• Archives of Pharmacal Research
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• v.29 no.10
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• pp.911-920
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• 2006

Phenolic antioxidant butylated hydroxyanisole (BHA) is a commonly used food preservative with broad biological activities, including protection against chemical-induced carcinogenesis, acute toxicity of chemicals, modulation of macromolecule synthesis and immune response, induction of phase II detoxifying enzymes, as well as its undesirable potential tumor-promoting activities. Understanding the molecular basis underlying these diverse biological actions of BHA is thus of great importance. Here we studied the pharmacokinetics, activation of signaling kinases and induction of phase II/III drug metabolizing enzymes/transporter gene expression by BHA in the mice. The peak plasma concentration of BHA achieved in our current study after oral administration of 200 mg/kg BHA was around $10\;{\mu}M$. This in vivo concentration might offer some insights for the many in vitro cell culture studies on signal transduction and induction of phase II genes using similar concentrations. The oral bioavailability (F) of BHA was about 43% in the mice. In the mouse liver, BHA induced the expression of phase II genes including NQO-1, HO-1, ${\gamma}-GCS$, GST-pi and UGT 1A6, as well as some of the phase III transporter genes, such as MRP1 and Slco1b2. In addition, BHA activated distinct mitogen-activated protein kinases (MAPKs), c-Jun N-terminal kinase (JNK), extracellular signal-regulated protein kinase (ERK), as well as p38, suggesting that the MAPK pathways may play an important role in early signaling events leading to the regulation of gene expression including phase II drug metabolizing and some phase III drug transporter genes. This is the first study to demonstrate the in vivo pharmacokinetics of BHA, the in vivo activation of MAPK signaling proteins, as well as the in vivo induction of Phase II/III drug metabolizing enzymes/transporters in the mouse livers.

• BMB Reports
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• v.50 no.5
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• pp.263-268
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• 2017

${\beta}$-Agarase cleaves the ${\beta}$-1,4 linkages of agar to produce neoagarooligosaccharides (NAO), which are associated with various physiological functions. However, the immunological functions of NAO are still unclear. In this study, we demonstrated that ${\beta}$-agarase DagA-produced neoagarohexaose (DP6), an NAO product, promoted the maturation of dendritic cells (DCs) by Toll-like receptor 4 (TLR4). DP6 directly and indirectly enhanced the activation of natural killer (NK) cells in a TLR4-dependent manner in vitro and in vivo. Finally, the antitumor activity of DP6 against B16F1 melanoma cells was inhibited in NK cell-depletion systems by using NK-cell depleting antibodies in vivo. Collectively, the results indicated that DP6 augments antitumor immunity against B16F1 melanoma cells via the activation of DC-mediated NK cells in a TLR4-dependent manner. Thus, DP6 is a potential candidate adjuvant that acts as an immune cell modulator for the treatment of melanoma.

• IMMUNE NETWORK
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• v.7 no.3
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• pp.133-140
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• 2007

Background: It is well established that cross talk between natural killer (NK) cells and myeloid dendritic cells (DC) leads to NK cell activation and DC maturation. In the present study, we investigated whether type 1-polarized DC (DC1) matured in the presence of IFN-${\gamma}$ and type 2-polarized DC (DC2) matured in the presence of PGE2 can differentially activate NK cells. Methods: In order to generate DC, plastic adherent monocytes were cultured in RPMI 1640 containing GM-CSF and IL-4. At day 6, maturation was induced by culturing the cells for 2 days with cytokines or PGE2 in the presence or absence of LPS. Each population of DC was cocultured with NK cells for 24 h. The antigen expression on DC was analyzed by flow cytometry and cytokine production in culture supernatant was measured by ELISA or a bioassay for TNF-${\alpha}$ determination. NK cell-mediated lysis was determined using a standard 4h chromium release assay. Results: DC2, unlike DC1, had weak, if any, ability to induce NK cell activation as measured by IFN-${\gamma}$ production and cytolytic activity. DC2 were weakly stimulated by activated NK cells compared to DC1. In addition, IFN-${\gamma}$-primed mature DC appeared to be most resistant to active NK cell-mediated lysis even at a high NK cell/DC ratio. On the other hand, PGE2-primed DC were less resistant to feedback regulation by NK cells than IFN-${\gamma}$-primed mature DC. Finally, we showed that the differential effect of two types of DC population on NK cell activity is not due to differences in their ability to form conjugates with NK cells. Conclusion: These results suggest that different combinations of inflammatory mediators differentially affect the effector function of DC and, as a result, the function of NK cells, eventually leading to distinct levels of activation in adaptive immunity.

