• The Journal of Internal Korean Medicine
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• v.25 no.3
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• pp.461-472
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• 2004

Objectives : For the screening of neuroprotective effects of medicinal herbs, the complex system of animal models suffer some disadvantages in controlling critical parameters such as blood pressure and body temperature. Additionally, application of drugs to the appropriate brain area sometimes is difficult, due to poor permeability though the blood brain barrier, and so potential protective effects might be masked. Methods : Organotypic hippocampal slice culture (OHSC) method has the advantages of being relatively easy to prepare and of maintaining the general structure, including tissue integrity and the connections between cells. Drugs can easily be applied and neuronal damage can easily be quantified by using tissues and culture media. This study demonstrates neuroprotective effects of Puerariae radix (葛根, PR), Salviae miltiorrhizae radix (丹蔘, SR), Rhei rhizoma (大黃, RR), and Bupleuri radix (柴胡, BR). These were screenedand compared to MK-801, antagonist of NMDA receptors, by using OHSC of 1 week-old Sprague-Dawley rats. Oxygen/glucose deprivation (OGD) were conducted in an anaerobic chamber $(85%\;N_2,\;10%\;CO_2\;and\;5%\;H_2)$ in a deoxygenated glucose-free medium for 60 minutes. Water extracts of each herbs were treated to culture media with $5\;{\mu}g/ml$ for 48 hours. Results : Neuronal cell death in the cultures was monitored by densitometric measurements of the cellular uptake of propidium iodide (PI). PI fluorescence images were obtained at 48 hours after the OGD and medicinal herb treatment. Also TUNEL-positive cells in the CAI and DG regions and LDH concentrations in culture media were measured at 48 hours after the OGD. According to measured data, MK-801, PR, SR and BR demonstrated significant neuroprotective effect against excessive neuronal cell death and apoptosis induced by the OGD insult. Especially, PR revealed similar neuroprotective effect to MK-801 and RR demonstrated weak neuroprotective effect. Conclusions : These results suggest that OHSC can be a suitable method for screening of neuroprotective effects of medicinal herbs. (This work was supported by the research program of Dongguk University and Grant 01-PJ9-PG1-01CO03-0003 from Ministry of Health & Welfare.)

• The Journal of Korean Medicine
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• v.24 no.1
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• pp.29-40
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• 2003

Object : This research was performed to investigate the protective effect of Aurantii Immaturus Fructus against ischemic damage using PC12 cells and global ischemia in gerbils. Methods : To observe the protective effect of Aurantii Immaturus Fructus on ischemia damage, viability and changes in activities of superoxide dismutase (SOD), glutathione peroxidase (GPx), catalase and production of malondialdehyde (MDA) were observed after treating PC12 cells with Aurantii Immaturus Fructus during ischemic insult. Gerbils were divided into three groups : a normal group, a 5-min two-vessel occlusion (2VO) group, and an Aurantii Immaturus Fructus administered after 2VO group. The CCAs were occluded by microclip for 5 minutes. Aurantii Immaturus Fructus was administered orally for 7 days after 2VO. The histological analysis was performed at 7 days after the surgery. For histological analysis, the brain tissue was stained with 1% cresyl violet solution. Results : The results showed that 1. Aurantii Immaturus Fructus had a protective effect against ischemia in the CAI area of the gerbil hippocampus 7 days after 5-minute occlusion, 2. In the hypoxia/reperfusion model using PC12 cells, the Aurantii Immaturus Fructus had a protective effect against ischemia in the dose of $0.2{\;}\mu\textrm{g}/ml,{\;}2{\;}\mu\textrm{g}/ml{\;}and{\;}20{\;}\mu\textrm{g}/ml$ 3. Aurantii Immaturus Fructus increased the activities of glutathione peroxidase and catalase, 4. The activity of superoxide dismutase (SOD) was increased by ischemic damage, which might represent self protection. This study suggests that Aurantii Immaturus Fructus has some neuroprotective effect against neuronal damage following cerebral ischemia in vivo with a widely used experimental model of cerebral ischemia in Mongolian gerbils, and it also has protective effects on a hypoxia/reperfusion cell culture model using PCq2 cells. Conclusions : Aurantii Immaturus Fructus has protective effects against ischemic brain damage at the early stage of ischemia.

