• Korean Journal of Plant Resources
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• v.28 no.3
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• pp.297-304
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• 2015

The flower buds of Sophora japonica L (SF), as a well-known traditional Chinese medicinal herb, have been used to treat bleeding-related disorders such as hematochezia, hemorrhoidal bleeding, dysfunctional uterine bleeding, and diarrhea. However, no specific anti-cancer effect and its molecular mechanism of SF have been described. Thus, we performed in vitro study to investigate if treatment of SF affects activating transcription factor 3 (ATF3) expression and ATF3-mediated apoptosis in human colorectal cancer cells. The effects of SF on cell viability and apoptosis were measured by MTT assay and Western blot analysis against cleaved poly (ADP-ribose) polymerase (PARP). ATF3 activation induced by SF was evaluated using Western blot analysis, RT-PCR and ATF3 promoter assay. SF treatment caused decrease of cell viability and increase of apoptosis in a dose-dependent manner in HCT116 and SW480 cells. Exposure of SF activated the levels of ATF3 protein and mRNA via transcriptional regulation in HCT116 and SW480 cells. Inhibition of extracellular signal-regulated kinases (ERK) 1/2 by PD98059 and p38 by SB203580 attenuated SF-induced ATF3 expression and transcriptional activation. Ectopic ATF3 overexpression accelerated SF-induced cleavage of PARP. These findings suggest that SF-mediated apoptosis may be the result of ATF3 expression through ERK1/2 and p38-mediated transcriptional activation.

• Journal of Veterinary Clinics
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• v.25 no.5
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• pp.355-358
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• 2008

The aim of this study was to investigate whether trophic factor supplementation (TFS) enhance the survival of kidney cell during cold ischemic storage and rewarming. The effect of TFS on the phosphorylation of p44/42 and p38 mitogen activated protein kinases (MAPK) signaling pathway was determined by Western blot. Apoptotic changes after cold ischemic storage and rewarming was determined by 4',6'-diamino-2-phenylindole (DAPI) staining. The cell viability was evaluated by live assay. TFS significantly decreased p44/42 and p38 MAPK activity induced by cold ischemic injury and rewarming (p < 0.05). The number of apoptotic cells was decreased after 5 minute rewarming in the presence of TFS. TFS significantly increased the cell viability after 5 minute rewarming (p < 0.05). Therefore, it was concluded that trophic factor supplementation protects kidney tubule cells from cold ischemic and rewarming injury via the inhibition of p44/42 and p38 MAPK activation and reducing apoptotic change.

• Animal cells and systems
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• v.16 no.4
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• pp.274-279
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• 2012

Thymoquinone (TQ), a drug extracted from the black seeds of Nigella sativa, has been shown to exhibit anti-inflammatory, anti-oxidant, and anti-neoplastic effects in numerous cancer cells. The effects of TQ on cyclooxygenase-2 (COX-2) expression and prostaglandin $E_2$ ($PGE_2$) production in MDA-MB-231, however, remain poorly understood. Western blot analysis and immunofluorescence staining were performed to study the expression levels of inflammation regulatory proteins in MDA-MB-231. $PGE_2$ assay was conducted to explore the TQ-induced production of $PGE_2$. In this study, we investigated the effects of TQ on COX-2 expression and $PGE_2$ production in MDA-MB-231. TQ significantly induced COX-2 expression and increased $PGE_2$ production in a dose-dependent manner, as determined by a Western blot analysis and $PGE_2$ assay. Furthermore, the activation of Akt and p38 kinase, respectively, was up-regulated in TQ treated cells. Inhibition of p38 kinase with SB203580 and PI3kinase (PI3K) with LY294002 abolished TQ-caused COX-2 expression and decreased $PGE_2$ production. These results collectively demonstrate that TQ effectively modulates COX-2 expression and $PGE_2$ production via PI3K and p38 kinase pathways in the human breast cancer cell line MDA-MB-231.

• Journal of Microbiology
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• v.42 no.2
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• pp.117-125
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• 2004

Propionibacterium acnes (P. acnes) plays an important role in the disease pathogenesis of acne vulgaris, a disorder of pilosebaceous follicles, seen primarily in the adolescent age group. In the present study, the presence of antibodies against P. acnes (MTCC1951) were detected in acne patient (n=50) and disease free controls (n=25) using dot-ELISA and Western blot assay. The ability of P. acnes to induce pro-inflammatory cytokines by human peripheral blood mononuclear cells (PBMCs), obtained from acne patients and healthy subjects, were also analysed. The patients (n=26) who were culture positive for skin swab culture, were found to have a more advanced disease and higher antibody titres (1:4000 to >1:16000) compared to the P. acnes negative patients (n=24) and normal controls (n=25). An analysis of patients' sera by western blot assay recognized a number of antigenic components of P. acnes, rang-ing from 29 to 205 kDa. The major reactive component was an approximately 96 kDa polypeptide, which was recognised in 92％ (24 of 26) of the patients sera. Further, the P. acnes culture supernatant, crude cell lysate and heat killed P. acnes whole cells, obtained from 72-h incubation culture, were observed to be able to induce significant amounts of IL-8 and tumor necrosis factor alpha (TNF-${\alpha}$) by the PBMCs in both the healthy subjects and patients, as analysed by cytokine-ELISA. The levels of cytokines were significantly higher in the patients than the healthy subjects. A major 96 kDa polypep-tide reactant was eluted from the gel and was found to cause dose dependent stimulation of the pro-ductions of IL-8 and TNF-${\alpha}$. Thus, the above results suggest that both humoral and pro-inflammatory responses play major roles in the pathogenesis of acne.

