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Proteins altered by elevated levels of palmitate or glucose implicated in impaired glucose-stimulated insulin secretion
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
2009 (English)In: Proteome Science, ISSN 1477-5956, E-ISSN 1477-5956, Vol. 7, 24- p.Article in journal (Refereed) Published
Abstract [en]

Background

Development of type 2 diabetes mellitus (T2DM) is characterized by deranged insulin secretory patterns, where elevated insulin levels at non-stimulatory basal conditions and reduced hormonal levels at stimulatory conditions are major components. To delineate mechanisms responsible for these alterations we cultured INS-1E cells for 48 hours at 20 mM glucose in absence or presence of 0.5 mM palmitate, when stimulatory release was reduced or basal secretion was elevated, respectively.

Results

After culture, cells were protein profiled by SELDI-TOF MS and 2D-PAGE. Differentially expressed proteins were discovered and identified by peptide mass fingerprinting. Complimentary protein profiles were obtained by the two approaches with SELDI-TOF MS being more efficient in separating proteins in the low molecular range and 2D-PAGE in the high molecular range. Identified proteins included alpha glucosidase, calmodulin, gars, glucose-6-phosphate dehydrogenase, heterogenous nuclear ribonucleoprotein A3, lon peptidase, nicotineamide adenine dinucleotide hydrogen (NADH) dehydrogenase, phosphoglycerate kinase, proteasome p45, rab2, pyruvate kinase and t-complex protein. The observed glucose-induced differential protein expression pattern indicates enhanced glucose metabolism, defense against reactive oxygen species, enhanced protein translation, folding and degradation and decreased insulin granular formation and trafficking. Palmitate-induced changes were related to altered exocytosis.

Conclusions

The identified altered proteins indicate mechanism important for deranged b-cell function in T2DM.

Place, publisher, year, edition, pages
2009. Vol. 7, 24- p.
Keyword [en]
insulin release, proteome profiling, SELDI-TOF MS, two-dimensional gel electrophoresis, type 2 diabetes mellitus
National Category
Medical and Health Sciences
Research subject
Medical Cell Biology
Identifiers
URN: urn:nbn:se:uu:diva-98362DOI: 10.1186/1477-5956-7-24ISI: 000269306000001OAI: oai:DiVA.org:uu-98362DiVA: diva2:174251
Available from: 2009-02-19 Created: 2009-02-19 Last updated: 2017-12-13Bibliographically approved
In thesis
1. Glucose, Palmitate and Apolipoprotein CIII-induced Effects on Insulin-Producing β-cells
Open this publication in new window or tab >>Glucose, Palmitate and Apolipoprotein CIII-induced Effects on Insulin-Producing β-cells
2009 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Background and aims: Type 2 diabetes mellitus results from complex interplay between genetic predisposition and environmental factors that together promote impairment of insulin-producing β-cells. Elevated levels of glucose, fatty acid palmitate and apolipoprotein CIII (apoCIII) are implicated in this process. To delineate effects of these factors, the role of enhanced carnitine palmitoyltransferase 1 (CPT1) expression in glucolipotoxic cells, glucose-dependency of the unfolded protein response (UPR) in palmitate-induced apoptosis and activation of mitogen activated protein kinases (MAPKs) in apoCIII-induced apoptosis were evaluated. In addition, protein profiles of β-cell exposed to elevated levels of glucose or palmitate were generated to identify proteins regulated by these nutrients.

Methodology: INS-1E cells were cultured at different glucose concentrations in the absence or presence of palmitate or apoCIII for up to 48 hours. CPT1 was over-expressed with a Tet-ON regulated adenovirus. In cells exposed to apoCIII, inhibitors of MAPKs p38 or ERK1/2 were included during culture. After culture, apoptosis, insulin secretion, expression of UPR-markers and MAPKs and protein profiles were determined.

Results: INS-1E cells exposed to elevated levels of glucose and palmitate showed deranged insulin secretion with increased insulin secretion at non-stimulatory glucose level, enhanced apoptosis and induced expression of UPR-markers. Over-expression of CPT1 reduced basal insulin secretion and attenuated apoptosis. Palmitate-induced apoptosis was accentuated by increasing the culture glucose concentration. Markers of UPR were not modulated by the glucose concentration in INS-1E cell exposed to palmitate, however. ApoCIII-induced apoptosis in INS-1E cells was accompanied by activation of p38 and ERK1/2. Protein profiling of INS-1E cells exposed to elevated levels of glucose or palmitate revealed changes in expression of multiple β-cell proteins implicated in glucose metabolism, defence against reactive oxygen species, protein translation/folding/degradation and insulin granular trafficking.

Conclusions: Over-expression of CPT1 counteracts β-cell glucolipotoxicity. Activation of UPR is not a major determinant for palmitate-induced β-cell apoptosis. ApoCIII-induced β-cell apoptosis involves activation of MAPKs. The identified differentially expressed proteins indicate a central role of altered glucose metabolism and protein synthesis in gluco- and lipotoxic β-cells and may provide specific molecular mechanisms offering new ways of treating the disease.

 

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2009. 48 p.
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Medicine, ISSN 1651-6206 ; 429
National Category
Cell and Molecular Biology
Research subject
Medical Cell Biology
Identifiers
urn:nbn:se:uu:diva-98367 (URN)978-91-554-7440-9 (ISBN)
Public defence
2009-04-03, B22, BMC, Husargatan 3, Uppsala, 13:15 (English)
Opponent
Supervisors
Available from: 2009-03-13 Created: 2009-02-19 Last updated: 2009-05-07Bibliographically approved

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Sol, E-ri Maria

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