• Asian Pacific Journal of Cancer Prevention
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• 제15권14호
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• pp.5667-5672
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• 2014

Folate receptor alpha ($FR{\alpha}$) mediates folate uptake by endocytosis, and while folate is essential to DNA methylation and synthesis and may have an important role in proliferating cells. $FR{\alpha}$ is known to be expressed in rapidly proliferating cells, including many cancer cell lines, but there has been no systematic assessment of expression in cervical cancer cell lines. The aim of the present study was to evaluate the effects of $FR{\alpha}$ on proliferation and apoptosis of cervical cells and correlation mechanism. In this study, we investigated the biological function of $FR{\alpha}$ in Hela cells using RNA interference. Cell proliferation was evaluated by Cell Counting Kit-8 (CCK8) assay, while cell cycling and apoptosis were assessed by flow cytometry, mRNA levels by real time-PCR and protein levels of $FR{\alpha}$, c-Fos and c-Jun by Western blotting. The results revealed that $FR{\alpha}$ was highly expressed in Hela cells and its silencing with a small interfering RNA (siRNA) inhibited cell proliferation and induced cell apoptosis, arresting the cell cycle in G0/G1 stages while decreasing the proportion in S and G2/M stages, and suppressed the expression levels of c-Fos and c-Jun. In conclusion, the results of this study indicated that $FR{\alpha}$ down-regulation might be capable of suppressing cervical cancer cell proliferation and promoting apoptosis. It suggested that $FR{\alpha}$ might be a novel therapeutic target for cervical cancer.

• 한국임상수의학회지
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• 제34권3호
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• pp.172-177
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• 2017

Fucoidan, a cell wall polysaccharide found in the brown seaweed, is reported to have broad-spectrum biological activities. The objectives of this study were to examine the effect of fucoidan on prostaglandin $E_2$ ($PGE_2$) and cyclooxygenase-2 (COX-2) expression in lipopolysaccharide (LPS)-stimulated porcine peripheral blood mononuclear cells (PBMCs) and to determine whether these effects are involved in Akt activation. The levels of $PGE_2$ production in the culture supernatants from PBMCs were determined by the enzyme-linked immunosorbent assay (ELISA) kit and the levels of COX-2 mRNA were measured by real time polymerase chain reaction (RT-PCR). Akt activity was determined by Western blot analysis. Fucoidan in LPS-$na{\ddot{i}ve}$ PBMCs has no effect on $PGE_2$ production and COX-2 mRNA expression. Furthermore, fucoidan does not affect Akt activation in LPS- $na{\ddot{i}ve}$ PBMCs. However, $PGE_2$ production and COX-2 mRNA expression on PBMCs were remarkably enhanced by LPS stimulation. Akt activity was also increased by LPS. Increasing effects of $PGE_2$ production and COX-2 mRNA expression in PBMCs induced by LPS were suppressed by addition of fucoidan. In addition, fucoidan reduced an increase in Akt activity in LPS-stimulated PBMCs. These results suggested that fucoidan exerts potent anti-inflammatory properties by suppression of $PGE_2$ production, COX-2 mRNA expression and Akt activation in LPS-stimulated PBMCs.

• Journal of Radiation Protection and Research
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• 제37권2호
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• pp.56-62
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• 2012

