This study was undertaken to find out the effects of acetylpuerarin on hippocampal neurons and intracellular free calcium in primary culture subjected to oxygen-glucose deprivation/reperfusion. Methods: According to different reperfusion time (1h, 6h, 12h, 24h), three concentrations (1.6 μmol l−1, 0.4 μmol l−1, 0.1 μmol l−1) of acetylpuerarin, and MK-801 (10 μmol l−1), a positive control drug, neurons were randomly divided into 21 groups. Each group was observed by inverted phase contrast microscope; neuron viability was measured by the reduction of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT); intracellular Ca2+ was observed by Fura-2/AM ester through fluorospectrophotometer. Results: The injured neurons were protected and degeneration and necrosis were alleviated in treatment groups of acetylpuerarin and MK-801. Acetylpuerarin increased the neuron viability at high, middle and low concentrations. Fluorescence detection results showed that the calcium concentration in the group treated with acetylpuerarin and MK-801 was lowered in each reperfusion time. Conclusion: Our results demonstrated that acetylpuerarin could protect the hippocampal neurons from ischemia-reperfusion injury in rats by alleviating the morphological damage, increasing neuron viability and decreasing calcium concentration in neuron.

Our present study was performed to investigate whether hydroxyethylpuerarin (HEP) has a neuroprotective effect on brain injury after focal cerebral ischemia/reperfusion by middle cerebral artery occlusion (MCAO) in adult male Wistar rats. Animals were subjected to one hour of middle cerebral artery occlusion and 48 hours of reperfusion with the pretreatment of drugs (HEP 15, 30, 60 mg/kg or nimodipine 0.4 mg/kg i.v.) or vehicle. The behavioral tests were used to evaluate the damage to central nervous system. The percentage of brain infarct area was assessed in the brain slices stained with 2% solution of 2, 3, 5-triphenyl tetrazolium chloride (TTC). The pathologic histological changes were observed by H&E staining and the occurrence of apoptosis was determined by flow cytometry. The results showed that pretreatment with HEP at doses of 15, 30, 60 mg/kg exhibited significant neuroprotective effects on rats against focal cerebral ischemia-reperfusion injury by markedly decreasing neurological deficit scores and the percentage of infarct area, reducing necrosis and apoptosis of neurons. All these findings suggest that HEP might provide neuroprotection against focal cerebral ischemia/reperfusion injury probably through its antioxidant and anti-inflammatory property.

Mounting evidence has suggested that paeonol possesses plenty of pharmacologic actions. Our research is to determine if paeonol can protect cultured rat hippocampal neurons from oxygen-glucose deprivation(OGD)-induced injury and elucidate the underlying mechanism. We cultivated the rat hippocampal neurons as the object of study and then established the model of oxygen-glucose deprivation. Neuronal viability was measured by the reduction of 3-(4,5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT), while intracellular Ca2+ concentration was observed by fluorospectrophotometer. The binding force of N-methyl-D-aspartate (NMDA) receptor was evaluated by liquid scintillation counting. Compared with oxygen-glucose deprivation group, paeonol treatment obviously increased cell survival rate and reduced the activity of the binding force of NMDA receptors, reversing the overload of intracellular Ca2+. These results demonstrate that paeonol protected rat neurons from oxygen-glucose deprivation-induced injury, resulting in alleviating the morphological damage and increasing neuron viability and suggest that paeonol may exhibit its protective effect against oxygen-glucose deprivation-induced injury by targeting on NMDA receptors.

The present study investigated the protective effects of scutellarin on cobalt chloride (CoCl2)- induced apoptosis in PC12 cells. Incubation of PC12 cells with 500 μM CoCl2 for 24 h resulted in significant apoptosis as evaluated by the crystal violet, electron microscopy and flow cytometry assays. The increase of caspase-3 activity, decrease of Bcl-XL expression, phosphorylation of p38 mitogenactivated protein kinase (MAPK) and accumulation of intracellular reactive oxygen species (ROS) were also seen in CoCl2-treated PC12 cells. Scutellarin at 0.1, 1 and 10 μM significantly protected against the apoptotic cell death induced by CoCl2. Scutellarin decreased caspase-3 activity, increased Bcl-XL expression, inhibited p38 phosphorylation and attenuated ROS production. These results demonstrate that scutellarin can protect PC12 cells from cobalt chloride induced apoptosis by scavenging ROS, inhibiting p38 phosphorylation, up-regulating Bcl-XL expression and decreasing caspase-3 activity, and may reduce the cellular damage in pathological conditions associated with hypoxia-mediated neuronal injury.

Microvascular changes in the brain are significant causes of cerebral edema and ischemia injury. A number of studies suggest that angiotensin (Ang) Ⅱ may be involved in the initiation and regulation of processes occurring in brain ischemia. We recently reported that Ang Ⅱ injures brain microvascular endothelial cells (BMEC) partially via stimulating intercellular adhesion molecule-1 (ICAM-1) expression. However, the signaling cascade leading to Ang Ⅱ-induced ICAM-1 expression in BMEC was unclear. The present study tested the hypothesis that Ang Ⅱ induces ICAM-1 expression via an AT1 receptor/nuclear factor-кB (NF-кB) pathway in BMEC. Ang Ⅱ directly stimulated the expression of ICAM-1 mRNA and protein in primary cultured BMEC. Ang Ⅱ treatment also resulted in the degradation of InBa and increase of NF-кB p65 subunit in the nucleus as well as the DNA binding activity of nuclear NF-кB. These effects were abolished by pretreatment with the selective AT1 receptor antagonists, losartan and compound EXP-2528, or losartan plus the AT2 receptor antagonist PD123319, but not by PD123319 alone. Moreover, there were no significant differences between the losartan and losartan plus PD123319 groups. These findings indicate that Ang ⅡⅡ-induced ICAM-1 upregulation in brain microvascular endothelial cells may be mediated via an AT1 receptor/NF-nB pathway.