• Journal of Life Science
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• v.31 no.2
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• pp.237-247
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• 2021

Immune responses in the central nervous system (CNS) function as the host's defense system against pathogens and usually help with repair and regeneration. However, chronic and exaggerated neuroinflammation is detrimental and may create neuronal damage in many cases. The NOD-, LRR-, and pyrin domain―containing 3 (NLRP3) inflammasome, a kind of NOD-like receptor, is a cytosolic multiprotein complex that consists of sensors (NLRP3), adaptors (apoptosis-associated speck like protein containing a caspase recruitment domain, ASC) and effectors (caspase 1). It can detect a broad range of microbial pathogens along with foreign and host-derived danger signals, resulting in the assembly and activation of the NLRP3 inflammasome. Upon activation, NLRP3 inflammasome leads to caspase 1-dependent secretion of the pro-inflammatory cytokines IL-1β and IL-18, as well as to gasdermin D-mediated pyroptotic cell death. NLRP3 inflammasome is highly expressed in CNS-resident cell types, including microglia and astrocytes, and growing evidence suggests that NLRP3 inflammasome is a crucial player in the pathophysiology of several neuroinflammatory and psychiatric diseases, such as Alzheimer's disease, Parkinson's disease, multiple sclerosis, stroke, traumatic brain injury, amyotrophic lateral sclerosis, and major depressive disorder. Thus, this review describes the molecular mechanisms of NLRP3 inflammasome activation and its crucial roles in the pathogenesis of neurological disorders.

• Biomolecules & Therapeutics
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• v.30 no.6
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• pp.529-539
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• 2022

Rheumatoid arthritis (RA) is a multifactorial immune-mediated disease, the pathogenesis of which involves different cell types. T-cell activation plays an important role in RA. Therefore, inhibiting T-cell activation is one of the current therapeutic strategies. Cytotoxic T-lymphocyte antigen 4-immunoglobulin (CTLA4-Ig), also known as abatacept, reduces cytokine secretion by inhibiting T-cell activation. To achieve a homeostatic therapeutic effect, CTLA4-Ig has to be administered repeatedly over several weeks, which limits its applicability in RA treatment. To overcome this limitation, we increased the number of sialic acid-capped antennas by genetically engineering the CTLA4 region to increase the therapeutic effect of CTLA4-Ig. N-acetylglucosaminyltransferase (GnT) and α2,6-sialyltransferase (α2,6-ST) were co-overexpressed in Chinese hamster ovary (CHO) cells to generate a highly sialylated CTLA4-Ig fusion protein, named ST6. The therapeutic and immunogenic effects of ST6 and CTLA4-Ig were compared. ST6 dose-dependently decreased paw edema in a mouse model of collagen-induced arthritis and reduced cytokine levels in a co-culture cell assay in a similar manner to CTLA4-Ig. ST6- and CTLA4-Ig-induced T cell-derived cytokines were examined in CD4 T cells isolated from peripheral blood mononuclear cells after cell killing through irradiation followed by flow- and magnetic-bead-assisted separation. Interestingly, compared to CTLA4-Ig, ST6 was substantially less immunogenic and more stable and durable. Our data suggest that ST6 can serve as a novel, less immunogenic therapeutic strategy for patients with RA.

• Clinical and Experimental Reproductive Medicine
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• v.50 no.1
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• pp.44-52
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• 2023

