• 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.

• 사상체질의학회지
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• 제24권1호
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• pp.54-65
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• 2012

1. Objective : The purpose of this study is to investigate the effects of TAM (Taraxacum mongolicum) on Th2 cytokine production in MC/9 mast cells. 2. Methods : The effects of TAM was analyzed by ELISA and Real-time PCR in MC/9 mast cells. Levels of IL-5, IL-13 were measured using enzyme-linked immunosorbent assays(ELISA). mRNA levels of IL-4, IL-5, IL-6, IL-13 were analyzed with Real-time PCR. 3. Results : 1) TAM inhibited the IL-4 production significantly in comparison to PI-control group at concentration of $50{\mu}g/ml$, $100{\mu}g/ml$, $200{\mu}g/ml$. 2) TAM inhibited the IL-13 production significantly in comparison to PI-control group at concentration of $50{\mu}g/ml$, $100{\mu}g/ml$, $200{\mu}g/ml$. 3) TAM inhibited the IL-4 mRNA expression significantly in comparison to PI-control group at concentration of $100{\mu}g/ml$. 4) TAM inhibited the IL-5 mRNA expression significantly in comparison to PI-control group at concentration of $50{\mu}g/ml$, $100{\mu}g/ml$. 5) TAM inhibited the IL-6 mRNA expression significantly in comparison to PI-control group at concentration of $100{\mu}g/ml$. 6) TAM inhibited the IL-13 mRNA expression significantly in comparison to PI-control group at concentration of $100{\mu}g/ml$. 4. Conclusions : These results indicate that TAM (Taraxacum mongolicum) has the effect of decreasing the Th2 cytokine production in the MC/9 mast cell.

• 사상체질의학회지
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• 제24권3호
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• pp.80-92
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• 2012

Objectives The purpose of this study is to investigate the effects of korean herbal bathing extracts composition 1 and composition 2 on Th2 cytokine production in MC/9 mast cells. Methods The effects of composition 1, 2 was analyzed by ELISA and Real-time PCR in MC/9 mast cells. Levels of IL-5, IL-13 were measured using enzyme-linked immunosorbent assays(ELISA). mRNA levels of IL-4, IL-5, IL-6, IL-13 were analyzed with Real-time PCR. Results Composition 1, 2 inhibited the IL-5, IL-13 production significantly(p<.001) in comparison to PI-control group at concentration of 100 ${\mu}g/mL$, 200 ${\mu}g/mL$. Composition 1, 2 inhibited the IL-4, IL-5, IL-13 mRNA expression significantly in comparison to PI-control group at concentration of 100 ${\mu}g/mL$, 200 ${\mu}g/mL$. Composition 1 inhibited the IL-6 mRNA expression significantly in comparison to PI-control group at concentration of 200 ${\mu}g/ml$. Composition 2 inhibited the IL-6 mRNA expression significantly in comparison to PI-control group at concentration of 100 ${\mu}g/mL$, 200 ${\mu}g/mL$. Conclusions These results indicate that composition 1, 2 has the effect of decreasing the Th2 cytokine production in the MC/9 mast cell.

• 생명과학회지
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• 제18권10호
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• pp.1348-1354
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• 2008

mi transcription factor (MITF)는 비만세포의 분화를 조절하는 중요한 전사인자이다. 특히 MITF는 결합조직형 비만세포에서 일반적으로 발현하는 비만세포 특이적 세린 단백분해효소의 일종인 mMCP-6 유전자의 전사를 조절한다. 본 연구는 마우스 골수유래 배양비만세포에서 mMCP-6 유전자의 전사를 조절하는 MITF이형체를 규명하였다. MITF 이형체들의 발현은 RT-PCR로 확인하였다. IL-3존재 하에서 배양한 점막형 비만세포들은 MITF-A,-E, -H, -Mc 등이 발현하였다. 반면에 SCF존재 하에서 배양한 결합조직형 비만세포들은 MITF-A가 발현하였다. MITF이형체를 과발현시키면 NIH-3T3 세포에서 mMCP-6 promoter를 통한 luciferase 활성을 증가시키고, MC/9 비만세포주에서는 증가된 mMCP-6발현을 유도하였다. 더불어 비만세포에서의 mMCP-6 발현은 MITF-A 고갈로 인하여 유의적으로 억제되었다. MITF-A의 전사활성과 DNA결합은 MITF-E, -H, -Mc 등의 타 이형체들의 결과와 유사하였다. 따라서 본 연구의 결과들은 MITF-A가 마우스 결합조직형 비만세포에서 발현하여 mMCP-6 전사를 조절하는 중요한 이형체임을 제시한다.

