• Journal of the Korean Society of Food Science and Nutrition
• /
• v.46 no.3
• /
• pp.306-313
• /
• 2017

Purple bamboo salt (PuBS) is commonly used as a medicinal food in Korea and has beneficial potentials such as antioxidant and anti-inflammatory effects. Rubus coreanus is called Bokbunja, which is used as a traditional medicine for treating asthma, impotence, and allergic diseases in Korea. The aim of present study was to investigate the immunostimulatory effect of PuBS composed with Rubus coreanus (PuBS-R). We performed comparative analysis between PuBS and PuBS-R in Raw264.7 cells, which is a mouse macrophage cell line, and peritoneal macrophages isolated primarily from the mouse peritoneal cavity. We evaluated cytotoxicity and the immune cytokine response in PuBS- and PuBS-R-treated cells. Both PuBS and PuBS-R did not have any cytotoxicity in Raw264.7 cells up to $500{\mu}g/mL$. Gene and protein levels of immune cytokines such as tumor necrosis $factor-{\alpha}$ ($TNF-{\alpha}$), $interferon-{\gamma}$ ($IFN-{\gamma}$), interleukin (IL)-10, and IL-12 were significantly elevated by PuBS-R more than PuBS in Raw264.7 cells. Moreover, we evaluated the immunostimulatory effects of PuBS-R on mouse primary peritoneal macrophages. Protein levels of inducible nitric oxide synthase, $TNF-{\alpha}$, $IFN-{\gamma}$, IL-10, and IL-12 were significantly higher in PuBS-R-treated peritoneal macrophages than PuBS-treated peritoneal macrophages. These results suggest the potential immunostimulatory effect of PuBS-R for immunity against harmful infection.

• Korean Journal of Food Science and Technology
• /
• v.47 no.1
• /
• pp.119-125
• /
• 2015

The aim of this study was to investigate the synergistic anti-inflammatory effect of rosmarinic acid (RA) and luteolin from perilla (Perilla frutescens L.) leaves in lipopolysaccharide (LPS)-stimulated RAW264.7 macrophages. A combination of RA and luteolin more strongly inhibited the production of nitric oxide (NO), inducible NOS (iNOS), prostaglandin $E_2$ ($PGE_2$), and COX-2 than higher concentrations of RA or luteolin alone in LPS-stimulated RAW264.7 macrophages. The combined RA and luteolin synergistically inhibited the production of pro-inflammatory cytokines, such as tumor necrosis factor-${\alpha}$ (TNF-${\alpha}$), interleukin-6 (IL-6), and interleukin-$1{\beta}$ (IL-$1{\beta}$), in LPS-stimulated RAW264.7 macrophages. Furthermore, combined RA and luteolin more strongly suppressed NF-${\kappa}B$ activation than RA or luteolin alone, by inhibiting the degradation of inhibitor of NF-${\kappa}B(I{\kappa}B)$-${\alpha}$ and nuclear translocation of the p65 subunit of NF-${\kappa}B$ in LPS-stimulated RAW264.7 macrophages. Collectively, these results suggest that RA and luteolin in combination exhibit synergistic effects in suppression of LPS-induced inflammation in RAW264.7 macrophages.

• Tuberculosis and Respiratory Diseases
• /
• v.80 no.1
• /
• pp.77-82
• /
• 2017

Background: Delayed hypersensitivity plays a large role in the pathogenesis of tuberculous pleural effusion (TPE). Macrophages infected with live Mycobacterium tuberculosis (MTB) increase the levels of adenosine deaminase2 (ADA2) in the pleural fluid of TPE patients. However, it is as yet unclear whether ADA2 can be produced by macrophages when challenged with MTB antigens alone. This study therefore evaluated the levels of ADA2 mRNA expression, using monocyte-derived macrophages (MDMs) stimulated with MTB antigens. Methods: Purified monocytes from the peripheral blood mononuclear cells of healthy volunteers were differentiated into macrophages using granulocyte-macrophage colony-stimulating factor (GM-CSF) or macrophage colony-stimulating factor (M-CSF). The MDMs were stimulated with early secretory antigenic target protein 6 (ESAT6) and culture filtrate protein 10 (CFP10). The mRNA expression levels for the cat eye syndrome chromosome region, candidate 1 (CECR1) gene encoding ADA2 were then measured. Results: CECR1 mRNA expression levels were significantly higher in MDMs stimulated with ESAT6 and CFP10, than in the unstimulated MDMs. When stimulated with ESAT6, M-CSF-treated MDMs showed more pronounced CECR1 mRNA expression than GM-CSF-treated MDMs. Interferon-${\gamma}$ decreased the ESAT6- and CFP10-induced CECR1 mRNA expression in MDMs. CECR1 mRNA expression levels were positively correlated with mRNA expression of tumor necrosis factor ${\alpha}$ and interleukin 10, respectively. Conclusion: ADA2 mRNA expression increased when MDMs were stimulated with MTB antigens alone. This partly indicates that pleural fluid ADA levels could increase in patients with culture-negative TPE. Our results may be helpful in improving the understanding of TPE pathogenesis.

