• Animal Bioscience
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• v.34 no.3_spc
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• pp.463-470
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• 2021

Objective: This experiment was conducted to find out the immunological effects of wheat phytase when long-chain inorganic polyphosphate (polyP) treated with wheat phytase was added to a macrophage cell line, Raw 264.7, when compared to intact long-chain polyP. Methods: Nitric oxide (NO) production of Raw 264.7 cells exposed to P700, a long-chain polyP with an average of 1,150 phosphate residues, treated with or without wheat phytase, was measured by Griess method. Phagocytosis assay of P700 treated with or without phytase in Raw 264.7 cells was investigated using neutral red uptake. The secretion of tumor necrosis factor α (TNF-α) by Raw 264.7 cells with wheat phytase-treated P700 compared to intact P700 was observed by using Mouse TNF-α enzyme-linked immunosorbent assay kit. Results: P700 treated with wheat phytase effectively increased NO production of Raw 264.7 cells by 172% when compared with intact P700 at 12 h exposure. At 5 mM of P700 concentration, wheat phytase promoted NO production of macrophages most strongly. P700, treated with wheat phytase, stimulated phagocytosis in macrophages at 12 h exposure by about 1.7-fold compared to intact P700. In addition, P700 treated with wheat phytase effectively increased in vitro phagocytic activity of Raw 264.7 cells at a concentration above 5 mM when compared to intact P700. P700 dephosphorylated by wheat phytase increased the release of TNF-α from Raw 264.7 cells by 143% over that from intact P700 after 6 h exposure. At the concentration of 50 μM P700, wheat phytase increased the secretion of cytokine, TNF-α, by 124% over that from intact P700. Conclusion: In animal husbandry, wheat phytase can mitigate the long-chain polyP causing damage by improving the immune capabilities of macrophages in the host. Thus, wheat phytase has potential as an immunological modulator and future feed additive for regulating immune responses caused by inflammation induced by long-chain polyP from bacterial infection.

• The Korea Journal of Herbology
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• v.29 no.3
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• pp.79-85
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• 2014

Objectives : In this study, we attempted to evaluate the effects of Careswell on human mast cell-mediated allergy inflammation in vitro and pruritogen-induced scratching behavior in vivo. Method : The Careswell was extract by distilled water. The anti-itching effects of Careswell were investigated on the compound 48/80 ($50{\mu}g/kg$) or histamine ($100{\mu}g/kg$) induced scratching behavior male ICR mice for 30 min by an observer blind. Terfenadine (10 mg/kg) was used as a positive control drug. The cell toxicity of Careswell was determined by 3-(4,5-dimethylthiazole-2-yl)-2, 5-diphenyl-tetrazolium bromide (MTT) assay. The regulatory effect of Careswell on interleukin (IL)-6 and tumor necrosis factor (TNF)-${\alpha}$ levels was determined by enzyme-linked immunosorbent assay (ELISA) in phorbol 12-myristate 13-acetate plus calcium ionophore A23187 (PMACI) stimulated human mast cells (HMC-1). Also, we evaluated the effect of Careswell on PMACI induced the activation of Nuclear factor-kappa B (NF-${\kappa}B$) into nucleus by Western blot analysis. Result : The results revealed that the oral administration of Careswell (200 mg/kg, p.o.) attenuated the compound 48/80 or histamine-induced scratching behavior in mice. We showed that Careswell significantly reduced the PMACI-induced the production of IL-6 (0.5-1 mg/ml) and TNF-${\alpha}$ (0.1-1 mg/ml). Additionally, Careswell significantly inhibited the activation of NF-${\kappa}B$ in PMACI-stimulated HMC-1. Conclusion : Collectively, the findings of this study provide us with a novel insight into the pharmacological actions of Careswell as a potential molecule for use in the treatment of allergic inflammation diseases.

