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1
Barlow, Jonathan. "Mitochondrial involvement in pancreatic beta cell glucolipotoxicity." Thesis, University of Plymouth, 2015. http://hdl.handle.net/10026.1/3314.
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High circulating glucose and non-esterified free fatty acid (NEFA) levels can cause pancreatic β-cell failure. The molecular mechanisms of this β-cell glucolipotoxicity are yet to be established conclusively. In this thesis by exploring mitochondrial energy metabolism in INS-1E insulinoma cells and isolated pancreatic islets, a role of mitochondria in pancreatic β-cell glucolipotoxicity is uncovered. It is reported that prolonged palmitate exposure at high glucose attenuates glucose-stimulated mitochondrial respiration which is coupled to ADP phosphorylation. These mitochondrial defects coincide with an increased level of mitochondrial reactive oxygen species (ROS), impaired glucose-stimulated insulin secretion (GSIS) and decreased cell viability. Palmitoleate, on the other hand, does not affect mitochondrial ROS levels or cell viability and protects against the adverse effects of palmitate on these phenotypes. Interestingly, palmitoleate does not significantly protect against mitochondrial respiratory or insulin secretion defects and in pancreatic islets tends to limit these functions on its own. Furthermore, strong evidence suggests that glucolipotoxic-induced ROS are of a mitochondrial origin and these ROS are somehow linked with NEFA-induced loss in cell viability. To explore the mechanism of glucolipotxic-induced mitochondrial ROS and associated cell loss, uncoupling protein-2 (UCP2) protein levels and activity were probed in NEFA exposed INS-1E cells. It is concluded that UCP2 neither mediates palmitate-induced mitochondrial ROS production and the related cell loss, nor protects against these deleterious effects. Instead, UCP2 dampens palmitoleate protection against palmitate toxicity. Collectively, these data shed important new light on the area of glucolipotoxicity in pancreatic β-cells and provide novel insights into the pathogenesis of Type 2 diabetes.
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Cui, Ju, and 崔菊. "Kinesin-1 in pancreatic beta cell and renal epithelial cell." Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2011. http://hdl.handle.net/10722/197835.
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Hanna, Katie. "Novel mechanisms of glucolipotoxic pancreatic beta cell death." Thesis, Nottingham Trent University, 2018. http://irep.ntu.ac.uk/id/eprint/35356/.
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Glucolipotoxicity (GLT) is the term given to the combined and damaging effect of increased glucose and fatty acid levels on pancreatic beta cells (β-cells) (Poitout et al, 2010). There is mounting evidence that glucolipotoxicity is the cause of the decline in β-cell function found in type 2 diabetes (T2D). T2D is a chronic metabolic disorder characterised by sustained elevated blood glucose and free fatty acids, with a continuously increasing prevalence (Olokoba et al, 2012). It is estimated 415 million people currently are living with diabetes and 193 million are undiagnosed, of those 90% are T2D cases. (chatterjee et al, 2017). There are multiple aims in this thesis including the identification of GLT-induced inflammatory pathways of the pancreatic β-cell resulting from NF-κB activation. To identify novel transcription factors associated with GLT-induced reduction in insulin secretion and insulin gene expression and whether their expression is associated with the presence CD40. To observe whether the addition of carnosine to cultured cells can prevent/reverse the up-regulation in GLT-induced factors which potentially result in β-cell damage. Finally, to observed whether GLT can induce histone modifications resulting from disruption in the TCA cycle. To mimic GLT conditions INS-1 rat pancreatic β-cells were cultured in media supplemented with 28mM glucose, 200µM palmitic acid and 200µM oleic acid. The results showed following 5-day incubation ±GLT, there was an increase in TNF receptor CD40 and a CD40-dependent increase in NF-κB. Further to this exposure of INS-1 cells to GLT conditions resulted in a 3.7-fold increase in iNOS mRNA and increased 4-HNE and 3-NT adduct formation (43.4% and 33% respectively) indicating potential GLT-induced β-cell damage. The addition of 10mM carnosine was able to prevent/reverse the up-regulation of GLT-induced NF-κB activity, iNOS protein expression and 4-HNE and 3-NT adduction, identifying it as a potential therapeutic strategy for T2D. GLT-induced up-regulation of CD40 is also shown to be involved in the modulation of various genes, including insulin. siRNA down-regulation of CD40 resulted in increased insulin gene expression via modulation of ID4. Independent of CD40, a protein usually associated with MODY is observed. GLT results in 33.3% down-regulation of HNF4α, which has a knock-on effect on Rab protein expression resulting in down-regulation of insulin secretion. There by indicating that HNF4α is important in normal insulin secretion. This research found that GLT can result in acetylation of histones H3 and H4, subsequent to TCA cycle dysregulation and disruption to fatty acid synthesis and cholesterol biosynthesis pathways, indicating that GLT can affect gene transcription.
4
Hill, Jennifer. "Bacterial Regulation of Host Pancreatic Beta Cell Development." Thesis, University of Oregon, 2018. http://hdl.handle.net/1794/23140.
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Diabetes is a metabolic disease characterized by the loss of functional pancreatic beta cells. The incidence of diabetes has risen rapidly in recent decades, which has been attributed at least partially to alterations in host-associated microbial communities, or microbiota. It is hypothesized that the loss of important microbial functions from the microbiota of affected host populations plays a role in the mechanism of disease onset. Because the immune system also plays a causative role in diabetes progression, and it is well documented that immune cell development and function are regulated by the microbiota, most diabetes microbiota research has focused on the immune system. However, microbial regulation is also required for the development of many other important tissues, including stimulating differentiation and proliferation. We therefore explored the possibility that the microbiota plays a role in host beta cell development. Using the larval zebrafish as a model, we discovered that sterile or germ free (GF) larvae have a depleted beta cell mass compared to their siblings raised in the presence of bacteria and other microbes. This dissertation describes the discovery and characterization of a rare and novel bacterial gene, whose protein product is sufficient to rescue this beta cell developmental defect in the GF larvae. Importantly, these findings suggest a possible role for the microbiota in preventing or prolonging the eventual onset of diabetes through induction of robust beta cell development. Furthermore, the loss of rare bacterial products such as the one described herein could help to explain why low diversity microbial communities are correlated with diabetes.
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Westermark, Pål. "Models of the metabolism of the pancreatic beta-cell." Doctoral thesis, KTH, Numerical Analysis and Computer Science, NADA, 2005. http://urn.kb.se/resolve?urn=urn:nbn:se:kth:diva-408.
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The pancreatic β-cell secretes insulin in response to a raised blood glucose level. Deficiencies in this control system are an important part of the etiology of diabetes. The biochemical basis of glucose-stimulated insulin secretion is incompletely understood, and a more complete understanding is an important component in the quest for better therapies against diabetes.
In this thesis, mathematical modeling has been employed in order to increase our understanding of the biochemical principles that underlie glucosestimulated insulin secretion of the pancreatic β-cell. The modeling efforts include the glycolysis in theβ-cell with particular emphasis on glycolytic oscillations. The latter have earlier been hypothesized to be the cause of normal pulsatile insulin secretion. This model puts this hypothesis into quantitative form and predicts that the enzymes glucokinase and aldolase play important roles in setting the glucose concentration threshold governing oscillations. Also presented is a model of the mitochondrial metabolism in the β-cell, and of the mitochondrial shuttles that connect the mitochondrial metabolism to the glycolysis. This model gives sound explanations to what was earlier thought to be paradoxical behavior of the mitochondrial shuttles during certain conditions. Moreover, it predicts a strong signal from glucose towards cytosolic NADPH formation, a putative stimulant of insulin secretion. The model also identifies problems with earlier interpretations of experimental results regarding the β- cell mitochondrial metabolism. As an aside, an earlier proposed conceptual model of the generation of oscillations in the TCA cycle is critically analyzed.
Further, metabolic control analysis has been employed in order to obtain mathematical expressions that describe the control by pyruvate dehydrogenase and fatty acid oxidation over different aspects of the mitochondrial metabolism and the mitochondrial shuttles. The theories developed explain recently observed behavior of these systems and provide readily testable predictions.
The methodological aspects of the work presented in the thesis include the development of a new generic enzyme rate equation, the generalized reversible Hill equation, as well as a reversible version of the classical general modifier mechanism of enzyme action.
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Pinnick, Katherine Elizabeth. "Pancreatic fat accumulation and effects on beta cell function." Thesis, University of Oxford, 2009. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.492051.
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Type 2 Diabetes Mellitus (T2DM) is characterised by impaired pancreatic 13-cell function resulting in inadequate insulin secretion. The mechanisms involved in 13-cell dysfunction are largely unknown. Elevated fasting plasma non-esterified fatty acid (NEFA) concentrations have been identified as a risk factor for the development of T2DM. The work in this thesis investigates functional effects of NEFA on the 13-cell. Prolonged exposure to elevated NEFA has previously been associated with impaired insulin secretion, reduced insulin content and altered gene expression and lipid metabolism in the 13-cell. Determining the reversibility of these defects may lead to a greater understanding of the underlying mechanisms. Increased pancreatic fat content is positively associated with body mass index in humans and this may expose the 13-cell to high NEFA concentrations. However, the in vivo concentration and composition of NEFA in the pancreas is not known. An in vitro model of 13-cell 'recovery' from the deleterious effects of fatty acids is presented. The longterm culture (>48h) of mouse islets and INS-1 cells with NEFA (0.5mM) impaired glucose and tolbutamide-stimulated insulin secretion, but this was partially reversed by culture for 24h in the absence of exogenous fatty acids. Culture with oleic acid led to the accumulation of triacylglycerol (TAG) in cytosolic lipid droplets. The protein ADFP was found in close association with these droplets. In contrast, culture with palmitic acid produced large cytoplasmic 'splits'. The removal of exogenous fatty acids from the culture media led to a visible reduction in these morphological features. Extraction of the cellular lipids confirmed an increase in the TAG content following culture with NEFA and demonstrated the incorporation of the experimental fatty acid into the TAG and phospholipid (PL) fractions. Following removal of the fatty acids for 24h, TAG content was reduced and NEFA-induced changes in TAG and PL fatty acid composition were partially reversed. A reduction in TAG content in 'recovering' cells indicated the presence of active Iipases. Culture with NEFA increased lipolysis as shown by the measurement of glycerol in the culture media, but this was reduced in 'recovering' cells. Lipase inhibitors inhibited glycerol release but failed to inhibit a reduction in TAG content, and did not confirm a role for Iipases in the recovery of stimulated insulin secretion. Exposure of INS-1 cells to NEFA increased their oxidative capacity for fatty acids and this remained elevated in 'recovering' cells. Treatment with the CPT-1 inhibitor, etomoxir (10I-lM), impaired the fatty acid oxidative capacity of the 13-cell but did not affect the recovery of insulin secretion. A number of genes were upregulated following prolonged culture with NEFA, these included insulin I and II, CPT-1 and UCP2. These genes all displayed reduced expression in cells cultured further in the absence of exogenous fatty acids. The content and composition of fat in tissues from mice was investigated. The TAG composition reflected the major fatty acids found in the diet, with elevated proportions of palmitic and palmitoleic acid indicating the contribution of de novo lipogenesis and desaturase activity to this fatty acid pool. Pancreatic PL were highly unsaturated compared to liver PL, with arachidonic acid accounting for -25% of the PL fatty acids. In mice fed a high-fat (40%) diet (HFD) which was compositionally matched to a control (5%) diet, a 20-fold increase in pancreatic fat was found by 15 weeks. Adipocytes, which were positively labeled for perilipin were observed in the exocrine tissue of the pancreas in HFD mice and lipid droplets labeled for ADFP were identified in the cytoplasm of exocrine cells. By 15 weeks, the fatty acid composition of the TAG, PL and NEFA fractions showed significant differences between HFD and control mice. Perilipin-positive adipocytes were also identified in human pancreas samples and the percentage adipocyte area in histological sections positively correlated (r=0.64) to total pancreatic TAG content. In conclusion, the in vitro findings show the deleterious effects of fatty acids are not permanent. However, increased fat accumulation in the pancreas, as seen in obesity, could expose the 13-cell to elevated NEFA concentrations which, over many years, may lead to irreversible 13-cell failure.
