• The Korean Journal of Zoology
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• v.27 no.2
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• pp.103-116
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• 1984

The structural gene of rabbit hemoglobin was cloned into Pst I site of pBR322 in E. coli. The complementary DNA (cDNA) was synthesized from rabbit globin mRNA with avian myeloblastosis viral reverse transcriptase, and then RNA was destroyed at pH 11. The double stranded cDNA was synthesized with both Klenow fragment of E. coli DNA polymerase I and reverse transcriptase and then the hairpin loop was opened with Sl nuclease. Double stranded cDNA was subsequently tailed with dCTP and annealed to dGMP-tailed vector DNA. After transformation and initial screening of appropriate clones by plasmid size, the cloned colonies were identified by in situ colony hybridization using by plasmid size, the cloned colonies were identified by in situ colony hybridization using $[^32P]$-labeled cDNA probes and characterized the inserts with restriction endonucleases. The expression of cloned globin gene was investigated by standard radioimmunoassay using rat anti-rabbit Hb serum as primary antibody and goat antirat IgG serum as secondary antibody. The result suggested that the chimeric proteins (the part of $\\beta$-lactamase from the vector pBR322 and globin from rabbit) were supposedly produced in E. coli and the product had the antigenic determinant of rabbit hemoglobin.

• Toxicological Research
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• v.14 no.2
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• pp.205-209
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• 1998

To evaluate immunotoxicity of skin decontamination kit(SDK) newly-developed in Agency for Defense Development(ADD), delayed contact hypersensitivity (maximization) test and passive cutaneous anaphylaxis(PCA) test of SDK were performed and the results were compared with those of M 291. In maximization test, sensitization reaction was induced by id injection (2.5 mg / 0.1 $\textrm{m}{\ell}$/ guinea pig or 2.5 mg+CFA/0.1 $\textrm{m}{\ell}$/guinea pig) and topical application (2.5 mg/$\textrm{m}{\ell}$/guinea pig) with SDK or M291 at an interval of 1 week, and 2 weeks later, challenged by topical application with 25 mg/$\textrm{m}{\ell}$/guinea pig. SDK and M291 did not induce any reactions, showing 0 point of sensitization score and 0% of sensitization rate. In conclusion, it is suggested that SDK and M291 do not induce delayed contact hypersensitivity. In PCA test, rats were administered id with mouse anti-SDK serum and challenged iv with a mixture of antigen SDK and Evan's blue. SDK did not induce blue spots at the injection sites of both high (2.5 mg/mouse) and low (1.25 mg/mouse) dose-induced antisera. In contrast, BSA, positive control produced spots larger than 5 mm in diameter at the injection sites of BSA-induced antiserum up to $2^2$ ~ $2^4$dilution. In conclusion, it is suggested that SDK do not induce IgE production and is not a PCA-reaction inducer.

• The Journal of Pediatrics of Korean Medicine
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• v.26 no.1
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• pp.36-45
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• 2012

Objectives Perillae Folium (PF) has been widely used in Korean herbal medicines used for treatment of acute and chronic inflammatory diseases, such as rhinitis, asthma, and enteritis. In this study, to investigate the protective effect of PF on type 1 allergic response, we determined whether PF inhibits early or late allergic responses. Methods The effect of PF was analyzed by ELISA,. RT-PCR and Western blot in RBL-2H3 cells. Levels of ${\beta}$-hexosaminidase, interleukin (IL)-4 and TNF-${\alpha}$ were measured using enzyme-linked immunosorbent assays (ELISAs). mRNA levels of cytokines and enzymes were analyzed with RT-PCR. Signal transduction was analyzed with Western blot. Results We found that PF suppressed ${\beta}$-hexosaminidase release in RBL-2H3 by the IgE-DNP-HSA stimulation. PF also significantly inhibited enzymes level, such as COX-1, COX-2, iNOS, and HDC2, along with reduced cytokine levels, such as IL-2, IL-3, IL-4, IL-6, IL-13, and TNF-${\alpha}$ in RBL-2H3. In addition, PF suppressed the phospholyation of ERK1/2, JNK1/2, and $I{\kappa}B{\alpha}$. Conclusions Our results indicate that PF protects against type 1 allergic response and exert an anti-inflammatory effect through the inhibition of degranulation and expression of cytokines and enzymes via the suppression of signal transduction.