• Journal of Dental Anesthesia and Pain Medicine
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• v.19 no.2
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• pp.91-99
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• 2019

Background: The imbalance between osteoblasts and osteoclasts can lead to pathological conditions such as osteoporosis. It has been reported that opioid adversely affect the skeletal system, but it is inconsistent. Remifentanil is currently used as an adjuvant analgesic drug in general anesthesia and sedation. The aim of the present study was to investigate the effect of remifentanil on the osteoblast differentiation and mechanism involved in this effect. Methods: The C2C12 cells (mouse pluripotent mesenchymal cell line) were used as preosteoblast. Osteoblastic differentiation potency was determined by alkaline phosphatase (ALP) staining. C2C12 cell migration by remifentanil was evaluated using Boyden chamber migration assay. The expression of Runx2 and osterix was evaluated by RT-PCT and western blot analysis to investigate the mechanism involved in remifentanil-mediated osteoblast differentiation. Results: ALP staining showed that remifentanil increased significantly osteoblast differentiation. In Boyden chamber migration assay, C2C12 cell migration was increased by remifentanil. RT-PCR and western blot analysis showed that the expression of Runx2 and osterix was upregulated by remifentanil. Conclusions: We demonstrated that remifentanil increased osteoblast differentiation in vitro by upregulation of Runx2 and osterix expression. Therefore, remifentanil has the potential for assisting with bone formation and bone healing.

• The Korea Journal of Herbology
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• v.36 no.1
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• pp.41-49
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• 2021

Objective : Citrus unshiu peel (Citri Unshius Pericarpium) has been prescribed to suppress coughing and phlegm in Korean medicine. In this study, the effect of ethanol extract of Citrus unshiu peel (CEE) on apoptosis was investigated using cadmium chloride (CdCl2) treated HepG2 cells. Methods : CEE was prepared by extracting 300 g of Citri Unshius Pericarpium in 3 L of ethanol for 72 h. Apoptosis was determined by the TUNEL assay. The mitochondrial membrane potential (MMP) was monitored using the membrane-permeable fluorescent dye Rh123. The expression level of each protein was monitored by Western blot analysis. Results : CEE protected HepG2 cells from apoptosis as determined by the TUNEL assay. A decrease in MMP was observed in cells exposed to cadmium, indicating that mitochondria are involved in the induction of apoptosis. However, CEE recovered the reduction in MMP caused by cadmium. In addition, decreased expression of B-cell lymphoma 2 (Bcl-2), procaspase, and poly(ADP-ribose) polymerase (PARP) by cadmium was increased by CEE. The anti-apoptotic effect of CEE was found to be associated with inhibition of JNK and p38 phosphorylation when examining the expression of phosphorylated MAPK by Western blot. Conclusion : This study showed that CEE exerted anti-apoptotic effects in cadmium-induced HepG2 cells by inhibiting the reduction of MMP and changes in the expression level of apoptotic proteins. These results suggest the potential for CEE to be used for heavy metal-induced liver damage.

• The Korea Journal of Herbology
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• v.33 no.3
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• pp.19-24
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• 2018

Objectives : Hedgehog skin is one of the animal medicines in Traditional Korean Medicine for hematochezia and hemorrhoids. In this study, we examined cytotoxicity and anti-inflammatory effects. Methods : Cytotoxicity of hedgehog skin extracts was measured by MTT assay in vitro. We investigated the inhibition of lipopolysaccharide (LPS) stimulated nitric oxide (NO) production in RAW 264.7 cells. The phosphorylation of mitogen-activated protein kinases (MAPKs) was measured by western blot. And we observed the effect of hedgehog skin extracts on the expression of IL-6 genes using real time PCR. Results : As a result of MTT assay for cytotoxicity, there were no significant differences between non-treatment group and hedgehog skin extracts treatment groups. $500{\mu}g/m{\ell}$ of hedgehog skin extracts treatment significantly decreased nitric oxide production in comparison with non-treatment in LPS-induced RAW 264.7 cells. In measurement of the phosphorylation of MAPKs using western blot analysis, LPS stimulation increased the phosphorylation of MAPKs and $500{\mu}g/m{\ell}$ of hedgehog skin extracts treatment decreased the phosphorylation of ERK1, ERK2 and p38 significantly. But there were no significant differences the phosphorylation of JNK1 and JNK2. As a result of confirmation of the IL-6 mRNA gene expression using real time PCR, IL-6 mRNA gene expressions were significantly decreased in $50{\mu}g/m{\ell}$, $100{\mu}g/m{\ell}$ and $500{\mu}g/m{\ell}$ hedgehog skin extracts treated groups by comparison with non-treatment group. Conclusion : These results could provide a mechanistic explanation for the anti-inflammatory effects of the hedgehog skin.