To determine the biological effects of low-dose-rate radiation ($^{137}Cs$, 2.95 mGy/h) on EL4 lymphoma cells during 24 h, we investigated the expression of genes related to apoptosis, cell cycle arrest, DNA repair, iron transport, and ribonucleotide reductase. EL4 cells were continuously exposed to low-dose-rate radiation (total dose: 70.8 mGy) for 24 h. We analyzed cell proliferation and apoptosis by trypan blue exclusion and flow cytometry, gene expression by real-time PCR, and protein levels with the apoptosis ELISA kit. Apoptosis increased in the Low-dose-rate irradiated cells, but cell number did not differ between non- (Non-IR) and Low-dose-rate irradiated (LDR-IR) cells. In concordance with apoptotic rate, the transcriptional activity of ATM, p53, p21, and Parp was upregulated in the LDR-IR cells. Similarly, Phospho-p53 (Ser15), cleaved caspase 3 (Asp175), and cleaved Parp (Asp214) expression was upregulated in the LDR-IR cells. No difference was observed in the mRNA expression of DNA repair-related genes (Msh2, Msh3, Wrn, Lig4, Neil3, ERCC8, and ERCC6) between Non-IR and LDR-IR cells. Interestingly, the mRNA of Trfc was upregulated in the LDR-IR cells. Therefore, we suggest that short-term Low-dose-rate radiation activates apoptosis in EL4 lymphoma cells.

• 대한한방부인과학회지
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• 제24권2호
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• pp.13-32
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• 2011

Objectives: This study was performed to elucidate the effects of Danchisoyo-san (DS) on cell damage and gene expression in UVB-exposed dermal fibroblast. Methods: To demonstrate the inhibitory effects of DS on aging of the skin, we used human dermal fibroblast(F6) and UVB light(30 mJ/$cm^2$) was used to damage to dermal fibroblast. We measured the nitrite production, LDH release, and gene expression in UVB-irradiated dermal fibroblast to elucidate the actionmechanism of DS. Also, we evaluated the amount of increased PICP, TIMP-1 in dermal fibroblast. PICP, TIMP-1 concentration was measured using EIA kit, and gene expression (MMP-1, procollagen, c-fos, c-jun, NF-kB, Bcl-2, Bcl-xL, iNOS) were determined using real-time PCR. Results: 1. DS inhibited LDH-release, nitrite production in UVB-irradiated dermal fibroblast. 2. DS suppressed the gene expression of MMP-1 in UVB-irradiated dermal fibroblast. 3. DS increased the gene expression of procollagen in UVB-iradiated dermal fibroblast. 4. DS suppressed the gene expression of c-jun, c-fos, NF-kB, iNOS in UVBirradiated dermal fibroblast. 5. DS increased the gene expression of Bcl-2 in UVB-iradiated dermal fibroblast. 6. DS increased the cell proliferation of dermal fibroblast. Conclusions: From the results, we concluded DS increases the cell proliferation and collagen synthesis in dermal fibroblast. So we suggest that DS has the antiwrinkle effects.

• 대한본초학회지
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• 제28권1호
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• pp.33-42
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• 2013

Objective : Morus alba Linne Root Bark (MRAL) is a medicinal herb in Korean Medicine, known for its anti-inflammatory and anti-allergic properties. However, its mechanisms of action and the cellular targets have not yet been found and the study was developed to investigate the allergic suppressive effect of MRAL. The purpose of this study is to investigate the allergic suppressive effects of MRAL on activation of MC/9 mast cells. Methods : Cytotoxic activity of MRAL (50, 100, 200, 400 ${\mu}g/mL$) on MC/9 mast cells measured using EZ-Cytox cell viability assay kit (WST reagent). The levels of interleukin-5 (IL-5), IL-13 and IL-4, IL-5, IL-6, IL-13 mRNA expression were measured by enzyme-linked immunosorbent assay (ELISA) and real-time PCR respectively. The expression of transcription factors such as GATA-1, GATA-2, NFAT, AP-1 and NF-${\kappa}B$ p65 DNA binding activity were measured by western blot and electrophoresis mobility shift assay (EMSA). Results : Our results indicated that MRAL (50 ${\mu}g/mL$, 100 ${\mu}g/mL$) significantly inhibited PMA/Ionomycin-induced production of IL-5 and IL-13 and the expression of IL-4, IL-5, IL-6 and IL-13 mRNA in MC/9 mast cells. Moreover, MRAL (50 ${\mu}g/mL$, 100 ${\mu}g/mL$) inhibited PMA/Ionomycin-induced GATA-1, GATA-2, NFAT-1, NFAT-2, c-Fos protein expression and NF-${\kappa}B$ p65 DNA binding activity in MC/9 mast cells. Conclusions : In conclusion, we suspect the anti-allergenic activities of MRAL, may be related to the regulation of transcription factors GATA-1, GATA-2, NFAT-1, NFAT-2, c-Fos and NF-${\kappa}B$ p65 DNA binding assay causing inhibition of Th2 cytokines IL-5 and IL-13 in mast cells.

