• Animal cells and systems
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• 제14권4호
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• pp.275-282
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• 2010

Chunghyul-dan (CHD) is a combinatorial drug known to exert anti-inflammatory effects in endothelial cells. In this study, we employed global transcriptional profiling using cDNA microarrays to identify molecular mechanisms responsible for the anti-inflammatory activity of CHD in endothelial cells. An analysis of the microarray data revealed that transcript levels of monocyte chemotactic protein-1 (MCP-1), vascular cell-adhesion molecule-1 (VCAM-1) and activated leukocyte cell-adhesion molecule were dramatically altered in CHD-treated endothelial cells. These changes in gene expression were confirmed by RT-PCR, Western blotting and ELISA. Chronic CHD treatment also appeared to decrease MCP-1 secretion, probably as a result of decreased MCP-1 expression. In addition, we determined that chronic CHD treatment inhibited lipopolysaccharide-stimulated adhesion of THP-1 leukocytes to endothelial cells. The inhibitory effect of CHD on LPS-stimulated adhesion resulted from downregulation of VCAM-1 expression. Transmigration of THP-1 leukocytes through endothelial cells was also inhibited by chronic CHD treatment. In conclusion, CHD controls a variety of inflammatory activities by regulating MCP-1 and VCAM-1 gene expression.

• 대한본초학회지
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• 제35권4호
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• pp.51-60
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• 2020

Objective : The objective of this study was to demonstrate the effect of Persicae Semen (PS) in DNCB-induced atopic dermatitis mouse and HaCaT cell. Methods : The BALB/c mice were divided into four groups. To develop atopic dermatitis, 200 ㎕ of 1 and 0.5% DNCB solution was put on the back of mice in the Control group, the PS-Low group and the PS-High group once a day. After application of DNCB, 200 ㎕ of the PS extract was also treated. The Normal group was given PBS. The mice dorsal skin was stained with Masson's trichrome, H&E, and toluidine blue to evaluate the thickness of the epidermis and dermis, infiltration of eosinophils and mast cells respectively. ELISA was applied to measure the serum level of IgE and IL-6. Toxicity of PS was measured by MTS assay in HaCaT cell. To investigate the effects of PS on HaCaT cells, cells were pre-treated with PS for 1h, and then stimulated with TNF-α and IFN-γ. After 24 hours, the expression of TARC was analyzed using RT-PCR. Results : PS not only significantly diminished the thickness of the epidermis and dermis, but also reduced the infiltration of eosinophil and mast cell in skin lesion. PS also reduced the serum IgE and IL-6 level which plated important roles in the atopic dermatitis. The expression of TARC was decreased significantly in TNF-α/IFN-γ stimulated HaCaT cell. Conclusion : These results suggest that PS may be effective in alleviating the atopic dermatitis induced by DNCB and inflammation by TNF-α/IFN-γ.

• 한방안이비인후피부과학회지
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• 제25권3호
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• pp.1-12
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• 2012

Objectives : Fagopyrum esculentum(FE) has been used for removal of inflammation of internal organs and treatment of sore and ulcer by heat toxin in Korean herbal medicines. In this study, To investigated the protective effect of FE on allergic response, we determined whether FE inhibits allergic response. Methods : The effect of FE was analyzed by ELISA, RT-PCR and Western blot in RBL-2H3 cells. We investigated cell viability, ${\beta}$-hexosaminidase, as a marker of degranulation, cytokne, and intracellular ROS and MAPK and NF-${\kappa}B$ signaling. Results : We found that FE suppressed ${\beta}$-hexosaminidase release, the production of IL-4 and TNF-${\alpha}$ and intracellular ROS level in RBL-2H3 by the anti-DNP IgE plus DNP-HSA stimulation. FE also significantly inhibited cytokine mRNA expressions, such as IL-$1{\beta}$, IL-2, IL-3, IL-4, IL-5, IL-6, IL-13, TNF-${\alpha}$ and GM-CSF in RBL-2H3. In addition, PF suppressed the phospholyation of ERK1/2, JNK1/2, p38 and $I{\kappa}B{\alpha}$ and NF-${\kappa}B$ signal transduction pathway. Conclusions : Our results indicate that FE protects against allergic response and exerts an anti-inflammatory effect through the inhibition of degranulation and production of cytokines and ROS via the suppression MAPK and NF-${\kappa}B$ of signal transduction. Abbrevations : FE, Fagopyrum esculentum; RBL-2H3, rat basophilic leukemia cell line; ROS, reactive oxygen species; MAPK, Mitogen-activated protein kinase; $NF{\kappa}B$, nuclear factor ${\kappa}B$; $TNF{\alpha}$, Tumor necrosis factor alpha; GM-CSF, Granulocyte macrophage colony-stimulating factor; ERK, extracellular-signal-regulated kinase; JNK, c-Jun NH2-terminal kinase; p38, p38 MAP kinase; $I{\kappa}B{\alpha}$, inhibitory-kappa B alpha.

