• Tuberculosis and Respiratory Diseases
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• v.85 no.2
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• pp.147-154
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• 2022

Background: The expression of calcium signaling pathway molecules is altered in various carcinomas, which are related to the proliferation and altered characteristics of cancer cells. However, changes in calcium signaling in anti-cancer drug-resistant cells (bearing a T790M mutation in epidermal growth factor receptor [EGFR]) remain unclear. Methods: Afatinib-mediated changes in the level of store-operated Ca²⁺ entry (SOCE)-related proteins and intracellular Ca²⁺ level in non-small cell lung cancer cells with T790M mutation in the EGFR gene were analyzed using western blot and ratiometric assays, respectively. Afatinib-mediated autophagic flux was evaluated by measuring the cleavage of LC3B-II. Flow cytometry and cell proliferation assays were conducted to assess cell apoptosis and proliferation. Results: The levels of SOCE-mediating proteins (ORAI calcium release-activated calcium modulator 1 [ORAI1], stromal interaction molecule 1 [STIM1], and sarco/endoplasmic reticulum Ca²⁺ ATPase [SERCA2]) decreased after afatinib treatment in non-small cell lung cancer cells, whereas the levels of SOCE-related proteins did not change in gefitinib-resistant non-small cell lung cancer cells (PC-9/GR; bearing a T790M mutation in EGFR). Notably, the expression level of SOCE-related proteins in PC-9/GR cells was reduced also responding to afatinib in the absence of extracellular Ca²⁺. Moreover, extracellular Ca²⁺ influx through the SOCE was significantly reduced in PC-9 cells pre-treated with afatinib than in the control group. Additionally, afatinib was found to decrease the level of SOCE-related proteins through autophagic degradation, and the proliferation of PC-9GR cells was significantly inhibited by a lack of extracellular Ca²⁺. Conclusion: Extracellular Ca²⁺ plays important role in afatinib-mediated autophagic degradation of SOCE-related proteins in cells with T790M mutation in the EGFR gene and extracellular Ca²⁺ is essential for determining anti-cancer drug efficacy.

• Herbal Formula Science
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• v.25 no.1
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• pp.51-68
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• 2017

Objectives : Oxidative stress due to excessive accumulation of reactive oxygen species (ROS) is one of the risk factors for the development of several chronic diseases, including neurodegenerative diseases. Methods : In the present study, we investigated the protective effects of cheongnoemyeongsin-hwan (CNMSH) against oxidative stress‑induced cellular damage and elucidated the underlying mechanisms in neuronal-derived SH-SY5Y cells. Results : Our results revealed that treatment with CNMSH prior to hydrogen peroxide (H2O2) exposure significantly increased the SH-SY5Y cell viability, indicating that the exposure of the SH-SY5Y cells to CNMSH conferred a protective effect against oxidative stress. CNMSH also effectively attenuated H2O2‑induced comet tail formation, and decreased the phosphorylation levels of the histone ${\gamma}H2AX$, as well as the number of apoptotic bodies and Annexin V‑positive cells. In addition, CNMSH exhibited scavenging activity against intracellular ROS generation and restored the mitochondria membrane potential (MMP) loss that were induced by H2O2, suggesting that CNMSH prevents H2O2‑induced DNA damage and cell apoptosis. Moreover, H2O2 enhanced the cleavage of caspase-3 and degradation of poly (ADP-ribose)-polymerase, a typical substrate protein of activated caspase-3, as well as DNA fragmentation; however, these events were almost totally reversed by pretreatment with CNMSH. Furthermore, CNMSH increased the levels of heme oxygenase-1 (HO-1), which is a potent antioxidant enzyme, associated with the induction of nuclear factor-erythroid 2-related factor 2 (Nrf2). According to our data, CNMSH is able to protect SH-SY5Y cells from H2O2-induced apoptosis throughout blocking cellular damage related to oxidative stress through a mechanism that would affect ROS elimination and activating Nrf2/HO-1 signaling pathway. Conclusions : Therefore, we believed that CNMSH may potentially serve as an agent for the treatment and prevention of neurodegenerative diseases caused by oxidative stress.

