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Abstract: Melanoma is an aggressive form of skin cancer due to its rapid metastasis. Recently, several studies have reported that selenium can prevent metastasis of melanoma cells, but the mechanism of this anti-metastatic ability is not fully understood. In this study, we investigated the effect of selenium on cell migration in melanoma and on tumor metastasis in mice. Interestingly, tumor metastasis was suppressed by selenium in a mouse model. Cell migration was measured by a wound-healing assay using selenium-treated melanoma cells. Treatment with a non-cytotoxic concentration of selenium suppressed migration of melanoma cells in a dose-dependent manner. In addition, we found decreased HIF-1α and VEGF expression in selenium-treated melanoma cells as compared to non-treated control cells. Mechanistically, our studies show that selenium inhibits IL-18 gene expression in a dose-dependent manner. IL-18 protein level was suppressed by treatment with selenium. The wound-healing assay revealed that the anti-metastatic effect of selenium was abrogated by treatment with exogenous IL-18. These results suggest that selenium might be a potent inhibitor of the metastatic capacity of melanoma cells, via down-modulation of IL-18 expression. [Copyright &y& Elsevier]

Abstract: Thymosin beta 4 (Tβ4) is the major G-actin sequestering molecule and is abundant in lymphoid tissues. However, it is not clear what regulates Tβ4 expression and what its function is on natural killer (NK) cells. We investigated whether interleukin-18 (IL-18) has a role in Tβ4 expression and if enhanced Tβ4 influences IL-18-mediated interferon-gamma (IFN-γ) secretion. In this study, recombinant human IL-18 (rhIL-18) enhanced the endogenous level of Tβ4 through p38MAPK and JNK signaling pathway in the human NK cell line, NK-92MI. Overexpression of endogeneous Tβ4 stimulated IFN-γ expression and secretion. Additionally, pretreatment with an inhibitor for Tβ4 decreased IL-18-enhanced IFN-γ secretion, and transfection with Tβ4-specific short hairpin RNA resulted in reduction of IFN-γ production in primary NK cells as well as in the human NK cell line. Taken together, these data indicated that Tβ4 is regulated by IL-18 and is involved in IL-18-enhanced IFN-γ secretion in NK cells. [Copyright &y& Elsevier]

Erythroid differentiation regulator (Erdr1) was first discovered in mouse leukemia cell lines and functions as a stress-related survival factor. This study investigated whether Erdr1 regulates murine melanoma progression, as well as the mechanism involved in Erdr1-regulated metastasis. The expression of Erdr1 is negatively correlated with IL-18 expression, which has a pro-cancer effect in melanoma. To study the role of Erdr1 as an anti-cancer factor, cell migration, invasion, and proliferation were measured. Erdr1 overexpression markedly inhibited the level of cell migration, invasion, and proliferation in B16F10 cells in vitro. In addition, Erdr1 overexpression significantly suppressed melanoma lung colonization, metastasis, and tumor growth in vivo. To identify the factors involved in Erdr1-reduced metastasis, heat shock protein 90 (HSP90), a well-known stress protein and contributor to tumor metastasis, was examined. We found that HSP90 was significantly decreased in Erdr1-overexpressing cells. Functional analysis demonstrated that HSP90 small-interfering RNA transfection reduced the migration ability and metastasis of melanoma. In conclusion, Erdr1 shows a powerful anti-metastasis effect that leads to the ability to reduce the metastatic potential of murine malignant melanoma cells. Erdr1 is an anti-metastatic factor that may be a possible therapeutic target for treatment of melanoma. [ABSTRACT FROM AUTHOR]

Abstract: Natural Killer cells are known to play a major role in the innate immune response against viral infections and tumor cells. Several viruses, such as CMV, EBV and HIV-1, have acquired strategies to escape elimination by NK cells. In this study, we observed that EBV infection increased expression of IDO on B cells. To evaluate the function of IDO associated with EBV infection, we investigated whether EBV-induced IDO could modulate expression of NK cell-activation receptor, NKG2D. When NK cells were co-incubated with EBV transformed B cells, surface expression of NKG2D was significantly reduced in NK cells. Incubation with L-kynurenine, an IDO metabolite, down-modulated NKG2D expression in NK cells in a dose- and time-dependent manner. Incubation with the JNK inhibitor SP600125 also inhibited NKG2D expression in NK cells. In addition, we observed that the effect of L-kynurenine was blocked by JNK agonist, anisomycin, suggesting the involvement of the JNK pathway in the signal transduction of L-kynurenine-reduced NKG2D expression. Furthermore, IL-18 significantly reduced L-kynurenine-induced down-regulation of NKG2D expression in NK cells. Taken together, these data indicate that down-regulation of NKG2D by EBV-induced IDO metabolite provides a potential mechanism by which EBV escapes NKG2D-mediated attack by immune cells. [Copyright &y& Elsevier]

Abstract: Heat-killed lactic acid bacteria (LAB) are known to be important immunomodulators that stimulate tumor necrosis factor-α (TNF-α) and nitric oxide (NO) production as well as increase phagocytic activity in macrophages. NK cells play a critical role in innate immune response and induce spontaneous killing of tumor cells and virus-infected cells. However, the effect of heat-killed LAB on NK cells is still unclear. In this study, we investigated the effect of heat-killed Lactobacillus acidophilus La205 (La205) on NK cytolytic activity. We found that heat-killed La205 directly stimulated NK cytolytic activity in dose- and time-dependent manners. To determine the mechanism underlying heat-killed La205-enhanced NK cytotoxicity, the expression of NK activating receptors was tested. Heat-killed La205 did not affect the expression of NK activating receptors. To investigate whether NK degranulation is related to heat-killed La205-enhanced NK cytotoxicity, NK degranulation inhibitor concanamycin A (CMA) was used. CMA effectively blocked heat-killed La205-induced NK cytotoxicity, and an assay for detection of a degranulation marker, CD107a, showed that heat-killed La205 increased granule exocytosis approximately 2-fold in comparison to non-treated NK cells. In addition, heat-killed La205 dramatically elevated mRNA expression of granulysin, a component of the cytolytic granule contents, in NK cells. However, other granule contents, including perforin and granzymes, were not changed by heat-killed La205. From these data, we concluded that heat-killed La205 stimulated NK cytolytic activity through enhancement of granule exocytosis, and granulysin may be a critical mediator in heat-killed La205-induced granule exocytosis. [Copyright &y& Elsevier]

