Thesis (Ph. D.)--Ohio State University, 2006. / Title from first page of PDF file. Includes bibliographical references (p. 87-97).

(Uncorrected OCR) Abstract of thesis entitled NOVEL ASPECTS OF AUTOCRINEIP ARACRINE REGULATION OF GROWTH HORMONE SECRETION AND SYNTHESIS IN GRASS CARP PITUITARY CELLS Submitted by ZHOUHONG for the degree of Doctor of Philosophy at The University of Hong Kong in March 2003 In this study, autocrine/paracrine regulation of growth hormone (GH) synthesis and secretion by local interactions of gonadotrophs and somatotrophs was examined in vitro in pituitary cells prepared from Chinese grass carp (Ctenopharyngodon idellus). Treatment with exogenous OH and gonadotropin (OTH) resulted in a dose-dependent increase in basal GH release, GH production, and GH mRNA levels. However, the opposite effects were observed by removing endogenous OR and OTH using immunoneutralization. Furthermore, GR and OTH immunoneutralizations at the pituitary level were effective in blocking the stimulatory influence on GH mRNA expression induced by GH-releasing factors in fish, including GnRH, dopamine, and PACAP38�Apparently" GH-induced GH gene expression was mediated by increasing the T1/2 ofGH mRNA in the cytoplasm and enhancing the production of GH primary transcripts in the nucleus. Since GH-induced OR mRNA gene expression could be blocked by inhibiting JAK2, P42144MAPK, P38MAPK, and PI3K, it is likely that the JAK/MAPK and JAK/PI3K pathways are involved in the GH receptor signaling. Similarly, exogenous GTH increased the production ofGH primary transcripts. However, it did not improve OR mRNA stability but rather enhanced the turnover of GH transcripts. GTR also increased cAMP production in carp pituitary cells. GTH-induced GH mRNA expression Was mimicked by activating cAMP synthesis and blocked by inhibiting adenylate cyclase (AC) and PKA.. GTH-induced OR mRNA expression was also sensitive to inhibition of JAKz, P42/44MAPK, P3SM.AP1C and PI3K. Similar inhibitions, except for PI3K, were all effective in blocking OR mRNA expression induced by activation of cAMP synthesis. These results indicate that GTH may induce GR gene expression through the AC/ cAMP/PKA pathway secondary coupled to JAK.2 andlor MAPK. Apparently, a cAMP-independent PI3K component is also involved in the post-receptor signaling. Using a colunm perifusion approach, the dynamic interactions between somaotrophs and gonadotrophs were examined. In this case, exogenous OTR induced a rapid rise in basal GH secretion, whereas exogenous GR was found to inhibit basal GTR release. In parallel studies, GTHinduced OR mRNA expression was abolished by OR immunoneutralization. Similarly, GTR immunoneutralization blocked GR-induced OR mRNA expression in carp pituitary cells. These results, as a whole, indicate that endogenously secreted OH and GTR, besides their functions as endocrine hormones, serve as novel autocrine/paracrine factors at the pituitary level to modulate GH secretion, OH production, OH gene expression, and somatotroph sensitivity to stimulation by hypothalamic regulators. These stimulatory influences of GH and GTR on OR gene expression axe exerted at the level of GR rnRNA stability and OH gene transcription, presumably via a direct coupling to the JAK/MAPK and JAKiPI3K cascades or an indirect coupling via the AC/cAMP/PKA pathway. Apparently, a local il1trapituitary feedback loop is present. In this case, GTH released from gonadotrophs stimulates GH secretion in neighboring somatotrophs. GR release from somatotrophs is essential to maintain basal GH synthesis and secretion and also exerts a negative feedback on basal GTB release. This intrapituitary feedback loop formed by local interactions between gonadotrophs and somatotrophs may represent a novel mechanism to control OR gene expression in lower vertebrates. / abstract / toc / Zoology / Doctoral / Doctor of Philosophy

Thesis (Ph. D.)--University of Hawaii at Manoa, 1995. / Includes bibliographical references (leaves 116-131). / Microfiche. / xvi, 131 leaves, bound ill. (some col.) 29 cm

No description available.