• 한국자원식물학회지
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• 제25권5호
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• pp.578-585
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• 2012

홍삼의 6주간의 복강투여가 생체 내 면역계에 미치는 영향을 알아보기 위하여 면역계를 구성하는 주요 장기인 비장과 흉선의 무게변화와 성숙된 B세포와 T세포가 많이 분포되어 있는 비장세포를 배양하여 mitogen에 대한 비장세포의 세포증식능력을 연구하였다. 또한, 생체내 투여되는 홍삼이 면역반응의 매개역할을 하는 분비성 사이토카인의 조절 양상을 확인하기 위하여 생쥐의 혈청에서 분리한 사이토카인의 생성변화를 측정하였다. 연구결과 홍삼의 투여는 비장과 흉선의 무게를 증가시켰고, 비장세포내의 B세포와 T세포의 증식능력에도 유의적인 효과를 나타내었다. 혈청 내 분비된 T세포, B세포 및 대식세포가 분비하는 사이토카인의 농도변화에서도 홍삼 투여군은 면역계를 활성화시키는 IFN-${\gamma}$, TNF-${\alpha}$, IL-2, IL-6 및 IL-12의 분비량을 모두 증가시켰으며, 면역억제성 사이토카인으로 알려진 IL-10의 분비변화에는 영향을 미치지 않았다. 이와 같은 결과는 홍삼의 투여가 면역 반응계 전반을 조절하는 사이토카인의 생성에 영향을 미치며 모두 면역계를 활성화시키는 방향으로 작용시키는 것을 의미한다. 특히, 흉선과 비장의 무게지수가 증가되었다는 것은 세포증식 등의 변화에 영향을 미침으로써 면역계를 활성화시키는 것으로 생각되며, 특히 흉선의 무게 증가와 ConA에 대한 T세포 증식능력의 유의성 변화 및 T세포가 분비하는 IL-2, IFN-${\gamma}$ 등의 사이토카인의 증가는 홍삼의 생체 내 투여가 T세포의 활성화에 크게 영향을 미치는 것으로 사료된다.

• Biomolecules & Therapeutics
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• 제3권4호
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• pp.266-272
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• 1995

The analysis of membrane receptors for hormones and neurotransmitters has progressed considerably by pharmacological and biochemical means and more recently through the use of specific antibodies. To generate and characterize a moloclonal antibody against $\beta$-adrenergic receptor, a synthetic $\beta$2-adrenergic receptor peptide (Phe-Gly-Asn-Phe-Trp-Cys-Phe-Trp-Thr-Ser-lle-Asp-Val-Leu) which may comprise part of $\beta$-adrenergic receptor ligand binding pocket was coupled to Keyhole Limpet Hemocyanin (KLH) and used as an immunogen. Male BALB/C mice were immunized with this antigen and the immunized spleen was fused with myeloma SP2/0-Ag14 cells to produce monoclonal antibodies. Two clones were obtained but one of monoclonal antibodies, mAb5G09, was used throughout in this study because the other clone, mAb5All showed weak immunoreactivity against KLH as well. The mouse monoclonal antibody mAb5G09 produced in this study showed immunoreactivity to peptide-KLH conjugates and also to human A43l cells and guinea pig lung $\beta$2-adrenergic receptor as revealed by ELISA and western blot. In the course of determination of the effects of mAb5G09 on $\beta$-receptor ligand binding, it was observed that mAb5G09 specifically bound $\beta$-adrenergic radioligand [$^3$H]dihydroalprenolol (DHA) with a dissociation constant (Kd) of 60 nM. The [$^3$H]DHA binding activity of mAb5G09 had characteristics of immunoglobulins and the binding activity was not observed in the control anti-KLH monoclonal antibody. The monoclonal antibody, mAb5G09 produced in this study may provide useful models for the study of the structure of receptor binding sites.