7
Yang, Yu Hsuan Carol. "Identification and characterization of pancreatic beta-cell survival factors." Thesis, University of British Columbia, 2014. http://hdl.handle.net/2429/46424.
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Hughes, Jonathan Martyn. "Streptozotocin and sugar transport in pancreatic beta cell lines." Thesis, University of Bath, 1993. https://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.386772.
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Duffy, Joan. "Effects of insulin sensitising agents on pancreatic beta cell function." Thesis, University of Ulster, 2004. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.399052.
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Halvorsen, Tanya L. "Growth regulation and differentiation in the human pancreatic beta cell /." Diss., Connect to a 24 p. preview or request complete full text in PDF format. Access restricted to UC campuses, 2001. http://wwwlib.umi.com/cr/ucsd/fullcit?p3000408.
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Cosentino, C. "ROLE OF TRNA MODIFYING ENZYMES IN PANCREATIC BETA CELL DEMISE." Doctoral thesis, Università degli Studi di Milano, 2015. http://hdl.handle.net/2434/335205.
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Transfer RNAs (tRNAs) are small molecules of 70-80 nucleotides with a crucial role in protein synthesis. tRNAs once transcribed are highly modified and the methylation is the most common modification. Several enzymes are responsible of tRNA modification and their function is necessary to regulate the stability, the aminoacylation and the rigidity of the structure of tRNAs. De-aminoacylated or degraded tRNAs can act as important signal molecules in the cells, activating different pathways of stress response. For this reason is not surprising that mutations in genes codifying for tRNA modifying enzymes have been associated to many human diseases. Polymorphisms in the gene CDKAL1, codifying the Cdk5 regulatory associated protein 1, have been linked to the development of type 2 diabetes (T2D) in human. CDKAL1 is a methyl-thio transferase that modifies the residue in position 37 of tRNAs, which recognize the codon UUU for lysine. The absence of CDKAL1, and consequently of the modification catalyzed by the enzyme, was shown to induces a decrease of incorporation of lysine residues in proinsulin at the level of pancreatic beta cells. Lysine residues are crucial for the correct maturation of proinsulin. It was shown that the absence of CDKLA1 mediated modifications leads to defects in the processing of proinsulin to produce insulin and c-peptide and to impaired glucose-stimulated insulin secretion. Furthermore in CDKAL1 knock out beta cells it’s observed an increase of markers of endoplasmic reticulum (ER) stress. The chronic activation of ER stress processes decreases the general protein synthesis and the chronic activations triggers pro-apoptotic pathways. These events have been linked to the development of T2D.
The aim of the present work is to study the role of tRNA modifying enzymes in pancreatic beta cells and to investigate the consequences of mutations in these genes on cell function and survival.
A Whole Exome Sequencing study performed previously from my group produced a list of candidate genes for Congenital Hyperinsulinism (CHI). CHI is a rare disease, characterized by inappropriate insulin secretion leading to hypoglycemia. Mutations in nine genes are already known to be causative of the disease, but in 50% of patients the genetic cause is unknown. Using bioinformatics tool I identified CDKAL1 as one of the most promising candidate genes. The mutation identified leads to the substitution of a Serine with a Phenylalanine in position 561, with probable consequences on the transmembrane domain that ensures the correct localization of the protein in the membrane of the ER. In order to study the consequences of S561F CDKAL1 variant in beta cells, I used molecular biology techniques inducing the overexpression of wild type and mutated gene in INS-1E cell line, derived from rat insulinoma. The localization of CDKAL1 was analyzed by immunofluorescence microscopy: the S561F variant affect the localization of the protein that, although still inserted in the ER, tends to accumulate in vesicular structures in some regions of the ER membrane. I also studied the impact of S561F CDKAL1 overexpression on the beta cell function, by measuring the content and the release of insulin in basal growing conditions. I observed an increase of insulin content induced by the overexpression of the wild type protein while the insulin release was not changed. On the other hand, the S561F variant doesn’t affect the insulin content that doesn’t change compared to not-transfected cells, but induces an increase in insulin release. These preliminary results suggest that the S561F CDKAL1 variant could have a role in the development of beta cell dysfunction leading to an inappropriate insulin secretion.
The second part of my project regards the methyl-transferase TRMT10A. A mutation in this gene - the insertion of a stop codon and consequent absence of mature protein - was identified in patients affected by microcephaly and young onset diabetes. It was demonstrated that TRMT10A modifies guanine residues, but its role in tRNA modification in human is still not demonstrated. The absence of the protein leads to an increase of cell death in basal conditions and sensitizes cells to ER stress-induced apoptosis. My work aimed at the characterization of the consequences of TRMT10A deficiency on tRNA modification and stability. I used lymphoblast cells derived from controls subjects and patients to investigate this tRNAs molecules specific for glutamine and methionine were identified to be modified by the enzyme, and the development of a northern blot technique allowed me to obtain preliminary on the aminoacylation and stability of these molecules in TRMT10A deficiency conditions. Furthermore I investigated the mechanisms that lead to beta cell death, triggered by the absence of the protein. With this purpose I induced the silencing of the gene in two different cell lines: INS-1E and EndoC- βH1 (human beta cell line). Results obtained demonstrated that TRMT10A deficiency triggers the activation of the intrinsic pathway of apoptosis through the modulation of Bim expression, a proapoptotic protein of the BH3-only family. The results obtained highlighted the importance of TRMT10A for the survival of the beta cells. Furthermore the activation of the intrinsic pathway of apoptosis is one of the events observed in the development of type 2 diabetes. These findings can give additional proves that the monogenic forms of diabetes can be used as model for the study of mechanisms involved in type 2 diabetes.
Even if further investigations on the complex processes involved are needed, the present work provides important evidences of the role of tRNA modifying enzyme in beta cell homeostasis. Moreover recent reports about the role of tRNAs in signalling pathways support the hypothesis that these molecules can be important mediators of stress response in beta cells, and the tRNA modifying enzymes may act as activators or inhibitors of these responses.
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Tsang, Siu-wai. "Involvement of Pdzd2 in the regulation of pancreatic beta-cell functions." View the Table of Contents & Abstract, 2007. http://sunzi.lib.hku.hk/hkuto/record/B39716430.
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Tsang, Siu-wai, and 曾少慧. "Involvement of Pdzd2 in the regulation of pancreatic beta-cell functions." Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2007. http://hub.hku.hk/bib/B39793746.
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Nishi, Kiyoto. "Nardilysin Is Required for Maintaining Pancreatic β-Cell Function." 京都大学 (Kyoto University), 2017. http://hdl.handle.net/2433/225463.
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Cromwell, Diane. "Pancreatic beta-cell actions of nutrients and metabolizable nutrient ester derivatives." Thesis, University of Ulster, 2006. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.494335.
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Tym, Amy. "Effect of protein glycation by methylglyoxal on pancreatic beta cell function." Thesis, University of Warwick, 2014. http://wrap.warwick.ac.uk/61717/.
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Methylglyoxal is a physiological dicarbonyl metabolite and potent argininedirected glycating agent. It often modifies proteins at functional sites producing loss of positive charge, structural distortion and inactivation. Plasma methylglyoxal is increased in hyperglycaemia associated with diabetes and is linked to the development of vascular complications of diabetes – particularly nephropathy, retinopathy and neuropathy. The effects of dicarbonyl glycation on beta cells and involvement in early stage dysfunction and development of type 2 diabetes mellitus are not known. The aim of this project was to investigate the effect of dicarbonyl protein glycation on beta cell function and related involvement in the development of diabetes. Studies were performed in an in vitro model of beta cell dysfunction - MIN6 insulinoma cells incubated under low and high glucose concentrations, and in a pre-clinical in vivo model of decline of glucose tolerance preceding development of type 2 diabetes - high fat diet-induced insulin resistant mice. Dicarbonyl metabolism and protein damage by glycation and oxidation were studied by stable isotopic dilution analysis liquid chromatography-tandem mass spectrometry. Localisation of methylglyoxal glycation adducts within the pancreas were visualised by immunostaining. Interactions between the extracellular matrix protein, collagen IV, and MIN6 cells in vitro were investigated and impairments in adhesion were assessed following glycation with methylglyoxal. Impairments in adhesion of MIN6 cells to methylglyoxal-glycated collagen IV were assessed using atomic force microscopy force spectroscopy. The results show that MIN6 cells were resistant to accumulation of methylglyoxal when incubated in high glucose concentration although the flux of methylglyoxal was increased 41%. Glycation of collagen IV by methylglyoxal impairs binding to MIN6 cells in vitro resulting in a 91% decrease in the energy necessary to detach cells from the extracellular matrix protein. In high fat diet fed mice the concentration of methylglyoxal in the pancreas was increased. Visualisation of MG-H1 adduct residues in the pancreas showed they were predominantly on the extracellular matrix. In conclusion, protein glycation by methylglyoxal occurs in MIN6 cells in vitro and in the mouse pancreas in vivo. Although the methylglyoxal concentration in the pancreas of high fat diet fed, insulin resistant mice was increased, the lack of a concurrent increase in methylglyoxal protein glycation adducts suggests there may be increased turnover of methylglyoxal-modified proteins. Impairment of beta cell attachment to the extracellular matrix protein, collagen IV, by methylglyoxal and increased protein turnover stimulated by an increased rate of methylglyoxal glycation may impair beta cell function in pre-diabetes in vivo. Glycation by methylglyoxal may contribute to beta cell glucotoxicity and dysfunction with progression to type 2 diabetes mellitus.