• 디지털융복합연구
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• 제13권8호
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• pp.503-514
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• 2015

은행잎 유래 플라보놀 성분이 갖는 항아토피 활성에 대해 입증된 바는 드물다. 이 논문에서는 MC/9 비만세포에서의 은행잎 플라보놀의 항아토피 효과를 조사하기 위하여 ELISA와 Real-time PCR, western blot으로 은행잎 플라보놀을 분석하였다. 분석 결과 IL-13, MIP-1a의 생성량은 농도 의존적으로 현저하게 감소되었으며 IL-4, IL-5, IL-13, TNF-a 유전자 발현량은 25, 50, $100{\mu}g/m{\ell}$의 농도에서 효과적으로 억제되었다. western blot 분석 결과 c-jun과 NFAT-1단백질 발현이 감소되었음을 확인하였다. 이러한 결과들은 은행잎 유래 플라보놀 성분이 NFAT-1, c-jun 전사인자의 전사를 억제함으로써 MC/9 비만세포에서의 Th2 싸이토카인의 생성을 감소시키는 효과를 갖는 것을 나타낸다. 따라서 은행잎 유래 플라보놀 성분이 아토피 피부염 치료제로서 활용가능성이 있음을 보고하고자 한다.

• 대한한방소아과학회지
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• 제28권2호
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• pp.23-39
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• 2014

Objectives PRAL (Prunus armniaca Linne Var) is a herbal formula in Oriental Medicine, known for its anti-inflammatory and anti-allergenic properties. However, its mechanism of action and the cellular targets have not yet been found enough. The purpose of this study is to investigate the effects of PRAL on Th2 cytokines expression in MC/9 mast cells. Methods The effect of PRAL was analyzed by ELISA, Real-time PCR, Western blot in MC/9 mast cells. mRNA levels of GM-CSF, IL-4, IL-5, IL-6, IL-13, TNF-${\alpha}$ were analyzed with Real-time PCR. Levels of IL-13, MIP-$1{\alpha}$ were measured using enzyme-linked immunosorbent assays (ELISA). NFAT, AP-1 and NF-${\kappa}B$ p65 were examined by Western blot analysis. Results PRAL inhibited GM-CSF, IL-4, IL-5, IL-6, IL-13, TNF-${\alpha}$ mRNA expression in a dose dependent manner. GM-CSF, IL-4, IL-5 mRNA expression were inhibited significantly in comparison to DNP-IgE control group at concentration of 100 ${\mu}g/ml$ and IL-6, IL-13, TNF-${\alpha}$ mRNA expression were inhibited at concentration of 50 ${\mu}g/ml$, 100 ${\mu}g/ml$. PRAL also inhibited the IL-13, MIP-$1{\alpha}$ production significantly in comparison to DNP-IgE control group in a dose dependent manner. IL-13 production was inhibited at a concentration of 200 ${\mu}g/ml$, 400 ${\mu}g/ml$ and MIP-$1{\alpha}$ was inhibited at a concentration of 100 ${\mu}g/ml$, 200 ${\mu}g/ml$, 400 ${\mu}g/ml$. Western blot analysis of transcription factors involving Th2 cytokines expression revealed prominent decrease of the mast cell specific transcription factors including NFAT-1, c-Jun as well as NF-${\kappa}B$ p65 but not NFAT-2 and c-Fos. Conclusion These results indicate that PRAL has the effect of suppressing Th2 cytokines production in the MC/9 mast cells. These data represent that PRAL potentiates therapeutic activities to the allergic disease by regulating Th2 cytokines in the MC/9 mast cells.