IL-32 was recently identified as a proinflammatory cytokine that is induced by IL-18 in natural killer (NK) cells and is highly correlated with inflammatory disorders. However, the relationship between IL-32 and tumor progression is still unknown. In this study, we investigated whether overexpression of IL-32 affects susceptibility of chronic myeloid leukemia (CML) cells to NK cells. Interestingly, IL-32α-overexpressing CML cell lines, K562, Kcl22, and BV173, showed higher NK cell-mediated killing. Flow cytometry analysis revealed that overexpression of IL-32α induced increased expression of Fas and UL16-binding protein 2 (ULBP2) in CML cells. The direct relationship between overexpression of surface molecules by IL-32α and increased NK cell-mediated killing was confirmed by Fas or ULBP2 siRNA transfection. IL-32α-induced Fas and ULBP2 expression are regulated p38 MAPK. In addition, the transcription factor Etsi plays a key role in ULBP2 specific expression by IL-32α overexpression in ULBP family members. Taken together) these data show that IL-32α stimulates Fas and ULBP2 expression via activation of p38 MAPK, which increases NK sus- ceptibility of CML cells. Enhanced NI( cell susceptibility of CML cells by IL-32α overexpression may improve the efficiency ófNK cell-based immunotherapy. [ABSTRACT FROM AUTHOR]

Abstract: Thymosin beta 4 (Tβ4), which is the major G-actin sequestering protein, has been shown to have ubiquitous distribution and multiple biological activities. However, Tβ4''s functions in relation to natural killer (NK) cells are still unknown. In this study, we show that synthetic Tβ4 peptide increases NK cell cytotoxicity mediated by intercellular adhesion molecule-1 (ICAM-1) through the secretion of cytolytic granules to target cells. This suggests that Tβ4 is a key activator of NK cell cytotoxicity. [Copyright &y& Elsevier]

Erythroid differentiation regulator 1 (Erdr1) has previously been reported to control thymocyte selection via TCR signal regulation, but the effect of Erdr1 as a TCR signaling modulator was not studied in peripheral T cells. In this report, it was determined whether Erdr1 affected TCR signaling strength in CD4 T cells. Results revealed that Erdr1 significantly enhanced the anti-TCR antibody-mediated activation and proliferation of T cells while failing to activate T cells in the absence of TCR stimulation. In addition, Erdr1 amplified Ca2+ influx and the phosphorylation of PLCγ1 in CD4 T cells with the TCR stimuli. Furthermore, NFAT1 translocation into nuclei in CD4 T cells was also significantly promoted by Erdr1 in the presence of TCR stimulation. Taken together, our results indicate that Erdr1 positively modulates TCR signaling strength via enhancing the PLCγ1/Ca2+/NFAT1 signal transduction pathway. [ABSTRACT FROM AUTHOR]

Abstract: T helper type 1 (Th1) cell-mediated immune responses play various roles in cellular immunity, including inducing cytotoxic T lymphocytes (CTLs) and they have been shown to be crucial in cancer immunotherapy. Previously, we found that aminoacyl-tRNA synthetase-interacting multifunctional protein 1 (AIMP1) stimulated antigen-presenting cells to secrete IL-12, leading to enhanced Th1 cell responses. In this study, as a way of enhancing antigen-specific Th1 responses, mouse fibroblasts (H-2b) were genetically modified to express an AIMP1 and a costimulatory B7.1 (Fb/AIMP1/B7.1). Fb/AIMP1/B7.1 cells were then loaded with an ovalbumin epitope as a model antigen (Fb/AIMP1/B7.1/OVA), and tested to determine if they induced OVA-specific CTLs in C57BL/6 mice (H-2b). Immunization with Fb/AIMP1/B7.1/OVA cells induced strong cytotoxic activities against OVA-expressing EG7 tumor cells, but not against other H-2b tumor cells. The levels of the cytotoxic response in the immunized mice with Fb/AIMP1/B7.1/OVA cells were significantly higher than the responses in mice immunized with other cell constructs. CD8+ T cells were a major cell-type of OVA-specific antitumor immunity induced by Fb/AIMP1/B7.1/OVA cells. Furthermore, treatment with Fb/AIMP1/B7.1/OVA cells significantly prolonged the survival period of EG7 tumor-bearing mice. These results indicate that AIMP1-secreting, epitope-loaded fibroblasts efficiently induce antigen-specific CTL responses in mice. [Copyright &y& Elsevier]

Abstract: Expression of UL16-binding proteins (ULBPs) has been reported in various cancers, such as leukemia and melanoma, and also in some other cancer cell lines. However, the factors that modulate the expression of ULBPs are not well defined. In this study, we investigated the effects of IL-18 on the expression of NKG2D ligands in leukemia cells. IL-18 treatment increased ULBP2 expression in leukemia cells at the mRNA and protein levels. In addition, PD98059 (an ERK1/2 MAPK inhibitor) and SP600125 (a JNK inhibitor) attenuated IL-18-induced ULBP2 expression in a dose-dependent manner. We observed that ERK1/2 and JNK MAPK phosphorylation increased upon treatment with IL-18. IL-18 elevated CD107a expression in cancer cells and increased the cytotoxic activity of NK cells; therefore, we propose that IL-18 increases the susceptibility of target cells by inducing surface expression of ULBP2. Taken together, these findings suggest that IL-18 may play a critical role in regulating ULBP2 expression via the ERK1/2 and JNK MAPK pathways in leukemia cells. [Copyright &y& Elsevier]

Abstract: Induction of differentiation is a new and promising approach to cancer therapy, well illustrated by the treatment of acute myeloid leukemia with all-trans retinoic acid (ATRA). Using combination of ATRA and chemotherapy, adverse effects such as retinoic acid syndrome have decreased, and long-term survival has improved. In this study, we demonstrated that the indeno[1,2-c]isoquinolines markedly enhanced differentiation of human myeloid leukemia HL-60 and NB4 cells when simultaneously combined with a low dose of ATRA. Of the tested compounds, 6-(4-methoxybenzyl)-2,11-dimethyl-6H,11H-indeno[1,2-c]isoquinolin-5-one (IIQ-16), an indeno[1,2-c]isoquinoline derivative, showed the highest differentiation-enhancing activity via a pathway involved with protein kinase C, extracellular signal-regulated kinase, and c-Jun N-terminal kinase. The ability to enhance the differentiation potential of ATRA by IIQ-16 may improve outcomes in the therapy of acute promyelocytic leukemia. [Copyright &y& Elsevier]