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Zhang, Wen. "Mechanism of genistein in the regulation of pancreatic beta-cell proliferation." Thesis, Virginia Tech, 2007. http://hdl.handle.net/10919/35772.
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This study was designed to examine the effect of genistein, a botanical derived primarily from legumes, on pancreatic β-cell proliferation and the related molecular mechanisms. Diabetes mellitus is a major and growing public health problem worldwide. Both in type 1 (T1D) and type 2 diabetes (T2D), the deterioration of glycemic control over time is primarily caused by an inadequate mass and progressive dysfunction of β-cells. Therefore, the search for novel, safe and cost-effective agents that can enhance islet β-cell proliferation, thereby preserving β-cell mass, could be one of the essential strategies to prevent diabetes, given that β-cells have the potential to regenerate by proliferation of pre-existing b-cells in both physiological condition and after onset of diabetes. Genistein has various biological actions. However, studies on whether genistein has an effect on pancreatic β-cell function are very limited. Our laboratory recently found that genistein activates cAMP/protein kinase A (PKA) signaling in both clonal β-cells and mouse islets. Here I present evidence that genistein induced cellular proliferation of clonal rat pancreatic β-cells (INS1) and human islets following 24 h of incubation. This effect was dose-dependent with 5 µM genistein inducing a maximal 41% increase. The effect of genistein on cell proliferation was not dependent on estrogen receptors because this effect was not blocked by the estrogen receptor inhibitor ICI182,780. In addition, the genistein effect on β-cell proliferation was not shared by 17-β-estradiol or a host of structurally related flavonoid compounds, suggesting that this genistein action is structure-specific. Pharmacological or molecular intervention of PKA or MEK1/2, the upstream kinase of p42/44 mitogen activated protein kinases (ERK1/2), completely abolished the genistein-stimulated proliferation of INS1 cells and human islets, suggesting that both molecules are essential for genistein action. Consistent with its effect on cell proliferation, genistein increased intracellular cAMP and subsequently activated PKA in human islets. Genistein also caused a rapid and sustained phosphorylation of ERK1/2 with a maximal increase of 185% at 5 µM genistein. The genistein-induced ERK1/2 activation was completely ablated by inhibition of PKA in INS1 cells and human islets. Furthermore, I found that genistein induced protein expression of cyclin D1, a nuclear target of PKA and ERK1/2 activation and a major cell-cycle regulator essential for ï ¢-cell growth. These findings demonstrated that genistein may be a plant-derived growth factor for pancreatic β-cells involving induction of cyclin D1 via activation of the cAMP/PKA-dependent ERK1/2 signaling pathway, thereby providing a novel role for genistein in the regulation of pancreatic β-cell function.Master of Science
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Yan, Zhongyu. "Charaterization of Chlorpyrifos Toxicity on the Pancreatic Beta Cell Line RINm5f." Wright State University / OhioLINK, 2010. http://rave.ohiolink.edu/etdc/view?acc_num=wright1290111576.
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Weng, Chen. "SINGLE-CELL TRANSCRIPTOMICS OF HUMAN PANCREATIC ISLETS IN DIABETES AND ΒETA CELL DIFFERENTIATION." Case Western Reserve University School of Graduate Studies / OhioLINK, 2021. http://rave.ohiolink.edu/etdc/view?acc_num=case1612882103714773.
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Watson, Maria. "The role of palmitate in skeletal muscle cell insulin resistance and pancreatic beta cell dysfunction." Thesis, University of Dundee, 2009. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.505620.
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Jeffrey, Kristin Danielle. "Novel pathways in fatty-acid induced apoptosis in the pancreatic beta-cell." Thesis, University of British Columbia, 2007. http://hdl.handle.net/2429/31378.
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Pancreatic β-cell death is a critical event in the pathogenesis of all forms of
diabetes. Type 2 diabetes is caused by the combination of acquired factors such as
elevated circulating fatty acids as well as genetic factors. In this study, we show that the
free fatty acid palmitate increases markers of endoplasmic stress and apoptosis in
pancreatic β-cells. Carboxypeptidase E (CPE), an enzyme involved in the processing of
insulin, was identified as the major down-regulated protein spot during palmitate-induced
apoptosis using Cy-dye 2D gel proteomics in both the MIN6 β-cell line and human islets.
Using MIN6 cells treated with 1.5 mM palmitate complexed to BSA (6:1), a significant
decrease in total carboxypeptidase E protein was confirmed through Western blots. The
decrease in CPE was seen in the presence of palmitate at both low and high glucose and
was not affected in high glucose alone or with the ER-stress inducer, thapsigargin.
Palmitate-induced changes in carboxypeptidase E were present after 2 hours, while
CHOP, a marker of ER-stress, was not expressed until after 6 hours of incubation,
suggesting that the decrease in CPE occurs before ER-stress. This finding, together with
experiments using protein synthesis inhibitors and RT-PCR suggested that CPE was
likely regulated at the post-translational level. Treatment with the non-metabolizable
palmitate, 2-bromopalmitate, did not decrease CPE expression and delayed β-cell death.
Addition of the L-type Ca2 + channel blocker nifedipine to palmitate-treated MIN6 cells
restored CPE protein levels, reduced ER-stress, and rescued β-cells from cell death. The
calpain inhibitor E64D also reversed the palmitate-induced decrease in CPE, further
implicating Ca2+-dependent proteolysis pathways. This inhibitor however increased ERstress
on its own and did not prevent ER-stress or β-cell death induced by palmitate.
Interestingly, islets from CPE mutant mice exhibited increased TUNEL labeling,
suggesting elevated apoptosis in vivo. Isolated CPE-deficient islets demonstrated
increased CHOP and cleaved caspase-3 levels compared to control mice. The effects of
palmitate on ER-stress and apoptosis were not additive to those of CPE deficiency. On
the other hand, β-cells over-expressing CPE were resistant to palmitate-induced ERstress
and apoptosis. Together, these results show that palmitate markedly lowers CPE
protein in the cell in a Ca2+-dependent manner, potentially through Ca2+-dependent
proteases, such as the calpains. In addition, a lack of CPE appears to increase the
susceptibility of the β-cell to CHOP induction and apoptosis. Conversely, overexpression
of CPE protects β-cells from ER-stress and apoptosis induced by palmitate.
These findings suggest that CPE represents a novel link between hyperlipidemia and β-cell
death in diabetes.Medicine, Faculty of
Cellular and Physiological Sciences, Department of
Graduate
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Stokesberry, Susan Anne. "Functional effects of temperature on pancreatic beta-cell insulin secretion and integrity." Thesis, University of Ulster, 2005. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.422895.
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Rodrigues, Costa Ana, Celia M. Antunes, and Júlio Cruz-Morais. "Abnormal regulation of pancreatic beta cell Na,K-ATPase on glucose intolerant rats." Bachelor's thesis, Springer Berlin, 2010. http://hdl.handle.net/10174/3307.
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Background and aims: Glucose (G) is the most important physiological insulin secretagogue. It is widely accepted that, in pancreatic -cell, G evoked early ionic events such as membrane depolarization and Ca2+ influx through voltage dependent Ca2+ channels triggers insulin exocytosis. However, the role of other electrogenic systems, namely ionic pumps, to these events remains essentially uninvestigated. It is known that the activity of Na,K-ATPase is modified in type 2 diabetes (T2D). The pump is responsible for maintaining Na+ and K+ gradients across the plasma membrane and generates a net outward current as a result of 3Na+/2K+ exchange. It remains elusive whether Na,K-ATPase activity is regulated by G in pancreatic β-cell and/or this current contributes to the ionic events regulating insulin secretion.
The aim of this work was to assess G evoked regulation of Na,K-ATPase activity in intact -cells of normal and G intolerant rats.
Materials and methods: Pancreatic -cells, from normal (controls) or glucose-intolerant Wistar rats (GIR), were isolated and cultured (48h). Cell batches were pre-incubated (30min) with 2.1mM G to reach basal. Afterwards cells were challenged with [G] in the interval 0-11.1mM for 60min, for dose-dependence evaluation, or with 8.4mM G for 5-120min, for time-dependence evaluation. ATPase activity was assessed in intact cells by colorimetric quantification of Pi formed in 30min. Na,K-ATPase activity was calculated by the difference between the activities obtained in the absence and in presence the of 1mM ouabain.
Results: G evoked both time- and dose-dependent regulation of Na,K-ATPase. In β-cells from controls, G induced a bimodal regulation of Na,K-ATPase. In the absence of G, Na,K-ATPase activity was 0.056±0.015U/mg. Raising [G] to 2.1mM induced a ≈3 fold increase of Na,K-ATPase activity whereas a further increase in [G] in the interval of 5.6-11.1mM evoked a significant reduction of Na,K-ATPase activity to the levels observed in the absence of the secretagogue. Compared to 2mM G, the activity was reduced in 68%, 55% and 66% when [G] was increased to 5.6, 8.4 and 11.1mM, respectively (n=3-12). GIR β-cells exhibit an altered profile of response to the secretagogue; In the absence of G, Na,K-ATPase activity was ≈4 fold the activity observed in the controls (0.202±0.036U/mg; n=3). The pump activity remained unchanged for 2.1-5.6mM G and similar to maximal activity observed in the controls (0.188±0.035 U/mg, for 2.1mM G; n=4). A significant reduction of the pump activity in GIR β-cells was induced by 8.4mM G (0.118±0.018 U/mg).
G (8mM) induced a time-dependent inhibition of Na,K-ATPase with a biphasic profile. Pump activity decreased to a minimum value (32%) after 20min exposure to G, showing a partial recovery to 45%, 46% and 47% for 30, 60 and 120min, respectively (n=5-12). GIR β-cells showed an attenuated response to G (59% activity after 20min) without any recovery (n=5-11).
Conclusions: This work demonstrates that Na,K-ATPase is finely regulated by G in pancreatic β-cell from normal subjects. This regulation is impaired in GIR where desensitization and an attenuation of the inhibitory action of G were observed. In summary, Na,K-ATPase contribution to G-induced ionic events and insulin secretion might be relevant in T2D development.