• The Korean Journal of Physiology and Pharmacology
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• 제27권3호
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• pp.197-208
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• 2023

Dysregulation of certain long non-coding RNAs may facilitate tumor initiation and progression. However, numerous carcinogenesis-related long noncoding RNAs have not been characterized. The goal of this study was to elucidate the role of LINC00562 in gastric cancer (GC). The expression of LINC00562 was analyzed using real-time quantitative PCR and Western blotting. The proliferative capacity of GC cells was determined using Cell Counting Kit-8 and colony-formation assays. The migration of GC cells were evaluated using wound-healing assays. The apoptosis of GC cells was assessed by measuring the expression levels of apoptosis-related proteins (Bax and Bcl-2). Xenograft models in nude mice were constructed for in vivo functional analysis of LINC00562. The binding relationship between miR-4636 and LINC00562 or adaptor protein complex 1 sigma 3 (AP1S3), obtained from public databases, was confirmed using dual-luciferase and RNA-binding protein immunoprecipitation experiments. LINC00562 was expressed in GC cells at high levels. Knockdown of LINC00562 repressed GC cell growth and migration, promoted apoptosis in vitro, and inhibited tumor growth in nude mouse models. LINC00562 directly targeted miR-4636, and miR-4636 depletion restored the GC cell behavior inhibited by LINC00562 absence. AP1S3, an oncogene, binds to miR-4636. MiR-4636 downregulation increased AP1S3 level, restoring GC cell malignant behaviors inhibited by AP1S3 downregulation. Thus, LINC00562 exerts carcinogenic effects on GC development by targeting miR-4636-mediated AP1S3 signaling.

• 대한화장품학회지
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• 제43권2호
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• pp.157-164
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• 2017

스테비아(Stevia rebaudiana)는 남아메리카 지역이 원산지인 국화과 스테비아 속의 다년생 식물로 스테비올(steviol)을 기본 구조로 하는 다양한 배당체가 존재하며 스테비오사이드(stevioside)와 리바우디오사이드(rebaudioside) A 등이 주성분이다. 스테비올 배당체들은 설탕보다 단맛이 월등히 뛰어나 감미료로 널리 사용되어지고 있다. 최근 여러 논문들에서 스테비올 배당체들이 미백 및 항염 효과 뿐 아니라 피부장벽 타이트정션 단백질 조절에 연관되어 있다는 보고가 있었다. 따라서 본 연구에서는 스테비올 배당체인 리바우디오사이드 A의 항염 효과 연구를 통해 향후 아토피 피부염 개선 화장품 원료 개발 가능성을 확인하고자 하였다. 항염 연구를 위해 마우스 대식세포인 RAW264.7 세포를 이용하여 cell viability 및 염증 유발 사이토카인 mRNA 발현량을 분석하였다. 우선 cell viability 측정을 위해 cell counting kit-8 (CCK-8) assay를 수행하였고 세포독성이 없는 최대 농도를 $250{\mu}M$로 설정하여 이후 모든 실험을 진행하였다. 리바우디오사이드 A의 염증 조절 기능 연구는 주로 정량적 real-time RT-PCR 방법을 이용하였다. LPS에 의해 활성화된 RAW264.7 대식세포에서 리바우디오사이드 A 처리 결과 LPS 처리군 대비 iNOS 발현량은 약 47% 감소하였고, COX-2 또한 41% 감소하였다. 생성된 NO의 양 또한 농도 의존적으로 감소하였다. 대식세포를 LPS로 활성화시킨 조건에서 염증 관련 사이토카인 유전자인 interleukin (IL)-$1{\alpha}$, IL-$1{\beta}$, IL-6의 발현량 조절을 확인한 결과 사이토카인(IL-$1{\alpha}$, $1{\beta}$, 6) 발현이 LPS 처리군 대비 40%, 45%, 59%로 농도 의존적 유의성 있게 감소하였다. 결론적으로 스테비올 배당체인 리바우디오사이드 A는 NO 생성 및 사이토카인 분비 억제를 통해 염증 반응을 저해하였다. 이러한 리바우디오사이드 A의 신규 항염증 조절 기능을 통해 아토피성피부염 개선 소재로의 개발이 기대된다.