It is well known that UVB (290–320 nm) induces inflammation in skin by the transcription and release of cytokines and chemokines from skin keratinocytes. In addition, it is considered that intracellular reactive oxygen species (ROS) plays an important role in UVB-induced inflammatory response in the skin. Therefore, we investigated the effect of vitamin C, a potent antioxidant, on the regulation of UVB-induced skin inflammation via the modulation of chemokines production. Vitamin C uptake into keratinocytes is increased by UVB irradiation in a time- and dose-dependent manner through the translocation of sodium-dependent vitamin C transporter-1 (SVCT-1), a vitamin C-specific transporter, from the cytosol to the membrane. To evaluate the effect of vitamin C on the chemokine mRNA expression, we performed RNase protection assay. As a result, there was a remarkable change in chemokine mRNA expression, especially IL-8 and monocyte chemoattractant protein (MCP)-1 expression. In addition, increased IL-8 and MCP-1 mRNA expressions were suppressed by vitamin C treatment. We also confirmed the results of protein levels measured by ELISA. Taken together, vitamin C uptake is increased in UVB-irradiated keratinocytes through the translocation of SVCT-1 and regulates inflammatory response in the skin via the downregulation of IL-8 and MCP-1 production.Journal of Investigative Dermatology (2007) 127, 698–706. doi:10.1038/sj.jid.5700572; published online 28 September 2006 [ABSTRACT FROM AUTHOR]

In patients with autoimmune disorders such as rheumatoid arthritis (RA), delayed wound healing is often observed. Timely and effective wound healing is a crucial determinant of a patient's quality of life, and novel materials for skin wound repair, such as bioactive peptides, are continuously being studied and developed. One such bioactive peptide, AESIS-1, has been studied for its well-established anti-rheumatoid arthritis properties. In this study, we attempted to use the anti-RA material AESIS-1 as a therapeutic wound-healing agent based on disease-modifying antirheumatic drugs (DMARDs), which can help restore prompt wound healing. The efficacy of AESIS-1 in wound healing was assessed using a full-thickness excision model in diabetic mice; this is a well-established model for studying chronic wound repair. Initial observations revealed that mice treated with AESIS-1 exhibited significantly advanced wound repair compared with the control group. In vitro studies revealed that AESIS-1 increased the migration activity of human dermal fibroblasts (HDFs) without affecting proliferative activity. Moreover, increased HDF cell migration is mediated by upregulating chemokine receptor expression, such as that of CXC chemokine receptor 2 (CXCR2). The upregulation of CXCR2 through AESIS-1 treatment enhanced the chemotactic reactivity to CXCR2 ligands, including CXC motif ligand 8 (CXCL8). AESIS-1 directly activates the ERK and p38 mitogen-activated protein kinase (MAPK) signaling cascades, which regulate the migration and expression of CXCR2 in fibroblasts. Our results suggest that the AESIS-1 peptide is a strong wound-healing substance that increases the movement of fibroblasts and the expression of CXCR2 by turning on the ERK and p38 MAPK signaling cascades.

Activation of microglia, which is the primary immune cell of the central nervous system, plays an important role in neuroinflammation associated with several neuronal diseases. Aminoacyl tRNA synthetase (ARS) complex-interacting multifunctional protein 1 (AIMP1), a structural component of the multienzyme ARS complex, is secreted to trigger a pro-inflammatory function and has been associated with several inflammatory diseases. However, the effect of AIMP1 on microglial activation remains unknown. AIMP1 elevated the expression levels of activation-related cell surface markers and pro-inflammatory cytokines in primary and BV-2 microglial cells. In addition to the AIMP1-mediated increase in the expression levels of M1 markers [interleukin (IL)-6, tumor necrosis factor-α, and IL-1β], the expression levels of CD68, an M1 cell surface molecule, were also increased in AIMP-1-treated microglial cells, while those of CD206, an M2 cell surface molecule, were not, indicating that AIMP1 triggers the polarization of microglial cells into the M1 state but not the M2 state. AIMP1 treatment induced the phosphorylation of mitogen-activated protein kinases (MAPKs), while MAPK inhibitors suppressed the AIMP1-induced microglial cell activation. AIMP1 also induced the phosphorylation of the nuclear factor-kappa B (NF-κB) components and nuclear translocation of the NF-κB p65 subunit in microglial cells. Furthermore, c-Jun N-terminal kinase (JNK) and p38 inhibitors markedly suppressed the AIMP1-induced phosphorylation of NF-κB components as well as the nuclear translocation of NF-κB p65 subunit, suggesting the involvement of JNK and p38 as upstream regulators of NF-κB in AIMP1-induced microglial cell activation. The NF-κB inhibitor suppressed the AIMP1-induced M1 polarization of the microglial cells. Taken together, AIMP1 effectively induces M1 microglial activation via the JNK and p38/NF-κB-dependent pathways. These results suggest that AIMP1 released under stress conditions may be a pathological factor that induces neuroinflammation., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Oh, Jung, Hong, Cho, Park, Cho and Kim.)

Acne is a chronic skin disease of the pilosebaceous unit resulting from Propionibacterium acnes (P. acnes), a commensal microorganism. Although numerous therapies are available for acne, there is still a need for the development of effective therapies. Erythroid differentiation regulator 1 (Erdr1) has been suggested to be beneficial during inflammatory skin diseases. In the current study, we first showed that Erdr1 expression level was lower in inflammatory acne skin compared to the normal skin, suggesting that Erdr1 was negatively regulated in acne skin. To evaluate the effect of Erdr1 further, Erdr1 was injected subcutaneously in the acne mouse model. Results revealed that the necrotic lesions by inflamed acne were dramatically decreased and collagen synthesis and fibroblasts activation were induced by Erdr1. In addition, Erdr1 reduced the infiltration of inflammatory cells in vivo and accelerated collagen production in P. acnes-treated human dermal fibroblasts through TGF-β/Smad signaling. Taken together, Erdr1 enhanced wound healing through acceleration of collagen synthesis and activation of fibroblasts in acne skin, suggesting its potential use for acne improvement.