• Korean Journal of Veterinary Research
• /
• v.42 no.3
• /
• pp.351-362
• /
• 2002

Newcastle disease (ND) is a highly contagious infection of poultry, Two pathology-based techniques, in situ RT-PCR and in situ hybridization (ISH) were applied to formalin-fixed, paraffin-embedded tissues from chickens naturally infected with velogenic ND virus (VNDV). Two pairs of primers and a probe for ISH and in situ RT-PCR, respectively, were selected from highly conserved region of matrix gene of NDV. The ISH experiment was carried out using MicroProbe$^{TM}$ capillary action system within 2 hours. In situ RT-PCR was performed using MicroProbe$^{TM}$ capillary action system and GeneAmp In Situ PCR system. With ISH and in situ RT-PCR, viral nucleic acid was detected in the central nervous system of chickens from infected with neurotropic velogenic Newcastle disease virus (NVNDV), whereas viral nucleic acid was detected in various organs or tissues of chickens from infected with viscerotropic velogenic Newcastle disease virus (VVNDV). In the NVND group, positive signals were characteristically defined in the cytoplasm of neuron, vascular endothelial cells, and perivascular mononuclear macrophages in the central nervous system. One of NVND group, chicken from one farm exhibited positive signals in the bronchial epithelium. The VVND group chickens showed positive reaction in the macrophages, vascular endothelium, and bronchiolar epithelium. Markedly, viral nucleic acid was detected in the macrophages of morphologically normal tissues which were peripheral or located in distant areas from lesions. The central nervous system of chickens infected with VVND virus had positive signals in the vascular endothelial cell, perivascular mononuclear macrophages and some neuron. The number and intensity of the positive cells by in situ RT-PCR were more and stronger, respectively, in comparison with those by ISH. Particularly, positive reaction was detected in macrophages infiltrating in cardiac muscle by in situ RT-PCR, but not obtained by ISH. Therefore, these results demonstrated that ISH is a rapid diagnostic method for detection of NDV and in situ RT-PCR can be used as an efficient method for detection of low viral load infection or subclinical viral infection of NDV.

• Journal of fish pathology
• /
• v.7 no.2
• /
• pp.127-149
• /
• 1994

Histopathological studies on the two lymphomyeloid organs of spleen and head kidney in tilapia, Oreochromis niloticus, were carried out to clarify the significance on the morphological characteristics of melano - macrophage centers (MMCs) which are varied in different physiological and pathological conditions of teleosts. To examine the histological changes by the artificial modification of the immunological states, tilapia were treated intraperitoneally with FKC and LPS of Edwardsiella tarda, and orally with dexamethasone, and then followed by the intraperitoneal injection of colloidal carbon for chasing the macrophages. There were marked differences in phagocytic avidity of macrophages, and accumulating patterns of carbon - ladening macrophages into the MMCs among the test groups. In the non - pretreated control group, carbon - ladening macrophages were densely accumulated at 12th and 20th day within the MMCs of head kidney and spleen, respectively. And, in the groups treated with bacterial antigens (FKC & LPS), the macrophages were more rapidly and densely aggregated within MMCs. But in the group with dexamethasone, only a few carbon particles were detected in both organs. Any compactly isolated form of particles was not found in this group. From the present results, it was strongly suggested that certain changes in immunological states of tilapia influence on the morphology of MMCs including the frequency of appearance, sizes, aggregating patterns or outlines. Therefore, morphology of MMCs would be very important in the interpretation for histopathological findings seen in the teleost's lymphomyeloid organs.

