ABSTRACT
Oxidative stress-induced cell damage is involved in many neurological diseases. Interferon regulatory factor 6 (IRF6), a member of the IRF family of transcription factors, is required for the differentiation of skin, breast epithelium, and oral epithelium. However, the regulation and function of IRF6 in central nervous system remain unknown. This study aimed to investigate the role of IRF6 in hydrogen peroxide (H2O2)-induced oxidative neuronal injury in HT22 mouse hippocampal cells. Treatment with H2O2 significantly increased the expression of IRF6 at both mRNA and protein levels, and knockdown of IRF6 using specific small interfering RNA reduced H2O2-induced cytotoxicity, as evidenced by increased cell viability and decreased apoptosis. Knockdown of IRF6 attenuated intracellular reactive oxygen species (ROS) generation and lipid peroxidation, and also preserved endogenous antioxidant enzyme activities. The inhibitory effect of IRF6 knockdown on mitochondrial dysfunction was demonstrated by reduced mitochondrial oxidative level, preserved mitochondrial membrane potential (MMP) and ATP generation, as well as attenuated mitochondrial swelling. In addition, down-regulation of IRF6 inhibited the activation of mitochondrial apoptotic factors, whereas IRF6 knockdown together with caspase inhibitors had no extra effect on cell viability and LDH release. These results suggest that knockdown of IRF6 has protective effects against H2O2-induced oxidative stress by reducing ROS accumulation and apoptosis, and these protective effects are dependent on preservation of mitochondrial function.
Subject(s)
Hydrogen Peroxide/pharmacology , Interferon Regulatory Factors/metabolism , Membrane Potential, Mitochondrial/drug effects , Mitochondria/drug effects , Oxidative Stress/drug effects , Animals , Apoptosis/drug effects , Cell Line, Tumor , Cell Survival/drug effects , Cytochromes c/metabolism , Down-Regulation , Gene Knockdown Techniques/methods , Interferon Regulatory Factors/genetics , Lipid Peroxidation/drug effects , Mice , Mitochondria/metabolism , RNA, Small Interfering/metabolism , Reactive Oxygen Species/metabolismABSTRACT
The blood-brain barrier (BBB) is formed by brain endothelial cells, and decreased BBB integrity contributes to vasogenic cerebral edema and increased mortality after stroke. In the present study, we investigated the protective effect of perampanel, an orally active noncompetitive AMPA receptor antagonist, on BBB permeability in an in vitro ischemia model in murine brain endothelial cells (mBECs). The results showed that perampanel significantly attenuated oxygen glucose deprivation (OGD)-induced loss of cell viability, release of lactate dehydrogenase, and apoptotic cell death in a dose-dependent manner. Perampanel treatment did not alter the expression and surface distribution of various glutamate receptors. Furthermore, the results of calcium imaging showed that perampanel had no effect on OGD-induced increase in intracellular Ca(2+) concentrations. Treatment with perampanel markedly reduced the paracellular permeability of mBECs after OGD in different time points, as measured by transepithelial electrical resistance assay. In addition, the expression of claudin-5 at protein level, but not at mRNA level, was increased by perampanel treatment after OGD. Knockdown of claudin-5 partially prevented perampanel-induced protection in cell viability and BBB integrity in OGD-injured mBECs. These data show that the noncompetitive AMPA receptor antagonist perampanel affords protection against ischemic stroke through caludin-5 mediated regulation of BBB permeability.