Objective: The DNA integrity of spermatozoa that attach to fallopian tube (FT) cells is higher than spermatozoa that do not attach. FT epithelial cells can distinguish normal and abnormal sperm chromatin. This study investigated the effects of sperm with a high-DNA fragmentation index (DFI) from men with unexplained repeated implantation failure (RIF) on the Toll-like receptor (TLR) signaling pathway in human FT cells in vitro. Methods: Ten men with a RIF history and high-DFI and 10 healthy donors with low-DFI comprised the high-DFI (>30%) and control (<30%) groups, respectively. After fresh semen preparation, sperm were co-cultured with a human FT epithelial cell line (OE-E6/E7) for 24 hours. RNA was extracted from the cell line and the human innate and adaptive immune responses were tested using an RT2 profiler polymerase chain reaction (PCR) array. Results: The PCR array data showed significantly higher TLR-1, TLR-2, TLR-3, TLR-6, interleukin 1α (IL-1α), IL-1β, IL-6, IL-12, interferon α (IFN-α), IFN-β, tumor necrosis factor α (TNF-α), CXCL8, GM-CSF, G-CSF, CD14, ELK1, IRAK1, IRAK2, IRAK4, IRF1, IRF3, LY96, MAP2K3, MAP2K4, MAP3K7, MAP4K4, MAPK8, MAPK8IP3, MYD88, NFKB1, NFKB2, REL, TIRAP, and TRAF6 expression in the high-DFI group than in the control group. These factors are all involved in the TLR-MyD88 signaling pathway. Conclusion: The MyD88-dependent pathway through TLR-1, TLR-2, and TLR-6 activation may be one of the main inflammatory pathways activated by high-DFI sperm from men with RIF. Following activation of this pathway, epithelial cells produce inflammatory cytokines, resulting in neutrophil infiltration, activation, phagocytosis, neutrophil extracellular trap formation, and apoptosis.

• Journal of Life Science
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• v.23 no.1
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• pp.63-68
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• 2013

The effect of black garlic extract on the activation of spleen cells from a C57BL6 mouse was investigated to examine immune activities of of fermented black garlic containing a variety of bioactive substances. xtract obtained from the concentration of commercial Namhae black garlic was used for the analysis of immune activities. Treatment with the extract increased the expression of interleukin-2 (IL-2) cytokine. The simultaneous administration of the extract plus lipopolysaccharide (LPS) increased the expression of IL-2, tumor necrosis factor (TNF)-${\alpha}$, and interferon (IFN)-${\gamma}$ compared with that of a control group. This result suggests that cellular immunity can be induced by macrophages, resulting in the expression of T lymphocytes and T helper type 1 (Th1) cells. In addition, treatment with the extract increased the late response of IL-6 cytokines, and the extract plus LPS augmented the expression of IL-4 and IL-6 compared with that of an LPS-treated group. Meanwhile, the extract plus LPS decreased the late response of IL-10, suggesting that humoral immunity can be activated by stimulating B lymphocytes, suppressing cellular immunity, and effectively modulating the conversion into humoral immune responses. These findings demonstrate that the black garlic extract activates Th1 and Th2 cells by stimulating T lymphocytes in mouse spleen cells and leads to immunomodulation by activating cellular and humoral immune responses of the immune system.

• Journal of the Korean Society of Food Science and Nutrition
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• v.34 no.5
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• pp.605-612
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• 2005

In previous studies, we evaluated the effect of the 6 energy-tonic or blood-building prescriptions of traditional oriental medicine, and observed that Si-Wu-Tang and Bu-Zhong-Yi-Qi-Tang showed high activity in the protection of the gastrointestinal and hematopoietic organs in irradiated mice. But any of these prescriptions did not show a high activity in the activation of the immune cells. We performed this study to design an herb mixture which protects the self-renewal tissues and also promotes recovery of the immune system against radiation damage. In order to meet all the requirements, we designed a new mixture of 3 edible herbs listed in Korean Food Code. The mixture of Angelim gigas radix, Cnidium officinale rhizoma and Paeonia japonica radix was decocted with hot water, and the activities of the water extract (HIM-I) were evaluated. HIM-1 stimulated the immune cells in a much higher extent than the traditional prescriptions, and promoted dramatically the growth of bone marrow stem cells in vitro. Also, HIM-1 protected digestive and hematopoietic organs against radiation as effectively as the 2 prescriptions, Si-Wu-Tang and Bu-Zhong-Yi-Qi-Tang. On the other hand, it showed high in vitro antioxidative activity that might be considered as a mechanism of the protective effects against radiation. Although the detailed mechanisms of those effects remain to be elucidated, these results indicated that HIM-I might be a useful agent for protection and recovery of body from various risk factor as well as radiation, especially since it is a relatively nontoxic natural product.