Erythroid differentiation regulator 1 (Erdr1) has been identified as a stromal survival factor released under stressful conditions. Previously, Erdr1 was reported to be expressed highly in thymus, but roles of Erdr1 in thymus were not known. Here, the effects of Erdr1 on T cell development were investigated. The expression of Erdr1 was higher in thymus than bone marrow and Erdr1 was detected in both the cortex and medulla of thymus. Erdr1 treatment significantly induced the expression of activation marker, CD69, from thymocytes in the presence of TCR stimuli in vitro and the induction was dependent on increased Ca 2+ influx. In addition, in vivo administration of Erdr1 resulted in significant increase of total and positive selected thymocyte numbers, particularly in the number of CD3TCR hi CD69 + DP thymocytes. Taken together, our results show that Erdr1 enhances the strength of TCR signaling and cellularity of thymocytes by amplifying Ca 2+ influx in thymocytes receiving TCR signals., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Wound healing is an important issue that influences quality of life, and the need for products associated with wound healing is growing annually. New materials and therapies for skin wounds are being continuously researched and developed in order to increase treatment efficacy. Here, we show that the peptide AES16-2M comprised of five short amino acid sequences (REGRT) demonstrates efficacy in wound healing. AES16-2M exerted more effective healing than the control in an acute wound model, and tissue regeneration was similar to that of normal tissue in AES16-2M-treated skin. We found that the increase in re-epithelialization by AES16-2M early in wound development was due to migration of keratinocytes; a scratch assay using a human keratinocyte cell line (HaCaT) also demonstrated effective wound closure by AES16-2M. The migration of keratinocytes effected by AES16-2M was promoted through ERK phosphorylation and blocked with U0126, an ERK inhibitor. Moreover, AES16-2M treatment stimulated human dermal fibroblast (HDF) migration as well as keratinocyte. Taken together, these results suggest that AES16-2M can be an effective therapeutic agent for wound healing by promoting migration of keratinocytes and fibroblasts via ERK phosphorylation.

IL-33 is associated with a variety of autoimmune diseases, such as sclerosis, inflammatory bowel disease, and rheumatoid arthritis. Although IL-33 is mainly involved in the induction of Th2 cells, however, the relationship between IL-33 and Th17 cells is still largely unknown. In this study, we investigated the effects of IL-33 on DC-mediated CD4 + T cell activation and Th17 cell differentiation because DCs are essential cells for presenting self-antigens to CD4 + T cells in autoimmune disease conditions. OT-II mice were injected with IL-33-treated DCs or untreated DCs that were primed by OVA 323-339 peptide, and their Th17 cell responses were compared. Th17 cell population and IL-17 expression levels were significantly increased in draining lymph nodes of mice injected with IL-33-treated DCs, compared with those in mice injected with untreated DCs. IL-33 treatment maturated DCs to present self-antigens and to increase production of proinflammatory cytokines such as IL-1β and IL-6, which have a crucial role in Th17 cell differentiation. We found that the IL-33-matured DCs enhanced the expression of an early T cell activation marker (CD69) and the Th17 master transcription factor (RORγt), but IL-33 did not directly affect CD4 + T cell differentiation or increase Th17 polarization. Notably, neutralizing IL-1β and/or IL-6 significantly decreased IL-17 expression levels and Th17 cell population which were increased by the coculture of CD4 + T cells with IL-33-matured DCs, indicating that IL-33 may induce Th17 cell responses via IL-1β and IL-6 derived from IL-33-matured DCs., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Myeloid-derived suppressor cells (MDSCs) contribute to tumor-mediated immune escape by suppressing antitumor immune responses. Interleukin-33 (IL-33) is capable of regulating various immune cell populations; however, the effects of IL-33 on the differentiation of MDSCs have not been well characterized. In this study, we evaluated the effects of IL-33 on MDSCs and found that IL-33 significantly reduced the differentiation of lineage-negative bone marrow progenitor cells into granulocytic MDSCs (G-MDSCs). IL-33-treated MDSCs exhibited diminished immunosuppressive capacity; reduced inhibition on T-cell proliferation and interferon-γ production, and diminished production of reactive oxygen species. However, IL-33 treatment did not affect the frequency of monocytic MDSCs (M-MDSCs) or their production of nitric oxide and expression of arginase-1. Additionally, compared with control MDSCs, IL-33-treated MDSCs had reduced capacity to induce the differentiation or expansion of Treg cells. Moreover, in vivo IL-33 administration significantly decreased MDSCs and G-MDSCs accumulation in the spleen and tumor microenvironment. Also, despite increasing CD4 + and CD8 + T-cell infiltration, IL-33 administration markedly decreased Treg-cell population in tumor microenvironment. Taken together, our findings indicate that IL-33 reduces the frequency and immunosuppressive activity of G-MDSCs and ultimately the extent of tumor growth.