Orientador: Luiz Henrique de Araújo Machado / Resumo: O diabetes mellitus (DM) é uma das doenças que mais tem aumentado sua incidência na última década. As células-tronco mesenquimais derivadas de tecido adiposo (CTMs-TA) possuem características imunomoduladoras e reparadoras, sendo capazes de reverter efeitos sistêmicos da doença. Sendo assim, nosso trabalho teve como objetivo avaliar o impacto do transplante de CTMs-TA pelas vias intravenosa, intrapancreática e intra-capsular renal em ratos diabéticos e comparar a que melhor exerce efeito regenerador. Para o estudo, utilizamos 45 ratos, wistar, fêmeas, divididos em 5 grupos: GCS – controle sadio; GCD – controle diabético; GIV – diabético, via intravenosa; GIP – diabético, via intrapancreática; GIR – diabético, via cápsula renal. Para a indução, utilizamos uma única aplicação intraperitoneal (i.p) de Aloxana (120mg/kg), e após 3 dias, os animais foram submetidos ao transplante com as CTMs-TA (2x106cel/animal). Após 15 dias do transplante, os animais foram eutanasiados. O transplante de CTMs-TA pela cápsula renal foi capaz de reverter a hiperglicemia, além de restaurar o diâmetro das ilhotas e restabelecer a função das massa β-pancreática. Concluímos que a cápsula renal foi a que apresentou melhores resultados na regeneração morfofuncional do pâncreas diabético. As CTMs-TA utilizadas neste estudo foram capazes de exercer um importante efeito parácrino, podendo ser utilizadas como alternativa para a melhoria da qualidade de vida de indivíduos diabéticos. / Mestre

The TIR/BB-loop mimetic AS-1 has been reported to prevent cardiac hypertrophy by inhibiting interleukin-1 receptor (IL-1R)-mediated myeloid differentiation primary response gene 88 (MyD88)-dependent signaling. To date, it remains unknown whether and if so how AS-1 contributes to mechanical stress (MS)-induced cardiac fibroblast activation, a key process in pressure overload-induced cardiac remodeling and heart failure. Here, we show that phosphorylation and expression of large tumor suppressor kinase 1 (LATS1), a key molecule in the Hippo-Yes associated protein (YAP) signaling pathway, were down-regulated in primary neonatal rat cardiac fibroblasts (NRCFs) in response to MS and in the hearts of mice subjected to transverse aortic constriction (TAC) procedure; AS-1 treatment was able to restore LATS1 phosphorylation and expression both in vitro and in vivo. AS-1 treatment suppressed the induction of proliferation, differentiation and collagen synthesis in response to MS in NRCFs. AS-1 also ameliorated cardiomyocyte hypertrophy and apoptosis through dampening paracrine secretion of stretched cardiac fibroblasts. In mice, AS-1 treatment could protect against TAC-induced cardiac hypertrophy, myocardial fibrosis and heart failure. Of note, LATS1 depletion using siRNA completely abrogated the inhibitory effects of AS-1 on NRCFs under MS including accelerated proliferation, differentiation, enhanced ability to produce collagen and augmented paracrine secretion of tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β) to induce cardiomyocyte hypertrophy. Therefore, our results delineate a previously unrecognized role for LATS1 in cardiac fibroblast to mediate the beneficial effects of AS-1 in preventing pressure overload-induced cardiac remodeling and heart failure.