• Journal of Microbiology and Biotechnology
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• v.11 no.2
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• pp.186-192
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• 2001

A primary culture of human fetal hepatocytes was obtained through a therapeutic abortion process at 26 weeks of gestation period. More than $10^8$ cells were seeded on a plastic plate. These hepatocytes were infected with hepatitis B virus (HBV). The HBV was purified from serum of one chronic HBV carrier. Transformed hepatocytes were subcultured in a 10% FBS-supplemented medium. The morphology of the transformed cell was epithelial-like. The cells from the first pass showed signs of early proliferation and had a latent period of more than 3 months after 6-7 passages. After the rest period, the transformed cell proliferated actively and they were subcultured every three days. Transformed hepatocytes were characterized by detection of the HBV transcript by RT-PCR. The secretion of virions from transformed cells was investigated by PCR with the cell medium. Two types of virions secreted into the culture medium were examined by using the transmission electron microscope. Another approach to study the secretion of virions in to culture medium was carried out with HBV antibody. HBsAg was detected in the culture medium of transformed cells using ELISA and Western blot analyses. These data suggested that the human fetal hepatocyte cell line has been established by infection of HBV, in which this cell line secreted viral particles into the culture medium.

• Advances in Traditional Medicine
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• v.4 no.3
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• pp.171-178
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• 2004

Ginseng radix, the root of Panax ginseng C. A. Meyer (Araliaceae), is a medicinal plant used world-widely and has been reported to have various biological effects. To investigate the effects of Ginseng radix on HL-60 cell apoptosis, MTT assay, DNA fragmentation assay and flow cytometry were performed on HL-60 cells. Cells were treated with Ginseng radix at different concentrations $(10^{-4},\;10^{-3}\;and\;10^{-2};\;dilution\;rate)$. Ginseng radix significantly induced cells apoptosis with a time- and dose-dependent manner. To determine whether Ginseng radix-induced apoptosis is due to increase of tumor necrosis factor $(TNF-{\alpha})$ secretion, enzyme-linked immunosorbent assay was performed on HL-60 cells. Unexpectedly, Ginseng radix $(96\;{\pm}\;5\;pg/ml)$ significantly decreased the $TNF-{\alpha}$ secretion compared with control $(174\;{\pm}\;14\;pg/ml)$. Furthermore, Ginseng radix with $rIFN-{\gamma}$ synergistically increased nitric oxide production in mouse peritoneal macrophages. Taken together, our data indicate that Ginseng radix induce apoptosis on HL-60 cells without increase of $TNF-{\alpha}$ secretion and could be used for a supplementary remedy of cancer.

• Ocean and Polar Research
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• v.40 no.4
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• pp.249-257
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• 2018

Matrix metalloproteinases (MMPs) are associated with the invasion and metastasis of malignant tumors composed of cancer cells in an increased state of expression. This study evaluates the inhibitory effect of Carex pumila on MMP-2 and MMP-9 activity in phorbol-12-myristate-13-acetate (PMA)-stimulated HT-1080 human fibrosarcoma cells using gelatin zymography, MMPs enzyme-linked immunosorbent assay (ELISA), reverse transcription-polymerase chain reaction (RT-PCR) and Western blot assay. C. pumila was extracted twice with dichloromethane ($CH_2Cl_2$) and methanol (MeOH). Treatment with $CH_2Cl_2$ extract and MeOH extract in PMA-stimulated HT-1080 cells effectively reduced the production of MMP-2 and 9. Also, the combined crude extracts ($CH_2Cl_2$ and MeOH) significantly inhibited the enzymatic activities and the expression of MMP-2 and MMP-9 in mRNA and protein levels. The combined crude extracts were partitioned between $CH_2Cl_2$ and water. The organic layer was further fractionated with n-hexane, 85% aqueous methanol (85% aq.MeOH) and the aqueous layer was separated into n-butanol and water, successively. Of the fractions, 85% aq.MeOH fraction showed the highest inhibitory activity of MMP-2 and MMP-9 in gelatin zymography and MMP ELISA kit. Furthermore, 85% aq.MeOH fraction most significantly suppressed cell migration. In RT-PCR and Western blot assay, n-butanol and 85% aq.MeOH fractions exerted the greatest inhibition on mRNA and protein expression of MMP-2 and MMP-9, respectively. As a result, C. pumila can be used as a good anti-invasive agent source.

