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Quercetin is a bioactive flavonoid which abundantly exists in vegetables and fruits. Quercetin exerts a neuroprotective effect against cerebral ischemia. Thioredoxin acts as antioxidant by regulating redox signaling. This study investigated whether quercetin regulates thioredoxin expression in focal cerebral ischemia and glutamate-induced neuronal cell death. Male Sprague Dawley rats (210–230 g) were intraperitoneally injected with vehicle or quercetin (10 mg/kg) 1 h prior to middle cerebral artery occlusion (MCAO). Cerebral cortex was collected 24 h after MCAO. MCAO led to neurological movement deficits, brain edema, and serious histopathological damages in cerebral cortex, and quercetin alleviated these damages following MCAO. We observed the change of thioredoxin expression in MCAO animals with quercetin using proteomic approach, reverse-transcription PCR, and Western blot analyses. Thioredoxin expression decreased in vehicle-treated MCAO animals, while quercetin attenuated this decrease. Moreover, quercetin treatment alleviated the decrease in the number of thioredoxin-positive cells in cerebral cortex of MCAO animals. Furthermore, immunoprecipitation analysis demonstrated that interaction of apoptosis signal-regulating kinase 1 (ASK1) and thioredoxin was decreased in MCAO animals with vehicle, while quercetin prevented MCAO-induced decrease in these binding. In addition, quercetin also alleviated the reduction of cell viability and the decrease in thioredoxin expression in glutamate-treated hippocampal cell line and primary cultures of cortical neurons. However in thioredoxin-silenced cortical neuron, anti-apoptotic effect of quercetin was decreased. Thus, changes of thioredoxin expression by quercetin may contribute to the neuroprotective effect of quercetin in focal cerebral ischemia. Our findings suggest that quercetin mediates its neuroprotective function by regulation of thioredoxin expression and maintenance of interaction between ASK1 and thioredoxin.

Calcium acts as a second messenger that mediates physiologic functions, such as metabolism, cell proliferation, and apoptosis. Hippocalcin is a neuronal calcium sensor protein that regulates intracellular calcium concentration. Moreover, it prevents neuronal cell death from oxidative stress. Quercetin has excellent antioxidant properties and preventative effects. We studied modulation of hippocalcin expression by quercetin treatment in cerebral ischemic injury and glutamate-induced neuronal cell damage. Focal cerebral ischemia was induced by permanent middle cerebral artery occlusion (pMCAO). Male Sprague-Dawley rats were injected with vehicle or quercetin (10 mg/kg) 1 h prior to pMCAO, and cerebral cortical tissues were isolated 24 h after pMCAO. Quercetin improved pMCAO-induced neuronal movement deficit and infarction. pMCAO induced a decrease in hippocalcin expression in the cerebral cortex. However, quercetin treatment attenuated this pMCAO-induced decrease. In cultured hippocampal cells, glutamate excitotoxicity dramatically increased the intracellular calcium concentration, whereas quercetin alleviated intracellular calcium overload. Moreover, Western blot and immunocytochemical studies showed reduction of hippocalcin expression in glutamate-exposed cells. Quercetin prevented this glutamate-induced decrease. Furthermore, caspase-3 expression in hippocalcin siRNA transfection conditions is higher than caspase-3 expression in un-transfection conditions. Quercetin treatment attenuated the increase of caspase-3. Taken together, these results suggest that quercetin exerts a preventative effect through attenuation of intracellular calcium overload and restoration of down-regulated hippocalcin expression during ischemic injury.