A mitochondria-targeted mass spectrometry probe to detect glyoxals: implications for diabetes

• Published in: Free radical biology & medicine 2014-02, Vol.67 (100), p.437-450
• Main Authors: Pun, Pamela Boon Li, Logan, Angela, Darley-Usmar, Victor, Chacko, Balu, Johnson, Michelle S., Huang, Guang W., Rogatti, Sebastian, Prime, Tracy A., Methner, Carmen, Krieg, Thomas, Fearnley, Ian M., Larsen, Lesley, Larsen, David S., Menger, Katja E., Collins, Yvonne, James, Andrew M., Kumar, G.D. Kishore, Hartley, Richard C., Smith, Robin A.J., Murphy, Michael P.
• Format: Article
• Language: English
• Subjects: Animals; Cattle; Cell Line; Chromatography, Liquid; Diabetes Mellitus, Type 1 - diagnosis; Diabetes Mellitus, Type 1 - metabolism; Diabetes Mellitus, Type 1 - pathology; Disease Models, Animal; Endothelial Cells - metabolism; Endothelial Cells - pathology; Exomarker; Free radicals; Glyoxal; Glyoxal - analysis; Glyoxal - metabolism; Hyperglycemia; Hyperglycemia - diagnosis; Hyperglycemia - metabolism; Hyperglycemia - pathology; Methylglyoxal; Mice; Mitochondria; Mitochondria, Liver - metabolism; Mitochondria, Liver - pathology; MitoG; Molecular Probes - chemical synthesis; Myoblasts - metabolism; Myoblasts - pathology; Organophosphorus Compounds - chemistry; Original Contribution; Oxidative Stress; Phenylenediamines - chemistry; Pyruvaldehyde - analysis; Pyruvaldehyde - metabolism; Rats; Tandem Mass Spectrometry
• ISSN: 0891-5849; 1873-4596
• DOI: 10.1016/j.freeradbiomed.2013.11.025

The glycation of protein and nucleic acids that occurs as a consequence of hyperglycemia disrupts cell function and contributes to many pathologies, including those associated with diabetes and aging. Intracellular glycation occurs after the generation of the reactive 1,2-dicarbonyls methylglyoxal and glyoxal, and disruption of mitochondrial function is associated with hyperglycemia. However, the contribution of these reactive dicarbonyls to mitochondrial damage in pathology is unclear owing to uncertainties about their levels within mitochondria in cells and in vivo. To address this we have developed a mitochondria-targeted reagent (MitoG) designed to assess the levels of mitochondrial dicarbonyls within cells. MitoG comprises a lipophilic triphenylphosphonium cationic function, which directs the molecules to mitochondria within cells, and an o-phenylenediamine moiety that reacts with dicarbonyls to give distinctive and stable products. The extent of accumulation of these diagnostic heterocyclic products can be readily and sensitively quantified by liquid chromatography–tandem mass spectrometry, enabling changes to be determined. Using the MitoG-based analysis we assessed the formation of methylglyoxal and glyoxal in response to hyperglycemia in cells in culture and in the Akita mouse model of diabetes in vivo. These findings indicated that the levels of methylglyoxal and glyoxal within mitochondria increase during hyperglycemia both in cells and in vivo, suggesting that they can contribute to the pathological mitochondrial dysfunction that occurs in diabetes and aging. •A mitochondria-targeted mass spectrometric probe, MitoG, has been developed to measure glyoxal and methylglyoxal.•Using MitoG we show that mitochondrial glyoxal and methylglyoxal can be measured in hyperglycemic cells.•MitoG can also be used in vivo to infer mitochondrial glyoxal and methylglyoxal production in a mouse model of type I diabetes.•These findings suggest that the accumulation of glyoxal and methylglyoxal within mitochondria may contribute to mitochondrial dysfunction in diabetes.