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Role of Cell-cell Interactions and Palmitate on β-cells Function
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
2014 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

The islets of Langerhans secrets insulin in response to fluctuations of blood glucose level and efficient secretion requires extensive intra-islet communication. Secretory failure from islets is one of the hallmark in progression of type 2 diabetes.  Changes in islet structure and high levels of saturated free fatty acids may contribute to this failure. The aim of this thesis is to study the role of cell-cell interactions and palmitate on β-cells functions.

To address the role of cell-cell interactions on β-cells functions MIN6 cells were cultured as monolayers and as pseudoislets. Glucose stimulated insulin secretion was higher in pseudoislets compared to monolayers. Transcript levels of mitochondrial metabolism as well glucose oxidation rate was higher in pseudoislets. Insulin receptor substrate-1 (IRS-1) phosphorylation was altered when cells were grown as pseudoislets. Proteins expression levels related to glycolysis, cellular connections and translational regulations were up-regulated in pseudoislets. We propose the superior capacity of pseudoislets compared to monolayers depend on metabolism, cell coupling, gene translation, protein turnover and differential IRS-1 phosphorylation.

To address the role of palmitate on β-cells human islets were cultured in palmitate. Long term palmitate treatment decreased insulin secretion which is associated with up-regulation of suppressor of cytokine signaling-2 (SOCS2) and protein inhibitor of activated STAT-1 (PIAS1). Up-regulation of SOCS2 decreased phosphorylation of Akt at site T308, whereas PIAS1 decreased protein level of ATP- citrate lyase (ACLY) and ATP synthase subunit B (ATP5B). We propose long term palmitate treatment reduces phosphatidylinositol 3-kinase (PI3K) activity, attenuates formation of acetyl-CoA and decreases ATP synthesis which may aggravate β-cells dysfunction.  

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2014. , 42 p.
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Medicine, ISSN 1651-6206 ; 1025
Keyword [en]
Metabolism, PI3K, Pseudoislets, Human islets, SOCS, PIAS
National Category
Cell and Molecular Biology
Research subject
Biology with specialization in Molecular Biology
Identifiers
URN: urn:nbn:se:uu:diva-230841ISBN: 978-91-554-9021-8 (print)OAI: oai:DiVA.org:uu-230841DiVA: diva2:742080
Public defence
2014-10-17, B41, BMC, Husargatan 3, Uppsala, 09:15 (English)
Opponent
Supervisors
Available from: 2014-09-25 Created: 2014-08-30 Last updated: 2015-01-23
List of papers
1. Functional differences between aggregated and dispersed insulin-producing cells
Open this publication in new window or tab >>Functional differences between aggregated and dispersed insulin-producing cells
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2013 (English)In: Diabetologia, ISSN 0012-186X, E-ISSN 1432-0428, Vol. 56, no 7, 1557-1568 p.Article in journal (Refereed) Published
Abstract [en]

Beta cells situated in the islet of Langerhans respond more vigorously to glucose than do dissociated beta cells. Mechanisms for this discrepancy were studied by comparing insulin-producing MIN6 cells aggregated into pseudoislets with MIN6 monolayer cells and mouse and human islets. MIN6 monolayers, pseudoislets and mouse and human islets were exposed to glucose, alpha-ketoisocaproic acid (KIC), pyruvate, KIC plus glutamine and the phosphatidylinositol 3-kinase (PI3K) inhibitors LY294002 or wortmannin. Insulin secretion (ELISA), cytoplasmic Ca2+ concentration ([Ca2+](c); microfluorometry), glucose oxidation (radiolabelling), the expression of genes involved in mitochondrial metabolism (PCR) and the phosphorylation of insulin receptor signalling proteins (western blotting) were measured. Insulin secretory responses to glucose, pyruvate, KIC and glutamine were higher in pseudoislets than monolayers and comparable to those of human islets. Glucose oxidation and genes for mitochondrial metabolism were upregulated in pseudoislets compared with single cells and monolayers, respectively. Phosphorylation at the inhibitory S636/639 site of IRS-1 was significantly higher in monolayers and dispersed human and mouse cells than pseudoislets and intact human and mouse islets. PI3K inhibition only slightly attenuated glucose-stimulated insulin secretion from monolayers, but substantially reduced that from pseudoislets and human and mouse islets without suppressing the glucose-induced [Ca2+](c) response. We propose that islet architecture is critical for proper beta cell mitochondrial metabolism and IRS-1 signalling, and that PI3K regulates insulin secretion at a step distal to the elevation of [Ca2+](c).

