• Food Science and Preservation
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• v.31 no.4
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• pp.590-600
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• 2024

Angelica keiskei, a perennial herb from Apiaceae family, has been reported to improve diabetes, inhibit thrombosis, alleviate dyslipidemia, and prevent type 2 diabetes, obesity, and atherosclerosis. In this study, the protective effects of A. keiskei extract (AKE) against tumor necrosis factor-alpha (TNF-α)-induced oxidative stress and vascular inflammation in human umbilical vein endothelial cells (HUVECs) were investigated through cell viability analysis, antioxidant enzyme analysis, western blotting, and immunofluorescence staining. The results demonstrated that pretreatment of Angelica keiskei with AKE significantly inhibited the expression of key adhesion molecules such as E-selectin, ICAM-1 and VCAM-1 induced by TNF-α. AKE also showed a substantial reduction in intracellular reactive oxygen species levels and an increase in antioxidant enzyme activity, indicating potential antioxidant capabilities. This study further explained that AKE interfered with the nuclear factor-kappa B (NF-κB) pathway by inhibiting phosphorylation of IκBα and NF-κB, thereby preventing nuclear translocation. Additionally, AKE selectively inhibited the activation of c-Jun N-terminal kinase (JNK) within the mitogen-activated protein kinase (MAPK) pathway, revealing a specific action mechanism. These findings collectively suggest that AKE possesses multi-faceted protective properties, making it a potential therapeutic agent for inflammatory conditions and early atherosclerosis.

• Herbal Formula Science
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• v.32 no.2
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• pp.181-191
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• 2024

Objective : Inulae Flos(IF) has been used to treat arthritis, sever furuncle, fear and palpitation, vomiting, stroke, asthma and cough in Korean Medicine. Although the anticancer activity of IF has been reported, the molecular mechanism is still not well understood. In this study, we investigated the growth inhibitory activity of an ethanol extract of IF in HT-1080 human fibrosarcoma cells and its underlying mechanisms using two-dimensional (2D) and three-dimensional (3D) cell culture system. Methods : HT-1080 cells were cultured with IF for 9 days in 3D cell culture. To check an inhibition of cell prolifelation by IF, MTT assay was performed. DNA contents were measured using flow cytometry. Western blotting was used to evaluate the regulation of cell cycle- and autophagy-related proteins. Acridine orange staining was performed to confirm autophagy, and DCF-DA staining was performed to confirm the occurrence of ROS. Results : IF controlled a spheroid formation and decreased a cell viability in 3D cell culture. IF-induced cell proliferation inhibition was associated with a distinct increase of S and G2/M phase cell distribution in 2D cell cultre. In addition, IF significantly induced autophagy and generated reactive oxygen species(ROS). Interestingly, IF-induced cell cycle arrest and autophagy were recovered after pre-treatment of N-acetyl-L-cysteine, ROS scavenger. Conclusion : Our results indicate that IF induced ROS-mediated cell cycle arrest and autophagy and it may potentially useful for human fibrosarcoma treatment.

• Journal of the korean academy of Pediatric Dentistry
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• v.33 no.1
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• pp.13-24
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• 2006

Neuronal apoptotic events, consequently resulting in neuronal cell death, are occurred in hypoxic/ischemic condition. This cell death has been shown to be accompanied with the production of reactive oxygen species (ROS), which can attack cellular components such as nucleic acids, proteins and phospholipid. However, the underlying mechanisms of apoptosis induced in hypoxic/ischemic condition and its treatment methods are unsettled. Cobalt chloride $(CoCl_2)$ has been known to mimic hypoxic condition including the production of ROS. Epigallocatechin gallate (EGCG), a green tea polyphenol, has diverse pharmacologial activities in cell growth and death. This study was aimed to investigate the apoptotic mechanism by $CoCL_2$ and effects of EGCG on $CoCl_2-induced$ apoptosis in PC12 cells. Administration of $CoCl_2$ decreased cell survival in dose- and time-dependent manners and induced genomic DNA fragmentation. Treatment with $100{\mu}M$ EGCG for 30 min before PC12 cells were exposed to $150{\mu}M$ $CoCl_2$, being resulted in the cell viability and DNA fragmentation being rescued. $CoCl_2$ caused morphologic changes such as cell swelling and condensed nuclei whereas EGCG attenuated morphologic changes by $CoCl_2$. EGCG suppressed the apoptotic peak and a loss of ${\Delta}{\psi}_m$ induced by $CoCl_2$. $CoCl_2$ decreased Bcl-2 expression but Bax expression was not changed in $CoCl_2$- treated cells. EGCG attenuated the Bcl-2 underexpression by $CoCl_2$. $CoCl_2$ augumented the cytochrome c release from mitochondria into cytoplasm and increased caspase-8, -9 and caspase-3 activity a marker of the apoptotic executing stage. EGCG ameliorated the incruement in caspase-8, -9 and -3 activity, and cytochrome c release by $CoCl_2$ NAC (N-acetyl-cysteine), a scavenger of ROS, attenuated $CoCl_2-induced$ apoptosis in consistent with those of EGCG. These results suggest that $CoCl_2$ induces apoptotic cell death through both mitochondria- and death receptor-dependent pathway and EGCG has neuroprotective effects against $CoCl_2-induced$ apoptosis in PC12 cells.

