Thesis (M.S.)--University of Michigan, 1987.

Thesis (M.S.)--University of Michigan, 1987.

Thesis (Ph. D.)--University of Virginia, 1999. / Spine title: IRBP expression & turnover. Includes bibliographical references (p. 176-177). Also available online through Digital Dissertations.

Les oligosaccharides non digestibles (OND) occupent aujourd'hui une place de choix parmi les ingrédients fonctionnels du fait de leur potentielles propriétés prébiotiques. Les transgalacto-oligosaccharides (TOS), oligomères linéaires de galactose obtenus par fermentation du lactose, appartiennent à la famille des OND. L'objectif de ce travail, réalisé en étroite collaboration avec la société Blédina, était de caractériser les effets induits par la consommation de TOS sur la fonction intestinale. Ce travail incluait une première partie dédiée aux effets des TOS sur la muqueuse intestinale, une deuxième partie visant à étudier les conséquences de la consommation de TOS sur la composition de la flore fécale et les paramètres des réponses immunitaires innées et adaptatives au niveau intestinal et périphérique, et la troisième partie a consisté en une étude des effets des TOS sur les signes cliniques et biologiques d'allergie à l'arachide. Nous avons établi que la consommation de TOS induit une forte augmentation de la quantité de protéines totales contenues dans la muqueuse intestinale. Cependant, cet effet ne semble pas dû à une augmentation de la production des mucines, ni à un effet trophique des TOS. Par ailleurs, nous avons confirmé l'effet prébiotique des TOS en montrant que le nombre de bifidobactéries était augmenté après 4 semaines de consommation de TOS. Une analyse plus fine par DGGE nous a également permis de montrer que la diversité de la flore lactobacille est augmentée par la consommation de TOS, même si, d'un point de vue quantitatif, les lactobacilles sont très peu augmentés par les TOS. Certains paramètres immunitaires périphériques, et notamment la proportion de cellules CD4+ dans la rate, sont également modifiés lorsque les animaux consomment les TOS. Nous n'avons en revanche pas constaté d'effets au niveau intestinal. Enfin, lors de notre étude sur animaux sensibilisés à l'arachide, nous avons établi que la consommation de TOS permet de diminuer les immunoglobulines IgG1 spécifiques, tout en augmentant la prolifération des splénocytes cultivés en présence d'un mitogène non spécifique et leur production de cytokines. Cependant, si certains signes biologiques d'allergie sont diminués par la consommation de TOS, les signes cliniques d'allergie à l'arachide ne sont pas affectés. Nos résultats ont donc prouvé que les TOS induisent de réelles modifications de l'écosystème intestinal, mais aussi du système immunitaire périphérique. Des travaux supplémentaires seraient nécessaires afin de mieux comprendre les mécanismes sous-jacents.

L’oviducte joue un rôle central dans le transport et la préparation des gamètes, la fécondation et le développement précoce. Un dialogue embryo-maternel s’établit pour assurer le succès du développement de l’embryon et de son transport vers le site d’implantation. L’objectif principal de ce travail était de confirmer l’existence d’un dialogue moléculaire et fonctionnel précoce grâce à la coculture d’embryons bovins produits in vitro sur des cellules épithéliales tubaires bovines (BOEC). Nous avons montré que les BOEC ont un effet important sur le développement embryonnaire précoce, particulièrement pendant les 4 premiers jours. Cet effet ce traduit par un clivage accéléré, une modulation des gènes exprimés après l’activation du génome embryonnaire, un taux plus élevé de développement au stade de blastocyste et une adaptation de l’expression génique de ces blastocystes. En retour, les embryons induisent dans les BOEC, des changements d’expression de facteurs impliqués dans la réponse à l’interféron. Une spécificité régionale des profils d’expression a également été observée dans l’oviducte. / The oviduct plays a pivotal role in gametes transport and final capacitation, as well as in fertilization and early embryo development. An embryo-maternal communication takes place to ensure the successful early embryo development and transport towards its implantation site. The principal aim of this research was to confirm the existence of such early embryo-maternal molecular and functional dialogue using bovine oviduct epithelial cells (BOEC) as coculture to support the development of in vitro produced bovine embryos. We showed that BOEC had an important effect on early embryo development, especially during the first 4 days. This effect translates into accelerated cleavage kinetics, modulation of gene expression after embryonic genome activation, increased rate of embryo development to the blastocyst stage and improved gene expression profile. Moreover the embryos are triggering a BOEC response by upregulating genes related to interferon signaling. A regional specificity of gene expression profile in the oviduct has also been detected.

