos tratados com EPE a partir da detecção de folículo(s) entre 20-23mm de diâmetro. No experimento 2, a metodologia foi similiar ao experimento 1, contudo, as éguas receberam 12,5 mg de EPE, e o tratamento foi iniciado no sexto dia após a ovulação para o Grupo D6-12,5 mg (n=9), ou quando detectou-se folículo(s) entre 20-23mm, Grupo F20-23mm 12,5 mg (n=10) e F20-23mm Salina (Controle, n=10). Em todos os grupos (Exp. 1 e 2) o tratamento (EPE ou salina) foi mantido até 12 horas anterior a detecção de folículo(s) com diâmetro 35 mm, nesse momento a ovulação foi induzida com única dose de 2500 U.I. de hCG, i.v. (Vetecor®)... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: The present study aimed to evaluate the ovarian response and embrionary recovery in mares treated with doses of 6, 8 and 12.5 mg of EPE. The treatment was started after application of prostaglandin F2 for reduce the period and cost of treatment. Two experiments were conducted, for the experiment one, 40 estrous cycles of Mangalarga Marchador mares and ranged in age from 5 to 24 yrs, and for the second experiment 30 estrous cycles of crossbreed mares ranged in age from four to 12 yrs was used. In the eighth (experiment 1) or sixth (experiment 2) after ovulation was applied 7.5 mg i.m. of prostaglandin F2 (Dinoprost Trometamina). In this moment, all mares in experiment 1 was randomly assigned to treatment and control groups as follows: Groups 1 and 2 (n=8 cycles/group) received 6 mg of EPE, i.m. twice daily, beginning when largest follicle (s) was (were) 20-23 mm (Group F20-23mm 6mg) or regardless of follicle size on Day 8 (Group D8 - 6 mg); Groups 3 and 4 (n=8 cycles/group), received 8 mg of EPE, i.m. twice daily, beginning when largest follicle was 20-23 mm (Group F20-23 mm 8 mg) or regardless of follicle size on Day 8 (Group D8 - 8mg); Group F20-23mm Saline (Control, n=8 cycles) received saline, i.m. beginning when largest follicle (s) was (were) 20-23 mm in the same range of groups treated with EPE. In the second experiment was used the same methodology of the experiment 1, however, the mares received 12.5mg of EPE, and the treatment began in the sixth day after ovulation for the Group D6- 12.5mg (n=9), or when the largest follicle (s) was (were) 20-23 mm, Group F20-23mm 12,5 mg (n=10) e F20-23mm Saline (Control, n=10). Treatments with EPE or saline continued until 12 hours before detection of follicle (s) reached 35 mm, at which time a single dose of hCG (2500 U.I., i.v., Vetecor®) was given. Groups were compared using ANOVA and mean differences among groups were... (Complete abstract click electronic access below) / Mestre

Os hormônios tireoidianos (HTs) participam de diversos processos biológicos por meio de ações que exercem na expressão de genes que codificam proteínas envolvidas no metabolismo, crescimento e desenvolvimento. O objetivo desse trabalho foi caracterizar ações genômicas e não genômicas dos HT no eixo hipotálamo-hipófise-adrenal. Detectamos nos ratos tireoidectomizados (Tx) uma redução do conteúdo de mRNA da POMC, em paralelo a um aumento de sua cauda poli(A) e de sua taxa de tradução. T3 em dose suprafisiológica por 30 minutos ou 5 dias promoveu aumento na expressão desse mRNA e o conteúdo proteico de POMC/ACTH permaneceu inalterado. A enzima PC1 teve sua expressão aumentada em ratos Tx, fato revertido pelo tratamento com T3. As enzimas esteroidogênicas e a corticosterona sérica não apresentaram diferenças nos grupos estudados in vivo, apesar da redução sérica de ACTH em animais Tx, porém SF1 teve sua expressão aumentada in vitro após tratamento com T3. Na cultura de hipófises, o TSH se mostrou capaz de aumentar AMPc e Ca2+ citosólico promovendo a secreção de ACTH. / Thyroid hormones (TH) participate in many biological processes promoting the expression of genes that encode proteins involved in the metabolism, growth and development. The aim of this study was to characterize genomic and non genomic actions of TH in the hypothalamic-pituitary-adrenal axis. We detected in thyroidectomized (Tx) rats a reduction of POMC mRNA content in parallel to an increase of its poli(A) tail and its translation rate. Supraphysiological doses of T3 for 30 minutes or 5 days injections increased the expression of this mRNA, although POMC/ACTH protein content remained unchanged. PC1 enzyme had its expression increased in Tx rats, which was reversed back to control levels after T3 treatment. Steroidogenic enzymes and serum corticosterone did not show any differences in the groups studied in vivo, despite the reduction of serum ACTH in Tx animals, but SF-1 had its in vitro expression increased after treatment with T3. In pituitary primary culture, TSH promoted increase in cAMP and cytosolic Ca2+ concentration stimulating ACTH secretion.