• Journal of Haehwa Medicine
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• v.24 no.2
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• pp.35-57
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• 2016

Objectives : The purpose of this study is to prove the effect and mechanism of Gamikyejakjimogawusul-tang(GKHA) herbal acupuncture on induced rheumatoid arthritis model of DBA/1 mice. Methods : We check effect of GKHA extract on the AST, ALT, Creatinine, BUN of serum and cell viability of GK extract in RAW 264.7 cells to test the stability of this study. In vitro, we measure total phenol contents, total flavonoid contents, DPPH free radical scavenging activity, ABTS cation radical scavenging activity of Gamikyejakjimogawusul-tang, effect of GK extract on ROS(Reactive Ooxygen Species) production to estimate a anti-oxidant capacity, and we also measure effect of GK extract on NO (Nitric Oxid), IL-$1{\beta}$, IL-6, IL-17, IL-21, TNF-${\alpha}$, MCP-1, GM-CSF production in RAW 264.7 cells to estimate a anti-inflammatory efficacy. In vivo, we compare a rheumatoid arthritis manifestation between control and experimental group and estimate a AI. Then we check effect of GKHA on the level of WBC, neutrophil, lympocyte, monocyte in the blood to see the effect of immune cells in blood. In addition we measure effect of GKHA on the level of hs-CRP, IgM, IgG, IL-$1{\beta}$, IL-6, IL-17, IL-21, TNF-${\alpha}$, MCP-1, GM-CSF in serum. We observe effects of GKHA on imaging of cartilage degeneration using micro CT-arthrography in paw hind. And we calculate effects of GKHA that reduced BV ratio, BS/BV ratio using 3D Micro-CT. Lastly we observe effects of GKHA histopathologic examination analysis. Results : 1. The toxicity on liver and kidney was disregardable and the cytotoxicity against RAW 264.7 cells was also disregardable. 1. Total phenol contents and total flavonoid contents in GK extract were in high level. 2. DPPH free radical scavenging activity and ABTS cation radical scavenging activity were increased according to concentration of GK extract 3. ROS production was significantly decreased in GK extract (at 10, $100{\mu}g/ml$). 4. NO, IL-6, TNF-${\alpha}$, MCP-1 production were significantly decreased in GK extract(at 10, $100{\mu}g/ml$). IL-17, GM-CSF production were significantly decreased in GK extract(at 1, 10, $100{\mu}g/ml$). IL-$1{\beta}$, IL-21 production were also decreased but there was no statistical significance. 5. 25x observation after H&E and M-T staining, infiltration of immune cells and subsidence of the cartilage and damage to the synovial cells were decreased. Conclusions : This study showed that GKHA extract had anti-oxidant capacity, anti-inflammatory efficacy. GKHA extract also had inhibiting effect on the process of rheumatoid arthritis and can protect joint and cartilage. So we expect that GKHA extract can be a meaningful treatment to rheumatoid arthritis patients.

• KSBB Journal
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• v.16 no.5
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• pp.486-492
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• 2001

The antimutagenic mechanism of the fraction III(RG III)separated from the water extract of Rehmannia glutionosa was investigated by Escherichia. coli GW and B/r strains. RG-III treatment did not affect the ${\beta}$-galactosidase activity E. coli GW-1060, 1106, 1107 and 1105. These results indicated that RG-III did not induce RecA protein amplification and did not also prevent the proteolytic cleavage of LexA. The bio-antimutagenicity and survival effect of RG-III on 4-nitroquinoline 1-oxide(4NQO), N-methyl-N-nitor-N\`-nitrosoguanidine(MNING) were investigate by E. coli B/r strains with have different pathway of DNA repai. RG-III slightly increased the survival of 4NQO-treated WP2, WP2s, WP67, CM561, CM611 cells, but the reactivation of survival cannot ve explained by the repair mode. RG-III caused the decrease of mutagenicity and lethality treated with MNNG in ZA159 despite of the increase in WP2, WP2s, WP67, CW561, CM611. Compared with bio-antimutagenic effects of RG-III on 4NQO, greatly increased antimutagenic effects of RG-III were observed with all the E. coli B/r strains tested, but less active in ZA159. These results suggest that RG-III was identified as a blocking agent for preventing the 4NQO induced mutagenesis, and may act as chl-products.