• Tuberculosis and Respiratory Diseases
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• v.54 no.5
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• pp.551-562
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• 2003

Background : NF-${\kappa}B$ is a characteristic transcriptional factor which has been shown to regulate production of acute inflammatory mediators and to be involved in the pathogenesis of many inflammatory lung diseases. There has been some evidence that PI3K/Akt pathway could activate NF-${\kappa}B$ in human cell lines. However, the effect of PI3K/Akt pathway on the activation of NF-${\kappa}B$ varied depending on the cell lines used in the experiments. In this study we evaluated the effect of PI3K/Akt pathway on the activation of NF-${\kappa}B$ in human respiratory epithelial cell lines. Methods : BEAS-2B, A549 and NCI-H157 cell lines were used in this experiment. To evaluate the activation of Akt activation and I${\kappa}B$ degradation, cells were analysed by western blot assay using phospho-specific Akt Ab and $I{\kappa}B$ Ab. To block PI3K/Akt pathway, cells were pretreated with wortmannin or LY294002 and transfected with dominant negative Akt (DN-Akt). For IKK activity, immune complex kinase assay was performed. To evaluate the DNA binding affinity and transcriptional activity of NF-${\kappa}B$, electrophoretic mobility shift assay (EMSA) and luciferase assay were performed, respectively. Results : In BEAS-2B, A549 and NCI-H157 cell lines, Akt was activated by TNF-$\alpha$ and insulin. Activation of Akt by insulin did not induce $I{\kappa}B{\alpha}$ degradation. Blocking of PI3K/Akt pathway via wortmannin/LY294002 or DN-Akt did not inhibit TNF-$\alpha$-induced $I{\kappa}B{\alpha}$ degradation or IKK activation. Inhibition of PI3K/Akt did not affect TNF-$\alpha$-induced NF-${\kappa}B$ activation. Overexpression of DN-Akt did not block TNF-$\alpha$-induced transcriptional activation of NF-${\kappa}B$, but wortmannin enhanced TNF-$\alpha$-induced in NF-${\kappa}B$ transcriptional activity. Conclusion : PI3K/Akt was not involved in TNF-$\alpha$-induced $I{\kappa}B{\alpha}$ degradation or transcriptional activity of NF-${\kappa}B$ in human respiratory epithelial cell lines.

• Radiation Oncology Journal
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• v.25 no.3
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• pp.185-191
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• 2007

Purpose: This study was performed to investigate the effects of immune cell activation and the antitumor effect for the combination of treatment with X-irradiation and E/eutherococcus senticosus Maxim Root (ESMR) on mouse tumor cells. Materials and Methods: ESMR (250g) was extracted with 80% methanol, concentrated under decompression and lyophilized. To determine whether ESMR is able to activate the immune cells or not, the proliferation of splenocytes in vitro and the number of B cells and T cells in splenic lymphocytes in ESMR-pretreated mice were evaluated. X-irradiation was given to the mouse fibrosarcoma tumor cells (FSa II) by 250 kv X-irradiation machine. The cytotoxicity of ESMR was evaluated from its ability to reduce the clonogenecity of FSa II cells. In X-irradiation alone group, each 2, 4, 6 and 8 Gy was given to FSa II cells. In X-irradiation with ESMR group, 0.2 mg/ml of ESMR was exposed to FSa II cells for 1 hour before X-irradiation. Results: The proliferation of cultured mouse splenocytes and thymocytes were enhanced by the addition of ESMR in vitro. The number of B cells and T cells in mouse splenic lymphocytes was significantly increased in ESMR pretreated mice in vivo. In FSa II cells that received a combination of 0.2 mg/ml of ESMR with X-irradiation exposure, the survival fraction with a dose of 2, 4 and 6 Gy was $0.39{\pm}0.005$, $0.22{\pm}0.005$ and $0.06{\pm}0.007$, respectively. For FSa II cells treated with X-irradiation alone, the survival fraction with a dose of 2, 4 and 6 Gy was $0.76{\pm}0.02$, $0.47{\pm}0.008$ and $0.37{\pm}0.01$. The difference in the survival fraction of the mouse FSa II cells treated with and without ESMR was statistically significant (p<0.05). Conclusion: Treatment with ESMR increased cell viability of mouse splenocytes in vitro and especially the subpopulation of B cells and T cells in splenocytes in ESMR-pretreated mice. However, treatment with ESMR did not increase the level of Th and Tc subpopulations in the thymocytes. Treatment with the combination of ESMR and X-irradiation was more cytotoxic to mouse tumor cells than treatment with X-irradiation alone; this finding was statistically significant.