• Tuberculosis and Respiratory Diseases
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• 제64권1호
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• pp.22-27
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• 2008

연구배경: 본 연구에서는 황색포도알균이 숙주세포 내로 침입하여 증식하는 정도를 관찰하고자 한다. 이때 세포 외 바탕 단백질의 변화가 수반될 것으로 가설을 설정하고, 이러한 변화에 영향을 미칠 것으로 생각되는 matrix metalloproteinase (MMP)의 발현과 역할에 대해 연구하고자 하였다. 방법: 황색포도알균은 $10^6{\sim}10^7CFU/ml$을 $10^5$개의 기관지상피세포인 BEAS-2B 세포에 2시간 동안 침입시킨다. 이후 세척으로 세포 밖에 있는 황색포도알균을 제거한 후, BEAS-2B 세포를 다양한 시간 동안(4, 6, 8, 12 시간) 배양한 후 황색포도알균의 집락수(CFU/ml)를 측정하였고, 단백질을 분리하여 세포 외 바탕단백질의 발현 정도와 MMP의 활성도를 측정하였다. 또한 MMP 억제제인 GM6001을 전처치한 후 황색포도알균을 세포에 침입시킨 후 세포 내에서의 균의 집락수 및 세포 외 바탕단백질의 변화를 관찰하였다. 결과: 황색포도알균의 집락을 측정한 결과 4시간과 12시간을 비교해 볼 때 MOI가 증가할수록, 감염시킨 시간이 길수록 숙주세포 내로 침입이 유의하게 증가하였다. BEAS-2B 세포에서 황색포도알균을 침입시킨 시간이 길수록, MOI가 증가할수록 MMP 2 및 MMP 9의 활성도와 dysadherin의 발현은 증가하였고, 이와는 대조적으로 E-cadherin의 발현은 감소하였다. MMP억제제인 GM6001을 전 처치 한 결과 황색포도알균의 세포 내 침입을 유의하게 감소시켰다. 결론: 황색포도알균이 기관지 상피세포 내로 침입할 때 dysadherin 및 E-cadherin 같은 세포 외 바탕 단백질의 변화를 동반하며, MMP 활성도가 균의 세포 내 침입에 관여하는 것으로 보인다.

• Tuberculosis and Respiratory Diseases
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• 제78권2호
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• pp.99-105
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• 2015

Background: Aryl hydrocarbon receptor (AhR), a ligand-dependent transcription factor, binds to a wide variety of synthetic and naturally occurring compounds. AhR is involved in the regulation of inflammatory response during acute and chronic respiratory diseases. We investigated whether nuclear receptor coactivator 7 (NCOA7) could regulate transcriptional levels of AhR target genes and inflammatory cytokines in 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD)-treated human bronchial epithelial cells. This study was based on our previous study that NCOA7 was differentially expressed between normal and chronic obstructive pulmonary disease lung tissues. Methods: BEAS-2B and A549 cells grown under serum-free conditions were treated with or without TCDD (0.15 nM and 6.5 nM) for 24 hours after transfection of pCMV-NCOA7 isoform 4. Expression levels of cytochrome P4501A1 (CYP1A1), IL-6, and IL-8 were measured by quantitative real-time polymerase chain reaction. Results: The transcriptional activities of CYP1A1 and inflammatory cytokines were strongly induced by TCDD treatment in both BEAS-2B and A549 cell lines. The NCOA7 isoform 4 oppositely regulated the transcriptional activities of CYP1A1 and inflammatory cytokines between BEAS-2B and A549 cell lines. Conclusion: Our results suggest that NCOA7 could act as a regulator in the TCDD-AhR signaling pathway with dual roles in normal and abnormal physiological conditions.

• 대한본초학회지
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• 제25권3호
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• pp.65-71
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• 2010

Objectives：To develop and validate HPLC-PDA methods for simultaneous determination of seven constituents in Samchulkunbi-tang (SKT). Additionally, we investigated the cytotoxicity against BEAS-2B cell line and splenocytes of SKT. Methods：Reverse-phase chromatography using a Gemini $C_{18}$ column operating at $40^{\circ}C$, and photodiode array (PDA) detection at 230, 254 and 280 nm, were used for quantification of the three marker components of SKT. The mobile phase using a gradient flow consisted of two solvent systems. Solvent A was 1.0% (v/v) aqueous acetic acid and solvent B was acetonitrile with 1.0% (v/v) acetic acid. The cytotoxicity of SKT were measured by the CCK-8 assay method. Results：Calibration curves were acquired with $r^2$>0.9999, and the relative standard deviation (RSD) values (%) for intra- and inter-day precision were less than 6.0%. The recovery rate of each compound was in the range of 86.89-109.78%, with an RSD less than 4.0%. The contents of seven compounds in SKT were 1.39-6.84 mg/g. SKT had no cytotoxicity effect at 50-200 ${\mu}g$/mL concentrations. Conclusions：The established method will be helpful to improve quality control and in vitro efficacy study of SKT.

