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  • About
  • The Global ETD Search service is a free service for researchers to find electronic theses and dissertations. This service is provided by the Networked Digital Library of Theses and Dissertations.
    Our metadata is collected from universities around the world. If you manage a university/consortium/country archive and want to be added, details can be found on the NDLTD website.
1

Transporter gene expression in rat lactating mammary epithelial cells & primary organoid cultures using quantitative real-time reverse transcription polymerase chain reaction (QRT-PCR)

Gilchrist, Samuel Edward 30 January 2007
Transporters dynamically expressed at the mammary gland transport critical nutrients into the breast milk of nursing mothers to meet the nutritional demands of the suckling infant. However, xenobiotics may interact with these transporters to potentially alter the nutrient composition of milk and compromise neonatal nutrition. The aim of the present study was to quantitatively evaluate the constitutive expression of various nutrient transporters in whole mammary gland tissue and mammary epithelial organoids (MEO) isolated from female Sprague-Dawley rats at various stages of pregnancy, lactation, and involution. Furthermore, the studys aim was to determine if appropriately cultured mammary epithelial organoids (MEO) maintain in vivo transporter expression to lay down critical groundwork for the development of an in vitro screening tool assessing xenobiotic-nutrient transporter interactions. The following transporters were evaluated using quantitative real-time reverse transcription polymerase chain reaction (QRT-PCR): multidrug resistance protein (Mdr) 1a, 1b; multidrug resistance-like protein (Mrp) 1; organic cation transporter (Oct) 1; organic cation/carnitine transporter (Octn) 1, 2, and 3; concentrative nucleoside transporter (Cnt) 1, 2, and 3; equilibrative nucleoside transporter (Ent) 1, 2, and 3; nucleobase transporter (Ncbt) 1 and 2; oligopeptide transporter (Pept) 1 and 2; methotrexate carrier (Mtx) 1; divalent metal transporter (Dmt) 1; and the milk protein ?-casein. Transporter expression patterns in MEO differed from whole tissue for ?-actin, Mdr1a, Mdr1b, Oct1, Octn3, Ent3, Cnt1, Cnt3, Ncbt1, Pept2, Mtx1, and ?-casein. This brings into question whether whole mammary gland tissue is truly appropriate for an understanding of transporter expression in the mammary epithelium. Nevertheless, four general transporter expression patterns emerged in isolated MEO: decline throughout lactation (Mdr1a, Mdr1b, Mrp1 & Dmt1), increase throughout lactation (Cnt1 & Octn3), increase in early lactation (Oct1, Octn2, Ent1, Cnt2, Cnt3, Pept2 & Mtx1) and constant expression throughout lactation (Octn1, Ent2, Ent3, Ncbt1, Ncbt2 & Pept1). These expression patterns will provide insight into the critical windows of nutrient delivery to the breast milk to provide adequate nutritional stimuli to the suckling infant. Furthermore, MEO cultured in an extracellular matrix-rich environment maintained transporter expression at the mRNA level, which underscores the potential of the primary MEO in vitro model system as a screening tool for xenobiotic-transporter interactions at the mammary gland. Transporter expression patterns in MEO were unique for each transporter evaluated. This information accompanied by an in vitro screening tool may allow for predictions of xenobiotic interference with breast milk composition to help safeguard infant health.
2

Transporter gene expression in rat lactating mammary epithelial cells & primary organoid cultures using quantitative real-time reverse transcription polymerase chain reaction (QRT-PCR)

Gilchrist, Samuel Edward 30 January 2007 (has links)
Transporters dynamically expressed at the mammary gland transport critical nutrients into the breast milk of nursing mothers to meet the nutritional demands of the suckling infant. However, xenobiotics may interact with these transporters to potentially alter the nutrient composition of milk and compromise neonatal nutrition. The aim of the present study was to quantitatively evaluate the constitutive expression of various nutrient transporters in whole mammary gland tissue and mammary epithelial organoids (MEO) isolated from female Sprague-Dawley rats at various stages of pregnancy, lactation, and involution. Furthermore, the studys aim was to determine if appropriately cultured mammary epithelial organoids (MEO) maintain in vivo transporter expression to lay down critical groundwork for the development of an in vitro screening tool assessing xenobiotic-nutrient transporter interactions. The following transporters were evaluated using quantitative real-time reverse transcription polymerase chain reaction (QRT-PCR): multidrug resistance protein (Mdr) 1a, 1b; multidrug resistance-like protein (Mrp) 1; organic cation transporter (Oct) 1; organic cation/carnitine transporter (Octn) 1, 2, and 3; concentrative nucleoside transporter (Cnt) 1, 2, and 3; equilibrative nucleoside transporter (Ent) 1, 2, and 3; nucleobase transporter (Ncbt) 1 and 2; oligopeptide transporter (Pept) 1 and 2; methotrexate carrier (Mtx) 1; divalent metal transporter (Dmt) 1; and the milk protein ?-casein. Transporter expression patterns in MEO differed from whole tissue for ?-actin, Mdr1a, Mdr1b, Oct1, Octn3, Ent3, Cnt1, Cnt3, Ncbt1, Pept2, Mtx1, and ?-casein. This brings into question whether whole mammary gland tissue is truly appropriate for an understanding of transporter expression in the mammary epithelium. Nevertheless, four general transporter expression patterns emerged in isolated MEO: decline throughout lactation (Mdr1a, Mdr1b, Mrp1 & Dmt1), increase throughout lactation (Cnt1 & Octn3), increase in early lactation (Oct1, Octn2, Ent1, Cnt2, Cnt3, Pept2 & Mtx1) and constant expression throughout lactation (Octn1, Ent2, Ent3, Ncbt1, Ncbt2 & Pept1). These expression patterns will provide insight into the critical windows of nutrient delivery to the breast milk to provide adequate nutritional stimuli to the suckling infant. Furthermore, MEO cultured in an extracellular matrix-rich environment maintained transporter expression at the mRNA level, which underscores the potential of the primary MEO in vitro model system as a screening tool for xenobiotic-transporter interactions at the mammary gland. Transporter expression patterns in MEO were unique for each transporter evaluated. This information accompanied by an in vitro screening tool may allow for predictions of xenobiotic interference with breast milk composition to help safeguard infant health.
3

