• Journal of Physiology & Pathology in Korean Medicine
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• v.25 no.5
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• pp.799-806
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• 2011

This study was performed in order to evaluate the neuroprotective effect of Boyanghwano-tang (BYT) water extract on intracerebral hemorrhage (ICH). ICH was induced by the stereotaxic intrastriatal injection of bacterial collagenase type IV in Sprague-Dawley rats. BYT was orally given once a day for 3 days after ICH. Hematoma volume and percentage edema were examined. As imflammatory markers, myeloperoxidase (MPO)-positive neutrophils infiltration and iNOS expression in the peri-ICH regions were examined using immunohistochemistry. As cellular damage markers, c-Fos, Bax, and HSP72 positive cells in the peri-ICH regions were measured also. BYT significantly reduced the hematoma volume and percentage edema of the ICH-induced rat brain. In the peri-hematoma regions, BYT significantly reduced MPO-positive neutrophil infiltration and iNOS expression of the ICH-induced rat brain. Additionally, BYT significantly reduced c-Fos, Bax, and HSP72 positive cells in the peri-hematoma regions of the ICH-induced rat brain. These results suggest that BYT plays a neuroprotective role against ICH through suppression of inflammatory responses, apoptosis and cellular damage.

• The Journal of Internal Korean Medicine
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• v.31 no.4
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• pp.706-713
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• 2010

Objectives : The source is from the flower of Carthamus tinctorius L., family Compositae. It is used in clinical medicine to promote blood circulation, remove blood stasis, promote menstruation and alleviate pain. In the present study, we investigated the genome wide analysis of Carthami Flos on the intra-cranial hemorrhage(ICH) model. Methods : ICH in rat was induced by injection of collagenase type IV and Carthami Flos extract(CFe) was administered orally. The molecular profile of cerebral hemorrhage in rat brain tissue was measured using microarray technique to identify up- or down- regulated genes in brain tissue. Results : Expression profile showed that diverse genes were up- or down-regulated by ICH induction. Administration of CFe restored the expression level of some of altered genes by ICH to normal expressional level. Interestingly, these recovered genes by CFe were involved in the same biological pathways which were significantly activated or suppressed by ICH. Conclusion : The above results might explain the therapeutic mechanism of CFe on ICH. Further, by analyzing interaction network, core genes was identified which could be key molecular targets of CFe against ICH.

• Nutritional Sciences
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• v.7 no.3
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• pp.144-150
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• 2004

We have investigated the effect of calcium spirulan(Ca-SP) isolated from a blue-green alga Spirulina platensis, which is a sulfated polysaccharide chelating calcium and mainly composed of rhamnose and fructose, on invasion of both B16- BL6 melanoma cells, Colon 26 carcinoma and HT-1080 fibrosarcoma cells through reconstituted basement membrane (Matrigel). Ca-SP significantly inhibited the invasion of these tumor cells through Matrigel/fibronectin-coated filters in a concentration-dependent manner. Ca-SP also inhibited the haptotactic migration of tumor cells to laminin, but it had no inhibitory effect on tumor cell migration to fibronectin-coated filters. Ca-SP prevented the adhesion of B16-BL6 cells to Matrigel- and laminin-substrates but did not affect the adhesion to fibronectin. The pretreatment of tumor cells with Ca-SP inhibited the adhesion to laminin in a concentration-dependent fashion, while the pretreatment of laminin-substrates did not. Ca-SP had no effect on the production and activation of type IV collagenase in gelatin zymography. In contraset, Ca-SP significantly inhibited degradation of heparan sulfate by purified heparanase. The experimental lung metastasis was significantly reduced by co-injection of B16-BL6 cells with Ca-SP in a dose-dependent manner. Seven intermittent ⅰ.ⅴ. injection of 100$\mu\textrm{g}$ of Ca-SP caused a marked decrease of lung tumor colonization of B16-BL6 cells in a spontaneous lung metastasis model. These results suggest that Ca-SP, a novel sulfated polysaccharide, could reduce the lung colonization of B16-BL6 melanoma cells in experimental metastasis model, by inhibiting the tumor invasion of basement membrane Matrigel, probably through the prevention of the adhesion and migration of tumor cells to laminin-substrate and of the heparanase activity.