• 대한한방내과학회지
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• 제31권2호
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• pp.314-330
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• 2010

Objective : This study was performed to investigate the anti-fibrogenic effect and changes of inflammation-related genes by YBR I and YBR II (YBR I: Arteisiae Capillaris Herba, Atractylodis Rhizoma Alba, Hoelen/ YBR II: YBR I +Sanguisorbae Radix, Biotae Cacumen, Cirsii Japonici Herba) on HSC(hepatic stellate cells)-T6 and TAA-induced rat liver tissue. Materials and Methods : HSC-T6 were treated with various concentrations of distilled-water extract YBR I and YBR II extract for 24, 48 and 72 hours. After the treatment, cell viability, proliferation, procollagen levels and IL-6 levels were measured by using MTT Assay, BrdU Assay, Procollagen Type 1 C-peptide EIA kit, and Murine IL-6 ELISA Development kit. Rat liver fibrosis was induced by intraperitoneal TAA injection of 150mg/kg 3 times a week for 6 weeks. After the treatment, body weight, liver & spleen weights, liver function test, complete blood cell count and change of portal pressure were studied. In addition, gene expressions of ASMA, IL-6, MMP-2, TIMP-1 and TIMP-2, all of which are known to be associated with liver fibrosis, were analyzed by using Real-Time PCR. After YBR I and YBR IItreatment, percentages of collagen in TAA-induced rat liver tissue were measured. Results : The viability and proliferation of the HSC-T6 decreased as the concentration increased. The production of procollagen decreased as the concentration increased. The production of IL-6 was little influenced by YBR I and YBR II. There was no difference in rat body weight between the TAA-only group and the YBR groups. Compared with rat liver weight of TAA-only group, that of the YBR groups increased. In the YBR I group, the serum level of AST elevated by TAA injection significantly decreased and in the YBR I and II group, the serum level of ALP and ALT elevated by TAA injection decreased. In the YBR I group, white blood cell count elevated by TAA injection decreased but platelets increased. In the YBR I group, the portal pressure elevated by TAA injection significantly decreased. Decreases in the gene expression of ASMA and MMP-2 were observed in the YBR I group. The gene expression of IL-6 was little influenced by YBR I and YBR II -treated groups. In the histological finding, TAA injections caused severe fibrosis, but YBR I and YBR II treatment significantly reduced the amounts of hepatic collagens. Conclusions : These results suggest that YBR I and II have inhibitory effects on the hepatic fibrogenesis.

• 대한본초학회지
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• 제28권4호
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• pp.7-16
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• 2013

