Lightning Talks 29th Annual Lorne Proteomics Symposium 2024

Characterisation of cysteine redox post-translational modifications upon antioxidant N-propionylglycine treatment in the diabetic heart, identified using quantitative mass spectrometry. (#101)

Molly K Talbot 1 2 , Alexander W Rookyard 1 3 , Desmond K Li 3 4 , Stuart J Cordwell 1 3 5 , Melanie Y White 1 2
  1. Charles Perkins Centre, University of Sydney, Camperdown, NSW, Australia
  2. School of Medical Science, University of Sydney, Camperdown, NSW, Australia
  3. School of Life and Environmental Sciences, University of Sydney, Camperdown, NSW, Australia
  4. Heart Research Institute , Sydney , NSW, Australia
  5. Sydney Mass Spectrometry , University of Sydney, Camperdown, NSW, Australia

Overproduction of reactive oxygen species (ROS) is a key determinant in the pathogenesis of Type 2 Diabetes (T2D) and cardiovascular disease (CVD) linked to obesity, hyperglycaemia, and loss of insulin sensitivity. Cellular redox levels directly reduce antioxidant capacity promoting protein alterations, manifesting as cardiac functional deficiencies. Current pre-clinical evidence reveals the potential for antioxidant therapies to reduce or attenuate ROS derived damage and dysfunction (Ribeiro et al., 2018) and restore redox balance. Large human cohort studies have observed correlations between high antioxidant intake with decreased likelihood of T2D and CVD (Kwon et al.,2023;Wang and Yi, 2022). However, this remains to be translated into therapeutic outcomes in the general population.

 

Using an 8-week rat model of T2D that combines a low-dose streptozotocin (STZ) and high-fat diet to model the T2D phenotype, to create the T2D phenotype, with treatment controls and physiological setting for comparison, hearts are excised to undergo isolated ex vivo perfusion. For this study, we created two subsets, the first received 5-minute perfusion to define the native redox balance; and the second received 40-minute perfusion in the presence of a global antioxidant, N-propionylglycine (MPG; 1mM). In the presence of MPG, contractile performance was maintained in T2D myocardium, contrary to our previous studies which showed a gradual decline in function. To determine how MPG was affording such protection, we performed quantitative redox proteomics, using thiol-disulfide exchange to specifically identify and quantify cysteine-containing peptides using tandem mass spectrometry (MS/MS). To date we have identified that MPG treatment rescues reversibly modified cysteines in endogenous antioxidant mechanisms, mitochondrial proteins, and contractile proteins. Conversely, MPG showed little capacity to protect against redox changes to proteins regulating metabolic pathways. Metabolomics confirmed these observations with dysregulation of TCA intermediates unable to be salvaged by MPG-treatment. This suggests that redox changes in metabolic pathways occur during the development of the T2D phenotype (prior to ex vivo), while contractile dysfunction during ex vivo perfusion is specifically driven by redox alterations in the mitochondria and contractile apparatus.

  1. Kwon, Y. J., Park, H. M., & Lee, J. H. (2023). Inverse Association between Oxidative Balance Score and Incident Type 2 Diabetes Mellitus. Nutrients, 15(11), 2497. https://doi.org/10.3390/nu15112497
  2. Ribeiro, J. A., Benfeito, S., Cagide, F., Teixeira, J., Oliveira, P. J., Borges, F., Silva, A. F., & Pereira, C. M. (2018). Electrochemical Behavior of a Mitochondria-Targeted Antioxidant at an Interface between Two Immiscible Electrolyte Solutions: An Alternative Approach to Study Lipophilicity. Analytical chemistry, 90(13), 7989–7996.
  3. Wang, L., & Yi, Z. (2022). Association of the Composite dietary antioxidant index with all-cause and cardiovascular mortality: A prospective cohort study. Frontiers in cardiovascular medicine, 9, 993930. https://doi.org/10.3389/fcvm.2022.993930