by Yeung Sze Mang. / Thesis (M.Phil.)--Chinese University of Hong Kong, 1999. / Includes bibliographical references (leaves 123-133). / Abstracts in English and Chinese. / ACKNOWLEDGMENTS --- p.i / ABSTRACT --- p.ii / 槪論 --- p.iii / ABBREVIATIONS --- p.iv / AMINO ACIDS SHORTHAND --- p.vi / TABLE OF CONTENTS --- p.vii-x / Chapter CHAPTER 1 --- LITERATURE REVIEW / Chapter 1.1 --- Introduction --- p.1 / Chapter 1.2 --- Structural Analysis of SL --- p.1 / Chapter 1.3 --- Location of SL-producing cells and Expression of SL --- p.5 / Chapter 1.4 --- Possible Functions of SL --- p.9 / Chapter 1.4.1 --- Adaptation to various backgrounds and Intensities of Illuminations --- p.9 / Chapter 1.4.2 --- Control of Reproduction and Maturation --- p.10 / Chapter 1.4.3 --- Responses to Stress --- p.12 / Chapter 1.4.4 --- Regulation of P034- and Ca2+ Metabolism --- p.12 / Chapter 1.4.5 --- Acid - Base Balance --- p.14 / Chapter 1.4.6 --- Regulation of Energy Metabolism --- p.15 / Chapter 1.4.7 --- Regulation of Fat Metabolism --- p.15 / Chapter 1.5 --- Regulation of SL Gene Expression --- p.19 / Chapter 1.5.1 --- Pit-1 Related Gene Regulation --- p.19 / Chapter 1.5.2 --- Regulation of Hormone Secretion --- p.21 / Chapter 1.5.2.1 --- Hypothalamic Factors --- p.21 / Chapter 1.5.2.2 --- Steroids --- p.23 / Chapter 1.6 --- Aims of Thesis --- p.23 / Chapter 1.6.1 --- Identification of SLII from Goldfish (Carassius auratus) --- p.23 / Chapter 1.6.2 --- Aims --- p.27 / Chapter CHAPTER 2 --- PCR ANALYSIS OF GFSLII GENE AND ITS EXPRESSION IN GOLDFISH TISSUE / Chapter 2.1 --- Introduction --- p.28 / Chapter 2.2 --- Materials and Methods --- p.31 / Chapter 2.2.1 --- Materials --- p.31 / Chapter 2.2.2 --- Methods --- p.33 / Chapter 2.2.2.1 --- Subcloning and DNA Sequencing of the Goldfish SLII Amplified by PCR --- p.33 / Chapter 2.2.2.1.1 --- PCR Cloning of Goldfish SLII Gene --- p.33 / Chapter 2.2.2.1.2 --- Restriction Enzyme Digestion of the PCR Clones --- p.33 / Chapter 2.2.2.1.3 --- Subcloning of the Digested Fragments --- p.33 / Chapter 2.2.2.1.4 --- DNA Sequencing of the Subcloned Fragments --- p.34 / Chapter 2.2.2.2 --- Tissue Distribution Studies Using RNA Assay --- p.35 / Chapter 2.2.2.2.1 --- Tissue Preparation --- p.35 / Chapter 2.2.2.2.2 --- Total RNA Extraction --- p.35 / Chapter 2.2.2.2.3 --- Electrophoresis of RNA in Formadehyde Agarose Gel --- p.36 / Chapter 2.2.2.2.4 --- First Strand cDNA Synthesis --- p.37 / Chapter 2.2.2.2.5 --- Goldfish SLII Specific PCR --- p.37 / Chapter 2.2.2.2.6 --- PCR to Test DNA Contamination --- p.38 / Chapter 2.3 --- Results --- p.39 / Chapter 2.3.1 --- Subcloning and DNA Sequencing of the Goldfish SLII Amplified by PCR --- p.39 / Chapter 2.3.2 --- Tissue Distribution Studies Using RNA Assay --- p.40 / Chapter 2.4 --- Discussion --- p.45 / Chapter 2.4.1 --- Subcloning and DNA Sequencing of the Goldfish SLII Amplified by PCR --- p.45 / Chapter 2.4.2 --- Tissue Distribution Studies Using RNA Assay --- p.46 / Chapter CHAPTER 3 --- ANALYSIS OF GOLDFISH SLII GENE / Chapter 3.1 --- Introduction --- p.47 / Chapter 3.2 --- Materials and Methods --- p.49 / Chapter 3.2.1 --- Materials --- p.49 / Chapter 3.2.2 --- Methods --- p.54 / Chapter 3.2.2.1 --- Screening of Goldfish Genomic Library --- p.54 / Chapter 3.2.2.1.1 --- Preparation of the Plating Host --- p.54 / Chapter 3.2.2.1.2 --- Preparation of the Probe --- p.54 / Chapter 3.2.2.1.3 --- Primary Screening of Goldfish Genomic Library --- p.55 / Chapter 3.2.2.1.4 --- Isolation of the Positive Clones --- p.56 / Chapter 3.2.2.1.5 --- Phage Titering --- p.56 / Chapter 3.2.2.1.6 --- Purification of the Positive Clones --- p.57 / Chapter 3.2.2.1.7 --- Phage DNA Preparation --- p.57 / Chapter 3.2.2.1.8 --- Find out the Target Gene Size of the Positive Clones --- p.58 / Chapter 3.2.2.1.9 --- Cloning of the PCR Fragments into pUC18 Vector --- p.59 / Chapter 3.2.2.1.10 --- Checking the Cloned Insert Size --- p.60 / Chapter 3.2.2.1.11 --- Restriction Enzyme Digestion to Release the Inserts --- p.61 / Chapter 3.2.2.1.12 --- Mini prep of the Positive Clones for Further Investigations --- p.61 / Chapter 3.2.2.1.13 --- DNA Sequencing of the Positive Clones --- p.61 / Chapter 3.2.2.1.14 --- Restriction Enzyme Mapping of the Positive Clones --- p.62 / Chapter 3.2.2.1.15 --- Subcloning of Clone 2A and5A / Chapter 3.2.2.1.16 --- Determination of the Promoter Region of Clone 2A Using Universal Genome Walker Kit --- p.63 / Chapter 3.2.2.2 --- Southern Blot Analysis of Goldfish and Catfish Genomic DNA --- p.66 / Chapter 3.2.2.2.1 --- Genomic DNA Preparation from Goldfish and Catfish Tissues --- p.66 / Chapter 3.2.2.2.2 --- Restriction Enzyme Digestion of the Genomic DNA --- p.67 / Chapter 3.2.2.2.3 --- Alkaline Transfer of the Digested Genomic DNA --- p.67 / Chapter 3.2.2.2.4 --- Hybridization of the Digested Genomic DNA --- p.67 / Chapter 3.3 --- Results --- p.69 / Chapter 3.3.1 --- Screening of the Goldfish Genomic Library --- p.69 / Chapter 3.3.2 --- Mapping the Target Genes --- p.69 / Chapter 3.3.3 --- DNA Sequencing of the 2 Positive Clones --- p.69 / Chapter 3.3.4 --- Southern Blot Analysis of Goldfish and Catfish Genomic DNA --- p.81 / Chapter 3.4 --- Discussion --- p.83 / Chapter CHAPTER 4 --- EXPRESSION OF RECOMBINANT GOLDFISH SOMATOLACTIN IN ESCHERICHIA COLI (E. COLI) / Chapter 4.1 --- Introduction --- p.87 / Chapter 4.2 --- Materials and Methods --- p.89 / Chapter 4.2.1 --- Materials --- p.89 / Chapter 4.2.2 --- Methods --- p.96 / Chapter 4.2.2.1 --- Transformation of the Recombinant Protein Carrying Plasmid into E. coli. (BL21) --- p.96 / Chapter 4.2.2.2 --- Small Scale Expression of Recombinant Goldfish SLII Protein --- p.96 / Chapter 4.2.2.3 --- Large Scale Expression of Recombinant Goldfish SLII Protein --- p.97 / Chapter 4.2.2.4 --- Preparation of the Recombinant Protein for Purification --- p.99 / Chapter 4.2.2.5 --- Protein Purification Using Novagen His-Bind Resin Kit --- p.99 / Chapter 4.2.2.6 --- Production of Polyclonal Antibody in Rabbits --- p.100 / Chapter 4.2.2.7 --- Enzyme Linked Immunosorbant Assay (ELISA) --- p.101 / Chapter 4.2.2.8 --- Western Blot Analysis of the Recombinant Hormones --- p.103 / Chapter 4.3 --- Results --- p.105 / Chapter 4.3.1 --- Expression of the Recombinant Goldfish SLII --- p.105 / Chapter 4.3.2 --- Purification of the Recombinant Goldfish SLII --- p.105 / Chapter 4.3.3 --- ELISA Analysis --- p.105 / Chapter 4.3.4 --- Western Blot Analysis --- p.110 / Chapter 4.4 --- Discussion --- p.113 / Chapter 4.4.1 --- Expression of the Recombinant Goldfish SLII --- p.113 / Chapter 4.4.2 --- Purification of the Recombinant Goldfish SLII --- p.114 / Chapter 4.4.3 --- Analysis of the Recombinant Goldfish SLII --- p.114 / Chapter CHAPTER 5 --- GENERAL DISCUSSION AND CONCLUSIONS --- p.116 / REFERENCES --- p.123