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Manesso, Erica. "DYNAMICS OF PANCREATIC BETA CELLS: Evidence for Beta Cell Turnover and Attempted Regeneration in Diabetes from Sources of Beta Cells other than Beta Cell Replication in Rats, Monkeys, and Humans." Doctoral thesis, Università degli studi di Padova, 2010. http://hdl.handle.net/11577/3426591.
Full textAbstract:
Since the fundamental defect in both type 1 (T1DM) and type 2 diabetes (T2DM) is beta cell failure, there is increasing interest in the capacity, if any, for beta cell regeneration. In this context quantitative analysis of beta cell turnover becomes essential to permit investigation of the mechanisms that regulate it. For example, how does beta cell mass adapt to obesity?
How is beta cell mass preserved during aging? How does beta cell mass expand during childhood?
In collaboration with the Larry Hillblom Islet Research at David Geffen School of Medicine, University of California Los Angeles, we developed a dynamic model to estimate beta cell turnover. Assuming homogeneity of beta cells, the model describes beta cell mass as the balance between beta cell formation and loss. Beta cells are added either by replication of existing beta cells or by other sources of beta cells (OSB), and they are mainly lost through beta cell apoptosis. Since all the model parameters can be quantified with the exception of OSB, it was possible to solve for this unknown.
The resulting components of beta cell turnover, i.e. new beta cell formation (replication of existing beta cells plus other sources of beta cells rather than beta cell replication) and beta cell death (apoptosis), were used to develop a population model to assess the mean age of a beta cell as well as the mean beta cell lifetime. The resulting model is a variation of the classical McKendrick-von Foerster equation and describes beta cells as a variegate population of cells that differ each other by their own age.
The novel insights that emerge by applying the model to different species, i.e. rats, monkeys, and humans are: 1) there is ongoing beta cell turnover in non diabetic rats, monkeys, and humans through adult life; 2) formation and maintenance of the population of adult beta cells largely depend on OSB in non diabetic rats, monkeys, and humans; 3) the formation of new beta cells from other sources of beta cells increases substantially in the face of the increased beta cell apoptosis in the HIP rat model of type 2 diabetes, delaying the decline in beta cell mass. In contrast, beta cell turnover is low in the streptozotocin (STZ) monkey model of T1DM, compared to non diabetic controls. The extent that beta cells are formed in the STZ monkey is again primarily from OSB. 4) Beta cell replication is the primary mechanism subserving the postnatal expansion of beta cell mass in childhood, while in adulthood OSB is the principal mechanism for maintaining beta cell mass in face of an increased beta cell apoptosis; 5) beta cell mass and beta cell turnover increase in response to obesity in humans; 6) the estimated mean age of a beta cell (1-2 months in rats, 2-5 months in monkeys, and 6 months- 2 years in humans) and the mean beta cell lifetime (1-3 months in rats, 2-5 months in monkeys, and 6 months-2 years in humans) potentially permit endogenous regeneration of beta cell mass in diabetes if beta cell turnover could be altered therapeutically.
The presented models provide, for the first time, information about sources of beta cells other than those derived from beta cell replication and estimates of the mean age of a beta cell and the mean beta cell lifetime in rats, monkeys, and humans. The results have an impact from a clinical point of view considering that: a) the origin of beta cells is actively debated, i.e. some propose duplication of existing beta cells, and others suggest formation of new beta cells from a variety of sources; b) restoration of glycemic control in type 1 and type 2 diabetes through endogenous regeneration could be a potential alternative strategy to pancreas transplantation given the insufficient number of pancreases available for transplantation and the risks of prolonged immunosuppression; c) the unique experimental approach to identify other sources of beta cells is the cell-lineage tracing that is not available in humans. Furthermore, the results encourage: a) future studies on beta cell turnover in patients with diabetes; b) the development of ad hoc experiments that identify the possible other sources of beta cells rather than beta cell replication; c) to plan both experiments and mathematical models to establish the forms and the time required for endogenous regeneration of beta cell mass.L’interesse verso una potenziale rigenerazione della massa beta cellulare è in aumento poiché un difetto della stessa caratterizza sia il diabete di tipo 1 che quello di tipo 2. In questo contesto un’analisi quantitativa del turnover beta cellulare diviene essenziale per comprendere la varietà dei meccanismi che lo regolano. Per esempio, la massa beta cellulare si adatta all’obesità? Si preserva con l'età? Come si espande nell’infanzia? In collaborazione con il Larry Hillblom Islet Research at David Geffen School of Medicine, University of California Los Angeles, è stato sviluppato un modello dinamico per la stima del turnover beta cellulare. Assumendo un comportamento omogeneo delle beta cellule in termini di turnover, il modello riesce a descrivere la massa beta cellulare come il bilancio tra la formazione e la morte di beta cellule. Le beta cellule si formano o dalla duplicazione di beta cellule esistenti o da altre sorgenti (abbreviate con OSB, dall’inglese Other Sources of Beta cells) e muoiono principalmente per apoptosi. Dal momento che tutti i parametri del modello possono essere determinati ad eccezione di OSB, dal modello si determina questa quantità incognita. Le componenti del turnover beta cellulare, ovvero la formazione di nuove beta cellule (duplicazione di beta cellule esistenti sommata a OSB) e l’apoptosi, sono state impiegate nello sviluppo di un modello di popolazione per la stima dell’età e dell’aspettativa di vita medie di una beta cellula. Il modello risultante è una variazione della classica equazione di McKendrick-von Foerster e descrive le beta cellule come una popolazione di cellule che differiscono l’una dall’altra per la loro età. Gli innovativi risultati che emergono dall’applicazione dei modelli a specie differenti, ovvero ratti, scimmie e individui sono: 1) c’è turnover beta cellulare nei ratti, nelle scimmie e negli individui non diabetici in età adulta; 2) la formazione ed il mantenimento della massa beta cellulare dipendono maggiormente da OSB; 3) la formazione di nuove beta cellule da parte di OSB aumenta in modo sostanziale a fronte di un incremento di apoptosi nei ratti di tipo HIP, modello animale del diabete di tipo 2, rallentando in questo modo il declino della massa beta cellulare. In contrasto, il turnover beta cellulare è ridotto nelle scimmie di tipo STZ (ovvero scimmie trattate con streptozotocin), modello animale del diabete di tipo 1, rispetto alle scimmie non diabetiche di controllo. Inoltre la formazione di nuove beta cellule nelle scimmie di tipo STZ è dovuta in gran parte a OSB. 4) La duplicazione di beta cellule esistenti è il meccanismo primario che regola l’espansione della massa beta cellulare nell’infanzia, mentre in età adulta OSB è responsabile del mantenimento della massa beta cellulare a fronte di un incremento dell’apoptosi; 5) la massa ed il turnover beta cellulari aumentano in risposta all’obesità negli individui; 6) le stime ottenute per l’età media di una beta cellula (1-2 mesi nei ratti, 2-5 mesi nelle scimmie e 6 mesi-2 anni negli individui) e per la sua aspettativa di vita media (1-3 mesi nei ratti, 2-5 mesi nelle scimmie e 6 mesi-2 anni negli individui) sono potenzialmente compatibili con la rigenerazione endogena della massa beta cellulare nel diabete, qualora fosse possibile alterare il turnover beta cellulare in modo terapeutico. I modelli presentati forniscono per la prima volta informazioni sulla presenza di sorgenti di beta cellule diverse dalla duplicazione di beta cellule e stime dell’età e dell’aspettativa di vita medie di una beta cellula nei ratti, nelle scimmie e negli individui. I risultati ottenuti hanno un impatto dal punto di vista clinico considerando che: a) l’origine delle beta cellule è causa di accesi dibattiti: alcuni ricercatori suggeriscono come origine principale la duplicazione delle beta cellule esistenti, altri la formazione di nuove beta cellule da svariate sorgenti diverse dalla duplicazione beta cellulare; b) il ripristino del controllo glicemico sia nel diabete di tipo 1 sia in quello di tipo 2 attraverso una rigenerazione interna potrebbe essere una potenziale strategia alternativa al trapianto di pancreas, dati il numero insufficiente di pancreas disponibili per il trapianto e i rischi di una prolungata terapia immunosoppressiva; c) l’unico approccio sperimentale che consente di identificare le sorgenti di nuove beta cellule diverse dalla duplicazione beta cellulare è la cell-lineage tracing, non disponibile negli studi clinici. In aggiunta i risultati incoraggiano: a) studi futuri sul turnover beta cellulare nei pazienti diabetici; b) lo sviluppo di esperimenti ad hoc atti ad identificare OSB; c) la messa a punto di esperimenti e modelli matematici in grado di stabilire le modalità ed i tempi richiesti per la rigenerazione endogena della massa beta cellulare.
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Ullsten, Sara. "The Impact of Pancreatic Islet Vascular Heterogeneity on Beta Cell Function and Disease." Doctoral thesis, Uppsala universitet, Institutionen för medicinsk cellbiologi, 2017. http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-330805.
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Diabetes Mellitus is a group of complex and heterogeneous metabolic disorders characterized by hyperglycemia. Even though the condition has been extensively studied, its causes and complex pathologies are still not fully understood. The occurring damage to the pancreatic islets is strikingly heterogeneous. In type 1 diabetes, the insulin producing beta cells are all destroyed within some islets, and similarly in type 2 diabetes, some islets may be severely affected by amyloid. At the same time other islets, in the near vicinity of the ones that are affected by disease, may appear fully normal in both diseases. Little is known about this heterogeneity in susceptibility to disease between pancreatic islets. This thesis examines the physiological and pathophysiological characteristics of islet subpopulations. Two subpopulations of islets were studied; one constituting highly vascularized islets with superior beta cell functionality, and one of low-oxygenated islets with low metabolic activity. The highly functional islets were found to be more susceptible to cellular stress both in vitro and in vivo, and developed more islet amyloid when metabolically challenged. Highly functional islets preferentially had a direct venous drainage, facilitating the distribution of islet hormones to the peripheral tissues. Further, these islets had an increased capacity for insulin secretion at low glucose levels, a response that was observed abolished in patients with recent onset type 1 diabetes. The second investigated islet subpopulation, low-oxygenated islets, was found to be an over time stable subpopulation of islets with low vascular density and beta cell proliferation. In summary, two subpopulations of islets can be identified in the pancreas based on dissimilarities in vascular support and blood flow. These subpopulations appear to have different physiological functions of importance for the maintenance of glucose homeostasis. However, they also seem to differ in vulnerability, and a preferential death of the highly functional islets may accelerate the progression of both type 1 and type 2 diabetes.
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Yuan, Yuan. "Small-Molecule Modulators of Pancreatic Ductal Cells: Histone Methyltransferases and \(\beta\)-Cell Transdifferentiation." Thesis, Harvard University, 2012. http://dissertations.umi.com/gsas.harvard:10637.
