Rat brain endothelial cells are a target of manganese toxicity

Abstract

Manganese (Mn) is an essential trace metal, however exposure to high Mn levels can result in neurodegenerative changes resembling Parkinson´s disease (PD). Information on Mn´s effects on endothelial cells of the blood-brain barrier (BBB) is lacking. Accordingly, we tested the hypothesis that BBB endothelial cells are a primary target for Mn-induced neurotoxicity. The studies were conducted in an in vitro BBB model of immortalized rat brain endothelial (RBE4) cells. ROS production was determined by F2-Isoprostane (F2-IsoPs) measurement. The relationship between Mn toxicity and redox status was investigated upon intracellular glutathione (GSH) depletion with diethylmaleate (DEM) or L-buthionine sulfoximine (BSO). Mn exposure (200 or 800 µM MnCl2 or MnSO4) for 4 or 24h led to significant decrease in cell viability vs. controls. DEM or BSO pre-treatment led to further enhancement in cytotoxicity vs. exposure to Mn alone, with more pronounced cell death after 24h DEM pre-treatment. F2-IsoPs levels in cells exposed to MnCl2 (200 or 800 µM), were significantly increased after 4h and remained elevated 24h after exposure compared with controls. Consistent with the effects on cell viability and F2-IsoPs, treatment with MnCl2 (200 or 800 µM) was also associated with a significant decrease in membrane potential. This effect was more pronounced in cells exposed to DEM plus MnCl2 vs. cells exposed to Mn alone. We conclude that Mn induces direct injury to mitochondria in RBE4 cells. The ensuing impairment in energy metabolism and redox status may modify the restrictive properties of the BBB compromising its function.

Comparison Between 5-Aminosalicylic Acid (5-ASA) and Para-Aminosalicylic Acid (4-PAS) as Potential Protectors Against Mn-Induced Neurotoxicity

“…The present study confirms that in cells exposed to MnCl 2 alone, the increase in LDH release is concordant with the decrease in MTT levels, corroborating previous reports (Marreilha dos Santos et al 2008, dos Santos et al 2010). 5-ASA significantly decreased Mn cytotoxicity in RBE4 cells exposed to MnCl 2 for 24 h.…”

Effect of chronic glutathione deficiency on the behavioral phenotype of Gclm(−/−) knockout mice

“…It is also possible that humans with mutations that deplete GSH might be at higher risk to exposure from neurotoxic agents that increase ROS. For example, Gclm(−/−) mice are more susceptible to the pollutant 2,3,7,8-tetrachlorodibenzo- p -dioxin (Chen et al, 2011), and rats having decreased GSH are more susceptible to manganese neurotoxicity (dos Santos et al, 2010). …”

Silymarin, a natural antioxidant, protects cerebral cortex against manganese-induced neurotoxicity in adult rats

“…Cell dissociation was achieved with 0.05% trypsin-0.02% EDTA. Briefly, cells were seeded on 24-well culture plates in medium at an approximate density of 10 5 cells/cm 2 and, after 24 h stabilization, neuronal cells were co-cultured with medium containing various concentrations of MnCl 2 (200 and 800 lM) and silymarin (10, 50 and 100 lM) for 24 h. The concentration of MnCl 2 was selected based on previously reported cytotoxic levels in cultured cells (dos Santos et al 2010). For stock solution, MnCl 2 was dissolved in MilliQ Plus sterilized water at the concentration of 800 mM and silymarin was dissolved in dimethylsulfoxide (DMSO) to obtain a 100 mM.…”