• Molecular & Cellular Toxicology
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• 제5권2호
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• pp.172-178
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• 2009

Nanoparticles are small-scale substances (<100 nm) with unique properties, complex exposure and health risk implications. Aluminum oxide ($Al_2O_3$) nanoparticles (NP) have been widely used as abrasives, wear-resistant coatings on propeller shafts of ships, to increase the specific impulse per weight of composite propellants used in solid rocket fuel and as drug delivery systems to increase solubility. However, recent studies have shown that nano-sized aluminum (10 nm in diameter) can generate adverse effects, such as pulmonary response. The cytotoxicity and genotoxicity of $Al_2O_3$ NP were investigated using the dye exclusion assay, the comet assay, and the mouse lymphoma thymidine kinase (tk$^{+/-}$) gene mutation assay (MLA). IC$_{20}$ values of $Al_2O_3$ NP in BEAS-2B cells were determined the concentration of 273.44 $\mu$g/mL and 390.63 $\mu$g/mL with and without S-9. However IC$_{20}$ values of $Al_2O_3$ NP were found nontoxic in L5178Y cells both of with and without S-9 fraction. In the comet assay, L5178Y cells and BEAS-2B cells were treated with $Al_2O_3$ NP which significantly increased 2-fold tail moment with and without S-9. Also, the mutant frequencies in the $Al_2O_3$ NP treated L5178Y cells were increased compared to the vehicle controls with S-9. The results of this study indicate that $Al_2O_3$ NP can cause primary DNA damage and cytotoxicity but not mutagenicity in cultured mammalian cells.

• Tuberculosis and Respiratory Diseases
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• 제80권3호
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• pp.247-254
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• 2017

Background: Airway epithelial cells are the first line of defense, against pathogens and environmental pollutants, in the lungs. Cellular stress by cadmium (Cd), resulting in airway inflammation, is assumed to be directly involved in tissue injury, linked to the development of lung cancer, and chronic obstructive pulmonary disease (COPD). We had earlier shown that ACN9 (chromosome 7q21), is a potential candidate gene for COPD, and identified significant interaction with smoking, based on genetic studies. However, the role of ACN9 in the inflammatory response, in the airway cells, has not yet been reported. Methods: We first checked the anatomical distribution of ACN9 in lung tissues, using mRNA in situ hybridization, and immunohistochemistry. Gene expression profiling in bronchial epithelial cells (BEAS-2B), was performed, after silencing ACN9. We further tested the roles of ACN9, in the intracellular mechanism, leading to Cd-induced production, of proinflammatory cytokines in BEAS-2B. Results: ACN9 was localized in lymphoid, and epithelial cells, of human lung tissues. ACN9 silencing, led to differential expression of 216 genes. Pathways of sensory perception to chemical stimuli, and cell surface receptor-linked signal transduction, were significantly enriched. ACN9 silencing, further increased the expression of proinflammatory cytokines, in BEAS-2B after Cd exposure. Conclusion: Our findings suggest, that ACN9 may have a role, in the inflammatory response in the airway.

• Tuberculosis and Respiratory Diseases
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• 제54권5호
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• pp.551-562
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• 2003

