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PLoS One. 2012;7(5):e33023. doi: 10.1371/journal.pone.0033023. Epub 2012 May 14.

A novel high-throughput assay for islet respiration reveals uncoupling of rodent and human islets.

Author information

1
Department of Medicine, Boston University School of Medicine, Boston, Massachusetts, United States of America.

Erratum in

  • PLoS One. 2013;8(12). doi:10.1371/annotation/f21efac9-2906-4a2f-ad22-befcb7714dd0.

Abstract

BACKGROUND:

The pancreatic beta cell is unique in its response to nutrient by increased fuel oxidation. Recent studies have demonstrated that oxygen consumption rate (OCR) may be a valuable predictor of islet quality and long term nutrient responsiveness. To date, high-throughput and user-friendly assays for islet respiration are lacking. The aim of this study was to develop such an assay and to examine bioenergetic efficiency of rodent and human islets.

METHODOLOGY/PRINCIPAL FINDINGS:

The XF24 respirometer platform was adapted to islets by the development of a 24-well plate specifically designed to confine islets. The islet plate generated data with low inter-well variability and enabled stable measurement of oxygen consumption for hours. The F1F0 ATP synthase blocker oligomycin was used to assess uncoupling while rotenone together with myxothiazol/antimycin was used to measure the level of non-mitochondrial respiration. The use of oligomycin in islets was validated by reversing its effect in the presence of the uncoupler FCCP. Respiratory leak averaged to 59% and 49% of basal OCR in islets from C57Bl6/J and FVB/N mice, respectively. In comparison, respiratory leak of INS-1 cells and C2C12 myotubes was measured to 38% and 23% respectively. Islets from a cohort of human donors showed a respiratory leak of 38%, significantly lower than mouse islets.

CONCLUSIONS/SIGNIFICANCE:

The assay for islet respiration presented here provides a novel tool that can be used to study islet mitochondrial function in a relatively high-throughput manner. The data obtained in this study shows that rodent islets are less bioenergetically efficient than human islets as well as INS1 cells.

PMID:
22606219
PMCID:
PMC3351473
DOI:
10.1371/journal.pone.0033023
[Indexed for MEDLINE]
Free PMC Article

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