• The Journal of Korean Medicine
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• v.24 no.1
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• pp.110-121
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• 2003

Objective : This research was performed to investigate the protective effect of Angelicae Dahuri Radix against ischemic damage using PC12 cells and global ischemia in gerbils. Methods : To observe the protective effect of Angelicae Dahuri Radix on ischemia damage, viability and changes in activities of superoxide dismutase (SOD), glutathione peroxidase (GPx), catalase and production of malondialdehyde (MDA) were observed after treating PC12 cells with Angelicae Dahuri Radix during ischemic insult. Gerbils were divided into three groups : a normal group, a 5-min two-vessel occlusion (2VO) group, and an Angelicae Dahuri Radix administered after 2VO group. The CCAs were occluded by microclip for 5 minutes. Angelicae Dahuri Radix was administered orally for 7 days after 2VO. The histological analysis was performed at 7 days after surgery. For histological analysis, the brain tissue was stained with 1% cresyl violet solution. Results : 1. Angelicae Dahuri Radix has a protective effect against ischemia in the CA1 area of the gerbil hippocampus 7 days after 5-minute occlusion, 2. In the hypoxia/reperfusion model using PC12 cells, Angelicae Dahuri Radix has a protective effect against ischemia in the dose of $0.2\mu\textrm{g}/ml$, $2\mu\textrm{g}/ml$ and $20\mu\textrm{g}/ml$, 3. Angelicae Dahuri Radix increased the activities of glutathione peroxidase and catalase. 4. The activity of superoxide dismutase (SOD) was increased by ischemic damage, which might represent self protection. This study suggests that Angelicae Dahuri Radix has some neuroprotective effect against neuronal damage following cerebral ischemia in vivo with a widely used experimental model of cerebral ischemia in Mongolian gerbils, and it also has protective effects on a hypoxia/reperfusion cell culture model using PC12 cells. Conclusions : Angelicae Dahuri Radix has protective effects against ischemic brain damage at the early stage of ischemia.

• The Journal of Korean Medicine
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• v.23 no.4
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• pp.113-124
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• 2002

Objects: This research was conducted to investigate the protective effect of Bupleuri Radix against ischemic damage using PC12 cells and global ischemia in gerbils, Methods: To observe the protective effect of Bupleuri Radixon ischemic damage, viability and changes in activities of superoxide dismutase (SOD), glutathione peroxidase (GPx), catalase and production of malondialdehyde (MDA) were observed after treating PC12 cells with Bupleuri Radix during ischemic damage. Gerbils were divided into three groups: a normal group, a 5-minute two-vessel occlusion (2VO) group and a Bupleun Radix administered group after 2VO. The CCAs were occluded by microclip for 5 minutes, Bupleuri Radix was administered orally for 7 days after 2VO. Histological analysis was performed on the 7th day. For histological analysis, the brain tissue was stained with 1 % of cresyl violet solution. Results: 1. Bupleuri Radix has a protective effect against ischemia in the CA1 area of the gerbil's hippocampus 7 days after 5-minute occlusion. 2. In the hypoxia/reperfusion model using PC12 cells, the Bupleuri Radix has a protective effect against ischemia in the dose of 0.2{\;}\mu\textrm{g}/ml,2{\;}\mu\textrm{g}/ml{\;}and{\;} 20{\;}\mu\textrm{g}/ml$. 3. Bupleuri Radix increased the activities of glutathione peroxidase and catalase. 4. The increased activity of superoxidedismutase (SOD) by ischemic damage might have been induced as an act of self-protection. This study suggests that Bupleuri Radix has some neuroprotective effect against neuronal damage following cerebral ischemia in vivo with a widely used experimental model of cerebral ischemia in Mongolian gerbils. Bupleuri Radix also has protective effect on a hypoxia/reperfusion cell culture model using PC12 cells. Conclusions: Bupleuri Radix has protective effect against ischemic brain damage during the early stages of ischemia.