• Korean Journal of Pharmacognosy
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• v.48 no.1
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• pp.38-45
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• 2017

Black-red ginseng (BRGP) exhibits more potent biological activities than white and red ginseng. However, the effect of BRGP on allergic inflammation has not been studied extensively. In this study, we attempted to evaluate the effects of BRGP on allergic inflammation in vitro and in vivo. The effects of BRGP on pro-inflammatory mediators in phorbol myristate acetate (PMA) and A23187-stimulated human mast cells (HMC-1) and on trimellitic anhydride (TMA)-induced atopic dermatitis-like skin lesions in BALB/c mice were evaluated. BRGP suppressed the expression of $TNF-{\alpha}$, IL-6 and IL-8 from HMC-1 stimulated with PMA and A23187. Furthermore, the oral administration of BRGP markedly significantly reduced apparent severity of atopic dermatitis-like skin lesions, ear swelling, lymph node weight gains and serum IgE levels induced by TMA in mice. BRGP treatment also ameliorated epidermal hyperplasia and infiltration of inflammatory cells including mast cells. Taken together, BRGP possesses significant anti-atopic efficacy, suggesting that it could be used as a potential therapeutic agent for allergic inflammatory diseases, including atopic dermatitis.

• The Journal of Korean Medicine Ophthalmology and Otolaryngology and Dermatology
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• v.24 no.1
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• pp.86-95
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• 2011

Objectives : Ulmus davidiana (UD) has been widely used in Korean herbal medicines used for treatment of acute and chronic inflammatory diseases, such as rhinitis, asthma, and abscess. In this study, To investigated the protective effect of UD on type 1 allergic response, we determined whether UD inhibits early and late allergic response. Methods : The effect of UD was analyzed by ELISA and RT-PCR in RBL-2H3 cells. Levels of ${\beta}$ -hexosaminidase, interleukin (IL)-4 and TNF-${\alpha}$ were measured using enzyme-linked immunosorbent assays (ELISAs). mRNA levels of COX-2 and T-helper type 2(Th2) cytokines were analyzed with RT-PCR. Results : We found that UD suppressed ${\beta}$-hexosaminidase release in RBL-2H3 not only by the PMA plus A23187 stimulation, but also by the IgE-DNP-HSA stimulation at the antigen-antibody binding stage and antibody-receptor binding stage. UD also significantly inhibited COX2 level, along with reduced Th2 cytokine levels, such as IL-3, IL-4, IL-5, IL-13, GM-CSF, and TNF-${\alpha}$ in RBL-2H3. Conclusions : Our results indicate that UD protects against type 1 allergic response and exerts an anti-inflammatory effect through the inhibition of degranulation and expression of COX2 and Th2 cytokines.

• Journal of Haehwa Medicine
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• v.19 no.2
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• pp.85-100
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• 2011

In order to investigate the efficacy of HYGBH on atopic dermatitis, various immune related factors were studied. The results and conclusions are as follows. Atopic dermatitis symptoms were improved in HYGBH treated group and significant decrease in dermatitis index were observed in 12 and 14 weeks. HYGBH treated group showed significant decrease in CD4+, CD3+/CD69+ immune cell ratio in PBMC by 18% and 40.6% respectively. HYGBH treated group showed significant decrease in CD3+, CD11b+/Gr-1+ immune cell ratio in dorsal skin by 44.6% and 53.1% respectively. HYGBH treated group showed significant decrease in IL-4, IFN-${\gamma}$ in spleen by 29.5%, 7.7% respectively. HYGBH treated group showed decrease in the expression of IL-5, IL-13, IL-17 and histamine by 21%, 9.6%, 14%, and 32.2% respectively. Also the group showed decrease in the expression of IgE by 6.8% respectively. HYGBH treated group showed significant decrease in the transcription of IL-5 and IL-3 mRNA in skin by 35.5% and 23.2% respectively. The results above indicated that treatment of HYGBH improved atopic dermatitis symptoms by anti-oxidant activity as well as immune modulation activity as a clinical evidence. Also, to increase the application of fermented oriental medicine, different fermentation conditions using various microbial strains should be accumulated as the clinical evidence in the future.

• The Korean Journal of Physiology and Pharmacology
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• v.10 no.1
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• pp.19-24
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• 2006

The intermediate conductance $Ca^{2+}-activated$ $K^+$ channels (SK4, IKCa1) are present in lymphocytes, and their membrane expression is upregulated by various immunological stimuli. In this study, the activity of SK4 was compared between Bal-17 and WEHI-231 cell lines which represent mature and immature stages of murine B lymphocytes, respectively. The whole-cell patch clamp with high-$Ca^{2+}$ ($0.8{\mu}M$) KCl pipette solution revealed a voltage-independent $K^+$ current that was blocked by clotrimazole (1 mM), an SK4 blocker. The expression of mRNAs for SK4 was confirmed in both Bal-17 and WEHI-231 cells. The density of clotrimazole-sensitive SK4 current was significantly larger in Bal-17 than WEHI-231 cells ($-11.4{\pm}3.1$ Vs. $-5.7{\pm}1.15$ pA/pF). Also, the chronic stimulation of B cell receptors (BCR) by BCR-ligation (anti-IgM Ab, $3{\mu}g$/ml, 8∼12 h) significantly upregulated the amplitude of clotrimazolesensitive current from $-11.4{\pm}3.1$ to $-53.1{\pm}8.6$ pA/pF in Bal-17 cells. In WEHI-231 cells, the effect of BCR-ligation was significantly small ($-5.7{\pm}1.15$ to $-9.0{\pm}1.00$ pA/pF). The differential expression and regulation by BCR-ligation might reflect functional changes in the maturation of B lymphocytes.