• The Korea Journal of Herbology
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• v.34 no.6
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• pp.117-124
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• 2019

Objective : Broccoli is edible green plant that has a wide variety of health benefits including cancer prevention and cholesterol reduction. However, leaves of broccoli are not eaten and are mostly left as waste. This study was conducted to evaluate the effects of the broccoli leaf extract (BLE) on prostaglandin E₂ (PGE₂) production related to nuclear factor kappa B (NF-κB) signaling in lipopolysaccharide (LPS)-activated macrophages. Methods : BLE was prepared by extracting dried leaf with ethanol. Cell viability was determined by 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. PGE₂ and inflammatory cytokines were detected by enzyme-linked immunosorbent assay (ELISA). Expression level of each protein was monitored by Western blot analysis. Results : In LPS-activated Raw264.7 cells, PGE₂ release into culture medium was dramatically enhanced compared to control cells. However, increased PGE₂ was attenuated dose-dependently by treatment with BLE. Inhibition of PGE₂ production by BLE was due to the suppression of cyclooxygenase-2 (COX-2) expression determined by Western blot analysis. BLE also inhibited the production of inflammatory cytokines such as interleukin-6 (IL-6) and tumor necrosis factor-alpha (TNF-α). Inhibition at PGE₂ and cytokine was mediated from inhibition of nuclear translocation of NF-κB due to the repression of inhibitory kappa B alpha (IκBα) phosphorylation and degradation. Conclusion : This study showed that BLE exerted inhibitory activities against PGE₂, which is critical for the initiation and resolution of inflammatory responses, and that inhibition of PGE₂ was mediated by suppression of NF-κB signaling. These results suggest that the waste broccoli leaves could be used for controlling inflammation.

• The Korea Journal of Herbology
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• v.34 no.4
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• pp.19-25
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• 2019

Objectives : Thymol (2-isopropyl-5-methylphenol), a natural monoterpenoid phenol, is one of the major odorant constituents found in natural essential oils of various herbal plants, such as Thymus quinquecostatus and Thymus vulgaris. Multiple biological activities of thymol, including antioxidative, antimicrobial, and anti-inflammatory effects, have been reported in numerous in vitro studies, and recently it was suggested that thymol may could inhibit oxidization of L-dihydroxyphenylalanine (L-DOPA) to dopaquinone required in melanogenesis pathway, as an antioxidant. Methods : MTT assay was performed to test the cytotoxic effect of thymol in B16F10 cells. Inhibitory effect of thymol to tyrosinase activities were examined using both mushroom tyrosinase and intracellular tyrosinase. Expression level of tyrosinase in B16F10 cells were investigated by western blot analysis. Results : The cell viability was decreased by thymol treatment in dose-dependant manner, leading significant cytotoxicity in 500 and $1000{\mu}M$ thymol-treated groups. In the alpha-melanocyte stimulating hormone (${\alpha}$-MSH)-induced melanogenesis, administration of thymol significantly decreased extracellular (secreted) melanin content in dose-dependent manner. Cellular tyrosinase activity assay and western blot analysis of intracellular tyrosinase showed that thymol has a strong anti-melanogenic effect by inhibition of tyrosinase activity and by decreasing expression of tyrosinase that contribute to melanin synthesis in the B1610 cells. Conclusions : As the first functional study that prove anti-melanogenic effect of thymol and its underlying mechanism in the living cells, our study suggests the applicability of fragrance as the functional materials of cosmetics or health supplement, not as just an additive.

• Journal of Pharmacopuncture
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• v.25 no.4
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• pp.390-395
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• 2022

Objectives: Sweet bee venom (sBV) is purified from Apis mellifera, containing a high level of melittin-its main component. It has been used as a therapeutic agent for pain relief and anti-inflammation, as well as for treating neuronal abnormalities. Recently, there have been studies on the therapeutic application of sBV for anticancer treatment. In the present study, we investigated the pharmacological effect of sBV treatment in A549 human lung cancer cells. Methods: We used microscopic analysis to observe the morphological changes in A549 cells after sBV treatment. The MTT assay was used to examine the cytotoxic effect after dose-dependent sBV treatment. Molecular changes in sBV were evaluated by the expression of apoptosis marker proteins using western blot analysis. Results: Microscopic analysis suggested that the growth inhibitory effect occurred in a dose-dependent manner; however, cell lysis occurred at a concentration over 20 ㎍/mL of sBV. The MTT assay indicated that sBV treatment exhibited a growth inhibitory effect at a concentration over 5 ㎍/mL. On fluorescence activated cell sorting analysis, G0 dead cells were observed after G1 arrest at treatment concentrations up to 10 ㎍/mL. However, rapid cell rupture was observed at a concentration of 20 ㎍/mL. Western blot analysis demonstrated that sBV treatment modulated the expression of multiple cell death-related proteins, including cleaved-PARP, cleaved-caspase 9, p53, Bcl2, and Bax. Conclusion: sBV induced cell death in A549 human lung cancer cells at a pharmacological concentration, albeit causing hemolytic cell death at a high concentration.