• 대한한방소아과학회지
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• 제28권3호
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• pp.31-46
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• 2014

Objectives Forsythiae Fructus treatment has been used for inflammatory and allergic diseases in Korean Medicine. Nevertheless, the mechanism of action and the cellular targets are not understood well. The pathogenesis of allergic diseases are associated with Th2 cytokines such as IL-13, MIP-$1{\alpha}$, IL-13, IL-5, GM-CSF, IL-4, TNF-${\alpha}$ and IL-6, which are secreted by the mast cells. This study was conducted to investigate the effects of Forsythiae Fructus extracts (FF) on Th2 cytokines expression and signal transduction in MC/9 mast cells. Methods In the study, MC/9 mast cells were stimulated with DNP-IgE for 24 hours and then treated separately with CsA $10{\mu}g/m{\ell}$ and varying doses of FF for one hour. MC/9 mast cells stimulated with DNP-IgE was the control group, a treatment with CsA was the positive control group and a treatment with varying doses FF was the experimental groups. The mRNA levels of IL-13, IL-5, GM-CSF, IL-4, TNF-${\alpha}$, IL-6 were analyzed with Real-time PCR. The levels of IL-13, MIP-$1{\alpha}$ were measured using enzyme-linked immunosorbent assays(ELISA). NFAT, AP-1 and NF-${\kappa}B$ p65 were examined by Western blot analysis. Results 1. FF were observed to suppress the mRNA expression of IL-13, IL-5, GM-CSF, IL-4, TNF-${\alpha}$, IL-6 in comparison to DNP-IgE control group. 2. FF also has inhibited the IL-13, MIP-$1{\alpha}$ production significantly in comparison to DNP-IgE control group. 3. Western blot analysis of transduction factors involving Th2 cytokines expression has revealed a prominent decrease of the mast cell specific transduction factors including NFAT-1, NFAT-2, c-Jun, and NF-${\kappa}B$ p65 but c-Fos. Conclusions In conclusion, the anti-allergenic activities of FF may be strongly related to the regulation of transcription factors NFAT-1, NFAT-2, c-Jun, and NF-${\kappa}B$ p65 causing inhibition of Th2 cytokines in mast cells.

• 대한한방소아과학회지
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• 제29권4호
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• pp.39-66
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• 2015

Objectives The purpose of this study is to investigate the effects of Sesamum indicum extracted (SEI) on atopic dermatitis in an in-vitro and in-vivo experiment using a MC/9 murine mast cells and a NC/Nga mouse. Methods In-vitro experiment, IL-4, IL-5, IL-6, IL-13, TNF-${\alpha}$ and GM-CSF mRNA expression were evaluated by Real-time PCR, IL-13, MIP-$1{\alpha}$ production by ELISA and manifestations of NFAT-1, NFAT-2, c-jun, c-fos, NF-${\kappa}B$ p65 transcription factors by western blotting. In-vivo experiment, we measured WBC, Eosinophil, Neutrophil, and serum IL-5, IL-13 in NC/Nga atopic dermatitis mouse, IL-5, IL-13, IFN-${\gamma}$, IL-4 in the spleenocyte culture supernatant by ELISA, the absolute cell numbers of CD4+, CD8+, +Gr-1+CD11b, B220+CD23+ in the axillary lymph node (ALN), peripheral blood mononuclear cells (PBMCs) and dorsal skin tissue, IL-5, IL-13 by Real-time PCR, the distribution of tissue inflammation and cellular infiltration by H&E and toluidine blue. Results SEI decreased IL-4, IL-5, IL-6, IL-13, GM-CSF, TNF-${\alpha}$ mRNA expression, IL-13, MIP-$1{\alpha}$ production and the expression of transcription factors including NFAT-1, c-jun, NF-${\kappa}B$ p65 in MC/9 murine mast cells. SEI orally administration decreased cell number of WBC, Eosinophil, the level of serum IgE, total cell number of ALN and dorsal skin tissue, absolute cell number of CD4+, CD8+, B220+CD23+ in the ALN. SEI orally administration also increased absolute cell number of CD8+/CD3+ and decreased Gr-1+/CD11b+ in PBMCs, decreased CD4+ in dorsal skin tissue, inhibited IL-5, IL-13 mRNA expression. Infiltration levels of inflammatory immune cells, mast cells and thickness of epidermis decreased in dorsal skin tissue. Conclusions SEI can regulate allergic inflammatory response suppressed the gene expression and production of cytokines that mediate allergic reactions, and will be able to be effectively utilized in the treatment of atopic dermatitis future.