• Journal of dental hygiene science
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• v.20 no.4
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• pp.213-220
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• 2020

Background: The leaves of Perilla frutescens, commonly called perilla and used for food in Korea, contain components with a variety of biological effects and potential therapeutic applications. The purpose of this study was to identify the components of 70% ethanol extracted Perilla frutescens (EEPF) and determine its inhibitory effects on oral microbial activity and production of nitric oxide (NO) and prostaglandin E2 (PGE2) in lipopolysaccharides (LPS)-stimulated Raw264.7 macrophages, consequently, to confirm the possibility of using EEPF as a functional component for improving the oral environment and preventing inflammation. Methods: One kg of P. frutescens leaves was extracted with 70% ethanol and dried at -70℃. EEPF was analyzed using high-performance liquid chromatography analysis, and antimicrobial activity against oral microorganisms was revealed using the disk diffusion test. Cell viability was elucidated using a methylthiazolydiphenyl-tetrazolium bromide assay, and the effect of EEPF on LPS-induced morphological variation was confirmed through microscopic observation. The effect of EEPF on LPS-induced production of pro-inflammatory mediators, NO and PGE2 was confirmed by the NO assay and PGE2 enzyme-linked immunosorbent assay. Results: The main component of EEPF was rosemarinic acid, and EEPF showed weak anti-bacterial and anti-fungal effects against microorganisms living in the oral cavity. EEPF did not show toxicity to Raw264.7 macrophages and had inhibitory effects on the morphological variations and production of pro-inflammatory mediators, NO and PGE2 in LPS-stimulated Raw264.7 macrophages. Conclusion: EEPF can be used as a functional material for improving the oral environment through the control of oral microorganisms and for modulating inflammation by inhibiting the production of inflammatory mediators.

• Applied Chemistry for Engineering
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• v.33 no.6
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• pp.557-562
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• 2022

Hyaluronic acid (HA) is a mucopolysaccharide, occurring naturally in living organisms. It is one of the most hydrophilic molecules, so it has been known as being related to skin hydration and skin aging. The purpose of this study was to examine the effects of Cimicifuga heracleifolia ethanol extract on the hyaluronic acid synthesis and the inhibition of hyaluronidase activity. To determine cytotoxicity, hyaluronic acid synthase 2 (HAS2) gene expression, HA production and, hyaluronidase inhibitory effects, 3-(4,5-dimethylthiazol-2-ly)-2,5-diphenyl tetrazolium bromide (MTT) assay, real time - polymerase chain reaction (RT-PCR), hyaluronic acid enzyme linked immunosorbent assay (HA-ELISA), and hyaluronidase assay were used, respectively. When the Cimicifuga heracleifolia extract was treated in the HaCaT cells up to 500 ㎍/mL concentration, cytotoxicity was confirmed by the Cimicifuga heracleifolia extract at concentrations above 200 ㎍/mL. Therefore, the optimum concentration of all experiments used in this study was determined to be 200 ㎍/mL. HAS2 gene expression increased by Cimicifuga heracleifolia extract in a concentration-dependent manner at all treatment concentrations. The production rate of HA was tended to decrease at the highest concentration of 200 ㎍/mL. The hyaluronidase activity inhibition effect of Cimicifuga heracleifolia extract was very high compared to the control group. Based on these results, Cimicifuga heracleifolia extract was expected to have a moisturizing effect on human skin and special attention should be paid to the determination of the concentration of Cimicifuga heracleifolia when developing cosmetic materials using it.