• Journal of Acupuncture Research
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• v.36 no.3
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• pp.140-146
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• 2019

Background: This study investigated the effects of Vipera lebetina turanica snake venom (SV) on cerebral infarction induced by middle cerebral artery occlusion in mice. Methods: Following cerebral infarction, SV was injected intravenously or added to BV2 cell culture. Tissue injury was detected using triphenyltetrazolium chloride (TTC) staining, neurological deficit score, NO, ROS, and GSH/GSSG assays, qPCR, Western blot, and cell viability. Results: Cerebral infarction caused by middle cerebral artery occlusion as observed by TTC staining, showed SV inhibited cell death, reducing the number of brain cells injured due to infarction. SV treatment for cerebral infarction showed a significant decrease in abnormal behavior, as determined by the neurological deficit score. The oxidation and inflammation of the cells that had cerebral infarction caused by middle cerebral artery occlusion (NO assay, ROS, GSH/GSSG assay, and qPCR), showed significant protection by SV. Western blot of brain infarction cells showed the expression of iNOS, COX-2, p-IkB-${\alpha}$, P38, p-JNK, p-ERK to be lower in the SV group. In addition, the expression of IkB increased. BV2 cells were viable when treated with SV at $20{\mu}g/mL$ or less. Western blot of BV2 cells, treated with 0.625, 1.5, $2.5{\mu}g/mL$ of SV, showed a significant decrease in the expression of p-IkB-${\alpha}$, p-JNK, iNOS, and COX-2 on BV2 cells induced by LPS. Conclusion: SV showed anti-inflammatory and anti-oxidant effects against cerebral infarction and inflammation.

• Journal of Physiology & Pathology in Korean Medicine
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• v.21 no.6
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• pp.1401-1406
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• 2007

The present study investigated neuroprotective effects Rehmanniae Radix on PC12 cells and hippocampal neural cells. PC12 cells were damage by $H_2O_2$ and nitric oxide and organotypic hippocampal slice cultures were damaged by oxygen-glucose deprivation. Then methanol extract of Rehmanniae Radix was treated with 0.5, 5, and $50\;{\mu}g/ml$ in culture media. Effects of Rehmanniae Radix were evaluated with cell viability assay, PI-staining, and TUNEL-labeling. Treatment of Rehmanniae Radix ($with\;5\;and\;50\;{\mu}g/ml$) produced significant increase of cell viability of PC12 cells damaged by $H_2O_2$ and by SNP-induced nitric oxide. Treatment of Rehmanniae Radix produced significant decrease of PI-uptake % in CA1 ($with\;5\;and\;50\;{\mu}g/ml$) and DG ($with\;50\;{\mu}g/ml$) regions of organotypic hippocampal slice cultures damaged by oxygen-glucose deprivation. Moreover, treatment of Rehmanniae Radix produced significant decrease of TUNEL- positive cells in CA1 ($with\;5\;and\;50\;{\mu}g/ml$) and DG ($with\;50\;{\mu}g/ml$) regions of organotypic hippocampal slice cultures damaged by oxygen-glucose deprivation. These results suggest that methanol extract of Rehmanniae Radix has neuroprotective effects on PC12 cells damaged by oxidative stress and on organotypic hippocampal slice cultures damaged by oxygen-glucose deprivation.

• The Korea Journal of Herbology
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• v.23 no.3
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• pp.1-9
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• 2008

Objectives : This research was performed to investigate protective effect of Sophora Subprostrata against transient global ischemic damage after 5-min two vessel occlusion. Methods : Gerbils were divided into three groups: Normal group, 5-min two vessel occlusion (2VO) group, Sophora Subprostrata administrated group after 2VO. The CCAs were occluded by microclip for 5min. Sophora Subprostrata was administrated orally(12mg/ml) for 7 days after 2VO. The histological and immunohistochemistrical analysis was performed at 72 hours and 7 days after the surgery each. For histological analysis, the brain tissue was stained with 1% cresyl violet solution and Immunohistochemistry for BAX and Bcl-2 was carried out to examine effect of Sophora Subprostrata on ischemic brain tissue. Results : The results showed that (1) Sophora Subprostrata has the protective effect against ischemia in CA1 area of the gerbil hippocampus 7 days after 5-minute occlusion, (2) the treatment of Sophora Subprostrata inhibits the expression of Bax relatively after 2VO-induced ischemia. That protective effect of the Sophora Subprostrata seems to be performed by regulating the proportion of Bax and Bcl-2 protein, (3) in hypoxia/reperfusion model using PC12 cell, the Sophora Subprostrata extract has the protective effect against ischemia in the dose of $2{\mu}/m{\ell}$ and $20{\mu}/m{\ell}$.This study suggests that Sophora Subprostrata has neuroprotective effect against neuronal damage following cerebral ischemia in vivo with a widely used experimental model of cerebral ischemia in Mongolian gerbils and that Sophora Subprostrata regulates the proportion of Bax and Bcl-2 protein following ischemia. And, Sophora Subprostrata extract has protective effects also on a hypoxia/reperfusion cell culture model using PC12 cell. Conclusions : Sophora Subprostrata has protective effects against ischemic brain damage at the early stage of ischemia.

