Impaired anaplerosis and insulin secretion in insulinoma cells caused by small interfering RNA-mediated suppression of pyruvate carboxylase.
Open accessAbstract
Anaplerosis, the synthesis of citric acid cycle intermediates, by pancreatic beta cell mitochondria has been proposed to be as important for insulin secretion as mitochondrial energy production. However, studies designed to lower the rate of anaplerosis in the beta cell have been inconclusive. To test the hypothesis that anaplerosis is important for insulin secretion, we lowered the activity of pyruvate carboxylase (PC), the major enzyme of anaplerosis in the beta cell. Stable transfection of short hairpin RNA was used to generate a number of INS-1 832/13-derived cell lines with various levels of PC enzyme activity that retained normal levels of control enzymes, insulin content, and glucose oxidation. Glucose-induced insulin release was decreased in proportion to the decrease in PC activity. Insulin release in response to pyruvate alone, 2-aminobicyclo[2,2,1]heptane-2-carboxylic acid (BCH) plus glutamine, or methyl succinate plus beta-hydroxybutyrate was also decreased in the PC knockdown cells. Consistent with a block at PC, the most PC-deficient cells showed a metabolic crossover point at PC with increased basal and/or glucose-stimulated pyruvate plus lactate and decreased malate and citrate. In addition, in BCH plus glutamine-stimulated PC knockdown cells, pyruvate plus lactate was increased, whereas citrate was severely decreased, and malate and aspartate were slightly decreased. The incorporation of 14C into lipid from [U-14C]glucose was decreased in the PC knockdown cells. The results confirm the central importance of PC and anaplerosis to generate metabolites from glucose that support insulin secretion and even suggest PC is important for insulin secretion stimulated by noncarbohydrate insulin secretagogues.
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Subsequent recruitment of granules to the plasma membrane (second phase) appears to depend on mitochondrial metabolites produced by anaplerosis.
“Subsequent recruitment of granules to the plasma membrane (second phase) appears to depend on mitochondrial metabolites produced by anaplerosis (238).”
From The role of mitochondria in the pathogenesis of type 2 diabetes (2010) · cites this paper as [238]
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- Authors
- Hasan, N. M., Longacre, M. J., Stoker, S. W., Boonsaen, T., Jitrapakdee, S., Kendrick, M. A., Wallace, J. C., & MacDonald, M. J.
- Affiliations
- University of Wisconsin–Madison (US)
- Mahidol University (TH)
- The University of Adelaide (AU)
- Corresponding author
- MacDonald, Michael J.
- Venue
- Journal of Biological Chemistry
- Publisher
- Elsevier BV
- Published
- 2008-10
- Volume
- 283
- Issue
- 42
- Pages
- 28048-28059
- ISSN
- 0021-9258 (ISSN-L)
- PMC
- PMC2568907
- OpenAlex
- W2089250997
- Cited by
- 88
- References
- 57
- Citations by year
- 2026: 2 · 2025: 3 · 2024: 4 · 2023: 1 · 2022: 2 · 2021: 2 · 2019: 1 · 2018: 4 · 2017: 3 · 2016: 4 · 2015: 5 · 2014: 9 · 2013: 11 · 2012: 5
- FWCI
- 11.27
- Type
- journal article
- Language
- en
- Open access
- hybrid
- doi.org
- Licence
- Locations
- Journal of Biological Chemistry (publishedVersion, cc-by)
- PubMed Central (publishedVersion)
- PubMed (publishedVersion)
- Adelaide Research & Scholarship (AR&S) (University of Adelaide) (submittedVersion)
- Funders
- Mahidol University
- Thailand Research Fund
- National Institutes of Health (DK28348)
- Australian Research Council (DP0346807)
- Faculty of Science, Mahidol University
- National Institute of Diabetes and Digestive and Kidney Diseases (DK 28348, R01DK028348)
- Topics
- Health Sciences › Medicine › Surgery › Pancreatic function and diabetes
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Mitochondrial Function and Pathology
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Metabolism, Diabetes, and Cancer
- Keywords
- anaplerosis, pyruvate carboxylase, insulin secretion, pancreatic beta cells, insulinoma, INS-1 cells, shRNA knockdown, citric acid cycle intermediates, glucose-stimulated insulin secretion, beta-hydroxybutyrate, lactate, mitochondrial metabolism
- MeSH terms
- Insulin Secretion, Animals, Cell Line, Humans, Insulin, Insulinoma, Lipids, Models, Biological, Phosphorylation, Pyruvate Carboxylase, RNA, Messenger, Transfection, RNA, Small Interfering, Insulin-Secreting Cells, Mice, Rats
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