Environmental air pollution encompasses various particulate matters (PMs). The increased ambient PM from industrialization and urbanization is highly associated with morbidity and mortality worldwide, presenting one of the most severe environmental pollution problems. This article focuses on the correlation between PM and skin diseases, along with related immunological mechanisms. Recent epidemiological studies on the cutaneous impacts of PM showed that PM affects the development and exacerbation of skin diseases. PM induces oxidative stress via production of reactive oxygen species and secretion of pro-inflammatory cytokines such as TNF-α, IL-1α, and IL-8. In addition, the increased production of ROS such as superoxide and hydroxyl radical by PM exposure increases MMPs including MMP-1, MMP-2, and MMP-9, resulting in the degradation of collagen. These processes lead to the increased inflammatory skin diseases and skin aging. In addition, environmental cigarette smoke, which is well known as an oxidizing agent, is closely related with androgenetic alopecia (AGA). Also, ultrafine particles (UFPs) including black carbon and polycyclic aromatic hydrocarbons (PAHs) enhance the incidence of skin cancer. Overall, increased PM levels are highly associated with the development of various skin diseases via the regulation of oxidative stress and inflammatory cytokines. Therefore, anti-oxidant and anti-inflammatory drugs may be useful for treating PM-induced skin diseases., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Myeloid-derived suppressor cells (MDSCs) are one of the most important cell types that contribute to negative regulation of immune responses in the tumor microenvironment. Recently, aminoacyl-tRNA synthetase-interacting multifunctional protein 1 (AIMP1), a novel pleiotropic cytokine, was identified as an antitumor protein that inhibits angiogenesis and induces antitumor responses. However, the effect of AIMP1 on MDSCs in the tumor environment remains unclear. In the present study, we demonstrated that AIMP1 significantly inhibited tumor growth in 4T1 breast cancer-bearing mice and reduced MDSCs population of tumor sites and spleens of tumor-bearing mice. AIMP1 reduced expansion of MDSCs from bone marrow-derived cells in the tumor-conditioned media. AIMP1 also negatively regulated suppressive activities of MDSCs by inhibiting IL-6 and NO production, and Arg-1 expression. Furthermore, treatment of breast cancer-bearing mice with AIMP1 decreased the capacity of MDSCs to suppress T cell proliferation and Treg cell induction. Western blot and inhibition experiments showed that downregulation of MDSCs functions by AIMP1 may result from attenuated activation of STATs, Akt, and ERK. These findings indicate that AIMP1 plays an essential role in negative regulation of suppressive functions of MDSCs. Therefore, it has a significant potential as a therapeutic agent for cancer treatment.

The erythroid differentiation regulator 1 (Erdr1), which is a novel and highly conserved factor, was recently reported to be negatively regulated by IL-18 and to play a crucial role as an antimetastatic factor. IL-18 is a proinflammatory cytokine that functions as an angiogenic mediator in inflammation. Rosacea is a chronic inflammatory skin disorder that is characterized by abnormal inflammation and vascular hyperactivity of the facial skin. To determine whether Erdr1 contributes to the regulation of the chronic inflammatory process in the development of rosacea, an immunohistochemical analysis was performed in healthy donors and patients with rosacea. In this study, we showed that Erdr1 was downregulated, whereas IL-18 was upregulated, in patients with rosacea, which led us to question the role of Erdr1 in this disorder. Moreover, a rosacea-like BALB/c mouse model was used to determine the role of Erdr1 in rosacea in vivo. LL-37 injection induced typical rosacea features, including erythema, telangiectasia and inflammation. Treatment with recombinant Erdr1 (rErdr1) resulted in a significant reduction of erythema, inflammatory cell infiltration (including CD4(+) and CD8(+) T cells), and microvessel density with vascular endothelial growth factor (VEGF). Taken together, our findings suggest that rErdr1 may be involved in attenuating the inflammation and angiogenesis associated with the pathogenesis of rosacea. Thus, these results provide new insight into the mechanism involved in this condition and indicate that rErdr1 could be a potential target for therapeutic intervention of rosacea., (© 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Aims: Recently, immature adipocyte lineage cells have been suggested as a potential hair-growth stimulator. Diverse studies have been attempted to find methods for the preconditioning of immature adipocyte lineage cells. The present study investigates the effect of conditioned medium (CM) from vitamin D3 (Vd3) pre-activated preadipocytes on hair-growth ability., Main Methods: To test the effect of CM from Vd3 pre-activated preadipocytes on hair-growth efficiency in mice, we compared the differences in hair regenerated after injecting CM from mouse preadipocytes pre-activated with or without Vd3. Next, to determine the regulating factors, the VEGF level was measured by ELISA and angiogenesis level was evaluated by IHC. Finally, the signaling mechanism was investigated by inhibitor kinase assay and western blotting., Key Findings: The CM from Vd3 pre-activated preadipocyte injection markedly promoted the ability of hair regeneration in mice. The VEGF levels were increased by Vd3 treatment in vitro and the CM from Vd3 pre-activated preadipocytes significantly increased the angiogenesis in vivo, suggesting the involvement of angiognensis in the hair regeneration induced by CM from pre-activated preadipocytes. In signaling study, Vd3-enhanced VEGF production was reduced by an ERK1/2 inhibitor and the level of ERK1/2 phosphorylation was increased by treatment with Vd3., Significance: This has been the first report on CM from Vd3 pre-activated preadipocyte displaying stimulatory effects on hair growth via the enhancement of angiogenesis in a hairless-induced C57BL/6 mice., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Ceramides, lipid molecules located predominantly within the plasma membrane of a cell, can function as second messengers, and have been known to carry out a number of cellular functions. T helper type 1 (Th1) immune responses are known to be involved in the cellular immunity, which is crucial in the cancer and allergy immunotherapy. This study was designed to evaluate the effects of ceramides on T helper cell responses and their underlying mechanisms. We demonstrated that a cell-permeable C6-ceramide (C6) together with IL-12 enhanced Th1 cell differentiation, whereas C6 alone had no effects, as demonstrated by the increased populations of IFN-γ expressing CD4(+) T cells and the up-regulation of IFN-γ production from CD4(+) T cells. In contrast, C2-ceramide and long chain ceramides (C16 and C24) did not affect the Th1 responses. C6 treatment was shown to increase the expression of T-bet, a master transcription factor of Th1 responses, in a dose-dependent fashion. Furthermore, C6 increased the expression of cyclooxygenase-2 (COX-2) in CD4(+) T cells. The C6-mediated increase of IFN-γ production and IFN-γ expressing CD4(+) T cell populations were significantly suppressed by a COX-2 specific inhibitor (NS-398) in a dose-dependent manner. T-bet expression was also decreased by NS-398 treatment, thereby indicating that C6 ceramide enhances Th1 responses via a COX-2 dependent pathway. This result demonstrates that C6 may be utilized in therapies for the treatment of immune diseases such cancer and allergy by enhancing the Th1 activity., (Copyright © 2011 Elsevier GmbH. All rights reserved.)