<p>Skeletal muscle is dependent upon its microvasculature to deliver oxygen and substrates to support the metabolic demands of muscle contraction. Skeletal muscle capillary density is determined by a variety of factors including muscle fiber metabolic phenotype and mitochondrial volume as well as prior exercise training status. Additionally, muscle microvascular density and function can diminish with age, contributing to several age-related muscle dysfunctions. Skeletal muscle fibers regulate their surrounding microvasculature through the release of angiogenic and angiostatic signaling factors. A robust increase in angiogenic signaling from skeletal muscle facilitates increases in muscle capillarization following endurance exercise. Extracellular vesicles (EV) are membrane bound signaling factors secreted by every cell type. Skeletal muscle-derived EVs (SkM-EVs) may help facilitate numerous signaling functions of skeletal muscle including between skeletal muscle and its microvasculature.</p> <p>The primary aim of my dissertation research was to determine the signaling roles that SkM-EVs in regulating endothelial cell homeostasis and angiogenesis in states of aging and health. Chapter 1 provides an overview of the relevant literature. Chapter 2 represents an investigation into how age-related cellular senescence impacts the angiogenic potential of skeletal muscle progenitor cells. We found that stress-induced senescence increases release of small EVs and has pro-senescent and angiostatic effects on culture endothelial cells. In Chapter 3 we compared the release, contents, and angiogenic potential of SkM-EVs collected from primarily oxidative or primarily glycolytic skeletal muscle tissue in mice. We found that oxidative muscle tissue secretes more EVs than glycolytic muscle tissue, and the miR contents of EVs differ greatly between the two phenotypes. Additionally, EVs from oxidative tissue enhanced endothelial cell migration and tube formation compared to glycolytic tissue EVs, in a potentially nitric oxide mediated fashion. In Chapter 4, we tested how PGC-1α overexpression effected myotube EV release and angiogenic potential. We found that PGC-1α overexpression did not impact myotube EV release, but increased the angiogenic signaling potential of SkM-EVs. Chapter 5 is a brief summary of the results and limitations of the projects presented in Chapters 2-4, with a short discussion of potential future research directions.</p>

No description available.

Rationale: Autologous bone marrow- or cardiac-derived stem cell therapy for heart disease has demonstrated safety and efficacy in clinical trials but has only offered limited functional improvements. Finding the optimal stem cell type best suited for cardiac regeneration remains a key goal toward improving clinical outcomes. Objective: To determine the mechanism by which novel bone-derived stem cells support the injured heart. Methods and Results: Cortical bone stem cells (CBSCs) and cardiac-derived stem cells (CDCs) were isolated from EGFP+ transgenic mice and were shown to express c-kit and Sca-1 as well as 8 paracrine factors involved in cardioprotection, angiogenesis and stem cell function. Wild-type C57BL/6 mice underwent sham operation (n=21) or myocardial infarction (MI) with injection of CBSCs (n=57), CDCs (n=31) or saline (n=57). Cardiac function was monitored using echocardiography with strain analysis. EGFP+ CBSCs in vivo were shown to express only 2/8 factors tested (basic fibroblast growth factor and vascular endothelial growth factor) and this expression was associated with increased neovascularization of the infarct border zone. CBSC and CDC therapy improved survival, cardiac function, attenuated adverse remodeling, and decreased infarct size relative to saline-treated MI controls. CBSC treated animals showed the most pronounced improvements in all parameters. By 6 weeks post-MI, EGFP+ cardiomyocytes, vascular smooth muscle cells and endothelial cells could be identified on histology in CBSC-treated animals but not in CDC-treated animals. EGFP+ myocytes isolated from CBSC-treated animals were smaller, more frequently mononucleated, and demonstrated fractional shortening and calcium currents indistinguishable from EGFP- myocytes from the same hearts. Conclusions: CBSCs improve survival, cardiac function, and attenuate remodeling more so than CDCs and this occurs through two mechanisms: 1) secretion of the proangiogenic factors bFGF and VEGF (which stimulates endogenous neovascularization), and 2) differentiation into functional adult myocytes and vascular cells. / Physiology