The ovarian surface epithelium (OSE) is a monolayer of cells surrounding the ovary that is ruptured during ovulation. After ovulation the wound is repaired, however this process, and the mechanisms to maintain OSE homeostasis after the wound is repaired are poorly understood. We have shown the mouse OSE (mOSE) contains a stem cell population that is expanded by Transforming Growth Factor Beta 1 (TGFB1), a factor present in follicular fluid. These data suggest that components in the follicular fluid such as TGFB1 may promote wound repair and OSE homeostasis through maintenance of the OSE stem cell population. Additionally, TGFB1 may promote wound repair through induction of an epithelial-to-mesenchymal transition (EMT) and activation of pro-survival pathways, as seen in other tissues. To elucidate the mechanism for TGFB1-mediated ovulatory wound repair, mOSE cells were treated with TGFB1, which induced an EMT seen with increased Snai1 expression and cell migration. Snai1 overexpression also increased cell migration and sphere formation (a stem cell characteristic). RNA sequencing results suggest this is at least in part through elevated collagen deposition in SNAI1 overexpressing cells. A TGFB signalling targets array identified Cox2 induction following TGFB1 treatment. Constitutive Cox2 expression did not promote an EMT, but enhanced sphere formation and cell survival. Finally, TGFB1 treatment decreased Brca1 expression, which when deleted from mOSE cells also increased sphere formation. RNA sequencing results suggest that Brca1 deletion promotes stemness through activation of the stem cell genes Ly6a and Lgr5. RNA sequencing was also used to compare mOSE cells cultured as monolayers and as spheroids, with and without TGFB1. These results validate our findings that TGFB1 promotes an EMT partially through Snail induction and the upregulation of Cox2. mOSE cells cultured as spheroids acquire a mesenchymal transcriptional profile that is further enhanced with TGFB1 treatment. These data suggest that TGFB1 may promote ovulatory wound repair and maintain OSE homeostasis through the induction of an EMT, maintenance of the stem cell population and activation of a pro-survival pathway. Interestingly, mOSE spheroids also decrease Brca1 expression and upregulate cancer associated genes such as Pax8 and Greb1. The induction of survival pathways, while simultaneously increasing stemness and repressing Brca1 could render cells more susceptible to transformation. This work provides novel insights as to why ovulation is the primary non-hereditary risk factor for ovarian cancer.

La mucoviscidose est une maladie génétique dont l’atteinte respiratoire est responsable de 90 % de la morbidité et de la mortalité et est caractérisée par une infection chronique et une inflammation persistante. Cette inflammation non contrôlée participe de manière importante à la dégradation du tissu pulmonaire. Malgré les progrès récents, les thérapies actuelles ne permettent pas un traitement efficace de l’atteinte respiratoire. Il est donc indispensable d’identifier de nouveaux mécanismes moléculaires et cellulaires impliqués dans l’inflammation pulmonaire. Dans ce but, nous nous sommes intéressés au canal calcique "Transient Receptor Potential Vanilloid 4" (TRPV4) exprimé au niveau de l’épithélium respiratoire. A l’aide d’approches in vitro et in vivo, nous avons démontré que l’activation du TRPV4 déclenche la sécrétion de médiateurs inflammatoires cytokiniques et lipidiques et un recrutement leucocytaire dans les poumons. Nous avons également observé une altération de la signalisation dépendante du TRPV4 dans le contexte de la mucoviscidose, suggérant que le TRPV4 pourrait constituer une cible prometteuse pour le développement de nouvelles thérapies anti-inflammatoires applicables en santé respiratoire. / Cystic fibrosis (CF) is due to mutations in the gene encoding the Cystic Fibrosis Transmembrane conductance Regulator (CFTR). The pulmonary consequence of the disease accunts for over 90 % of the morbidity and mortality and is characterized by chronic infection and persistent inflammation. This uncontrolled inflammation participates significantly to the degradation of the lung tissue. Despite recent progress, current therapies do not allow effecgive treatment of CF lung disease. It is therefore necessary to characterize nex cellular and molecular mechanisms that could contribute to lung inflammation. In that purpose, we focused on the calcium channel "Transient Receptor Potential Vanilloid 4" (TRPV4) expressed by respiratory epithelium. Using in vitro and in vivo approaches, we found that TRP4 activation triggers the secretion of inflammatory mediators (including cytokines and lipids) and leukocytes recruitment into the lungs. We also observed a significant alteration of TRPV4-dependent signalling in the CF context, suggesting that TRPV4 could constitue a promising target for the development of new anti-inflammatory therapies in lung diseases such as CF.

