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

Abstract

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.

Mitochondria-Targeted Triphenylphosphonium-Based Compounds: Syntheses, Mechanisms of Action, and Therapeutic and Diagnostic Applications

“…337 To evaluate the importance of dicarbonyl-related glycation processes to mitochondrial function, a mitochondria-targeted mass spectrometry probe for glyoxals, MitoG (Chart 48), was developed by linking the TPP + cation with alkoxy-substituted o-phenylenediamine. 338 …”

“…338 The MitoG probe can be used to determine relative mitochondrial levels of methylglyoxal and glyoxal under hyperglycemia. 338 Although the proposed methodology of dicarbonyl detection reports on mitochondrial glyoxal and methylglyoxal exposure, it cannot indicate the source of those dicarbonyls, as they may be produced in the cytosol with subsequent diffusion into mitochondria. The MitoG probe was used to assess the changes in glyoxal and methylglyoxal production in a mouse model of type I diabetes.…”

“…The MitoG probe was used to assess the changes in glyoxal and methylglyoxal production in a mouse model of type I diabetes. 338 …”

Understanding and preventing mitochondrial oxidative damage

“…MitoB was also applied to mammalian systems and it was found that an intravenous injection of MitoB into a mouse enabled sufficient MitoB to be taken up into the heart to assess changes in mitochondrial ROS production during IR injury [37]. Therefore, MitoB can be used as a probe to assess mitochondrial hydrogen peroxide production in vivo , through generation of MitoP as an exomarker, and many other experimental models are currently being interrogated using MitoB and new types of probe designed to assess other aspects of mitochondrial function are also being investigated [38–40]. …”

The emerging theme of redox bioenergetics in health and disease

“…[62][63][64] The best understood of these targeting molecules is the triphenylphosphonium (TPP+) group which can be coupled to a broad range of pharmacophores and redox-sensitive probes. [16,65,66] These molecules have been used both in vivo and in cell culture to probe the mechanisms of redox signaling, and the effects they elicit are generally ascribed to the properties of the functional group and not the carrying molecule. However, it was recently shown that the TPP + group itself can modulate mitochondrial bioenergetics and the effects vary with the linker group used to attach the functional pharmacophore.…”