Objectives : Brassica campestris var narinosa (BCN), Canavalia gladiata DC semen (CGD) and their combinational prescription (BCN+CGD) have been use to demonstrate to regulate hematopoiesis. In the current study, we investigated whether Brassica campestris var narinosa, Canavalia gladiata DC semen and their combinational prescription is related to hemato-potentiating function using Sca-$1^+$ hematopoietic stem cells (Sca-$1^+HSCs$) as a testing system. Methods : Sca-$1^+HSCs$ isolated from femur in C57bl/6 mice with leukopenia and thrombocytopenia induced by cyclophosphamide (CTX). Then, Real-time PCR was performed to measure the mRNA expression, ELISA and haematopoiesis-related gene (EPO, TPO, IL-3, SCF, c-kit, GM-CSF), the phosphorylation of JAK2, GATA-1 and STAT-5a/b were observed by western blot, and the numbers of $CD117^+/Sca-1^+$ cell and the number of granulocyte erythrocyte monocyte macrophage colony-forming units (CFU-GEMM) and erythroid burst forming units (BFU-E), semisolid clonogenic assay was performed. Result : When Sca-$1^+HSCs$ were treated with Brassica campestris var narinosa, Canavalia gladiata DC semen and their combinational prescription with rIL-3/rSCF, the expression of haematopoiesis-related (EPO, TPO, IL-3, SCF, c-kit, and GM-CSF) were significantly increased at the levels of mRNA as well as production in Sca-$1^+HSCs$. Additionally, CGS enhanced phosphorylation of JAK2, GATA-1, and signal transducer and activator of transcription-5a/b (STAT-5a/b) in Sca-$1^+HSCs$. Furthermore, their combinational prescription (BCN+CGD) significantly enhanced the growth rate of granulocyte erythrocyte monocyte macrophage colony-forming units (CFU-GEMM) and erythroid burst forming units (BFU-E) in vitro. Conclusion : These result suggest that Brassica campestris var narinosa (BCN) and Canavalia gladiata DC have hematopoietic enhancement via hematopoietic cytokine-mediated JAK2/GATA-1/STAT-5a/b pathway, and their combinational prescription (BCN+CGD) has superior hematopoietic enhancement to those of individual extracts.

• Animal Bioscience
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• 제36권2호
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• pp.200-208
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• 2023

Objective: Muscle acetylcholine receptors have five alpha subunits (α, β, δ, ε, or γ), and cholinergic receptor nicotinic gamma subunit (CHRNG) is the γ subunit. It may also play an essential role in biological processes, including cell differentiation, growth, and survival, while the role of CHRNG has not been studied in the literature. Therefore, the purpose of this study is to clarify the effect of CHRNG on the proliferation and differentiation of bovine preadipocytes. Methods: We constructed a CHRNG overexpression adenovirus vector and successfully overexpressed it on bovine preadipocytes. The effects of CHRNG on bovine preadipocyte proliferation were detected by Edu assay, cell counting Kit-8 (CCK-8), real-time fluorescence quantitative polymerase chain reaction (RT-qPCR), Western blot and other techniques. We also performed oil red O, RT-qPCR, Western blot to explore its effect on the differentiation of preadipocytes. Results: The results of Edu proliferation experiments showed that the number of EDU-positive cells in the overexpression group was significantly less. CCK-8 experiments found that the optical density values of the cells in the overexpression group were lower than those of the control group, the mRNA levels of proliferating cell nuclear antigen (PCNA), cyclin A2 (CCNA2), cyclin B1 (CCNB1), cyclin D2 (CCND2) decreased significantly after CHRNG gene overexpression, the mRNA levels of cyclin dependent kinase inhibitor 1A (CDKN1A) increased significantly, and the protein levels of PCNA, CCNB1, CCND2 decreased significantly. Overexpression of CHRNG inhibited the differentiation of bovine preadipocytes. The results of oil red O and triglyceride determination showed that the size and speed of lipid droplets accumulation in the overexpression group were significantly lower. The mRNA and protein levels of peroxisome proliferator activated receptor gamma (PPAR class="checkNonKBPoint">γ), CCAAT enhancer binding protein alpha (CEBPα), fatty acid binding protein 4 (FABP4), fatty acid synthase (FASN) decreased significantly. Conclusion: Overexpression of CHRNG in bovine preadipocytes inhibits the proliferation and differentiation of bovine preadipocytes.