• Journal of Oriental Neuropsychiatry
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• v.22 no.2
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• pp.163-176
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• 2011

Objectives : BCS(Bambusae concretio silicae) is used as a traditional medicine in Korea for the incipient stroke. Recent reports indicated that BCS has a neuroprotective effect by anti-convulsion effect. However, it's mechanism is not well studied. The purpose of this study was to investigate into the molecular mechanism of BCS for neuroprotection in normoxia of cultured rat hippocampal neurons. Methods : BCS (1.0, 2.5, 5.0, and $10.0\;{\mu}g/m{\ell}$) was added to culture media (Neurobasal supplemented with B27) on DIV 0, given a normoxia, and the cell viability was measured by typical phase-contrast images of the cultures with 1.0, 2.5, 5.0, and $10.0\;{\mu}g/m{\ell}$ on DIV 21. Effects of BCS on the expression of various synaptic proteins ($GABA_B$ R1, $GABA_B$ R2, GlyR, PSD95) were observed by immunocytochemistry showing on DIV 3, 7 and 21. Results : Typical phase-contrast images of the cultures indicated that BCS has a protective effect of rat hippocampal cells in normoxia. The BCS upregulated $GABA_B$ R1 after normoxia on DIV 7, $GABA_A$ ${\beta}2/3$ on DIV 21 and $GABA_B$ R2 on DIV 21. And the BCS downregulated PSD95 after normoxia on DIV 7. Conclusions : The present study showed evidence for the efficacy of BCS in Typical phase-contrast images, upregulation of inhibitory neurotransmitter receptors($GABA_B$ R1) and downregulation of PSD95 which eventually protected neuronal cell death in normoxia.

• The Journal of Internal Korean Medicine
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• v.31 no.1
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• pp.54-65
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• 2010

Objective : The water extract of Chungsimyeonja-eum (CSYJE) has traditionally been used in treatments of heart diseases and brain diseases in Oriental medicine. However, little is known about the mechanism by which CSYJE protects neuronal cells from injury damages. Therefore, in this study we attempted to elucidate the mechanism of the cytoprotective effect of the CSYJE extract on glutamate-induced C6 glial cell death. Methods : Cultured cells were pretreated with CSYJE and exposed to glutamate, cell damage was assessed by using MTT assay and propidium iodide (PI), probe 2',7'-dichlorofluorescein diacetate (DCF-DA) staining. Western blotting was performed using anti-procaspase-3 and anti-PARP, respectively. Result : We determined the elevated cell viability by CSYJE extract on glutamate-induced C6 glial cell death. Glutamate induced DNA fragmentation on C6 glial cells but pre-treatment with CSYJE inhibited DNA fragmentation. One of the main mediators of glutamate-induced cytotoxicity was known to generation of reactive oxygen species (ROS). Pre-treatment with CSYJE inhibited this ROS generation from glutamate-stimulated C6 glial cells. Also, we identified that the ROS-induced DCF-DA green fluorescence was reduced by CSYJE pre-treatment. The critical markers of apoptotic cell death are the cleavages of procaspase-3 protease and PARP proteins, so we checked the expression level and cleavages of procaspase-3 protease and PARP proteins. Glutamate-treated C6 glial cells showed the cleavages of procaspase-3 protease and PARP proteins and followed the reduction of expression of these proteins. Conclusion : These findings indicate that CSYJE may prevent cell death from glutamate-induced C6 glial cell death by inhibiting the ROS generation and procaspase-3 and PARP expression.

• Toxicological Research
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• v.18 no.4
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• pp.355-362
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• 2002