Isoflavones are ubiquitous compounds in foods and in the environment in general. Daidzein and genistein, the best known of isoflavones, are structurally similar to 17β-estradiol and known to exert estrogenic effects. They also evidence a broad variety of biological properties, including antioxidant, anti-carcinogenic, anti-atherogenic and anti-osteoporotic activities. Previously, daidzein was reported to increase the phagocytic activity of peritoneal macrophages and splenocyte proliferation, and to inhibit nitric oxide (NO) production in macrophages. However, its potential impacts on immune response in dendritic cells (DCs), antigen-presenting cells that link innate and adaptive immunity, have yet to be clearly elucidated. In this study, we evaluated the effects of isoflavones on the maturation and activation of DCs. Isoflavones (formononetin, daidzein, equol, biochanin A, genistein) were found to differentially affect the expression of CD86, a costimulatory molecule, on lipopolysaccharide (LPS)-stimulated DCs. In particular, daidzein significantly and dose-dependently inhibited the expression levels of maturation-associated cell surface markers including CD40, costimulatory molecules (CD80, CD86), and major histocompatibility complex class II (I-A(b)) molecule on LPS-stimulated DCs. Daidzein also suppressed pro-inflammatory cytokine production such as IL-12p40, IL-6 and TNF-α, whereas it didn't affect IL-10 and IL-1β expression. Furthermore, daidzein enhanced endocytosis and inhibited the allo-stimulatory ability of LPS-stimulated DCs on T cells, indicating that daidzein treatment can inhibit the functional maturation of DCs. These results demonstrate that daidzein may exhibit immunosuppressive activity by inhibiting the maturation and activation of DCs., (Copyright © 2011 Elsevier Inc. All rights reserved.)

IL-32 was recently identified as a proinflammatory cytokine that is induced by IL-18 in natural killer (NK) cells and is highly correlated with inflammatory disorders. However, the relationship between IL-32 and tumor progression is still unknown. In this study, we investigated whether overexpression of IL-32 affects susceptibility of chronic myeloid leukemia (CML) cells to NK cells. Interestingly, IL-32α-overexpressing CML cell lines, K562, Kcl22, and BV173, showed higher NK cell-mediated killing. Flow cytometry analysis revealed that overexpression of IL-32α induced increased expression of Fas and UL16-binding protein 2 (ULBP2) in CML cells. The direct relationship between overexpression of surface molecules by IL-32α and increased NK cell-mediated killing was confirmed by Fas or ULBP2 siRNA transfection. IL-32α-induced Fas and ULBP2 expression are regulated p38 MAPK. In addition, the transcription factor Ets1 plays a key role in ULBP2 specific expression by IL-32α overexpression in ULBP family members. Taken together, these data show that IL-32α stimulates Fas and ULBP2 expression via activation of p38 MAPK, which increases NK susceptibility of CML cells. Enhanced NK cell susceptibility of CML cells by IL-32α overexpression may improve the efficiency of NK cell-based immunotherapy.

Intratumoral hypoxia has been correlated with distant metastatic potential. Two hypoxia inducible factors (HIFs), HIF-1α and HIF-2α, are induced by hypoxia, and high expression of these proteins has been correlated to angiogenesis and distant metastasis. Thymosin β4 (Tβ4) is frequently highly expressed in cancer, and this overexpression correlates with malignant progression. The objective of this study was to investigate the clinical correlation of HIF-α with Tβ4 and the intracellular functional roles of Tβ4 on HIF-α activation. We examined HIF-1α, HIF-2α and Tβ4 expressions in clinical human breast carcinoma (n=70) by immunohistochemistry. We show that high expression of HIF-1α and HIF-2α strongly correlates with Tβ4 expression (P≤0.0001) and overexpression of Tβ4 correlates significantly with patients with lymph node metastasis (P<0.05) of human breast cancer. Additionally, we demonstrate that hypoxia up-regulates intracellular Tβ4 protein, which then affects HIF-α activity, which is the key in regulating VEGF expression. We confirmed that hypoxia-induced intracellular Tβ4 and HIF-α activities were reduced by interference of Tβ4 expression using Tβ4 shRNA lentivirus. Vascular epidermal growth factor (VEGF)-A, a well-recognized lymphangiogenic cytokine, was also down-regulated, but VEGF-C and VEGF-D expressions were not affected. These findings suggest that the overexpression of Tβ4 is strongly associated with HIF-1α and HIF-2α expression and is also clinicopathologically involved with lymph node metastatic potential of breast cancer through the modulation of HIF-αactivation and induction of VEGF-A. Ultimately, these results highlight Tβ4 as a potentially therapeutic target in malignant cancers.

This study describes the immunotherapeutic properties of vaccines that encode tumor-associated calcium signal transducer-1 (Trop-1), a newly identified breast cancer antigen, in mice with breast cancer. Previously we found that Trop-1 was over-expressed in cellular breast cancer vaccines that were highly enriched for cells that induced therapeutic CTL-mediated immune responses in mice with breast cancer, as compared with non-enriched vaccines. In this study, to determine if the expression of Trop-1 by cells in the enriched vaccine was responsible for its therapeutic benefits, an expression plasmid that specified the Trop-1 gene was transfected into the LM fibroblast cells, which was then used as a vaccine. To augment their immunogenic properties, the fibroblasts were genetically modified before Trop-1 DNA-transfer to secrete IL-2 and to express allogeneic MHC class I H-2K(b)-determinants. Mice with established breast cancer treated solely by immunization with fibroblasts modified to express Trop-1 developed CD8(+) cell-mediated immunity to the breast cancer cells. The immunity was sufficient to prolong the survival of mice with established breast cancer. In some instances, the immunity was sufficient to result in rejection of the tumor; the mice remained tumor free more than 60 days., (Copyright © 2010 Elsevier Ltd. All rights reserved.)

In a previous study, we reported that a gene mutation of repeat in toxin E (RtxE), a transporter of cytotoxic factors, resulted in a significant impairment of epithelial cell cytotoxicity in Vibrio vulnificus, and that the expression of the rtxE gene was induced by the exposure to the host cells. In this study, we evaluated and compared the effects of co-culture supernatants from V. vulnificus-infected INT-407 cells and either the V. vulnificus wild-type or rtxE mutant on the production of interleukin (IL)-8, a pro-inflammatory cytokine, as well as its underlying mechanisms in human intestinal epithelial cells. INT-407 cells were co-cultured with the wild-type V. vulnificus or the rtxE mutant strain to obtain the conditioned supernatants. IL-8 production and nuclear factor (NF)-κB activation from the INT-407 cells treated with each supernatant, were investigated. The co-culture supernatants from the rtxE mutant V. vulnificus-infected INT-407 cells significantly induced lower levels of IL-8 production and promoter activation, NF-κB DNA binding activity, and NF-κB minimal promoter activation in human intestinal epithelial cells, than those from the wild-type V. vulnificus-infected INT-407 cells. Importantly, the reduced IL-8 production and NF-κB activity of the V. vulnificus rtxE mutant, were restored by co-culture supernatants from the rtxE-complemented V. vulnificus. On the whole, these results show that the rtxE gene of V. vulnificus performs a critical role in the secretion of factors from bacteria and host cells, which are involved in IL-8 production via the NF-κB activation pathway in host cells.