• Journal of Physiology & Pathology in Korean Medicine
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• v.21 no.1
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• pp.214-218
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• 2007

The water extracts of Samultang (Samul) has been used for treatment of ischemic heart and brain damage in Oriental traditional medicine. However, little is known about the mechanism by which the water extract of Samul rescues cells from oxidative damages in cisplatin-induced ototoxicity. Cisplatin is a widely used chemotherapeutic agent that is also highly ototoxic. This study was designed to investigate the protective effects of Samul on ciplatin-induced ototoxicity in HEI-OC1 auditory cells and organ of Corti explant culture. Cisplatin markedly decreased the viability of HEI-OC1 auditory cells. However, treatment of HEI-OC1 cells with Samul significantly reduced cisplatin-induced cell death and apoptotic characteristics through reduction of intracellular peroxide generation. Cisplatin induced cytotoxicity in isolated and cultured hair cell progenitors from postnatal rat cochleae. These progenitor cells are isolated from the lesser epithelial ridge (LER, or outer spiral sulcus cell) area of pre-plated neonatal rat cochlear segments. However, Samul completely protected the morphological changes of organ of Corti and LER. Taken together, these data suggest that the protective effects of the water extracts of Samul against cisplatin may be mediated by the reduction of intracellular peroxide generation.

• Journal of fish pathology
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• v.9 no.2
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• pp.169-176
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• 1996

An indirect double antibody enzyme-linked immunosorbent assay (ELISA) was developed for rapid detection of a new virus isolated from abnormally swimming salmonid fish, RVS (Retrovirus of salmonid). Results using brain tissue homogenates, and infected cell cultures are described. The sensitivity of the methods is $10^{2.6}$ $TCID_{50}/100{\mu}l$ of the examined cell culture fluid. The specificity was confirmed by the ELISA inhibition test and virological examinations. Viral antigen could be detected in artificially infected fish tissue homogenates. The assay will allow the diagnosis of RVS-infected fish within a day.

• The Korean Journal of Physiology and Pharmacology
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• v.4 no.4
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• pp.283-289
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• 2000

Natriuretic peptides comprise a family of three structurally related peptides; atrial natriuretic peptide (ANP), brain natriuretic peptide (BNP), and C-type natriuretic peptide (CNP). The present study was performed to investigate the effect of ANP on the proliferation and activity of ROS17/2.8 and HOS cells which are well-characterized osteoblastic cell lines. ANP dose-dependently decreased the number of ROS17/2.8 and HOS cells after 48-hour treatment. ANP generally increased the alkaline phosphatase activity of ROS17/2.8 and HOS cells after 48 hr treatment, regardless of the fact that basal activity of alkaline phosphatase was much lower in HOS cells compared to that of ROS17/1.8 cells. ANP increased the NBT reduction by ROS17/2.8 and HOS cells. ANP showed the variable but no significant effect on the nitric oxide production by ROS17/2.8 and HOS cells. ROS17/2.8 and HOS cells produced and secreted gelatinase into culture medium, and this enzyme was thought to be the gelatinase A type with the molecular weight determination. The gelatinase activity produced by ROS17/2.8 cells was increased by the treatment of ANP. However, the enzyme activity was not affected by ANP treatment in the HOS cell culture. In summary, ANP decreased the proliferation and increased the alkaline phosphatase activity and NBT reduction of osteoblasts. These results indicate that ANP is one of the important regulators of bone metabolism.