Keyword
Beta cell, Ca2+, Insulin secretion, IRS-1, Islets, Mitochondrial metabolism, PI3-kinase
National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:uu:diva-203520 (URN)10.1007/s00125-013-2903-3 (DOI)000319881300013 ()
Available from: 2013-07-16 Created: 2013-07-15 Last updated: 2017-12-06Bibliographically approved
2. Signaling in Insulin-Secreting MIN6 Pseudoislets and Monolayer Cells
Open this publication in new window or tab >>Signaling in Insulin-Secreting MIN6 Pseudoislets and Monolayer Cells
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2013 (English)In: Journal of Proteome Research, ISSN 1535-3893, E-ISSN 1535-3907, Vol. 12, no 12, 5954-5962 p.Article in journal (Refereed) Published
Abstract [en]

Cell cell interactions are of fundamental importance for cellular function. In islets of Langerhans, which control blood glucose levels by secreting insulin in response to the blood . glucose concentration, the secretory response of intact islets is c higher than that of insulin-producing beta-cells not arranged in the islet architecture. The objective was to define mechanisms by which cellular performance is enhanced when cells are arranged in a) three-dimensional space. The task was addressed by making a c comprehensive analysis based on protein expression patterns " generated from insulin-secreting MIN6 cells grown as islet-like c clusters, so-called pseudoislets, and in monolayers. After culture, glucose-stimulated insulin secretion (GSIS) was measured from monolayers and pseudoislets. GSIS rose 6-fold in pseudoislets but only 3-fold in monolayers when the glucose concentration was increased from 2 to 20 mmol/L. Proteins from pseudoislets and monolayers were extracted and analyzed by liquid-chromatography mass spectrometry, and differentially expressed proteins were mapped onto KEGG pathways. Protein profiling identified 1576 proteins, which were common to pseudoislets and monolayers. When mapped onto KEGG pathways, 11 highly enriched pathways were identified. On the basis of differences in expression of proteins belonging to the pathways in pseudoislets and monolayers, predictions of differential pathway activation were performed. Mechanisms enhancing insulin secretory capacity of the beta-cell, when situated in the islet, include pathways regulating glucose metabolism, cell interaction, and translational regulation.

Keyword
glucose-stimulated insulin secretion (GSIS), beta-cells, MIN6 cells, pseudoislets
National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:uu:diva-215288 (URN)10.1021/pr400864w (DOI)000328231300053 ()
Available from: 2014-01-13 Created: 2014-01-13 Last updated: 2017-12-06Bibliographically approved
3. GLP-1 recovers impaired insulin secretion from human islets treated with palmitate via down-regulation of SOCS2
Open this publication in new window or tab >>GLP-1 recovers impaired insulin secretion from human islets treated with palmitate via down-regulation of SOCS2
(English)Manuscript (preprint) (Other academic)
National Category
Cell and Molecular Biology
Identifiers
urn:nbn:se:uu:diva-230839 (URN)
Available from: 2014-08-30 Created: 2014-08-30 Last updated: 2015-01-23
4. Role of PIAS1 in palmitate-mediated beta-cell dysfunction
Open this publication in new window or tab >>Role of PIAS1 in palmitate-mediated beta-cell dysfunction
(English)Manuscript (preprint) (Other academic)
National Category
Cell and Molecular Biology
Identifiers
urn:nbn:se:uu:diva-230840 (URN)
Available from: 2014-08-30 Created: 2014-08-30 Last updated: 2015-10-23

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