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Small molecules are important not only for treating human diseases but also for studying disease-related biological processes. This dissertation focuses on the effects of small molecules on pancreatic ductal adenocarcinoma cells. Here, I describe the discovery of two small-molecule tool compounds and their applications for interrogating the biological processes related to two distinct diseases in the human pancreas. First, BRD4770 was identified as a histone methyltransferase inhibitor through a target-based biochemical approach, and was used as a probe to study the function of methyltransferases in cancer cells. Second, BRD7552 was discovered as an inducer of Pdx1 using a cell-based phenotypic screening approach, and was used to induce the expression of Pdx1, a master regulatory transcription factor required for \(\beta\)-cell transdifferentiation. This compound is particularly interesting for the study of type-1 diabetes (T1D). The histone methyltransferase G9a catalyzes methylation of lysine 9 on histone H3, a modification linked to aberrant silencing of tumor-suppressor genes. The second chapter describes the collaborative effort leading to the identification of BRD4770 as a probe to study the function of G9a in human pancreatic cancer cells. BRD4770 induces cellular senescence and inhibits both anchorage-dependent and -independent proliferation in PANC-1 cell line, presumably mediated through ATM-pathway activation. Chapter three describes the study of a natural product gossypol, which significantly enhances the BRD4770 cytotoxicity in p53-mutant cells through autophagic cell death. The up-regulation of BNIP3 might be responsible for the synergistic cell death, suggesting that G9a inhibition may help overcome drug resistance in certain cancer cells. Ectopic overexpression of Pdx1, Ngn3, and MafA can reprogram pancreatic exocrine cells to insulin-producing cells in mice, which sheds light on a new avenue for treating T1D. The fourth chapter focuses on a gene expression-based assay using quantitative real-time PCR technique to screen >60,000 compounds for induction of one or more of these three transcription factors. A novel compound BRD7552 which up-regulated Pdx1 mRNA and protein levels in PANC-1 cells was identified. BRD7552 induces changes of the epigenetic markers within the Pdx1 promoter region consistent with transcriptional activation. Furthermore, BRD7552 partially complements Pdx1 in cell culture, enhancing the expression of insulin induced by the introduction of the three genes in PANC-1 cells. In summary, the central theme of my dissertation is to identify novel bioactive small molecules using different screening approaches, as well as to explore their effects in pancreatic ductal cells.Chemistry and Chemical Biology
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Owen, R. A. "The role of transglutaminase in stimulus-secretion coupling in the pancreatic #beta#-cell." Thesis, Nottingham Trent University, 1988. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.384731.
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Zehri, Aqib Hyder. "Differential Effects of Pulsatile vs. Chronic Hyperglycemia on Fetal Pancreatic Beta Cell Population." Thesis, The University of Arizona, 2011. http://hdl.handle.net/10150/145129.
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Germanos, Mark. "A Cab for Insulin: Characterising Cab45 in Pancreatic β-Cells." Thesis, The University of Sydney, 2022. https://hdl.handle.net/2123/29871.
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The pancreatic β-cell is purpose-built for the production and secretion of insulin, the only hormone that can remove glucose from the blood. Insulin is kept inside miniature membrane-bound storage compartments known as secretory granules, and these specialised organelles can readily fuse with the plasma membrane upon cellular stimulation to release insulin into the circulation. At this point in time, the mechanisms that govern the generation of insulin secretory granules are poorly defined. This is pertinent to understanding the molecular pathogenesis of type 2 diabetes, where a reduced population and/or compromised quality of secretory granules can contribute to the failure of β-cells to release enough insulin for the control of circulating glucose. Recently, a soluble 45 kDa Ca2+-binding protein that localises to the Golgi apparatus, Cab45, has emerged as a promising candidate that could contribute to the initial stages of secretory granule biogenesis. In the constitutively secreting HeLa cell line, Cab45 sorts secretory cargo into vesicles from the Golgi apparatus. Its mode of action can be described as an aggregative mechanism that has preference for certain cargo, providing a means by which large volumes of protein can be directed through specific trafficking routes. This is a suitable mechanism for a cell such as the β-cell, which dedicates much of its energy to synthesising insulin and other secretory cargoes, but manages to consistently traffic this cargo with high accuracy. Presented here is the first investigation of Cab45 function in cell models that participate in regulated secretion. Cab45 expression is invariably correlated to β-cell function in both humans and in animal models of type 2 diabetes, and its experimental depletion impairs the function of all models that have been studied.
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Yeo, Wendy Wai Yeng. "Differentiation of skeletal muscle-derived stem cells into beta pancreatic lineage." Thesis, Montpellier, 2015. http://www.theses.fr/2015MONTS091.
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Le diabète de type 1 (DT1) est caractérisé par des niveaux élevés de glucose en raison de la destruction des cellules ß pancréatiques sécrétrices d'insuline. Cependant, les thérapies actuelles de remplacement des cellules bêta du pancréas impliquant la transplantation d'îlots pancréatiques sont techniquement difficiles et limitées par la disponibilité de don d'organes. Bien que les cellules souches embryonnaires et les cellules souches pluripotentes induites soient intensément étudiées, aucune de ces deux sources de cellules souches ne peut être utilisée directement sans le risque de développement de tumeurs. Les cellules souches dérivées du muscle squelettique (MDSC) sont une source de cellules alternative intéressante car elles sont multi-potentes et peuvent donc se différencier vers plusieurs lignages cellulaires tels que des cellules cardiaques à battement autonome “pacemaker-like” et des cellules neuronales. Par conséquent, nous avons émis l'hypothèse qu'elles pourraient se différencier en lignées de type pancréatique. Les objectifs de cette étude étaient donc d'étudier le potentiel des MDSC (1) à se différencier in vitro en cellules beta pancréatiques exprimant l'insuline et (2) à se différentier in vivo dans le pancréas et ainsi réduire l'hyperglycémie chez la souris modèle d'un diabète de type 1. Dans cette étude, les MDSC de muscle de souris ont été isolées via une série de passages des cellules les moins adhérentes en culture. Les cellules souches ainsi isolées peuvent adhérer sur une couche de cellules de types fibroblastes ou sur une matrice extra-cellulaire de type laminine pour ensuite se différentier in vitro ou bien être utilisées comme cellules souches MDSC non-adhérentes et non différentiées pour les études in vivo. In vitro, les MDSC peuvent se différencier spontanément en agrégats de cellules formant des îlots et exprimant des marqueurs de cellules bêta identifiés par immunofluorescence et analyse “PCR transcription inverse”. Ceci a été confirmé par immuno-analyse montrant l'expression des protéines nécessaires à la fonction des cellules ß, comme Nkx6.1, MafA et Glut2. Les MDSC différenciées en aggrégats cellulaires de type îlots pancréatiques montrent une sécrétion d'insuline en réponse au glucose in vitro. Cependant, dans des modèles murins de DT1 induit par la streptozotocine, l'injection intra-péritonéale des MDSC n'a pas permis de rétablir chez les souris diabétiques une normoglycémie du glucose sanguin en dépit d'un engreffement des MDSC dans les tissus pancréatiques. Ces données montrent que les MDSC peuvent constituer une source de cellules souches alternative intéressante pour le traitement du diabèteType 1 Diabetes (T1D) is characterized by high and poorly controlled glucose levels due to the destruction of insulin-secreting pancreatic ß-cells. However, current ß-cell replacement therapies, involving pancreas and pancreatic islet transplantation are technically demanding and limited by donor availability. While embryonic stem cells and induced pluripotent stem cells are intensely investigated, neither can be used due to safety issues. Skeletal muscle-derived stem cells (MDSC) are an attractive alternative cell source as they have the potential to undergo multilineage differentiation into beating pacemaker-like cells and neuronal cells. Hence, it is hypothesised that they can differentiate into pancreatic lineages. This led to the goals of this study, which were (1) to investigate the potential of MDSC to differentiate into mature insulin expressing cells in vitro and (2) to reduce hyperglycemia in mouse model type 1 diabetes. In this study, MDSC were isolated from mouse via a serial pre-plating based on the adhesive characteristics of cultured cells, in which the cells of interest adhered to plates at a later time for in vitro differentiation, while the non-adherence undifferentiated MDSC were used for in vivo study. The MDSC were found to spontaneously differentiate into islet-like aggregates and expressed ß-cell markers in vitro, as determined by immunofluorescence and reverse transcription PCR analyses. This was further confirmed by immunoblotting analysis showing expression of proteins required for ß-cell function, such as Nkx6.1, MafA and Glut2. The differentiation of MDSC into islet-like clusters demonstrated glucose responsiveness in vitro. In streptozotocin-induced T1D mouse models, intraperitoneal injection of the undifferentiated MDSC did not restore the blood glucose levels of the diabetic mice to normoglycemia despite successful engraftment of MDSC into the pancreatic tissues. Taken together, these data show that MDSC may serve as an alternative source of stem cells for the treatment of diabetes
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Hartman, Matthew G. "The roles of ATF3 in stress-regulated signal transduction and cell death in pancreatic beta-cells." Connect to resource, 2005. http://rave.ohiolink.edu/etdc/view?acc%5Fnum=osu1116425282.
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Thesis (Ph. D.)--Ohio State University, 2005.Title from first page of PDF file. Document formatted into pages; contains xxiv, 185 p.; also includes graphics. Includes bibliographical references (p. 164-185). Available online via OhioLINK's ETD Center
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Preston, Amanda Miriam Clinical School St Vincent's Hospital Faculty of Medicine UNSW. "The role of endoplasmic reticulum stress in beta-cell lipoapoptosis." Publisher:University of New South Wales. Clinical School - St Vincent's Hospital, 2008. http://handle.unsw.edu.au/1959.4/41231.
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Beta-cell failure is a key step in the progression from metabolic disorder to overt type 2 diabetes (T2D). This failure is characterised by both secretory defects and loss of beta-cell mass, the latter most likely through increases in the rate of apoptosis. Although the mechanisms underlying these beta-cell defects are unclear, evidence suggests that chronic exposure of beta-cells to elevated fatty acid (FA) plays a role in disease development in genetically susceptible individuals. Furthermore, it has been postulated that endoplasmic reticulum (ER) stress signalling pathways (the unfolded protein response; UPR) play a role in FA-induced beta-cell dysfunction. The broad aim of this thesis was to explore the nature of these relationships. Experiments detailed in this thesis demonstrate that MIN6 beta-cells mount a comprehensive ER stress response with exposure to elevated saturated fatty acid palmitate, but not the unsaturated fatty acid, oleate, within the low elevated physiological range. This response was time-dependent and involved both transcriptional and translational changes in UPR transducers and targets. The differential activation of ER stress in MIN6 beta-cells by saturated, but not unsaturated FA species may represent a mechanism of differential beta-cell death described in many studies with these FA. Furthermore, these experiments describe defects in ER to Golgi trafficking with chronic palmitate treatment, but not oleate or thapsigagin treatment, identifying this as a potential mechanism by which palmitate treatment induces ER stress. Moreover, these studies have shown the relevance to ER stress to a whole body model of T2D by demonstrating UPR activation in the islets of the db/db mouse. In conclusion, studies detailed in this thesis have demonstrated that ER stress occurs in in vitro and in vivo models of beta-cell lipotoxicity and apoptosis. In addition, these studies have identified defects in ER to Golgi trafficking as a mechanism by which palmitate treatment induces ER stress. These studies highlight the importance of ER stress in the development of T2D.