• Korean Journal of Microbiology
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• v.43 no.3
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• pp.159-165
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• 2007

For the development of basic genetic materials for specific and effective therapeutic approach to suppress multiplication of hepatitis C virus (HCV), HCV internal ribosome entry site (IRES)-targeting hammerhead ribozyme which activity is allosterically regulated by HCV regulatory protein, NS5B RNA replicase, was developed. The ribozyme targeted most effectively to +382 nucleotide (nt) site of HCV IRES RNA. The allosteric ribozyme was designed to be composed of sequence of RNA aptamer to HCV NS5B, communication module sequence which can transfer structural transition for inducing ribozyme activity upon binding NS5B to the aptamer, and sequence of ribozyme targeting +382 nt of HCV IRES. Noticeably, we employed in vitro selection technology to identify the most appropriate communication module sequence which can induce ribozyme activity depending on the US5B protein. We demonstrated that the ribozyme was nonfunctional either in the absence of any proteins or in the presence of control bovine serum albumin. In sharp contrast, the allosteric ribozyme can induce activity of cleavage reaction with HCV IRES RNA in the presence of the HCV NS5B protein. This allosteric ribozyme can be used as lead compound for specific and effective anti-HCV agent, tool for highthroughput screening to isolate lead chemicals for HCV therapeutics, and ligand for biosensor system for HCV diagnosis.

• THE JOURNAL OF KOREAN ORIENTAL ONCOLOGY
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• v.6 no.1
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• pp.81-97
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• 2000

Objectives: This experimental study was carried out to evaluate the effects of aqueous and methanol extracts of Hedyotis diffusa which has long been used for cancer treatment in oriental medicines on the induction of apoptotic cell death in human lymphoid leukemia cell line, HL-60. Methods: Cells were treated with various concentrations (200 to $0.4{\mu}g$) and periods (6 to 30 hr) of $H_2O$ and methanol extracts of Hedyotis diffusa. Then, cells were tested for viability by MTT assay. Cells wrere treated with $200{\mu}g/ml$ of methanol extract fork various periods. Genomic DNA was isolated, separated, on 1.5% agarose gels, stained with ethidium bromide and visualized under UV light. Cells were treated with $200{\mu}g/ml$ of each extract for 16 hr. Then, cells were treated with Hoechst dye 33342 and observed by fluorescence microscopy. Cells were treated with various doses of each for 12 hr and $100{\mu}g/ml$ of methanol extract for various periods. Lysate from the cells used to measure the activity of Caspase-1 and-3 proteases by using fluorogenic peptide substrates including acetyl-YVAD-AMC and acetyl-DEVD-AMC, respectively. Cells were treated with $200{\mu}g/ml$ of each extract for various periods. Cell lysates were immunoprecipated with anti-JNKl antibodies. The immune complex was reacted with $32^p-ATP$ and c-Jun as a substrate. The phosphotransferase activity of JNKI was measured by using PhosphoImage analyzer (Fuji Co., Japan). Nuclear extracts were isolated and incubated with oligonucleotide probe of $NF-{\kappa}B$. Transcriptional activation of ${\kappa}B$ was measured by using EMSA and visualized by PhosphoImage analyzer (Fuji Co, Japan). Cell lysates were prepared and analyzed by Western blotting with anti-Bc12 antibodies and anti-Bax antibodies. Cells were pretreated with various doses of methanol extract for 2 hr. Then, the extract was removed by centrifugation. Cells were resuspended with RPMI-1640 media containing 0.3% agarose, 10% FBS, overlayred onto bottom layer agarose and incubated at $CO_2$ incubator for 6 days. The number of colony was counted under light microscopy ($\time100$). Results: The death of HL-60 cells was markedly induced by the addition of methanol extract of Hedyotis diffusa in a dose and time-dependent manners. The apoptotic characteristic ladder pattern of DNA strand break was observed in death of HL-60 cells. In addition, it was shown nucleus chromatin condensation and fragmentation under Hoechst staining. Therefore, Hedyotis diffusa extract-induced death of HL-60 cells is mediated by apoptotic signaling processes. The activity of Caspase 3-like proteases remained in a basal level in HL-60 cells treated with aqueous extract of Hedyotis diffusa. However, it was markedly increased in HL-60 cells treated with methanol extract of Hedyotis diffusa. In addition, the phosphotransferase activity of JNKl was increased in HL-60 cells treated with methanol extract of Hedyotis diffusa. Furthermore, the activation of transcriptional activator, $NF-{\kappa}B$ was markedly induced by methanol extract of Hedyotis diffusa. Anti-apoptotic Bc12 was cleaved into 23Kda fragment by treatment of methanol extract of Hedyotis diffusa. However, expression of proapoptotic Bax protein was increased by treatment of methanol extract of Hedyotis diffusa in a time-dependent manner. Furthermore, methanol extract markedly inhibited the colony forming efficiency of HL-60 cells in semisolid agar culture. Conclusions: Above results suggest that methanol extract of Hedyotis diffusa induces the apoptotic death of human leukemic HL-60 cells via activations of Caspase-3 proteases, JNKI, transcriptional activator $NF-{\kappa}B$, In addition, our results also suggest that methanol extract of Hedyotis diffusa reduces the malignant potential of HL-60 cells via down regulation of colony forming effciency through cleavage of Bc12 as well as induction of Bax.