• Asian Pacific Journal of Cancer Prevention
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• v.17 no.3
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• pp.1399-1404
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• 2016

We investigated the urinary levels of 14-3-3 protein beta/alpha to evaluate their diagnostic significance with regard to clear cell renal cell carcinoma (ccRCC) and angiomyolipoma (AML). Urine samples from 91 patients with ccRCC, 16 patients with AML and 24 healthy volunteers were assessed. We used an enzyme-linked immunosorbent assay (ELISA) to quantify 14-3-3 protein beta/alpha levels in urine. Values were higher in patients with ccRCC than in those with AML and in healthy volunteers. High levels were associated with pathologic stage, lymph node status, distant metastasis and poor survival. Urinary levels of 14-3-3 protein beta/alpha were significantly increased in patients with small-sized carcinoma, irrespective of being less than 4.0 cm and 2.0 cm, compared with levels in patients with AML. This study is the first to report that increased expression of 14-3-3 protein beta/alpha in urine is associated with advanced stage and poor survival in patients with ccRCC. In addition, urinary 14-3-3 protein beta/alpha may differentiate AML from RCC, even when small sized. These results suggest that examination of urinary 14-3-3 protein beta/alpha could serve as a diagnostic and prognostic marker in patients with ccRCC.

• IMMUNE NETWORK
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• v.6 no.3
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• pp.138-144
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• 2006

Background: There are at least two different subsets of B cells, B-1 and B-2. The characteristic features and function of B-2 cells in addition to the effect of steroids on B-2 cells are well-known. Although B-1 cells have different features and functions from B-2 cells, the effect of steroids on B-1 cells is not completely understood. Therefore, this study examined the effects of dexamethasone on peritoneal (or B-1 cells) and splenic B cells (or B-2 cells). Methods: Purified B cells were obtained from the peritoneal fluid and the spleens of mice. The isolated B cells were cultured in a medium and after adding different concentrations of dexamaethasone. The cell survival rate was measured by flow cytometry using propidium iodide. The expression level of the B cell surface marker was analyzed by flow cytometry. During the culture of these cells, immunoglobulin secreted into the culture supernatants was evaluated by an enzyme-linked immunosorbent assay. Results: The survival rate of peritoneal and splenic B cells decreased with increasing dexamethasone concentration. However, the rate of peritofieal B cell apoptosis was lower than that of splenic B cells. CDS and B7.1 expression in peritoneal B cells and CD23 and sIgM expression in splenic B cells after the dexamethasone treatment were reduced. When B cells were treated with dexamethasone, the spontaneous IgM secretion decreased with increasing dexamethasone concentration. Conclusion: Dexamethasone induces apoptosis in peritoneal and splenic B cells. However, peritoneal B cells are less sensitive to dexamethasone. The dexamethasone suppressed expression of the surface markers in peritoneal B cells is different from those in splenic B cells.

• Asian Pacific Journal of Cancer Prevention
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• v.14 no.3
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• pp.1721-1724
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• 2013

Objective: To investigate the effect of epirubicin on soluble CD25 (sCD25) secretion by CD4+CD25+ regulatory T (Treg) cells isolated from diffuse large B-cell lymphoma (DLBCL) patients. Methods: Treg cells were isolated from the peripheral blood mononuclear cells isolated from the newly diagnosed DBLCL patients. The concentration of sCD25 in the supernatant was determined with a commercial sCD25 (IL-2R) enzyme-linked immunosorbent assay (ELISA) kit. The fluorescence intensity of CD25 was detected by flow cytometry. Results: Cell survival rate was significantly decreased along with the increase of epirubicin concentration after treatment for 24 h. There was also a significant difference in the concentration of sCD25 between the epirubicin group and the control group (P<0.01). A positive correlation between the Treg cells survival rate and the concentration of sCD25 was detected (r=0.993, P<0.01). When equal numbers of CD4+CD25+ Treg cells of the epirubicin group and the control group were cultured for another 24 h without epirubicin the CD25 fluorescence intensity on the surface of Treg cells was obviously higher in the epirubicin group than that in the control group (P<0.01), while the sCD25 concentration in the supernatant in the epirubicin group was significantly lower than that in the control group (P<0.05). Conclusion: Epirubicin may improve the body's immune functions by inhibiting the sCD25 secretion by Treg cells in DLBCL patients.