La thérapie cellulaire pour le traitement de l'insuffisance cardiaque post-infarctus semble prometteuse même si le bénéfice fonctionnel observé actuellement en recherche clinique reste souvent limité. Parmi les différent types cellulaires utilisables, les cellules souches mésenchymateuses (MSC) reconnues pour leur capacité d'immunomodulation, de transdifférenciation et de sécrétion paracrine représentent un outil intéressant pour la régénération myocardique.L'objectif de ce travail a été de mieux comprendre les mécanismes mis en place par les MSC pour réparer le myocarde lésé afin de développer ensuite une stratégie visant à optimiser les effets thérapeutiques de la greffe de MSCs dans le cadre expérimental de l'insuffisance cardiaque post-infarctus. Pour cette étude, nous avons réalisé des cocultures entre cardiomyocytes adultes et les MSC dérivées du tissu adipeux, les cellules hMADS (human Multipotent Adipose Derived Stem cells) afin de mimer le microenvironnement cardiaque in vitro. Des travaux antérieurs à ma thèse réalisés au laboratoire avaient montré que la communication intercellulaire entre ces deux types cellulaires grâce à des structures nanotubulaires aboutissait à la reprogrammation du cardiomyocyte vers le stade progéniteur. Durant ma thèse, nous avons ensuite pu montrer in vitro, toujours grâce au système de coculture, que ce meme type de communication hetérologue via des connexions nanotubulaires constituées de f-actine et de tubuline, modifiait la sécrétion paracrine des cellules souches hMADS. Les cellules souches ainsi reprogrammées, par les échanges intercellulaires de matériel cardiaque améliorent de façon significative leur potentiel angiogénique et de chémoattraction in vitro. Le bénéfice sur les MSCs de la coculture a été confirmé dans le traitement de l'insuffisance cardiaque post-infarctus chez la souris. Dans ce modèle nous avons pu montré que les cellules souches cocultivées avaient un capacité de régénération myocardique nettement supérieures aux cellules souches naives et que l'amélioration fonctionnelle était associée à une stimulation de la vascularisation et de la mobilisation des progéniteurs cardiaques endogènes. Enfin, des résultats similaires ont été observés dans notre modèle préclinique d'ischémie-reperfusion myocardique porcin encourageant la poursuite des travaux de recherche basés sur la communication intercellulaire afin d'optimiser l'efficacité thérapeutique des cellules souches dans la reconstruction cardiaque..En conclusion, nos travaux ont mis en évidence que la communication intercellulaire entre les cardiomyocytes souffrants et les cellules souches conditionnent de façon importante les effets thérapeutiques des cellules souches et que la manipulation ex vivo de ces phénomènes pourrait constituer une approche pour optimiser la thérapie cellulaire cardiaque chez l'homme. / Cell therapies represent one of the most promising approaches to rebuild damaged heart particularly those based on mesenchymal stem cells (MSC). These cells are known for their plasticity, immune privilege and strong self-renewal ability. Intramyocardial delivery of MSC ameliorates heart function after infarction in clinical studies but mechanisms by which MSC exert their therapeutic action is far from being understood and further investigations are required for improving the modest efficiency observed.The objective of this work was to better understand mechanisms by which MSC repair damaged myocardium in order to develop strategies optimizing their therapeutic effects. To mimic in vitro the microenvironment of an injured heart, we developed a species mismatch co-culture system consisting of terminally-differentiated cardiomyocytes (CM) and MSC from adipose tissue called hMADS for human Multipotent Adipose Derived Stem cells. Previous works in the laboratory showed that cell-to-cell communication processes between CM and hMADS involving tunnelling nanotubes (TNT) reprogram adult CM toward a progenitor-like state.During my PhD, we found that crosstalk between hMADS and CM through TNT altered the secretion by hMADS of cardioprotective soluble factors and thereby maximized the capacity of stem cells to promote angiogenesis and chemotaxis of bone-marrow multipotent cells. Additionally, engraftment experiments into mouse infracted hearts revealed that in vitro preconditioning of hMADS with CM increased the cell therapy efficacy of naive stem cells. Functional improvement was associated with higher angiogenesis and homing of bone marrow progenitor cells at the infarction site. Finally, similar results were observed in our preclinical study using a porcine model of myocardial infarction.In conclusion, our findings established the relationship between the paracrine regenerative action of MSC and the nanotubular croostalk with CM and emphasize that ex vivo manipulation of theses communication processes might be of interest for optimizing current cardiac cell therapies.