• Journal of Life Science
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• v.18 no.1
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• pp.120-128
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• 2008

The death ligand tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)/ Apo1L is a cytokine that activates apoptosis through cell surface death receptors. TRAIL has sparked growing interest in oncology due to its reported ability to selectively trigger cancer cell death. Euphorbiae humifusae Wind has been used in traditional Oriental medicine as a folk remedy used for the treatment of cancer. However, the mechanism responsible for the anticancer effects of E. humifusae not clearly understood. Here, we show that treatment with subtoxic doses of water extract of E. humifusae (WEEH) in combination with TRAIL induces apoptosis in TRAIL-resistant human gastric carcinoma AGS cells. Combined treatment with WEEH and TRAIL induced chromatin condensation and sub-G1 phase DNA content. These indicators of apoptosis were correlated with the induction of caspase activity that resulted in the cleavage of poly (ADP-ribose) polymerase. Combined treatment also triggered the loss of mitochondrial membrane potential. Furthermore, co-treatment with WEEH and TRAIL down-regulated the protein levels of the anti-apoptotic proteins such as Bcl-2, Bcl-xL, XIAP and cIAP-1. Although more study will be needed to examine the detailed mechanisms, this combined treatment may offer an attractive strategy for safely treating gastric adenocarcinomas and the results provide important new insights into the possible molecular mechanisms of the anticancer activity of E. humifusae.

• Economic and Environmental Geology
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• v.45 no.4
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• pp.385-396
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• 2012

The geochemical high-grade uranium anormal zone has been reported in the Shinbo mine and its eastern areas, Jinan-gun, Jeollabuk-do located in the southwestern part of Ogcheon metamorphic zone, Korea. In this paper is reported the time-relationship between deformation and growth of metamorphic minerals in the eastern area of Shinbo mine, which consists of the Precambrian metasedimentary rocks (quartzite, metapelite, metapsammite) and the age-unknown pegmatite and Cretaceous porphyry which intrude them, and is considered the relative mineralization time on the basis of the previous research's result. The D1 deformation formed the straight-type Si internal foliation which is defined mainly as the arrangement of elongate quartz, biotite, opaque mineral in andalusite porphyroblast. The D2 deformation, which is defined by the microfolding of Si foliation, formed S2 crenulation cleavage. It can be divided into two sub-phases, early crenulation and late crenulation. The former occurs as the curvetype Si foliation in the mantle part of andalusite. The latter occurs as S1-2 composite foliation which warps around the andalusite. The andalusite porphyroblast began to grow under non-deformation condition after the formation of S1 foliation which corresponds to the straight-type Si foliation. It continued to grow before the late crenulation phase. The age-unknown pegmatite intruded after the D2 deformation and grew the fibrous sillimanite which random masks the S1-2 composite foliation. The D3 deformation formed F3 fold which folded the S1-2 composite foliation, D2 crenulation, fibrous sillimanite. It means that the intrusion of pegmatite related to the growth of the fibrous sillimanite took place during the inter-tectonic phase of D2 and D3 deformations. The retrograde metamorphism is recognized by the chloritization of biotite and two-way cleavage lamellae which is parallel to the S1-2 composite foliation and the F3 fold axial surface in the andalusite porphyroblast. It occurred during the D2 late crenulation phase and D3 deformation. In considering of the previous research's result inferring the most likely candidate for the uranium source rock as pegamatite, it indicates that the age-unknown pegmatite intruded during the inter-tectonic phase of D2 and D3 deformations, i.e. during the retrograde metamorphism related to the uplifting of crust, and formed the uranium ore zone around the Shinbo mine.

• Journal of Dental Rehabilitation and Applied Science
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• v.26 no.2
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• pp.121-143
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• 2010

To investigate the effect of implant types and bone resorption on the fracture characteristics. 4 types of Osstem$^{(R)}$Implant were chosen and classified into external parallel, internal parallel, external taper, internal taper groups. Finite elements analysis was conducted with ANSYS Multi Physics software. Fatigue fracture test was performed by connecting the mold to the dynamic load fatigue testing machine with maximum load of 600N and minimum load of 60N. The entire fatigue test was performed with frequency of 14Hz and fractured specimens were observed with Hitachi S-3000 H scanning electron microscope. The results were as follows: 1. In the fatigue test of 2 mm exposed implants group, Tapered type and external connected type had higher fatigue life. 2. In the fatigue test of 4 mm exposed implants group, Parallel type and external connected types had higher fatigue life. 3. The fracture patterns of all 4 mm exposed implant system appeared transversely near the dead space of the fixture. With a exposing level of 2 mm, all internally connected implant systems were fractured transversely at the platform of fixture facing the abutment. but externally connected ones were fractured at the fillet of abutment body and hexa of fixture or near the dead space of the fixture. 4. Many fatigue striations were observed near the crack initiation and propagation sites. The cleavage with facet or dimple fractures appeared at the final fracture sites. 5. Effective stress of buccal site with compressive stress is higher than that of lingual site with tensile stress, and effective stress acting on the fixture is higher than that of the abutment screw. Also, maximum effective stress acting on the parallel type fixtures is higher. It is careful to use the internal type implant system in posterior area.