• BMB Reports
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• v.32 no.1
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• pp.60-66
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• 1999

Matrix metalloproteinase-2 (MMP-2; 72-kDa gelatinase; 72-kDa type IV collagenase; gelatinase A) plays an important role in normal physiological processes and in many pathologic processes such as arthritis and metastasis of cancer. Tissue inhibitor of metalloproteinases-2 (TIMP-2) binds to proMMP-2 or mature MMP-2 at a 1:1 ratio and inhibits the catalytic activity of MMP-2. We demonstrated that the baculovirus/insect cell system does not have TIMP-2 activity. The human proMMP-2 free of TIMP-2 was expressed in the expression system and purified by one-step affinity chromatography using gelatin-Sepharose. The free proMMP-2 was autoactivated to the mature MMP-2 and autodegraded into smaller molecular weight forms in the absence of external activator. The activation and autodegradation of the proMMP-2 was much more rapid in the presence of 4-aminophenylmercuric acetate (APMA). Addition of TIMP-2 inhibits both APMA-induced activation and autodegradation of the free proMMP-2. However, an increasing concentration of TIMP-2 more readily inhibited activation of the free proMMP-2 than autodegradation. These results demonstrate that TIMP-2 plays roles in inhibition of both activation and autodegradation of the free proMMP-2 in addition to inhibition of the catalytic activity of MMP-2.

• International Journal of Oral Biology
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• v.36 no.4
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• pp.195-201
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• 2011

Matrix metalloproteinases (MMPs) have been implicated in tissue development and re-modeling. Dynamic morphological changes of tooth germs reflect involvement of these enzymes during odontogenesis. The present study was performed to investigate expression and localization of MMP-2 and MMP-9, which have been known to have type IV collagenase activities, in rat tooth germs at different developmental stages. MMP-2 expression was increased gradually in the tooth germs from cap to crown staged germs at both transcription and translation levels. The localization of this molecule was detected in secretory ameloblasts and preameloblasts. The strong immunoreactivities were occasionally seen along the basement membrane between ameloblasts (or preameloblasts) and odontoblasts (preodontoblasts). However, weak reactivity was detected in odontoblasts and reduced enamel epithelium. The level of MMP-9 expression in the tooth germs was higher in cap stage than in crown staged germs at both transcription and translation levels. They were strongly expressed in both ameloblasts and odontoblasts. Even though reduced enamel epithelium after enamel formation and inner enamel epithelium at the cap stage exhibited weak reactivity, strong reactivity was detected in dental follicles and perifollicular tissues surrounding cap staged germs. These results suggested that MMP-2 may involve degradation of the basement membrane during hard tissue formation, whereas MMP-9 might be involved in remodeling of follicular tissues.

• Journal of Society of Preventive Korean Medicine
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• v.12 no.3
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• pp.81-90
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• 2008

Objectives : The pathophysiology of ICH is not fully understood, therefore, the fundamental therapeutic strategies for ICH also not well inspected either. The genetic profile for the effect of Carthami Flos extract on cerebral hemorrhage in rat brain tissue was measured using microarray technique. Genes displaying expressional change on brain damage were selected and the functional analysis on these genes was conducted. Methods : Rats were placed in a stereotaxic frame after intraperitoneal injection of chloralhydrate, and ICH was induced by injection of collagenase type IV and Carthami Flos extract was administered orally. The molecular profile of cerebral hemorrhage in rat brain tissue was measured using microarray technique to identify up- or down- regulated genes in brain tissue. Results : Upon treatment with Carthami Flos extract on the rat having brain damage, many genes show expressional change. The pattern of gene expressional change can be classified into 8 classes in which two types of classes were composed of recovered genes from up or down-regulation by brain damage, respectively. Conclusions : Further analysis using protein interaction database identified some key molecules that can be used for elucidation of therapeutical mechanism of Carthami Flos extract in future.

• The Korea Journal of Herbology
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• v.29 no.1
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• pp.19-26
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• 2014

Objectives : This study was performed in order to evaluate the effects of Salviae Miltiorrhizae Radix (SMR) water extract against the cerebral hemorrhage and the blood-brain barrier (BBB) impairment in the intracerebral hemorrhage (ICH). Method : ICH was induced by the stereotaxic intrastriatal injection of bacterial collagenase type IV in Sprague-Dawley rats. SMR was orally given three times every 20 hours during 3 days after the ICH induction. Hematoma volume, water content of brain tissue and volume of evans blue leakage were examined. Myeloperoxidase (MPO) positive neutrophils and tumor necrosis factor-${\alpha}$ (TNF-${\alpha}$) were observed with immunofluorescence labeling and confocal microscope. Results : SMR significantly reduced the hematoma volume of the ICH-induced rat brain. SMR significantly reduced the water content of brain tissue of the ICH-induced rat brain. SMR reduced the percentage of the evans blue leakage around the hematoma on the caudate putamen compared to the ICH group, especially on the cerebral cortex. SMR significantly reduced the volume of the evans blue leakage level in the peri-hematoma regions of the ICH-induced rat brain. SMR significantly reduced MPO positive neutrophils in the peri-hematoma regions of the ICH-induced rat brain. SMR reduced the TNF-${\alpha}$ expression in peri-hematoma regions of the ICH-induced rat brain. TNF-${\alpha}$ immuno-labeled cells were coincided with MPO immuno-labeled neutrophils in peri-hematoma regions of the ICH-induced rat brain. Conclusion : These results suggest that SMR plays a protective role against the blood-brain barrier impairment in the ICH through suppression of inflammation in the rat brain tissues.