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Mitchell, Ryan. "The effects of type 2 diabetes associated risk loci on pancreatic beta cell function." Thesis, Imperial College London, 2016. http://hdl.handle.net/10044/1/39040.
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The pancreatic islets of Langerhans play a fundamental role in the stabilisation of blood glucose levels. Pancreatic islets are spherical structures composed of multiple cell types, with each individual cell type secreting a peptide hormone, such as insulin and glucagon, which regulates whole body glucose homoeostasis. Defective hormone secretion from islet cells is a hallmark of certain metabolic diseases, including type 2 diabetes mellitus (T2D). The most abundant islet cell type is the pancreatic β-cell, a specialised cell type that secretes the hypoglycaemic hormone insulin in response to raised glucose levels. Alterations in both β-cell mass and function are the causative factor for the development of T2D. Both genetic and environmental factors are known to underlie the decline in β-cell function typical of T2D. Specifically, genome wide association studies (GWAS) have identified over 100 genomic loci that are associated with T2D risk. Among these loci, variants that lie within/near ADCY5, SLC30A8 and PAX6 show associations with both T2D and abnormal glycaemic parameters typical of a diabetic phenotype. Therefore, the aims of this thesis were to understand how variants associated with T2D manifest at the level of the pancreatic islet. The expression of these genes was therefore manipulated through the generation of tissue-specific transgenic and knockout mice and by RNA interference in human tissue. Reducing the expression of ADCY5, encoding adenylate cyclase five, in human islet tissue reduced glucose-stimulated insulin secretion. This was accompanied by impairments in the metabolic and non-metabolic parameters that govern the secretory response of islets to glucose and other secretagogue. Deleting ZnT8, a β-cell zinc transporter and the gene product of the Slc30a8 gene, in the mouse β-cell significantly impaired the ability of these animals to mount effective responses to glucose. Interestingly, the reverse phenotype, i.e. improved glucose tolerance, was seen in animal models that overexpress ZnT8 in the β-cell. Finally, deletion of Pax6 in the adult mouse resulted in a drastic diabetic phenotype accompanied with changes in the cellular architecture of the islet and alterations in β-cell glucose signalling. Therefore, ADCY5, SLC30A8/ZnT8 and PAX6 gene variants likely negatively impact upon β-cell mass and function leading to a diabetic phenotype. Furthermore, these genes highlight distinct pathways, intrinsic to the pancreatic β-cell, which could be therapeutically targeted in the treatment of T2D.
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Heister, Paula Maria. "The role of two pore channels (TPCs) in pancreatic beta cell stimulus-secretion coupling." Thesis, University of Oxford, 2012. http://ora.ox.ac.uk/objects/uuid:4bed27d2-e7e4-49ff-8168-aa02b6f9b613.
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This thesis presents an investigation into the role of the recently identified two pore channels (TPCs) in β-cell stimulus-secretion coupling. TPCs are the receptors for calcium mobilising messenger nicotinic acid adenine dinucleotide phosphate (NAADP) located in the membrane of acidic intracellular calcium stores. It is proposed that they are responsible for the ATP-sensitive potassium channel (Katp channel) independent pathway of stimulus-secretion coupling; and that this pathway is not subordinate to the KAT? channel dependent pathway; but an alternative explanation of stimulus-secretion coupling in its own right. The first section of this thesis presents a characterisation of sub-membrane cal- cium signals observed in primary mouse β-cells in response to glucose and the membrane-permeable acetoxymethyl ester form of NAADP (NAADP-AM) using the non-ratiometric fluorescent calcium indicator fluo-4 and total internal reflection (TIRF) microscopy. These are compared to global cytosolic calcium changes observed with epifluorescence microscopy. Factors affecting the shape and time course of re- sponses are investigated, and pharmacological tools used to provide evidence for the role of intracellular calcium release from acidic stores mediated by NAADP. Having characterised the calcium responses of β-cells using TIRF; the second part of the thesis examines the effects of knocking out TPC2 (single KO), or both TPC1 and TPC2 (DKO) on these responses; after an initial assessment of pancreatic islet and β-cell morphology using electron microscopy. Gender differences in β-cell responses to glucose and NAADP are assessed in both wild type and knockout animals. Finally, the third section presents the discovery of elementary calcium release events in pancreatic β-cells. The current project visualises what are likely the triggering events for the global calcium signals examined in sections one and two. They take the form of localised calcium release in response to NAADP-AM and glucose; akin to sparks and puffs observed by stimulation with cADPR and IP3. Optical quantal analysis demonstrates the quantal nature of the events and estimates the size of the unitary calcium release unit (CRU) for NAADP. .
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Naghiloo, Sheyda. "Proteomic Pathways to Type 2 Diabetes in the Pancreatic Islet." Thesis, The University of Sydney, 2022. https://hdl.handle.net/2123/29172.
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Unveiling proteomic changes that occur through stages of pathogenesis can provide unparalleled insights into the mechanisms underpinning disease. Using fluorescence-activated cell sorting (FACS) and liquid chromatography mass spectrometry/mass spectrometry (LC-MS/MS), I aimed to characterise the deep proteome of both whole islets and individual islet cell types (alpha, beta, gamma, delta and epsilon) from mice and humans in variable states of health and disease. Acquisition of individual islet cell type proteomes will serve as a window of enquiry into islet cell regulation and intra-cellular communication providing insight into how these factors affect systemic glucose regulation. A panel of diverse mice were phenotypically characterised to observe their responses to diet or genetically induced metabolic stress. Characterisation of the islet proteomes of these mice revealed background strain as the major determinant of islet capacity to maintain function under increased metabolic stress. Isolation of the individual islet cell types using FACS requires the targeting of specific markers for each cell type. Currently, specific surface markers for several islet cell types are not known and/or antibodies are not available, and the alternative of using internal markers requires fixation. Standardised methods of protein fixation have a variable extent of modifications, for example paraformaldehyde, and hence are incompatible with peptide identification after MS analysis. I have overcome these obstacles by using a reversible crosslinker, dithiobis(succinimidyl propionate), which specifically targets primary amines in proteins. Upon reversal of the crosslinker, it leaves modifications of a standard size, which can be incorporated into the peptide identification analysis as a protein modification. In doing so, internal markers specific to each islet cell type can be utilized for isolation and subsequent proteomic analysis, whilst retaining greater than 96% protein identification compared to non-fixed cells. Preliminary MS analysis of isolated human β-cells revealed 4,561 proteins representing the largest human β-cell proteome to date.
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Hartman, Matthew George. "The roles of ATF3 in stress-regulated signal transduction and cell death in pancreatic beta-cells." The Ohio State University, 2005. http://rave.ohiolink.edu/etdc/view?acc_num=osu1116425282.
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Kahve, A. "Biophysical and biochemical effects and distribution of fatty acids in pancreatic beta cells and microvascular endothelial cells." Thesis, University of Exeter, 2019. http://hdl.handle.net/10871/36684.
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The incidences of obesity and type 2 diabetes and their complications are increasing globally. The presence of elevated circulating free fatty acids has been associated with the initial dysfunction of pancreatic beta cells and microvascular endothelial cells followed later by their demise. The aim of this thesis was to investigate the mechanisms by which demise occurs, and how it may be prevented. Palmitate, a saturated fatty acid, caused cell death in both INS-1 beta cells and HCMec/D3 microvascular cells, whereas the unsaturated fatty acid oleic acid did not cause cell death, and also protected against palmitate-induced toxicity. Etomoxir, the mitochondrial CPT1 inhibitor did not rescue INS-1 or HCMec/D3 cells from palmitate-induced toxicity suggesting that palmitate-induced toxicity does not occur via entry into the mitochondria. Cells were exposed to 2-bromopalmitate, a non-metabolisable fatty acid used to reduce the pool of cytoplasmic CoA, to determine whether palmitate-induced toxicity might be mediated by its ability to be activated. Pre-incubation with 2-bromopalmitate in INS-1 cells significantly prevented palmitate-induced cell death. These data suggest that the activation of palmitate with CoA might mediate cell death. Cell cycle analysis found that neither oleic acid nor palmitate caused an increase or decrease in cell proliferation in both INS-1 and HCMec/D3 cells. The data suggest that the mechanism of oleic acid-induced cytoprotection might not be via a pro-proliferative mechanism. INS-1 cells were imaged using spontaneous Raman microspectroscopy after 24-hour exposure to esterified and non-esterified fatty acids. Uni- and multi-variate analysis and spectral decomposition were carried out using a methodology optimised and validated which is presented in this thesis. The aim was to quantify changes, if any, in lipid disposition: distribution, intensity (as a measure of concentration) and composition after exogenous exposure to these fatty acids. Exposure to 0.125 mM palmitate showed a significant decrease in the percentage of lipid within the cells and a corresponding increase in the intensity of this lipid. This suggests that palmitate, alone, might be shuttled into lipid droplets. This was not observed when the cells were exposed to oleic acid, whereby an increase in the intensity of lipid was observed even though no significant change was observed in the percentage of lipid within the cells. When palmitate and oleic acid were combined, the composition of the lipid droplets changed such that the levels of palmitate decreased and the levels of oleic acid increased. These data suggest that oleic acid does not shuttle palmitate into lipid droplets. These data do not support the hypothesis that oleic acid protects against palmitate-induced cytotoxicity by shuttling palmitate into lipid droplets. The methyl esters of palmitate and oleic acid were employed to determine whether they would affect lipid disposition. No change in lipid distribution or intensity was observed when the cells were exposed to these fatty acids, validating the requirement for the free carboxyl oxygen for the covalent binding to glycerol for the formation of lipid droplets. These data also suggest that INS-1 cells cannot de-esterify esterified fatty acids.
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Onyango, David J. "The effects of the adipocyte-secreted proteins resistin and visfatin on the pancreatic beta-cell." Thesis, University of Wolverhampton, 2009. http://hdl.handle.net/2436/89148.