• Current Research on Agriculture and Life Sciences
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• v.32 no.1
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• pp.1-9
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• 2014

In contrast to conventional microwaves, QRD (Quadratic Residue Diffusor) microwaves are a new energy-efficient technology that enhances the effect of sterilization based on changing the wavelength phase difference. Therefore, this study investigated the sterilization of wood using environmentally friendly and low energy consuming QRD microwaves. The results are as follows: for the QRD microwaves used in this study, the efficiency E = 5.75e0.32 S ($R^2$=0.908). Although the early water content was not constant, the average water content was 30.3% and after one week of natural drying, the water content was 22.6%, representing an average water content reduction of about 8%. When increasing the microwave level from 3 kW ~ 9 kW, the time taken for the temperature to increase was reduced. After the QRD microwave treatment, the wood samples showed no change in their flexural rigidity, compressive strength, or cleavage. The QRD microwave levels used in the experiments were 3, 5, 7, and 9 kW, where 9 kW was found to be the most efficient. Thus, for the purpose of eliminating nematodes and termites inside wood, a higher QRD microwave level was found to be more effective and energy efficient.

• The Journal of the Petrological Society of Korea
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• v.7 no.3
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• pp.190-206
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• 1998

The microstructures and time-relationship between deformation and growth of metamorphic minerals(metamorphism) of the Paleozoic metasedimentary rocks(Joseon Supergroup and Pyeongan Group) in the Janggunbong area at the central-south part in the North Sobaegsan Massif, Korea, have been analyzed in this paper. The first phase metamorphism (low-pressure type metamorphism), recognized as the crystallization of stack-type chloritoid and biotite and augen-type old andalusite, occurred under non-deformational condition before D1 deformation related to the formation of an E-W trending isocline-synclinal fold(Janggunbong fold) and associated its axial plane S1 foliation, and produced regional mineralogical zoning of E-W trend in the Paleozoic rocks. The second phase metamorphism(medium-pressure type metamorphism), related to the growth of staurolite and garnet porphyroblasts with straight or curved internal foliations(Si), occurred under non-deformational condition after D1 deformation related to the formation of E-W trending thrusts modifying the Janggunbong fold and during D2 deformation related to the formation of E-W trending Yecheon shear zone. This metamorphism also produced regional mineralogical zoning of E-W trend. After D2 deformation occurred the intrusion of Jurassic Chunyang granite and associated its contact metamorphism which crystallized patchy-type young andalusite and prismatic- or fibrous-type sillimanite and coarse-grained garnet. This metamorphism occurred under non-deformational condition before D3 deformation related to the formation of S3 crenulation cleavage and during early phase of D3 deformation, and formed narrow mineralogical zoning of N-S trend near Chunyang granite.

• Journal of Life Science
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• v.18 no.9
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• pp.1177-1185
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• 2008

In the present work, we show that spermine (spm)-induced cytotoxicity is due to the mitochondrial-dependent pathway triggered by the intracellular $Ca^{2+}$ increase in MCF-7 human breast cancer cells. Spm induced the intracellular $Ca^{2+}$ increase in a dose-dependent manner in the medium containing 1.5 mM $Ca^{2+}$. Even in the $Ca^{2+}$-free medium, spm could induce a minor $Ca^{2+}$ increase in a dose-dependent fashion, suggesting a probable leak from the internal storage. The cytotoxic effect of $Ca^{2+}$ could be further proved by using either BAPTA or ionophore. Spm-induced $Ca^{2+}$ increase led to the release of cytochrome c from mitochondria into the cytosol and the change of mitochondrial membrane potential. In MCF-7 cells, caspase-7 plays a key role in the downstream of apoptosis because caspase-3 is absent. In the cells treated with spm, the cleavage of caspase-7 and -12 was increased almost two-fold. The level of anti-apoptotic Bcl-2 protein decreased to 35% of the control; however, the cells showed increased expression of pro-apoptotic Bax protein about two-fold in response to spm. These results imply that the apoptotic signaling pathway activated by spm is likely to be mediated via the mitochondrial-dependent pathway.