Wound healing in the corneal epithelium is an essential process to maintain corneal clarity and organism health. The earliest events of cellular injury response include the release of nucleotides and the activation of P2 purinergic receptors. While the purinergic receptor P2X7 has been shown to promote cell migration, its role in corneal epithelial wound healing is still poorly understood. The goal of this work is to better understand the role of P2X7 in the injury response. We analyzed P2X7 expression after epithelial injury in rat corneal organ cultures and found that the receptor localizes to the leading edge of the corneal epithelium. However, overall mRNA and protein expression of P2X7 decreased after injury. Inhibition of P2X7 activation significantly delayed wound closure and prevented the leading edge-localization after injury. We found that P2X7 inhibition altered the wound-induced calcium wave in epithelial cells and altered the number and distribution of focal adhesions in the migrating cells. Live cell imaging of epithelial cells showed that P2X7 inhibition led to altered actin rearrangement, with thick actin bundles in the treated cells. In order to determine the importance of P2X7 in epithelial differentiation and stratified cell migration, we developed a stratified culture model. The cells in the stratified model expressed proliferative and differentiation markers similar to organ cultured corneas, as well as similar P2X7 expression and localization after injury. Together, these results show the importance of P2X7 in the overall purinergic response to injury, and provide tools to study P2X7 in stratified corneal cell migration. To determine if P2X7 may contribute to pathologic delayed wound healing in diseases such as type 2 diabetes, we analyzed P2X7 expression in diabetic human corneas and diabetic model rodent corneas. We showed that P2X7 expression is significantly elevated in unwounded diabetic corneas, and that wound healing is delayed in the rodent model. These data show that elevated P2X7 expression may contribute to the delayed healing in disease and may be a possible therapeutic target.

