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

Jakob D. Wikstrom, Samuel B. Sereda, Linsey Stiles, Alvaro Elorza, Emma M. Allister, Andy Neilson, David A. Ferrick, Michael B. Wheeler, Orian S. Shirihai

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Resumen

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.

Idioma originalInglés
Número de artículoe33023
PublicaciónPLoS ONE
Volumen7
N.º5
DOI
EstadoPublicada - 14 may 2012

Áreas temáticas de ASJC Scopus

  • Bioquímica, genética y biología molecular (todo)
  • Agricultura y biología (todo)
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    Wikstrom, J. D., Sereda, S. B., Stiles, L., Elorza, A., Allister, E. M., Neilson, A., Ferrick, D. A., Wheeler, M. B., & Shirihai, O. S. (2012). A novel high-throughput assay for islet respiration reveals uncoupling of rodent and human islets. PLoS ONE, 7(5), [e33023]. https://doi.org/10.1371/journal.pone.0033023