Research ReportDiphenyleneiodonium protects preoligodendrocytes against endotoxin-activated microglial NADPH oxidase-generated peroxynitrite in a neonatal rat model of periventricular leukomalacia

The contribution of microglial activation to preoligodendroglial (preOL) damage in the central nervous system (CNS) is considered to be one of the principal causes of periventricular leukomalacia (PVL) pathogenesis. The present study explores the effect of diphenyleneiodonium (DPI), a NADPH oxidase (NOX) inhibitor, on protection of preOLs from bacterial lipopolysaccharide (LPS)-induced microglial toxicity in vivo and in vitro. In vitro, preOLs co-cultured with microglia exhibited increased preOL apoptosis, accompanied by overproduction of superoxide anion (O2−) and the formation of peroxynitrite (ONOO−) after LPS exposure. LPS also significantly up-regulated accumulation of activated microglial NOX subunits p67-phox and gp91-phox in the plasma membrane. Diphenyleneiodonium (DPI) (10 μm) was found to significantly attenuate up-regulation of this NOX activity. In vivo, DPI was administered (1 mg/kg/day) by subcutaneous injection for 3 days to two-day-old neonatal Sprague-Dawley rats subjected to intracerebral injection of LPS. Treatment with DPI within 24 h of LPS injection significantly ameliorated white matter injury, decreasing preOL loss, O2− generation, and ONOO− formation, and inhibiting p67-phox, gp91-phox synthesis and p67phox membrane translocation in microglia. These results indicated that LPS-induced preOL apoptosis may have been mediated by microglia-derived ONOO−. DPI prevented this LPS-induced brain injury, most likely by inhibiting ONOO− formation via NOX, thereby preventing preOL loss and immature white matter injury.

► DPI treatment ameliorated LPS-induced cerebral white matter injury. ► DPI inhibited LPS-induced preOL apoptosis in vitro and ameliorated loss of preOLs in vivo. ► DPI inhibited LPS-induced the production of O2− in cultured microglia and in cerebral white matter. ► DPI inhibited LPS-induced NOX activation in cultured microglia and in cerebral white matter. ► DPI decreased LPS-induced the production of peroxynitrite in cultured microglia and inhibited the formation of nitrotyrosine in cerebral white matter.