Rationale: Probiotics have been shown to be effective in reducing allergic symptoms. However, there are few studies to evaluate the therapeutic effects of lactobacilli on allergen-induced airway inflammation., Objective: We investigated whether three heat-killed lactobacilli, Lactobacillus plantarum, Lactobacillus curvatus and Lactobacillus sakei subsp. sakei, isolated from kimchi, exerted inhibitory effects on airway hyper-responsiveness in a murine asthma model., Methods: Heat-killed lactic acid bacteria were orally administered into BALB/c mice, followed by challenge with aerosolized ovalbumin, after which allergic symptoms were evaluated., Results: Airway inflammation was suppressed in the L. plantarum- and L. curvatus-treated mice. Interleukin (IL)-4 and IL-5 levels were significantly lower in the L. plantarum- and L. curvatus-treated mice than in those treated with L. sakei subsp. sakei. Importantly, heat-killed L. plantarum administration induced Foxp3 expression in intestinal lamina propria cells, and heat-killed L. curvatus induced IL-10 as a way of inducing tolerance., Conclusion: Specific strains of lactobacilli isolated from kimchi can effectively suppress airway hyper-responsiveness.

Cancer cells metastasize to the other site after escaping from the immune system and CD70, CD44 and vascular endothelial growth factor (VEGF) play important roles in this process. It is recently reported that interleukin (IL)-18 is closely related with the pathogenesis of skin tumor. Therefore, we investigated the role of endogenous IL-18 from stomach cancer on the immune escape mechanism and metastasis via the regulation of CD70, CD44 and VEGF expression. IL-18 and IL-18R expressions were not only investigated on tumor tissues (n = 10), and sera (n = 20) from stomach cancer patients, but also on human stomach cancer cell lines. IL-18 and IL-18R expressions were found on stomach cancer cell lines and tumor tissues. In addition, IL-18 levels were elevated in sera from cancer patients (P < 0.05), compared with sera from normal individuals. Changes in CD70, CD44 and VEGF expression by flow cytometry, immunoblotting and enzyme-linked immunosorbent assay and immune susceptibility by (51)Cr-release assay were investigated, after silencing or neutralization of endogenous IL-18. CD70 expression was increased and it increases immune susceptibility of cancer cells. In contrast, CD44 and VEGF expression was decreased and it suppresses neovascularization and the metastasis of stomach cancer. After inoculation of IL-18 small interfering RNA (siRNA)-transfected stomach cancer cells into Balb/C (nu/nu) mice, regression of tumor mass was determined by measuring of tumor size. And the number and location of metastatic lesions were investigated by hematoxylin and eosin staining. The regression of tumor mass and the suppression of metastasis were observed in the mice, which are injected with IL-18 siRNA-transfected cell lines. Our data suggest that endogenous IL-18 might facilitate stomach cancer cell immune escape by suppressing CD70 and increasing metastatic ability by upregulating CD44 and VEGF.

Cytokines are a protein family of regulatory factors derived from tumors and their environmental components that contribute to the growth, invasion and metastasis of breast cancer. However, the way in which tumor progression and cytokines regulate each other is not well understood. In this study, we used an oligoDNA microarray to assess the kinetic expression profile of cytokine genes in tumor tissues and lymph nodes during the progression of tumor growth in mice that had been subcutaneously challenged with breast adenocarcinoma SB5b cells. Our results demonstrated that IL-15, IL-17, IL-18 and IL-18 binding protein (IL-18bp), which are associated with inflammation, were increased in tumor tissues. Conversely, chemokines and their receptors, including CXCR4/CXCL12, CCR7/CCL21, CCL9, CXCL9 and CCL12, were overexpressed in lymph nodes during tumor growth. Furthermore, RT-PCR and Western blot analysis revealed that IL-18, a pro-angiogenic factor in tumors, was up-regulated in tumor tissues. Interestingly, CCR3, IL-1R2, SOCS and IL-20 were up-regulated in tumor tissues, but down-regulated in lymph nodes during tumor growth. This result suggests that the expression of cytokines and cytokine-related genes was differentially regulated, which resulted in a beneficial effect for tumor progression.

Background: Recently, many plastic surgeons have been using adipogenic-differentiated cell implantation for remodeling scars in patients. However, this technique is not a long-term solution because implanted cells disappear gradually. Therefore, we investigated a method to increase the grafted cell preservation rate by using an effective adjuvant, botulinum toxin., Methods: The adipogenic-differentiated cells were subcutaneously injected in the dorsal area of C57/BL6 mice with or without botulinum toxin. Two and six weeks later we analyzed the residual volume and confirmed the characteristics of the implanted cells by real-time RT-PCR and immunohistochemistry., Results: Two and six weeks after transplantation we found that the residual volume of the transplantation site was higher in the botulinum toxin-treated group than in the untreated group. We also confirmed that the residual transplanted area has characteristics of adipogenic tissue by histological analysis. Next, to determine the mechanism related to the enhanced preservation rate of grafted cells via treatment with botulinum toxin, we performed immunohistochemical staining for the angiogenesis-related marker CD31. We found that CD31 expression was higher in the botulinum toxin-treated group than in the untreated group., Conclusion: We have shown that in vivo grafted adipocyte cell preservation can be enhanced by treatment with botulinum toxin as an adjuvant. We suggest that botulinum toxin further increases this graft preservation rate by enhancing angiogenesis.