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Adipose tissue secreted proteins (adipokines) have been proposed to form a link between obesity and type 2 diabetes (T2D). Resistin and visfatin are two adipokines which have been previously suggested as having roles in the pancreatic islet. The aim of this study was therefore to investigate the regulatory role of the adipokines resistin and visfatin in the pancreatic beta-cell. In order to do this, pancreatic β-cell lines from rat (BRIN-BD11) and mouse (βTC-6) were used to study the effect of exogenous incubation with physiological and pathological concentrations of resistin and visfatin on diverse elements of beta-cell biology including cell viability, gene expression and insulin secretion. In addition to this the expression levels of these two adipokines was also measured in the beta-cell. PCR array analysis showed that resistin and visfatin treatment resulted in significant changes in the expression of key beta-cell specific genes. Interestingly, both resistin and visfatin are highly expressed in the beta-cells. This suggests that the roles of these adipokines are not confined to adipose tissue but also in other endocrine organs. Resistin treatment significantly increased viability of the beta-cells at physiological concentrations however there was no increase with the elevated pathological concentrations. Resistin at elevated concentrations decreased insulin receptor expression in the beta-cells however there was no significant effect at lower concentrations. Both physiological and elevated resistin concentrations did not have any effect on glucose stimulated insulin secretion. Incubation of visfatin induced phosphorylation of insulin receptor and the intracellular signalling MAPK, ERK1/2. Visfatin treatment at 200ng/ml also significantly increased insulin secretion. These effects were replicated by incubation of beta-cells with the product of visfatin’s enzymatic action, nicotinamide mononucleotide and were reversed by visfatin inhibitor FK866. Visfatin treatment at low concentrations did not have any effect on cell viability however the elevated concentrations resulted in a decline. These data indicate that both resistin and visfatin potentially play important roles in beta-cell function and viability and that they form a significant link between adipose tissue and the pancreatic islet in type 2 diabetes.
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Turbitt, Julie Michelle. "The role of taurine in the regulation of insulin secretion and pancreatic beta-cell function." Thesis, University of Ulster, 2005. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.422896.
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Hamamatsu, Keita. "Establishment of non-invasive quantification of pancreatic beta cell mass in mice using SPECT/CT imaging with ¹¹¹In-labeled exendin-4 and its application to evaluation of diabetes treatment effects on pancreatic beta cell mass." Kyoto University, 2020. http://hdl.handle.net/2433/253199.
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Karlsson, Ella. "Studies of neuropeptides in pancreatic beta cell function with special emphasis on islet amyloid polypeptide (IAPP)." Doctoral thesis, Uppsala University, Department of Medical Cell Biology, 2000. http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-560.
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The presence of protein amyloid in pancreas and its association to diabetes was first described 100 years ago in 1901, but was not identified as Islet Amyloid Polypeptide (IAPP) until 1986. The aim of the present work was to determine the role of the beta cell hormone, IAPP, in normal pancreatic islet physiology and during early disturbances of islet function.
Intra-islet peptides, i.e. chromogranin peptides and an extra-islet peptide, i.e. leptin, were studied to identify possible endogenous regulators of IAPP and insulin secretion. Chromogranin-B, but not chromogranin-A or pancreastatin, had the ability to inhibit islet IAPP and insulin release, suggesting that chromogranin-B may serve as an autocrine regulator of IAPP and insulin secretion.
Leptin had a more potent effect on IAPP secretion than on insulin secretion, which was dissociated from effects on islet glucose metabolism. Glucose oxidation rates were increased at physiological leptin concentrations, whereas higher leptin concentrations showed an inhibitory effect and chronically high leptin concentrations had no effect.
Female NOD mice were studied to investigate the release of IAPP in the progression to type 1 diabetes. The release of IAPP was lower than that of insulin from immune cell infiltrated islets, indicating preferential insulin release during the early course of the disease.
IAPP is expressed at an early embryonic stage. The effect of IAPP on cell proliferation in neonatal rat islets was studied in the search for a physiological role of IAPP. IAPP concentrations of (1-1000) nM stimulated neonatal islet cell proliferation mostly in beta cells and to a lesser extent in alpha cells. IAPP did not have any marked effect on the islet cell death frequency. These data indicate a role for IAPP as a potential regulator of beta cell proliferation in neonatal pancreatic islet.
It is concluded that IAPP may be involved in regulation of pancreatic beta cell function both in fetal and adult life.
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Tatsuoka, Hisato. "Single-cell Transcriptome Analysis Dissects the Replicating Process of Pancreatic Beta Cells in Partial Pancreatectomy Model." Doctoral thesis, Kyoto University, 2021. http://hdl.handle.net/2433/263543.
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Åkerblom, Björn. "Frk/Shb Signalling in Pancreatic Beta-cells : Roles in Islet Function, Beta-cell Development and Survival as Implicated in Mouse Knockout Models." Doctoral thesis, Uppsala universitet, Institutionen för medicinsk cellbiologi, 2009. http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-89348.
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The adaptor protein Shb and the non-receptor tyrosine kinase Frk have been implicated in intracellular signalling in insulin-producing beta cells. In this thesis, knockout mice are used to further elucidate the role of Shb and Frk for beta cell number, cytokine-induced cell death, and glucose homeostasis. In addition, the effect of Shb deficiency upon tumour growth is studied in a mouse model of endogenous tumourigenesis. Previously, overexpression of Frk has been associated with increased beta cell replication, and increased susceptibility to cytokine induced beta cell destruction. To test whether Frk has a non-redundant role in regulating beta cell mass, beta cell number in Frk-/- mice was assessed at different stages of life. The results showed that Frk is involved in regulating beta cell number during embryonal and early postnatal life, but is probably redundant in the adult. An earlier study had suggested that Shb participates in cytokine-induced beta cell death, a model of autoimmune diabetes. To test this further, Shb-/- islets were exposed to cytokines, or to an ER-stress inducing agent. Shb knockout islets exhibited decreased cell death, and this effect appeared to be independent of NO, JNK, p38 MAP kinase, FAK and c-Abl, but may involve an augmented induction of Hsp70. Furthermore, glucose homeostasis in Shb-/- mice was impaired, with elevated basal blood sugar concentration and reduced glucose-induced insulin secretion. Previously Shb deficient mice had showed an impaired ability to sustain growth of implanted tumour cells, due to reduced angiogenesis. Tumour growth and angiogenesis were here assessed in an inheritable tumour model. Shb deficient mice exhibited fewer tumours, and reduced vessel density in small tumours, indicating impaired angiogenesis. However, a few large tumours developed in Shb-/- mice, suggesting that tumours can escape the angiogenic restriction caused by the absence of Shb.
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Herrero, Rodríguez Laura. "Implication of Long-Chain Fatty Acids in Glucose-Induced Insulin Secretion in the Pancreatic Beta-Cell." Doctoral thesis, Universitat de Barcelona, 2004. http://hdl.handle.net/10803/2999.
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INTRODUCTION Carnitine palmitoyltransferase I, which is expressed in the pancreas as the liver isoform (LCPTI), catalyzes the rate-limiting step in the transport of fatty acids into the mitochondria for their oxidation. To directly examine whether the availability of long-chain fatty acyl-CoA affects the regulation of insulin secretion in the Beta-cell, we infected INS(832/13) cells and rat islets with an adenovirus encoding a mutant form of LCPTI (Ad-LCPTI M593S) that is insensitive to its inhibitor malonyl-CoA. C75 is described as a potential drug for treatment of obesity and type 2 diabetes. First known as a synthetic inhibitor of fatty acid synthase, it has been also described as an activator of CPTI, increasing peripheral energy utilization and fatty acid oxidation in mice. To further investigate the C75/CPTI interaction, we have characterized the effects of C75 on CPTI in vitro and in vivo.OBJECTIVES 1) Study of the malonyl-CoA/CPTI interaction in the pancreatic Beta-cell and its involvement in glucose-stimulated insulin secretion (GSIS). 2) Construction of an INS stable cell line overexpressing LCPTI wt and LCPTI M593S. 3) Determine the effect of C75 on the CPTI activity and palmitate oxidation in pancreatic Beta-cells.
RESULTS. In Ad-LCPTI M593S infected INS(832/13) cells LCPTI activity increased six-fold. This was associated with enhanced fatty acid oxidation, at any glucose concentration, and a 60% suppression of GSIS. In isolated rat islets in which LCPTI M593S was overexpressed, GSIS decreased 40%. At high glucose concentration, overexpression of LCPTI M593S reduced partitioning of exogenous palmitate into lipid esterification products, and decreased PKC activation. Moreover, LCPTI M593S expression impaired KATP channel-independent GSIS in INS(832/13) cells.
INS-1 stable clones of LCPTIwt and LCPTImut were constructed, however none of them resulted in an increase in LCPTI protein expression compared to endogenous LCPTI nor in CPTI activity. Therefore, slight basal overexpression of LCPTI could probably be toxic for the cells, as a result of which only those cells that do not contain the LCPTI plasmids survived throughout cell passages.
When INS(823/13) cells are incubated with C75, CPTI activity is inhibited, as is fatty acid oxidation. In vivo, a single intraperitoneal injection of C75 to mice produces a short-term inhibition of CPTI activity in mitochondria from liver and pancreas.
DISCUSSION. The results with LCPTImut provide direct support for the hypothesis proposing that the malonyl-CoA/CPTI interaction is a component of a metabolic signalling network that controls insulin secretion. Overall, the findings with C75 provide compelling evidence that the drug is a potent inhibitor of CPTI.
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Miani, MICHELA. "The cross-talk between endoplasmatic reticulum stress and cytokines in pancreatic beta cell inflammation and apoptosis." Doctoral thesis, Universite Libre de Bruxelles, 2013. http://hdl.handle.net/2013/ULB-DIPOT:oai:dipot.ulb.ac.be:2013/209418.
