Can methylglyoxal be oxidized to CO2 in vascular smooth muscle cells?

Abstract

Miklós Péter Kalapos, Lingyun Wu

Aim: The aim of this study was to investigate the uptake of [2-14C]-methylglyoxal (MG), a glucose and fructose metabolite, and its possible oxidation to carbon dioxide (CO2 ) in cultured vascular smooth muscle cells (A-10). Materials and Methods: Cells were cultured to confluence and then treated with [2-14C]-MG at 37°C in a humidified atmosphere at 95% air and 5% CO2 . Conditioned medium and the wash, used to remove unbound [2-14C]-MG, were combined and referred to as the activity remaining in the medium. Afterwards the dishes were treated with 0.1% Triton X-100, the solutions were removed into eppendorf tubes and spun down. The supernatant was taken and referred to as cytosolic fraction, while the pellet was considered as membrane fraction. Sodium hydroxide was used to capture CO2 . Trypan blue dye uptake by the cells was used to follow cell viability. Measurement of proteins was performed by bicinchoninic acid assay. Results: Cells bound and took up [2-14C]-MG in a concentration-dependent manner. The uptake of [2-14C]-MG was a rapid process since it was detectable in the cells after 5 min of incubation, reaching a plateau by 30 min of incubation. The presence of fetal bovine serum in the incubation medium hampered the uptake of [2-14C]-MG, suggesting an interaction between proteins and the α-oxoaldehyde. After the incubation longer than 60 min a loss of cell bound radioactivity was detected, which was mainly due to CO2 formation from the α-oxoaldehyde. Conclusion: These results suggest that the generation of CO2 from MG may be substantial in cultured cells and that MG may contribute to the energy production below a certain limit. In A-10 cells, this limit proved to be 300 μM, since above this concentration of MG cell damage occurred.