Dissecting the effect of EGF starvation on the signaling and transcriptomic landscapes of the mouse intestinal epithelium

Hassanin, Ismail El-Shimy 17 January 2023 (has links)
Die EGFR-Signalübertragung steuert viele verschiedene zelluläre Prozesse in allen Arten von Epithelzellen, einschließlich des Darmepithels. Diese Prozesse reichen von Proliferation und Wachstum über Differenzierung bis hin zu Autophagie und Apoptose. Die vorliegende Studie zielt darauf ab, die Signalveränderungen zu charakterisieren, die im Darmepithel als Reaktion auf EGF-induzierten Hungerstress stattfinden. Kontraintuitiv führte eine 24-stündige EGF-Starre zu einer deutlichen Phosphorylierung von EGFR, MEK1/2 und ERK1/2, was auf eine Aktivierung dieser Signalachse in Darmzellen hindeutet. Diese Veränderungen waren am signifikantesten in den undifferenzierten CD44-reichen Krypta-Basiszellen. Interessanterweise war die EGF-Starvation-induzierte ERK1/2-Phosphorylierung mit der Hochregulierung einer Untergruppe von ERK-Zielgenen verbunden, bei denen es sich zumeist um primäre Zielgene handelt. Die Überexpression des EGFR-Liganden HBEGF und des FGFR-Liganden FGF1 in ausgehungerten Zellen könnte für die hungerbedingte Zunahme der MAPK-Aktivität verantwortlich sein, obwohl eine erhöhte Sekretion dieser Liganden durch ausgehungerte Organoide nicht bestätigt werden konnte. Dennoch wird die kompensatorische Ligandensekretion durch die Beobachtung gestützt, dass die erneute Zugabe von EGF zu ausgehungerten Organoiden die pERK1/2-Spiegel auf den Ausgangswert zurücksetzt, was bedeutet, dass EGF mit einem anderen von ausgehungerten Zellen sezernierten Liganden um den EGFR konkurriert. Zusätzlich zu HBEGF wurde festgestellt, dass andere Gene, die für den Schutz, das Überleben und die Regeneration des Darmepithels bekannt sind, in ausgehungerten Organoiden überexprimiert werden, wie z. B. Reg3b. Insgesamt können die in dieser Studie berichteten EGF-induzierten Veränderungen der MAPK-Signalübertragung und der globalen Genexpression als ein überlebensförderndes Programm interpretiert werden, das bevorzugt in Darmstammzellen und frühen Vorläuferzellen aktiviert wird. / EGFR signaling drives many different cellular processes in all kinds of epithelial cells including the intestinal epithelium. Such processes range from proliferation and growth to differentiation to autophagy and apoptosis. The present study aims to characterize signaling changes that take place in the intestinal epithelium in response to EGF starvation-induced stress using epithelial organoids derived from the mouse duodenum and human colorectal tumor tissue. Counterintuitively, 24 h EGF starvation induced a prominent phosphorylation of EGFR, MEK1/2 and ERK1/2 indicating an activation of this signaling axis in intestinal cells. These changes were most significant in the undifferentiated CD44-high crypt base cells. Interestingly, EGF starvation-induced ERK1/2 phosphorylation was associated with upregulation of a subset of ERK target genes that were mostly primary-response targets. Overexpression of the EGFR ligand HBEGF and the FGFR ligand FGF1 in starved cells may account for starvation-driven increase in MAPK activity, although an increased secretion of these ligands by starved organoids was not confirmed. Nevertheless, compensatory ligand secretion is still supported by the observation that EGF re-addition to starved organoids restores pERK1/2 levels to baseline which implies that EGF competes for EGFR with some other ligand secreted by starved cells. In addition to HBEGF, other genes known to promote protection, survival and regeneration of the intestinal epithelium were found to be overexpressed in starved organoids such as Reg3b. Collectively, EGF starvation-induced changes in MAPK signaling and global gene expression reported in this study can be interpreted as a pro-survival program that gets activated preferentially in intestinal stem cells and early progenitors.
4

Non-Pyroptotic Gasdermin-B (GSDMB) Regulates Epithelial Restitution and Repair, and is Increased in Inflammatory Bowel Disease

Rana, Nitish 23 May 2022 (has links)
No description available.

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