• The Journal of Korean Medicine
• /
• v.27 no.1 s.65
• /
• pp.184-195
• /
• 2006

Objectives : Betula Platyphylla(BP) is a traditional analgesic, anti-fungal, anti-inflammatory herb used in Chinese 1medicine. However, no information is available to explain its action. In this study. we investigated the anti-inflammatory 1effects of BP to elutidate the molecular pharmacological activity in the ethanol extract of BP(BPE). Methods : We performed WTS assay in lipopolysaccharide (LPS) stimulated RAW264.7 macrophages with BPE. Nitrite was measured by Griess assay, prostaglandin E2 (PGE2) by enzyme-linked immunosorbent assay (ELISA) in LPS induced RAW264.7 macrophages with BPE. Inducible nitric oxide synthase (iNOS) and cyclooxygenase (COX-2) were determined by Western blot. Activation of nuclear factor-kappaB (NF-kB) was measured by electrophoretic mobility shift assay (EMSA). Results : BPE significantly suppressed production of nitric oxide (NO) and PGE2 in LPS-induced RAW264.7 macrophages in a dose-dependent manner. The maximal inhibition rate of NO and PGE2 production by BPE was ca. 88.8% and 93% at the concentration of $100{\mu}g/ml$ (non-cytotoxic concentration), respectively. BPE also decreased iNOS protein and COX-2 protein in LPS-induced RAW264.7 macrophages. EMSA demonstrated that BPE inhibited the DNA binding activity of the NF-kB. Conclusions : These results suggest that BPE inhibits NF-${\kappa}B$-mediated gene expression and downregulates inflammatory mediator production in RAW264.7 macrophages.

• Biomedical Science Letters
• /
• v.20 no.4
• /
• pp.250-255
• /
• 2014

Mycobacterium tuberculosis (MTB) resides and replicates inside macrophages. In our previous report, we reported that CD8+ T cell-mediated immune responses specific for the peptide derived from MTB RNA polymerase beta-subunit ($RpoB_{127-135}$) could be induced in TB patients expressing HLA-$A^*0201$ subtype. In order to examine whether $RpoB_{127-135}$ specific CD8+ T cells can recognize MTB infected macrophages in vitro, CD8+ T cell lines specific for $RpoB_{127-135}$ peptide were generated from peripheral blood mononuclear cells (PBMCs) of healthy HLA-$A^*0201$ subjects by in vitro immunization technique. In this study, we observed $RpoB_{127-135}$ specific CD8+ T cells could recognize and destroy macrophages infected with MTB for 2 to 4 days. $RpoB_{127-135}$ specific CD8+ T cell immune response was inducible from PBMC of healthy subjects expressing HLA-$A^*0206$ subtype, one of HLA-A2 supertype members. Next, we investigated the HLA-I processing mechanism of $RpoB_{127-135}$ peptide in MTB infected macrophages. As a result, the presentation of the MTB derived epitope peptide, $RpoB_{127-135}$, to CD8+ T cells was not inhibited by the treatment with brefeldin-A (ER-Golgi transport inhibitor) or lactacystin (proteasome inhibitor), which blocks the classical HLA-I processing pathway. However, $RpoB_{127-135}$ specific CD8+ T cell activity was blocked either by the blocking agent for the endocytosis (cytochalasin D) or by the blocking antibody (W6/32) for HLA-I molecules. Therefore, the $RpoB_{127-135}$ peptide may be processed by accessing the alternate HLA-I processing pathway. Understanding the processing and presentation mechanisms of the MTB derived proteins will help to improve the efficacy of vaccines and the efficiency of therapeutic agents for TB.