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La prévalence de l’obésité et du diabète de type 1 (DT1) s’accroit dans le monde à une vitesse alarmante. L’augmentation du poids corporel, de la quantité d’acides gras libres (AGL) circulant et de la résistance à l’insuline peut induire un stress du réticulum endoplasmique (RE) dans les cellules beta du pancréas, ce qui pourrait favoriser l’inflammation. Afin de tester cette hypothèse, nous avons exposé des cellules beta à un léger stress chronique du RE induit par de l’acide ciclopiazonique (ACP) ou l’AGL palmitate et les avons ensuite traitées avec une faible dose d’interleukine 1β (IL-1β) ou de facteur de nécrose tumorale (TNF-α). L’addition d’IL- 1β, mais pas de TNF-α, a conduit à l’augmentation de la production de marqueurs pro-inflammatoires et de chimiokines. Cette différence de résultat en fonction de l’exposition à l’IL-1β ou au TNF-α peut-être au moins partiellement expliquée par un usage différentiel du complexe de la kinase IκB (IKK) et de la voie du facteur nucléaire κ-B (NF-κB), menant à terme à une activation plus élevée par l’IL-1β en comparaison à TNF-α. L’analyse des branches de la réponse de la protéine dépliée impliquées a révélé que la voie de l’Inositol-requiring enzyme 1 (IRE1) / X-box binding protein 1 spliced (XBP1s) est le médiateur clé dans le couplage avec la réponse inflammatoire déclenchée par l’IL-1β. En effet, le knockdown d’IRE1 ou de XBP1 a permis d’éviter l’exacerbation de l’activité du promoteur NF-κB et l’expression des gènes cibles de NF-κB. Les mécanismes impliqués dans la régulation XBP1-dépendante de la réponse pro-inflammatoire sont partiellement dépendant de la modulation de la Forkhead box O1 (FoxO1). Le stress du RE et l’IL-1β participent aussi à un autre évènement crucial dans le développement du DT1, à savoir la mort progressive des cellules beta, qui est également exacerbée par la combinaison ACP + IL-1β. Contrairement à l’inflammation, la voie IRE1/XBP1 n’est pas impliquée dans l’apoptose cellulaire induite par la combinaison d’ACP et d’IL-1β. Dans ce contexte, nous suggérons que le devenir de la cellule est décidé par la balance entre les protéines Bcl-2 anti-apoptotiques et pro-apoptotiques. Ainsi, nous avons montré que le gène A1 associé à Bcl-2 (A1) est négativement régulé par le stress du RE tandis que l’IL-1β active la protéine BH3-only Bim, aboutissant à une apoptose accrue des cellules beta. En conclusion, nos données suggèrent que le stress du RE est un facteur sensibilisation pour l’induction de l’inflammation des ilots pancréatiques et de l’apoptose des cellules beta. Ceci pourrait fournir une explication mécanistique à l’augmentation parallèle observée de l’obésité infantile et de la fréquence du DT1.\Doctorat en Sciences biomédicales et pharmaceutiques
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Du, Xiaoyu. "PLAGL1/ZAC, a transient neonatal diabetes mellitus locus gene, in pancreatic beta-cell development and function." Thesis, McGill University, 2011. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=96741.
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Studies on congenital disorders of the pancreas have contributed to the identification of genes that are critical in beta-cell development and function. Transient neonatal diabetes mellitus (TNDM) is a rare congenital disorder of the pancreas. It involves severe insulin deficiency at birth that reverses over weeks or months but may relapse with diabetes in later life. PLAGL1 (pleiomorphic adenoma gene-like 1, also known as ZAC, zinc finger protein that regulates apoptosis and cell cycle arrest, and LOT1, Lost On Transformation 1) is one of the two possible genes at the TNDM locus and the multiple functions of PLAGL1/ZAC strongly suggest it as the causative gene of TNDM. We hypothesized that double dose expression of PLAGL1/ZAC impairs both beta-cell development and function. To test this hypothesis, we began with the study of ZAC ontogeny. In developing human pancreas, ZAC is expressed with considerable specificity in differentiated beta-cells, and its expression decreases dramatically from the second trimester to adult. These results imply a role of ZAC in a critical time window in beta-cell development, supporting its role in TNDM and explaining the transient nature of the TNDM. In vitro, effects of ZAC overexpression on beta-cell function was observed in INS-1 cells by using tetracycline regulatable system. Overexpression of ZAC inhibits glucose-stimulated insulin exocytosis and proinsulin biosynthesis. Glucose was found to be able to downregulate Lot1/Zac1 expression in INS-1 cells and mouse islets. These data suggest ZAC as a negative regulator in some glucose-regulated pathways whose abnormally high level impairs beta-cell function that explains the relapse of diabetes in TNDM. Finally, a gene expression profile study of INS-1 cells with or without induced ZAC expression identified STC1, IGF1R, SNAP25, GRP78, and P58IPK as possible targets of ZAC that mediate beta-cell dysfunctions. CRABP2, strongly upregulated by ZAC, together with G0S2, GADD45alpha, and FHL2, may mediate ZAC function in both normal beta-cell development and TNDM pathophysiology. In general, these studies suggest that tightly controlled expression level of ZAC is critical for beta-cell development and function. It provides strong evidence that ZAC overexpression causes TNDM. In addition, two previously unstudied genes, STC1 and CRABP2, are suggested to play potential important roles in beta-cell function and development.Les études portant sur les désordres congénitaux du pancréas ont contribué à l'identification de gènes critiques pour le développement et la fonction des cellules bêta. Le diabète néonatal transitoire (DNNT) est une maladie héréditaire du pancréas. Elle est caractérisée par une déficience sévère en insuline à la naissance qui disparaît après quelques semaines/mois mais pouvant réapparaître plus tard au cours de la vie. PLAGL1 (pleiomorphic adenoma gene-like 1, aussi connu comme ZAC, zinc finger protein that regulates apoptosis and cell cycle arrest, et LOT1, Lost On Transformation 1) est un des deux gènes dans la région critique du DNNT et ses multiples fonctions en font le candidat causatif le plus probable. Notre hypothèse est que sa surexpression compromet la fonction et le développement des cellules bêta. Notre étude ontgénique de ZAC dans le pancréas en développement, démontre que ZAC était exprimé avec une spécificité considérable dans les cellules bêta, expression qui diminuait à partir du second trimestre. Ces résultats supportent l'existence d'une fenêtre temporelle critique pour la fonction de ZAC dans le développement des cellules bêta, compatible avec la nature transitoire du DNNT. In vitro, les effets de la surexpression de ZAC ont été observés dans les cellules bêta INS-1 en utilisant un système d'expréssion inductible par la tétracycline. L'exocytose glucose-dépendante de l'insuline et la biosynthèse de la proinsuline sont diminués par la surexpression de ZAC. Le glucose pouvait diminuer l'expression de Lot1/Zac1 dans les INS-1 et dans les îlots murins, une observation qui propose ZAC comme un régulateur négatif de voies métaboliques régulées par le glucose dont les niveaux anormalement élevés affectent la fonction des cellules bêta. Le profile d'expression génique sur les INS-1 après induction de ZAC a identifié STC1, IGF1R, SNAP25, GRP78 et P58IPK comme cibles potentielles de ZAC intervenant dans les dysfonctionnements des cellules bêta. CRABP2, qui est fortement augmenté, tout comme G0S2, GADD45alpha et FHL2, pourrait servir de médiateur de ZAC dans le développement des cellules bêta et dans le DNNT. Ces études indiquent qu'une expression minutieusement contrôlés de ZAC est critiques pour le développement et la fonction des cellules bêta et que sa surexpression peut causer le DNNT. Finalement, un rôle de STC1 et CRABP2 dans la fonction et le développement des cellules bêta est suggéré.
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Patterson, Steven. "Homocysteine and the effects of other amino thiols on pancreatic beta cell function and insulin secretion." Thesis, University of Ulster, 2003. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.398994.
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Watson, David. "Mechanisms of pancreatic beta cell death induced by cytokines and by reactive oxygen and nitrogen species." Thesis, Keele University, 2009. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.502942.
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O'Sullivan-Murphy, Bryan M. "Contribution of WFS1 to Pancreatic Beta Cell Survival and Adaptive Alterations in WFS1 Deficiency: A Dissertation." eScholarship@UMMS, 2012. https://escholarship.umassmed.edu/gsbs_diss/590.
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Diabetes mellitus comprises a cohort of genetic and metabolic diseases which are characterized by the hallmark symptom of hyperglycemia. Diabetic subtypes are based on their pathogenetic origins: the most prevalent subtypes are the autoimmune-mediated type 1 diabetes mellitus (T1DM) and the metabolic disease of type 2 diabetes mellitus (T2DM). Genetic factors are major contributory aspects to diabetes development, particularly in T2DM where there is close to 80% concordance rates between monozygotic twins. However, the functional state of the pancreatic β cell is of paramount importance to the development of diabetes. Perturbations that lead to β cell dysfunction impair insulin production and secretion and precede diabetes onset.
The endoplasmic reticulum (ER) is a subcellular organelle network of tubes and cisternae with multifaceted roles in cellular metabolism. Alterations to ER function such as those begotten by the accumulation of misfolded and unfolded ER client proteins upset the ER homeostatic balance, leading to a condition termed ER stress. Subsequent sensing of ER stress by three ER transmembrane proteins, initiates an adaptive reaction to alleviate ER stress: this is known as the unfolded protein response (UPR). Divergent cascades of the UPR attempt to mitigate ER stress and restore ER homeostasis: Failing that, the UPR initiates pro-apoptotic pathways. The demand of insulin production on the β cell necessitates the presence of a highly functional ER. However, the consequence of dependence on the ER for insulin synthesis and secretion portends disaster for the functional state of the β cell. Disturbances to the ER that elicit ER stress and UPR activation causes β cell dysfunction and may lead to apoptosis. There are numerous well-characterized models of ER stress-mediated diabetes, including genetic mutations in UPR transducers and insulin. Recently, polymorphisms in Wolfram syndrome 1 (WFS1), an ER transmembrane protein involved in the UPR, were suggested to contribute to T2DM risk.
In this thesis, one of the highlighted WFS1 polymorphism, H611R, was examined to identify its contribution to β cell function and viability, and hence, diabetes risk. It was revealed that augmentation of WFS1 expression increased insulin secretion and cellular content. In addition, WFS1 protected β cells against ER stress-mediated dysfunction, with a more pronounced effect in the WFS1-R611 protective allele. Subsequent gene expression analysis identified netrin-1 as a WFS1-induced survival factor.
As a contributory factor to diabetes progression, ER stress and UPR are potential drug and biomarker targets. In this dissertation, a novel UPR-regulating microRNA (miRNA) family was uncovered in ER stressed, WFS1-deficient islets. These miRNAs, the miR-29 family, are induced in WFS1 -/- islets as a possible adaptive alteration to chronic ER stress conditions, and indirectly decreases the expression of UPR transducers, while directly targeting downstream ER stress-related pro-apoptotic factors. Collectively, this work extends the function of WFS1 as a protective factor in the pancreatic β cell through the induction of netrin-1 signaling. Additionally, it further strengthens the role of miRNA as regulatory members of the UPR which contribute to cell survival.
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Brown, James. "Regulation of uncoupling protein-2 expression, cell function and viability in pancreatic islets and beta-cells." Thesis, University of Wolverhampton, 2005. https://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.419783.
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