Extracellular methylglyoxal; the passage across brain endothelial cells and the effect on barrier function

Background Methylglyoxal (MGO), a highly reactive by-product of glycolysis, has been associated with cognitive decline and Alzheimer’s disease, though the mechanistic role of MGO remains unclear. Moreover, conflicting findings exist regarding MGO’s toxicity on the blood-brain barrier (BBB). This study investigated whether MGO can cross the BBB under physiologically relevant conditions and whether MGO affects BBB permeability. Methods Mice were intravenously injected with highly purified home-made MGO or PBS, and MGO concentration was measured at five timepoints in the cerebral cortex up to 4 hours after injection. MGO toxicity was screened on a human brain endothelial cell line (hCMEC/D3) using a live/dead assay prior to the study of selected MGO concentrations and on hiPSC-derived brain microvascular endothelial cells (EECM-BMECs). EECM-BMECs were cultured on Transwell® inserts, and barrier function was assessed by sodium fluorescein permeability and transendothelial passage of 13 C3-MGO quantified by UPLC-MS/MS. Results MGO levels in the mouse cortex did not increase post-injection. MGO was not toxic to hCMEC/D3 cells, and it had no impact on barrier properties of EECM-BMECs. After 1-hour exposure, ∼13% of total 13 C3-MGO was recovered in its free form, and only ∼1% of supplemented MGO was recovered from the abluminal side. Conclusion MGO does not cross the BBB in vivo and does not affect barrier properties of a human in vitro model of the BBB. In vitro MGO passage across the BBB is minimal. These findings suggest that circulating MGO is unlikely to directly affect neuronal function via BBB disruption or enter the brain in its free from.