• 대한한의학방제학회지
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• 제26권4호
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• pp.295-306
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• 2018

Schisandra chinensis (SC) and Lycium chinense (LC) were widely distributed in Asia and the fruit has been used traditionally for medicinal herbs. The processing method was solid-state fermentation using Aspergillus oryzae for 48 h after stir-frying treatment at $220^{\circ}C$ for 12 min. In this study, in vitro the anti-inflammatory effect and in vivo hangover reduction were compared to unprocessed SC and LC water extract. Anti-inflammatory effects have been evaluated in pro-inflammatory mediators which were secreted by lipopolysaccharide (LPS)-induced RAW 264.7 macrophages. Nitric oxide (NO) was determined using Griess reaction. Proinflammatory cytokines such as tumor necrosis factor $(TNF)-{\alpha}$ and interleukin $(IL)-1{\beta}$ were measured by enzyme-linked immunosorbent assays (ELISA). Alcohol dehydrogenase (ADH) and acetaldehyde dehydrogenase (ALDH) activities were compared to processed SC or LC and mixtures thereof (1:1). In vivo study was compared to hangover relief in alcohol-fed mice. After administering a mixture of SC and LC (300 mg/kg) water extract (1:1), mice were fed 3 g/kg of ethanol. Serum was collected at 1, 3, and 5 h intervals to analyze ethanol and acetaldehyde levels using a colorimetric assay kit. The processed SC and LC water extracts compared to raw materials significantly inhibited LPS-induced NO and inflammatory cytokine production in RAW 264.7 cells. The results of the hangover mouse model are also consistent with anti-inflammatory effects. These results suggest that processed SC and LC extracts may be functional materials for the treatment of inflammation and hangover.

• 한국자원식물학회:학술대회논문집
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• 한국자원식물학회 2019년도 춘계학술대회
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• pp.54-54
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• 2019

Apple (Malus domestica) is one of the most economically important fruits in Korea. But virus infection has decreased sustainable production of apple and caused the serious problems such as yield loss and poor fruit quality. Virus or viroid infection including Apple chlorotic leaf spot virus (ACLSV), Apple stem pitting virus (ASPV), Apple stem grooving virus (ASGV), Apple mosaic virus (ApMV) and Apple scar skin viroid (ASSVd) has been also reported in Korea. In many cases, apple is infected with virus and viroid with no specific symptoms, the damage caused by the virus are unaware significantly. In our research, we tried to eliminate viruses in the rootstock for the disease-free seedlings of the apple dwarfing rootstock M.9 and M.26. The method of virus elimination was meristem culture, heat($37^{\circ}C$, 6weeks) treatment and chemistry($Ribavirin^{(R)}$) treatment. The analytical methods commonly used for the detection of virus is Enzyme-linked Immuno-Sorbent Assay(ELlSA) and Reverse Transcription-polymerase Chain Reaction(RT-PCR). RT-PCR method was more 30% sensitive than ELISA method. Efficiency of method eliminate virus appeared meristem method > heat treatment > chemistry treatment. The higher acquisition rate of disease-free seedlings is 30~40% on meristem treatment. In meristem treatment, the apple dwarfing rootstock M.9 gained infection ratio of ACLSV, ASPV and ASGV were 45%, 60% and 50% respectively. In the apple dwarfing rootstock M.26, infection ratio of ACLSV, ASPV and ASGV were 40%, 55%, 55%, respectively. Based on our results, it was found that most effective method of disease-free seedlings apple dwarfing rootstocks was by meristem treatment than heat method and chemistry treatment.

• 대한한방부인과학회지
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• 제32권2호
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• pp.1-17
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• 2019

Objectives: This study was performed to identify the anti-inflammatory effects of Salvia miltiorrhizae radix Water extract (SMW) on lipopolysaccharide (LPS) induced inflammation. Methods: RAW 264.7 cells were treated with 500 ng/ml of LPS. SMW (0.1, 0.25, 0.5 mg/ml) was treated 1 h prior to LPS. Cell viability was measured by MTT assay. Levels of nitric oxide (NO) were measured with Griess reagent and pro-inflammatory cytokines were measured by enzyme-linked immunosorbent assay (ELISA) and real-time polymerase chain reaction (PCR). We also examined molecular mechanisms such as mitogen-activated protein kinases (MAPKs) and nuclear factor-kappa B ($NF-{\kappa}B$) activation by western blot. In addition, we observed mice survival rate after LPS and examined their cytokine levels of serum and liver tissue. Results: SMW itself did not have cytotoxic effects in RAW 264.7 cells less than 0.5 mg/ml. SMW treatment inhibited the production of NO, and interleukin $(IL)-1{\beta}$ which is pro-inflammatory cytokine. And SMW treatment inhibited the LPS-induced activation of MAPKs such as extracellular signal-regulated kinase1/2 (ERK1/2), p38 kinases (p38), c-Jun NH2-terminal kinase (JNK) and $NF-{\kappa}B$. In addition, it also showed reducing the level of $IL-1{\beta}$ on the serum and liver tissue of mice. Also, death of LPS-induced mice was inhibited by SMW. Conclusions: The result suggests that treatment of SMW could reduce the LPS-induced inflammation. Thereby, SMW could be used as a protective agent against inflammation. Also, this study could give a clinical basis that SMW could be a drug or agent to prevent inflammatory diseases.