• Parasites, Hosts and Diseases
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• v.52 no.1
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• pp.41-46
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• 2014

The mature domain of a cysteine protease of Spirometra erinacei plerocercoid larva (i.e., sparganum) was expressed in Escherichia coli, and its value as an antigen for the serodiagnosis of sparganosis was investigated. The recombinant protein (rSepCp-1) has the molecular weight of 23.4 kDa, and strongly reacted with the sparganum positive human or mice sera but not with negative sera by immunoblotting. ELISA with rSepCp-1 protein or sparganum crude antigen (SeC) was evaluated for the serodiagnosis of sparganosis using patient's sera. The sensitivity and specificity of ELISA using rSepCp-1 protein were 95.0% (19/20) and 99.1% (111/112), respectively. In contrast, the sensitivity and specificity of ELISA with SeC were 100% (20/20) and 96.4% (108/112), respectively. Moreover, in experimentally infected mice, the sensitivity and specificity of both ELISA assays were 100% for the detection of anti-sparganum IgG. It is suggested that the rSepCp-1 protein-based ELISA could provide a highly sensitive and specific assay for the diagnosis of sparganosis.

• Journal of Ginseng Research
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• v.45 no.1
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• pp.176-182
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• 2021

Background: Atopic dermatitis (AD) is associated with chronic skin inflammatory reactions. p-coumaric acid (pCA) is an active ingredient of Panax ginseng Meyer (Araliaceae). Methods: Here, we estimated an anti-AD effect of pCA on activated mast cells, activated splenocytes, and a mouse model of AD. Cytokines levels were measured by ELISA and protein activation was analyzed by Western blotting. 2,4-dinitrofluorobenzene (DNFB) was used to induce AD-like skin lesions. Results: The treatment with pCA suppressed the productions and mRNA expressions of thymic stromal lymphopoietin (TSLP), TNF-α, IL-6, and IL-1β in HMC-1 cells. pCA downregulated the expressions of RIP2 and caspase-1, phosphorylated-(p)p38/pJNK/pERK, and pIKKβ/pIkBα/NF-κB in HMC-1 cells. pCA also decreased the productions of TSLP, TNF-α, IL-6, IL-4, and IFN-γ in the supernatant of stimulated splenic cells. Comparing to DNFB-sensitized control group, pCA-treated group alleviated pathological changes of AD-like lesions. pCA decreased the proteins and mRNA expressions levels of TSLP, IL-6, and IL-4 in the skin lesions. Caspase-1 activation was also downregulated by pCA treatment in the AD-like lesions. The serum levels of histamine, IgE, TSLP, TNF-α, IL-6, and IL-4 were suppressed following treatment with pCA. Conclusion: This study suggests that pCA has the potential to improve AD by suppressing TSLP as well as inflammatory cytokines via blocking of caspase-1/NF-κB signal cascade.

• IMMUNE NETWORK
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• v.20 no.5
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• pp.38.1-38.12
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• 2020

Dendritic cells (DCs) are professional antigen-presenting cells (APCs) that initiate both T-cell responses and tolerance. Tolerogenic DCs (tDCs) are regulatory DCs that suppress immune responses through the induction of T-cell anergy and Tregs. Because lactoferrin (LF) was demonstrated to induce functional Tregs and has a protective effect against inflammatory bowel disease, we explored the tolerogenic effects of LF on mouse bone marrow-derived DCs (BMDCs). The expression of CD80/86 and MHC class II was diminished in LF-treated BMDCs (LF-BMDCs). LF facilitated BMDCs to suppress proliferation and elevate Foxp3⁺ induced Treg (iTreg) differentiation in ovalbumin-specific CD4⁺ T-cell culture. Foxp3 expression was further increased by blockade of the B7 molecule using CTLA4-Ig but was diminished by additional CD28 stimulation using anti-CD28 Ab. On the other hand, the levels of arginase-1 and indoleamine 2,3-dioxygenase-1 (known as key T-cell suppressive molecules) were increased in LF-BMDCs. Consistently, the suppressive activity of LF-BMDCs was partially restored by inhibitors of these molecules. Collectively, these results suggest that LF effectively causes DCs to be tolerogenic by both the suppression of T-cell proliferation and enhancement of iTreg differentiation. This tolerogenic effect of LF is due to the reduction of costimulatory molecules and enhancement of suppressive molecules.