Submitted by Tatiane Pereira Scarpelli (tati.scarpelli@gmail.com) on 2018-08-15T01:03:56Z No. of bitstreams: 1 Tatiane- dissertação final.pdf: 1984978 bytes, checksum: 95d87d58bd4bea79691e4ec03d530a18 (MD5) / Approved for entry into archive by Elza Mitiko Sato null (elzasato@ibilce.unesp.br) on 2018-08-15T12:42:09Z (GMT) No. of bitstreams: 1 scarpelli_tp_me_sjrp.pdf: 1984978 bytes, checksum: 95d87d58bd4bea79691e4ec03d530a18 (MD5) / Made available in DSpace on 2018-08-15T12:42:09Z (GMT). No. of bitstreams: 1 scarpelli_tp_me_sjrp.pdf: 1984978 bytes, checksum: 95d87d58bd4bea79691e4ec03d530a18 (MD5) Previous issue date: 2018-08-07 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / O consumo excessivo de lipídios na dieta constitui um problema na alimentação da sociedade atual, contribuindo para o desenvolvimento de doenças e de cânceres como o de mama, de próstata e de cólon. Se tratando da próstata, os lipídios podem ter ação direta, ou por mediadores celulares deles derivados e desequilibrar os níveis de esteroides sexuais. Contudo, alguns componentes dietéticos exercem papel benéfico para a saúde prostática ou mesmo propriedades terapêuticas, como a curcumina. Existe, portanto, interesse crescente na ação da curcumina sobre a próstata, e seus análogos sintéticos vem sendo testados para aplicação no tratamento de doenças prostáticas. Sabendo-se que componentes da dieta materna podem afetar a próstata masculina permanentemente, por programação do desenvolvimento, este estudo avaliou as consequências da dieta materna enriquecida com lipídios saturados da banha de porco, durante a gestação e lactação de ratos Wistar, associada ou não à suplementação alimentar com curcumina, além da ingestão do óleo de milho comercial, sobre a histofisiologia da próstata ventral da prole na idade adulta. Ratos Wistar foram divididos em cinco grupos (n=12 por grupo; 12 semanas de idade), segundo o regime alimentar ao qual as mães foram submetidas da 8ª sem. de idade até o desmame dos filhotes: C - dieta padrão; O- dieta padrão e óleo de milho (100µL em dias alternados, via gavagem); Cm - dieta padrão e óleo de milho contendo curcumina (100mg/kg de peso corporal, em dias alternados, via gavagem); L- dieta enriquecida com banha de porco (31% de banha de porco) e LCm - dieta enriquecida com banha de porco e óleo de milho/curcumina. Após eutanásia dos animais e realizada coleta dos materiais biológicos, como a próstata ventral e o sangue, realizaram-se análises das quais se obteve aumento nos depósitos de gorduras epididimal e visceral e, consequentemente, aumento no índice de adiposidade dos grupos L e O. Os animais dos grupos O e LCm tiveram aumento nos níveis de triglicerídeos e o grupo O apresentou aumento no nível de colesterol HDL. Os animais de O, L e LCm apresentaram atrofia epitelial, evidenciada tanto pela análise da frequência de ácinos atróficos, pela altura do epitélio e pela diminuição da frequência relativa do epitélio acinar. Ainda, os animais do grupo L apresentaram aumento da frequência relativa do compartimento luminal e de fibras colágenas. Não houve alterações nos níveis de proliferação celular. A expressão de receptores de andrógenos (AR) no epitélio foi maior no grupo L e um pouco menor para os grupos O e LCm. A incidência (número de animais acometidos/número total de animais) de focos inflamatórios aumentou nos grupos O e L (C- 29%; O – 40%; L – 78%) e foi reduzida pelo consumo de curcumina (Cm- 8%; LCm- 38%). Os dados obtidos indicam que, as dietas maternas enriquecidas com lipídios causam respostas na morfologia e na fisiologia da próstata ventral, em diferentes níveis. A banha pareceu alterar a estrutura e a expressão de AR de maneira acentuada, além de propiciar inflamação na glândula, enquanto o óleo de milho altera em menor grau. Além disso, a curcumina sozinha responde positivamente na próstata. / Excessive consumption of lipids in the diet is a problem in the feeding of current society, contributing to the development of diseases and cancers such as breast, prostate and colon. Lipids can have direct action on the prostate gland, or by cellular mediators derived from them and unbalance levels of sexual steroids. However, some dietary components play a beneficial role in prostate health or even therapeutic properties, such as curcumin. There is, therefore, growing interest in the action of curcumin on the prostate, and its synthetic analogues have been tested for application in the treatment of prostatic diseases. Knowing that components of the maternal diet can permanently affect the male prostate gland by developmental programming, this study evaluated the consequences of the maternal diet enriched with lard saturated lipids during gestation and lactation of Wistar rats, associated or not with dietary supplementation with curcumin, in addition to the commercial corn oil intake, on the histophysiology of the ventral prostate of the offspring in adulthood. Wistar rats were divided into five groups (n = 12 per group, 12 weeks old), according to the diet to which the mothers were submitted from the 8th week of age until the weaning of the pups: C - standard diet; O- standard diet and corn oil (100 μL on alternate days, via gavage); Cm - standard diet and corn oil containing curcumin (100mg / kg body weight, every other day, via gavage); L - diet enriched with lard (31% of lard) and LCm - diet enriched with lard and corn oil / curcumina. After the euthanasia of the animals and the collection of biological materials, such as the ventral prostate and blood, analyzes were carried out, which resulted in an increase in the deposits of epididimal and visceral fat and, consequently, an increase in the adiposity index of the L and O groups. The animals in the O and LCm groups had an increase in triglyceride levels and the O group had an increase in the level of HDL cholesterol. The O, L and LCm animals presented epithelial atrophy, evidenced both by the analysis of the atrophic acini frequency, by the height of the epithelium and by the decrease of the relative frequency of the acinar epithelium. Still, the animals in the L group presented increased relative frequency of the luminal compartment and collagen fibers. There were no changes in the levels of cell proliferation. The expression of receptor androgen (AR) in the epithelium was higher in the L group and somewhat lower in the O and LCm groups. The incidence (number of affected animals / total number of animals) of inflammatory foci increased in the O and L groups (C - 29%, O - 40%, L - 78%) and was reduced by the consumption of curcumin (C - 8% (LC-38%). The data obtained indicate that maternal diets enriched with lipids cause responses in the morphology and physiology of the ventral prostate at different levels. The lard appeared to alter the structure and expression of AR very markedly, in addition to providing inflammation in the gland, while the corn oil changes to a lesser degree. In addition, curcumin alone responds positively in the prostate.

1. A brief historical introduction is given. It deals with the general features of wound healing in which epithelial participation is the dominant feature. 2. In Section A the influence of ascorbic acid on healing of skin wounds in rats is discussed. 3. Second wounds were inflicted at the same site as the initial wounds one month after the latter had healed. 4. In Section B the influence of ascorbic acid on healing of skin wounds in guinea pigs is investigated. 5. In order to discover the action ascorbic acid has on ear wounds, the tips of guinea pigs' ears were cut off. 6. The influence of ascorbic acid on the regeneration of corneal epithelium in guinea pigs is discussed. 7. The effect of ascorbic acid on healing of muco-periosteum was studied. 8. The influence of pantothenic acid on healing of skin wounds in rats was studied. 9. Pantothenic acid (Bepanthen-Salbe) ointment was applied directly to skin wounds.