• 대한한방소아과학회지
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• 제28권4호
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• pp.1-29
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• 2014

Objectives Dictamni Radicis Cortex extracts (DRC) has been known to suppress allergic reaction, however the cellular target of DRC and its mode of action remain unclear. The purpose of this study is to investigate the effects of Dictamni Radicis Cortex extracts on DNCB induced atopic dermatitis-like skin lesions of NC/Nga mouse. Methods This study was designed to investigate the effects of DRC extract in the DNP-IgE-induced activation of MC/9 murine mast cell lines in vitro and in the DNCB-induced activation of NC/Nga mouse in vivo. For this investigation, We examined IL-4, IL-5, IL-6, IL-13, TNF-${\alpha}$ and GM-CSF mRNA expression by Real-time PCR, IL-13, MIP-$1{\alpha}$ production by ELISA analysis and manifestations of NFAT1, NFAT2, AP-1 and NF-${\kappa}B$ p65 transcription factors by western blotting in vitro. Then, we examined WBC, eosinophil and neutrophil in NC/Nga mouse, IL-5, IL-13 in serum, IFN-${\gamma}$, IL-4 in the spleenocyte culture supernatant, the absolute cell numbers of $CD4^+$, $CD8^+$, $^+Gr-1^+CD11b$, $B220^+CD23^+$ in the ALN, PBMCs and dorsal skin, IL-5, IL-13 in the dorsal skin by Real-time PCR and the distribution of mast cells by H&E and toluidine blue. Results In vitro the mRNA expression of IL-4, IL-5, IL-6, IL-13, TNF-${\alpha}$, GM-CSF and IL-13, MIP-$1{\alpha}$ production by ELISA analysis were completely abolished by DRC and the western blot analysis decreased the expression of mast cell-specific transcription factors including NFAT-1, NF-${\kappa}B$ p65. In vivo DRC oral adminstration also decreased the counts of WBC, eosinophils and inflammatory cytokines such as IL-13 and IgE in the serum. DRC oral adminstration elevated IL-4 level in the spleenocyte culture supernatant. DRC oral adminstration decreased total ALN cells, total skin cells, cell numbers of $CD4^+$, $B220^+CD23^+$ in the ALN, $^+Gr-1^+CD11b$ in the PBMCs and $CD4^+$, $CD8^+$ in the dorsal skin. The mRNA expression of IL-5, IL-13, thickness of epidermis, inflammation immune cells and mast cells were abolished by DRC in the dorsal skin. Conclusions Histological examination showed that infiltration levels of immune cells in the skin of AD-induced NC/Nga mouse were much improved by DRC oral adminstration. These results, therefore, suggest that DRC can regulate molecular mediators and immune cells that are functionally associated with atopic dermatitis induced in NC/Nga mouse, and may play an important role in recovering AD symptoms.

• 한국응용과학기술학회지
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• 제38권5호
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• pp.1292-1301
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• 2021

본 연구는 섬애약쑥(Artemisia argyi H.)의 항가려움증 활성 가능성을 확인하기 위한 연구로써 가려움증 관련인자 등을 측정하였다. 섬애약쑥은 열수로 추출(Artemisia argyi H. distilled water extract 이하, AAD)하여 MTT assay로 세포독성을 측정하였고 항 가려움증 활성을 확인하기 위하여 IL-4와 IL-31 관련 전사인자 발현 및 단백질 생성을 측정하였으며, 히스타민의 발현을 측정하였다. 그 결과 25, 50, 100 ㎍/㎖의 농도에서는 유의한 세포독성이 나타나지 않는 것을 확인하였다. 가려움증 연관 유전인자인 IL4의 경우 25 ㎍/㎖의 농도에서 약 12%, 50 ㎍/㎖의 농도에서 약 26%, 100 ㎍/㎖의 농도에서 약 61%로 유의하게 감소하였으며, IL31의 경우 50 ㎍/㎖의 농도에서 약 33%, 100 ㎍/㎖의 농도에서 약 33%로 유의하게 감소하였다. 연관된 단백질 측정의 경우 각각 50 ㎍/㎖와 100 ㎍/㎖의 농도에서 IL-4는 약 34% 및 약 69%, IL-31은 약 36% 및 약 37% 유의하게 감소하였다. 이 결과는 ADD가 항가려움증을 위한 소재로써 가능성을 보여 소재 개발을 위한 기초자료로 제공될 수 있을 것으로 사료된다.