연구배경 : NF-${\kappa}B$는 많은 염증 유발성 물질들을 발현시키는데 필요한 전사 인자로서, 염증성 폐질환 발병에 관여한다는 사실이 확인되었다. 이러한 NF-${\kappa}B$의 활성화에는 여러 신호전달 체계가 관여한다는 사실이 밝혀지고 있으며 최근 PI3K/Akt 경로도 NF-${\kappa}B$ 활성화에 관여한다는 연구 결과가 보고되고 있으나, 실험 대상 세포주마다 활성화 기전이 다르고 호흡기 상피세포에 대한 결과도 알려져 있지 않아 호흡기 상피세포에서의 NF-${\kappa}B$ 활성화에 PI3K/Akt 경로가 관여하는지를 밝히기 위하여 본 연구를 시행하게 되었다. 방법 : 인체 기관지 상피세포주인 BEAS-2B와 폐암 세포주인 A549, NCI-H157을 사용하여 Akt 활성화와 $I{\kappa}B{\alpha}$ 분해 여부를 확인하기 위해 western blot을 시행하였다. Wortmannin, LY294002 및 DN-Akt를 이용하여 Akt 경로를 억제하였고, NF-${\kappa}B$ 활성화와 전사 활성을 측정하기 위하여 각각 EMSA와 luciferase assay를 시행하였다. 결과 : BEAS-2B, A549 및 NCI-H157 세포주에 TNF-$\alpha$ 및 insulin을 처리한 경우 Akt 활성화가 유도되었다. Insulin 으로 Akt 경로를 활성화시킨 경우 $I{\kappa}B{\alpha}$ 분해가 일어나지는 않았다. Wortmannin, LY294002 및 DN-Akt 를 이용하여 Akt 경로를 억제한 경우 TNF-$\alpha$에 의한 $I{\kappa}B{\alpha}$ 분해 및 IKK 활성화가 억제되지는 않았으며, NF-${\kappa}B$ 활성화도 억제되지 않았다. Wortmannin을 처리한 경우 TNF-$\alpha$에 의한 NF-${\kappa}B$ 전사 활성이 오히려 증가하는 양상을 보였으나, DN-Akt 이입시킨 경우에는 관찰되지 않았다. 결론 : 인체 호흡기 상피세포에서는 $I{\kappa}B$/NF-${\kappa}B$ 경로의 활성화는 PI3K/Akt 경로와 무관한 것으로 판단된다.

• Environmental Analysis Health and Toxicology
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• 제30권
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• pp.7.1-7.7
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• 2015

Objectives The widely promising applications of graphene nanomaterials raise considerable concerns regarding their environmental and human health risk assessment. The aim of the current study was to evaluate the toxicity profiling of graphene family nanano-materials (GFNs) in alternative in vitro and in vivo toxicity testing models. Methods The GFNs used in this study are graphene nanoplatelets ([GNPs]-pristine, carboxylate [COOH] and amide [$NH_2$]) and graphene oxides (single layer [SLGO] and few layers [FLGO]). The human bronchial epithelial cells (Beas2B cells) as in vitro system and the nematode Caenorhabditis elegans as in vivo system were used to profile the toxicity response of GFNs. Cytotoxicity assays, colony formation assay for cellular toxicity and reproduction potentiality in C. elegans were used as end points to evaluate the GFNs' toxicity. Results In general, GNPs exhibited higher toxicity than GOs in Beas2B cells, and among the GNPs the order of toxicity was pristine > $NH_2$ > COOH. Although the order of toxicity of the GNPs was maintained in C. elegans reproductive toxicity, but GOs were found to be more toxic in the worms than GNPs. In both systems, SLGO exhibited profoundly greater dose dependency than FLGO. The possible reason of their differential toxicity lay in their distinctive physicochemical characteristics and agglomeration behavior in the exposure media. Conclusions The present study revealed that the toxicity of GFNs is dependent on the graphene nanomaterial's physical forms, surface functionalizations, number of layers, dose, time of exposure and obviously, on the alternative model systems used for toxicity assessment.

• Asian Pacific Journal of Cancer Prevention
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• 제16권11호
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• pp.4781-4786
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• 2015

Siberian ginseng (Eleutherococcus senticosus) is used primarily as an adaptogen herb and also for its immune stimulant properties in Western herbal medicine. Another closely related species used in East Asian medicine systems i.e. Kampo, TCM (Manchuria, Korea, Japan and Ainu of Hokkaido) and also called Siberian ginseng (Acanthopanax senticosus) also displays immune-stimulant and anti-cancer properties. These may affect tumour growth and also provide an anti-fatigue effect for cancer patients, in particular for those suffering from lung cancer. There is some evidence that a carbohydrate in Siberian ginseng may possess not only immune stimulatory but also anti-tumour effects and also display other various anti-cancer properties. Our study aimed to determine the inhibitory and also proliferative effects of a methanol plant extract of Siberan ginseng (E. senticosus) on various cancer and normal cell lines including: A-549 (small cell lung cancer), XWLC-05 (Yunnan lung cancer cell line), CNE (human nasopharyngeal carcinoma cell line), HCT-116 (human colon cancer) and Beas-2b (human lung epithelial). These cell lines were treated with an extract from E. senticosus that was evaporated and reconstituted in DMSO. Treatment of A-549 (small cell lung cancer) cells with E. senticosus methanolic extract showed a concentration-dependent inhibitory trend from $12.5-50{\mu}g/mL$, and then a plateau, whereas at 12.5 and $25{\mu}g/mL$, there is a slight growth suppression in QBC-939 cells, but then a steady suppression from 50, 100 and $200{\mu}g/mL$. Further, in XWLC-05 (Yunnan lung cancer cell line), E. senticosus methanolic extract displayed an inhibitory effect which plateaued with increasing dosage. Next, in CNE (human nasopharyngeal carcinoma cell line) there was a dose dependent proliferative response, whereas in Beas-2 (human lung epithelial cell line), an inhibitory effect. Finally in colon cancer cell line (HCT-116) we observed an initially weak inhibitory effect and then plateau.