• 대한본초학회지
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• 제28권5호
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• pp.7-11
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• 2013

Objectives : Seungmagalgeun-tang (SMGGT) is traditional medicine widely used for inflammatory disease and flu. But SMGGT exhibits potent anti-inflammatory activity with an unknown mechanism. To elucidate the molecular mechanisms of SMGGT water extract on pharmacological and biochemical actions in inflammation, we examined the effect of SMGGT on pro-inflammatory mediators in Phorbol-12-myristate-13-acetate (PMA)+A23187-stimulated mast cells. Methods : In the present study, pro-inflammatory cytokine production was determined by performing enzyme-linked immunosorbent assay (ELISA), reverse transcription polymerase chain reaction (RT-PCR), and western blot analysis to measure the activation of MAPKs. Cells were treated with SMGGT 1 h prior to the addition of 50 nM of PMA and $1{\mu}M$ of A23187. Cell viability was measured by MTS assay. The investigation focused on whether SMGGT inhibited the expressions of interleukin-6 (IL-6), interleukin-8 (IL-8) and mitogen-activated protein kinases (MAPKs) in PMA+A23187-stimulated mast cells. Results : SMGGT has no cytotoxicity at examined concentration (100, 250, and $500{\mu}g/ml$). Also, gene expression of IL-6 and IL-8 in HMC-1 cells stimulated by PMA+A23187 was down regulated by SMGGT. Furthermore, SMGGT suppressed the PMA+A23187-induced phosphorylation of extracellular signal-regulated kinase (ERK) and c-jun N-terminal Kinase(JNK). But, SMGGT could not regulate phosphorylation of p38 MAPK. Conclusions : These results suggest that SMGGT has inhibitory effects on PMA+A23187-induced IL-6 and IL-8 production. These inhibitory effects occur through blockades on the phosphorylation of ERK and JNK.

• 대한본초학회지
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• 제32권2호
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• pp.77-85
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• 2017

Objectives : This study aimed to investigate the unknown mechanisms behind the anti- inflammatory activity of Hyeonto-dan(HT) 70% ethanol extract on LPS-stimulated RAW 264.7 cells. Methods : Cells were treated with Hyeonto-dan 1 h prior to addition of 200 ng/mL of LPS. Cell viability was measured by the MTS assay. Nitric oxide levels were determined by the Griess assay. $PGE_2$ were measured using EIA kit. Pro-inflammatory cytokine production was measured by the enzyme-linked immunosorbent assay (ELISA). The expression of COX-2, iNOS, and MAPKs was investigated by Western blot, qRT-PCR. $NF-{\kappa}B$/p65 localization and interaction of the TLR-4 receptor with LPS was examined by immunofluorescence assays. Results : Hyeonto-dan had no cytotoxicity at the measured concentration. Hyeonto-dan inhibited NO production and pro-inflammatory cytokines such as IL-6, $TNF-{\alpha}$, and PGE2 as well as the protein and mRNA expression of iNOS and COX-2. Moreover, Hyeonto-dan inhibited the interaction between LPS and TLR-4 in murine macrophages. It suppressed phosphorylation of extracellular signal-regulated kinase (ERK 1/2), c-jun N-terminal kinase (JNK 1/2) and p38. Finally, it inhibited translocation of $NF-{\kappa}B$ in response to competitive LPS. Conclusions : Based on the results of this study, Hyeonto-dan inhibited the binding of TLR-4 receptor to LPS and inhibited the phosphorylation of extracellular signaling pathway MAPKs. These inhibitory effects are thought that the amount of $NF-{\kappa}B$ delivered to the nucleus was decreased and the inflammatory reaction was prevented by decreasing the production of LPS-induced $PGE_2$, NO, IL-6 and $TNF-{\alpha}$.