An in vitro study of m-dinitrobenzene toxicity on the cellular components of the blood-brain barrier, astrocytes and endothelial cells.

We have studied the effects of an industrial compound, m-dinitrobenzene (DNB), on cultured astrocytes and brain capillary endothelial cells in an in vitro blood-brain barrier (BBB) model. In single cultures, the threshold DNB concentration that induced cell death, as assessed by morphology and lactose dehydrogenase leakage into the culture medium, was 1 mM for both cell types after 1 day of incubation. In cocultures, astrocytes showed a dose-response curve similar to that obtained in single cultures, while endothelial cells showed an increased sensitivity to DNB cytotoxicity. DNB induced a dose-dependent increase in glucose consumption and lactate production in both cell types in single cultures, although astrocytes appeared to be more sensitive than endothelial cells. The role of oxidative stress was also studied using the reduction of nitroblue tetrazolium as an index of generation of active oxygen species. A dose-dependent increase was observed for both cell types in single cultures, although this could not be prevented by the addition of superoxide dismutase to the culture medium. Addition of 0.3 mM carmustine to the culture medium increased the cytotoxicity of 0.5 mM DNB in both astrocytes and endothelial cells, indicating a role of oxidative stress in DNB-induced damage. Desferrioxamine (20 mm) completely protected endothelial cells from damage by 1 mM DNB, suggesting that hydroxyl radicals mediated at least part of the DNB neurotoxicity. However, astrocytic damage by 2 mM DNB was only partially prevented by desferrioxamine. We conclude that our in vitro model is suitable for studying interactions between astrocytes and endothelial cells in toxicological situations.