7,8-Dihydroxy-4-Methylcoumarin Provides Neuroprotection Against Glutamate-induced Toxicity In HT-22 Cells And Ischemic Brain Injury In Neonatal Rats

Part Ⅰ: Protection and mechanism of 7,8-dihydroxy-4-methylcoumarin on glutamate-induced oxidative stress in HT-22 cellsObjectiveTo investigate the neuroprotective effects of 7,8-dihydroxy-4-methylcoumarin(Dhmc)on the oxidative stress injury induced by glutamate in HT-22 cells, and to investigate the mechanism of the neuroprotective effect of Dhmc.MethodsAfter HT-22 cells incubation for 24 hours,the glutamate(5mmol/L) was added for inducing neuronal injury. Culture medium with different concentrations of Dhmc was incubated for 12 hours. We observed the morphological changes of HT-22 cells under an microscope. Cell viability was detected by MTS assay, and GSH levels and ROS activity were determined by a biochemistry method. The levels of hippocalcin was detected by immunocytochemistry and western blot.ResultDhmc protected against glutamate-induced cell death in HT-22 cells effectively(P<0.05). The neurites of HT-22 cells were decreased after treated with 5 mmol/L glutamate for 12 h and Dhmc(25μmol /L) treatment completely inhibit this morphological changes,and attenuate glutamate-induced ROS production and GSH depletion(P<0.05).and inhibite the glutamate-induced decrease in hippocalcin levels.ConclusionDhmc has neuroprotective effects on glutamate-induced oxidative stress in HT-22 cells. The possible mechanism of Dhmc was related to the decrease of ROS content, to maintain the content of GSH and to inhibite the decrease in hippocalcin levels.Part II: Protection and mechanism of 7,8-dihydroxy-4-methylcoumarin on the cerebral hypoxia /ischemia injury in neonatal ratsObjectiveTo investigate the neuroprotective effects of 7,8-dihydroxy-4-methylcoumarin(Dhmc)on the cerebral hypoxia /ischemia injury in neonatal rats, and to investigate the mechanism of the neuroprotective effect of Dhmc.Methods7-day-old neonates of SD rats were randomly divided into Sham group, Sham+Dhmc group, hypoxia /ischemia(H/I) group and H/I+Dhmc group for setting up the neonatal rat hypoxia /ischemia(H/I) model. To observe the effect of the treatment of Dhmc on the infarct volume of cerebral ischemia in neonatal rats.The levels of hippocalcin was detected by western blot.ResultDhmc pretreatment reduced the infarct volume significantly compared with the control group(P <0.05), and Dhmc treatment still decreased the infarct volume when administered 4 h after cerebral hypoxia/ischemia injury and attenuated the hypoxia/ischemia injury-induced decrease of hippocalcin expression in neonatal rats(P<0.05).ConclusionDhmc has neuroprotective effects on the cerebral hypoxia/ischemia injury in neonatal rats. The possible mechanism of Dhmc was related to inhibite the decrease in hippocalcin levels.The experimental results show that Dhmc has good neuroprotective effect in vitro and in vivo, and it is a kind of drug which has a hope to treat cerebral ischemia injury.