• 대한한의학회지
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• 제24권1호
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• pp.74-83
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• 2003

Background : Production of cytokines by bronchial epithelial cells may contribute to the local accumulation of inflammatory cells in patients with bronchial asthma. In many recent studies, molecular biological methods have been used to investigate the role of cytokines in pathogenesis and new therapeutic targets of asthma. Objective : We aimed to identify the dose-dependent inhibitory effects of Socheongryong-tang and Socheongryong-tang plus Sasam (Adenophorae Radix) on the mRNA expressions of Interleukin (IL)-6, IL-8 and granulocyte macrophage colony stimulating factor (GM-CSF) involved in the asthma model. Materials and Methods : In this study, BEAS-2B cell lines, human epithelial cells, were used. These cells were stimulated by tumor necrosis factor $(TNF)-{\alpha},{\;}IL-1{\beta}$ and histamine for artificial inflammatory expression. ${\beta}-actin$ messenger RNA (mRNA) was used for the internal standard. After each 24 hours of the Socheongryong-tang (小靑龍湯) and Socheongryong-tang plus Sasam (小靑 龍湯加沙蔘) treatment, total cellular RNAs were collected by applying RNAzol directly to the living cells. Then the transcriptional activities of IL-6, IL-8 and GM-CSF were measured by RT-PCR with electrophoresis. Results : In the Socheongryong-tang (小靑龍湯) study, the mRNA expressions of IL-6, IL-8 and GM-CSF were significantly inhibited compared to that of the control group (p<0.05). In the Socheongryong-tang plus Sasam (小靑龍湯加沙蔘) study, the mRNA expressions of IL-6, IL-8 and GM-CSF were significantly inhibited compared to that of the control group (p<0.05). Conclusions : This study shows that Socheongryong-tang (小靑龍湯) and Socheongryong-tang plus Sasam (小靑龍湯加沙蔘) have dose-dependent inhibitory effects on the mRNA expressions of IL-6, IL-8 and GM-CSF in human epithelial cells, so these herbal medicines may inhibit the inflammatory process of asthma. Advanced studies are required to investigate the mechanisms of inhibition by herbal medicine in the asthma model.

• 대한한의학회지
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• 제23권1호
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• pp.11-23
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• 2002

Objectives: We aimed to identify the dose-dependent inhibitory effects of Maekmundongcheongpye-eum and Liriopis Tuber on the mRNA expression of IL-6, IL-16, GM-CSF involved in the asthma model. Methods: In the study BEAS-2B cell lines, human epithelial cells were used. These cells were stimulated with tumor necrosis factor $(TNF)-{\alpha}$ for artificial inflammatory expression. ${\beta}-actin$ messenger RNA (mRNA) was used by internal standard. After 24 hours of Maekmundongcheongpye-eum, Liriopis Tuber-treatment, total cellular RNAs were collected, treating RNAzol directly on the alive cells. Then the transcriptional activities of IL-6, 16, GM-CSF were measured by RT-PCR with electrophoresis. Results: In the Maekmundongcheongpye-eum study, the mRNA expression of IL-6 showed 48% transcriptional inhibitory effect compared to the control group in the $100{\;}{\mu}l/ml$ category (P<0.001). In the IL-16, there was 53% and 57% transcriptional inhibitory effect compared to the control group in the $20{\;}{\mu}l/ml$ and $100{\;}{\mu}l/ml$ categories (P<0.001). In the GM-CSF, there was no inhibitory effect. In the Liriopis Tuber study, the mRNA expression of IL-6 showed 43% transcriptional inhibitory effect compared to the control group in the $100{\;}{\mu}l/ml$ category (p<0.005). In the IL-16, 34% and 26% of transcriptional inhibitory effect was shown compared to the control group in the $20{\;}{\mu}l/ml$ and $100{\;}{\mu}l/ml$ categories, respectively (P<0.05). In the GM-CSF, there was no inhibitory effect. Conclusions: This study shows that Maekmundongcheongpye-eum and Liriopis Tuber have dose-dependent inhibitory effects on the mRNA expression of IL-6 and IL-16 in BEAS-2B cell lines, human epithelial cells. Advanced studies are required to investigate the mechanisms of inhibition by herbal medicine in the asthma model.