Pathophysiological elevation of intracellular calcium concentration ($[Ca^{2+}]_1$) in the neuron has been considered as an important responsible factor in the neuronal cell damages. However the mechanism of increase of $[Ca^{2+}]_1$ and the relationship between $[Ca^{2+}]_1$ level and cytotocixity have not been fully demonstrated. In the present study, real-time alteration of $[Ca^{2+}]_1$and cellular response (cell damages) in the pheochromocytoma cells (PC12) stimulated by glutamate were investigated. Glutamate dose dependently decreased cell viability determined propidium iodide fluorescence method and morphology change. Conversely related with cell damages, glutamate dose dependently increased the level of［Ca$^{2+}$］$_{i}$ . To investigate the mechanism of glutamate-induced increase of $[Ca^{2+}]_1$,$[Ca^{2+}]_1$, was first measured in the cell cultured in calcium free media and in the presence of dantrolene, an inhibitor of calcium release from ryanodine receptor located in endoplasmic reticulum (ER). Similar to the increase$[Ca^{2+}]_1$ in the calcium-containing media, glutamate dose dependently increased $[Ca^{2+}]_1$ in the cell cultured in free calcium media. However pretreatment (2 hr) with 20~50 $\mu\textrm{M}$ dantrolene substantial lowered glutamate-induced increase of $[Ca^{2+}]_1$, suggesting that release of calcium from ER may be major sourse of increase of $[Ca^{2+}]_1$ in PC12 cells. Dantrolene-induced inhibition of $[Ca^{2+}]_1$ resulted in recovery of cytotoxicity by glutamate. Relevance of N-methy-D-aspartate (NMDA) receptor, a type of glutamte receptor on glutamate-induced incense of $[Ca^{2+}]_1$,$[Ca^{2+}]_1$ was also determined in the cells pretreated (2 hr) with NMDA receptor antagonist MK-80l. Glutamate-induced increase of $[Ca^{2+}]_1$ was reduced by MK-801 dose dependently. Furthermore, glutamate-induced cytotoxicity was also prevented by MK-80l. These results demonstrate that glutamte increase $[Ca^{2+}]_1$ dose dependently and thereby cause cytotoxicity. The increase of $[Ca^{2+}]_1$ may release from ER, especially through ryanodine receptor and/or through NMDA receptor Alteration of calcium homeostasis through disturbance of ER system and/or calcium influx through NMDA receptor could contribute glutamate-induced cell damages.s.

• Asian Pacific Journal of Cancer Prevention
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• v.15 no.7
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• pp.3093-3097
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• 2014

Background: Cancer is a major threat to the public health whether in developed or in developing countries. As the most common primary malignant tumor, the morbidity and mortality rate of lung cancer continues to rise in recent ten years worldwide. Chemotherapy is one of the main methods in the treatment of lung cancer, but this is hampered by chemotherapy drug resistance, especially MDR. As a component of the 60S large ribosomal subunit, ribosomal protein L39-L gene was reported to be expressed specifically in the human testis and human cancer samples of various tissue origins. Materials and Methods: Total RNA of cultured drug-resistant and susceptible A549 cells was isolated, and real time quantitative RT-PCR were used to indicate the transcribe difference between amycin resistant and susceptible strain of A549 cells. Viability assay were used to show the amycin resistance difference in RPL39-L transfected A549 cell line than control vector and null-transfected A549 cell line. Results: The ribosomal protein L39-L transcription level was 8.2 times higher in drug-resistant human lung cancer A549 cell line than in susceptible A549 cell line by quantitative RT-PCR analysis. The ribosomal protein L39-L transfected cells showed enhanced drug resistance compared to plasmid vector-transfected or null-transfected cells as determined by methyl tritiated thymidine (3H-TdR) incorporation. Conclusions and Implications for Practice: The ribosomal protein L39-L gene may have effects on the drug resistance mechanism of lung cancer A549 cells.

• Asian Pacific Journal of Cancer Prevention
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• v.14 no.10
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• pp.5825-5831
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• 2013

Background/Objectives: Maintenance of cellular function in culture is vital for transfer and development following adoptive immunotherapy. Dual properties of IL-21 in activating T cells and reducing activation induced cell death led us to explore the mechanism of action of IL-21 enhanced proliferation and cytotoxic potential of CIK cells. Method: CIK cells cultured from PBMCs of healthy subjects were stimulated with IL-21 and cellular viability and cytotoxicity to K562 cells were measured. To elucidate the mechanism of action of IL-21, mRNA expression of cytotoxic factors was assessed by RT-PCR and protein expression of significantly important cytotoxic factors and cytokine secretion were determined through flow cytometry and ELISA. Western blotting was performed to check the involvement of the JAK/STAT pathway following stimulation. Results: We found that IL-21 did not enhance in vitro proliferation of CIK cells, but did increase the number of cells expressing the CD3+/CD56+ phenotype. Cytotoxic potential was increased with corresponding increase in perforin ($0.9831{\pm}0.1265$ to $0.7592{\pm}0.1457$), granzyme B ($0.4084{\pm}0.1589$ to $0.7319{\pm}0.1639$) and FasL ($0.4015{\pm}0.2842$ to $0.7381{\pm}0.2568$). Interferon gamma and TNF-alpha were noted to increase ($25.8{\pm}6.1ng/L$ to $56.0{\pm}2.3ng/L$; and $5.64{\pm}0.61{\mu}g/L$ to $15.14{\pm}0.93{\mu}g/L$, respectively) while no significant differences were observed in the expression of granzyme A, TNF-alpha and NKG2D, and NKG2D. We further affirmed that IL-21 signals through the STAT-3 and STAT-5b signaling pathway in the CIK cell pool. Conclusion: IL-21 enhances cytotoxic potential of CIK cells through increasing expression of perforin, granzyme B, IFN-gamma and TNF-alpha. The effect is brought about by the activation of STAT-3 and STAT-5b proteins.