• Journal of Pharmacopuncture
• /
• v.17 no.1
• /
• pp.7-12
• /
• 2014

Objectives: The objective of this study is to investigate the effects of Salviae Miltiorrhizae Radix hot aqueous extract on nitric oxide (NO) and prostaglandin $E_2$ ($PGE_2$) production and on 1,1-diphenyl-2-picryl hydrazyl (DPPH) free-radical scavenging in macrophages. Methods: Salviae Miltiorrhizae Radix (300 g) was heated at $100^{\circ}C$ with distilled water (2 L) for 4 hours. The extract was filtered and concentrated to 100 mL by using a rotary evaporator, was frozen at $-80^{\circ}C$, and was then freeze-dried by using a freezing-drying system. The RAW 264.7 macrophage was subcultured by using $10-{\mu}g/mL$ lipopolysaccharide (LPS). In order to evaluate cytotoxicity, we performed 3-(4,5-dimrthylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assays and measured the cell viability. The NO production was measured by using Griess assays, and the $PGE_2$ production was measured by using enzyme immunoassays. The antioxidant activity, the 1,1-diphenyl-2-picryl hydrazyl (DPPH) free-radical scavenging capability, was measured by using the DPPH method. Results: Cell viability with the 1-, 5-, 25-, 125- and $625-{\mu}g/mL$ Salviae Miltiorrhizae Radix hot aqueous extract was not significantly decreased compared to the cell viability without the extract. When 125 and $625{\mu}g/mL$ of Salviae Miltiorrhizae Radix hot aqueous extract were used, nitric oxide (NO) production in LPS-stimulated RAW 264.7 macrophages was significantly inhibited compared to that in the control group. When 25, 125, and $625{\mu}g/mL$ of Salviae Miltiorrhizae Radix hot aqueous extract were used, $PGE_2$ production in LPS-stimulated RAW 264.7 macrophages was significantly inhibited compared to that in the control group. The 125- and $625-{\mu}g/mL$ Salviae Miltiorrhizae Radix hot aqueous extracts had high DPPH free-radical scavenging capabilities in RAW 264.7 macrophages. Conclusion: This study indicates that Salviae Miltiorrhizae Radix hot aqueous extract suppresses NO and $PGE_2$ production and improves DPPH free-radical scavenging capability. Thus, it seems that Salviae Miltiorrhizae Radix hot aqueous extract may have an anti-inflammation effect and antioxidant activity.

• Herbal Formula Science
• /
• v.19 no.1
• /
• pp.207-217
• /
• 2011

Objectives : In a previous study, we have shown that Gagam-Gongjin-Dan(GGD) has an inhibitory effect on the ovalbumin-induced immune responses and a hepatoprotective effect on actaminophen-induced liver injury in Balb/c Mice. However, the possible anti-inflammatory effect of GGD extract for inflammatory mediators was not reported. Therefore, the purpose of this study was to investigate an inhibitory effects of GGD extract against lipopolysaccharides(LPS) induced inflammatory mediators in mouse peritoneal macrophages. Methods : GGD extract was prepared by extracting with methanol for 7 days. The extract was freeze-dried following filtration through vacuum distillation system. Accumulated nitrite, an oxidative product of nitric oxide(NO), was measured in the culture medium by the Griess reaction. The levels of prostaglandin $E_2(PGE_2)$, interleukin-$1{\beta}$(IL-$1{\beta}$), tumor necrosis factor-${\alpha}$(TNF-${\alpha}$) were measured by enzyme-linked immunosorbent assay. The expressions of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2(COX-2) were measured by Western blot analysis. Results : GGD extract (50-$400\;{\mu}g$/ml) per se had no cytotoxic effect in LPS-stimulated peritoneal macrophages. GGD extract dose-dependently reduced NO, $PGE_2$, IL-$1{\beta}$ and TNF-${\alpha}$ production and COX-2 activity caused by stimulation of LPS. The levels of iNOS and COX-2 protein expressions were markedly suppressed by the treatment with GGD extract in a dose dependent manner. Conclusions : These results suggest that GGD extract has an anti-inflammatory effect against LPS-induced inflammatory mediators in peritoneal macrophages, these properties may contribute to inflammation disease care.

• Journal of Gastric Cancer
• /
• v.5 no.3 s.19
• /
• pp.206-212
• /
• 2005

Purpose: Angiogenesis has a critical role in tumor proliferation, invasion, and metastasis. In gastric cancer, tumor-associated macrophages and mast cells produce angiogenic factors such as VEGF, that inhibit the functional maturation of dendritic cells. The aim of this study is to identify tumor-associated macrophages, mast cells, dendritic cell infiltrations, and microvessel densities (MVD) to investigate the relationship between them and the prognosis for gastric-cancer patients. Materials and Methods: The subjects were 79 patients selected from those who had undergone a curative gastric resection for stomach cancer. With them, Immune-histochemical staining was done using CD34 for the MVD, CD68 antigen for macrophages, and S-100 protein for dendritic cells, and toluidine blue staining was done for mast cells. Results: Macrophage infiltration showed a statistically significant positive correlation with histologic differentiation and a negative correlation with invasion depth, nodal metastasis, and stage. S-100 (+) dendritic cells and mast cells had no significant correlations with histologic differentiation, invasion depth, nodal metastasis, distant metastasis, stage, and MVD. As survival, no statistically significant differences were seen between the variables. Conclusion: Tumor-associated macrophages should be evaluated as possible prognostic markers in gastric-cancer patients.