• The Korea Journal of Herbology
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• v.29 no.2
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• pp.33-38
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• 2014

Objectives : Inflammation is mediated by cellular components, such as leukocytes and microglia, and molecular components, including cytokines, extracellular proteases, and reactive oxygen species. Cnidium Rhizoma effects the anti-inflammatory, antioxidant, suppression of the microglia activation and protection of the nerve cell injury. For this reason, we investigated the anti-inflammatory effects of water extracts of Cnidium Rhizoma on intracerebral hemorrhage (ICH). Method : ICH was induced by the stereotaxic intracerebral injection of bacterial collagenase type IV (0.23 $U/{\mu}{\ell}$, 0.1 ${\mu}{\ell}/min$) in Sprague-Dawley rats. We orally administrated once 3 hours after ICH, then 2 times at 24-hour intervals the water extracts of Cnidium Rhizoma (500 mg/kg), myeloperoxidase (MPO) was observed by using immunofluorescense and expression of inducible nitric oxide synthase (iNOS), tumor necrosis factor-${\alpha}$ (TNF-${\alpha}$) and microglia were observed by using immunohistochemistry. Results : Infiltration of MPO expressing neutrophil, expression of iNOS and TNF-${\alpha}$ and activated microglia were significantly reduced in peri-hematoma of the rats fed with water extracts of Cnidium Rhizoma. Conclusion : These results demonstrated that water extracts of Cnidium Rhizoma suppressed an inflammatory reaction through inhibition of MPO, iNOS and TNF-${\alpha}$ positive cell and activated microglia number in peri-hematoma of ICH-induced rats.

• Radiation Oncology Journal
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• v.12 no.1
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• pp.27-31
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• 1994

There are a number of reports suggesting that there may be a correlation between the clinical response to radiotherapy in various tumors and the clonogenic survival of cell lines derived from these tumors following exposure to 2 Gy(SF2). Authors conducted this study to determine SF2 for cells in primary culture from surgical specimens. The tumor tissues with squamous cell carcinoma of uterine cervix and head and neck were obtained. The tumor tissues were disaggregated to single cells by incubating with collagenase type w for 2 hours with constant stirring. Single cell suspensions were inoculated in four 24-well plates precoated with cell adhesive matrix. After 24 hours of incubation at 37$ ^{\circ}C $, rows of four wells were then irradiated, consisting of control set and five other sets each receiving doses of 1,2,3,4, and 6 Gy. After incubation for a total of 13 days, the cultures were stained with crystal violet and survival at each dose was determined by quantitative image analysis system, To determine whether cell growth was of epithelial origin, immunocytochemical staining with a mixture of cytokeratin and epithelial monoclonal antibodies were performed on cell cultures. During the period of this study, we received 5 squamous cell carcinoma specimens of head and neck and 20 of uterine cervical carcinoma. Of these, 15 yielded enough cells for radiosensitivity testing. This resulted an overall success rate of 60$ \% $. The mean SF2 value for 15 tumours was 0.55$\pm$0.17 ranging from 0.20 to 0.79. These results indicate that there is a broad range of sensitivities to radiation in same histologic type. So with a large patient population, we plan to determine whether a different SF2 value is associated with tumours that are controlled with radiotherapy than those that are not.

• The Korea Journal of Herbology
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• v.22 no.4
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• pp.253-260
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• 2007

Objectives : Intracerebral hemorrhage (ICH) is characterized by breakdown of blood vessels within the brain parenchyma. Fundamental therapeutic strategies for ICH, particularly those aimed at neuroprotection, have to be established. So in this experiment, the effects of Woowhangchongshim-won, a traditional prescription formula for treating Cerebral Apoplexy in Asian countries, were investigated. Methods : After intraperitoneal injection of chloralhydrate, rats were placed in a stereotaxic frame. ICH was induced by injection of 1 U collagenase type IV and drug was administered orally for 10 days. The molecular profile of cerebral hemorrhage in rat brain tissue was measured using micro array technique to identify up- or down- regulated genes in brain tissue. These genes induced by brain damage were mainly concerned with general metabolic process such as primary metabolic process, cellular metabolic process, macromolecule metabolic process, and biosynthetic process. Results : The number of genes increased in control and not-changed in experiment was 374, and decreased in control and not-changed in experiment was 527. We are concerned with genes that can be recovered by treatment with medicine, it is especially interesting to above types of genes. Conclusions : Upon medicine treatment to the rat having cerebral hemorrhage, expressions of some genes were restored to normal level. Further analysis using protein interaction database identified some key molecules that can be used for elucidation of therapeutical mechanism of medicine in future.