To investigate the expression levels of S100A4 in human colorectal carcinoma (CC) and its relationship with clinicopathological parameters and metastatic potential, 73 pathological specimens from patients with CC were examined for S100A4 expression by RT-PCR and immunohistochemistry. An increase of S100A4 mRNA was observed in 19/23 (82.6%) CC specimens, and S100A4 was up-regulated in 40/73 (54.7%) CC cases compared with non-neoplastic mucosal tissues. Upregulation of S100A4 was significantly related to invasion, nodal status, distant metastasis and p53 expression. Next, we investigated whether S100A4 could affect p53 transactivation and stability. Interestingly, it was revealed that treatment with exogenous S100A4 protein reduced transcriptional activity of p53 and abrogated the modification of calcium binding affinity of S100A4 protein. These findings suggested that S100A4 might be involved in the progression and metastasis of human CC, presumably via modulation of the wild-type p53 protein.

Although there is known to be a marked concentration of reactive microglia in the substantia nigra pars compacta (SNpc) of patients with Parkinson's disease (PD), a disorder in which alpha-synuclein plays a key pathogenic role, the specific roles of alpha-synuclein and microglia remains poorly understood. In this study, we investigated the effects of alpha-synuclein and the mechanisms of invasive microglial migration into the SNpc. We show that alpha-synuclein up-regulates the expressions of the cell adhesion molecule CD44 and the cell surface protease membrane-type 1 matrix metalloproteinase through the extracellular regulated kinases 1/2 pathway. These concurrent inductions increased the generation of soluble CD44 to liberate microglia from the surrounding extracellular matrix for migration. The effects of alpha-synuclein were identical in BV-2 murine microglial cells subjected to cDNA transfection and extracellular treatment. These inductions in primary microglial cultures of C57Bl/6 mice were identical to those in BV-2 cells. alpha-Synuclein-induced microglial migration into the SNpc was confirmed in vivo using a 6-hydroxydopamine mouse model of PD. Our data demonstrate a correlation between alpha-synuclein-induced phenotypic changes and microglial migration. With the recruitment of the microglial population into the SNpc during dopaminergic neurodegeneration, alpha-synuclein may play a role in accelerating the pathogenesis of PD.

IL-18 production may enhance immune system defense against KG-1 cells ; NB4 cells, which are associated with good prognosis, do not produce IL-18. In this study, we treated KG-1 cells with IL-18 and used microarray technology to assess subsequent effects on gene expression. In UniGene-array of 7488 human genes, expression of 57 genes, including stress related genes, increased at least 2-fold, whereas expression of 48 genes decreased at least 2-fold. Following exogenous exposure of KG-1 cells to IL-18, expression of CRYGC, NF(kappa)BIA and NACA gene were monitored. The latter is a transcriptional coactivator potentiating c-Jun-mediated transcription. NF(kappa)BIA is an inhibitor of NF(kappa)B, and affects growth regulation, apoptosis and hypoxic stress. Studies, such as this one, are beginning to clarify the differences between cells associated with good and bad cancer prognoses, which may ultimately assist in medical treatment for acute myeloid leukemia.

CD4+CD25+ regulatory T cells (Treg cells) are negative regulator of the immune system and main obstacles to cancer immunotherapy in tumor-bearing hosts. Resveratrol is a natural product found in grapes with both immunomodulatory and anticancer effects, which can be controlled by Treg cells. Therefore, to determine whether resveratrol performs these actions via Treg cells, we investigated changes in Treg cell population and immunomodulatory cytokines in EG7 tumor-bearing C57BL/6 mice. In the present study, CD4+CD25+ cell population among CD4+ cells was inhibited ex vivo by resveratrol treatment in a dose-dependent manner. FoxP3+ expressing cells among CD4+CD25+ population were significantly reduced after resveratrol treatment ex vivo in intracellular FACS analysis. Single intraperitoneal administration of 4 mg/kg resveratrol suppressed the CD4+CD25+ cell population among CD4+ cells and downregulated secretion of TGF-beta, an immunosuppressive cytokine, measured from the spleens of tumor-bearing mice. Furthermore, resveratrol enhanced IFN-gamma expression in CD8+ T cells both ex vivo and in vivo,leading to immune stimulation. Taken together, these results suggest that resveratrol has a suppressive role on CD4+CD25+ cell population and makes peritumoral microenvironment unfavorable to tumor in tumor-bearing mice. Thus, resveratrol can be considered as possible adjuvant material for vaccination-based cancer therapy.

Interleukin-18 (IL-18) was discovered as an interferon-gamma-inducing factor and had a critical role in inflammatory and immune response. It stimulates natural killer (NK) and T cells and enhances Th1 immune response. These activated immune cells eliminate cancer cells and virus-infected cells effectively. However, IL-18 has also been found to promote tumor progression. Higher expression or secretion level of IL-18 is detected in various cancer cells in comparison with normal control, and IL-18 is able to induce angiogenesis, migration/metastasis, proliferation and immune escape. These dual effects and the mechanism of IL-18 need to be investigated further as it relates to cancer.

It is well known that UVB (290-320 nm) induces inflammation in skin by the transcription and release of cytokines and chemokines from skin keratinocytes. In addition, it is considered that intracellular reactive oxygen species (ROS) plays an important role in UVB-induced inflammatory response in the skin. Therefore, we investigated the effect of vitamin C, a potent antioxidant, on the regulation of UVB-induced skin inflammation via the modulation of chemokines production. Vitamin C uptake into keratinocytes is increased by UVB irradiation in a time- and dose-dependent manner through the translocation of sodium-dependent vitamin C transporter-1 (SVCT-1), a vitamin C-specific transporter, from the cytosol to the membrane. To evaluate the effect of vitamin C on the chemokine mRNA expression, we performed RNase protection assay. As a result, there was a remarkable change in chemokine mRNA expression, especially IL-8 and monocyte chemoattractant protein (MCP)-1 expression. In addition, increased IL-8 and MCP-1 mRNA expressions were suppressed by vitamin C treatment. We also confirmed the results of protein levels measured by ELISA. Taken together, vitamin C uptake is increased in UVB-irradiated keratinocytes through the translocation of SVCT-1 and regulates inflammatory response in the skin via the downregulation of IL-8 and MCP-1 production.

The IgJ gene is expressed in the plasma cell stage. However, its neighboring charged amino acid-rich leucine zipper 1 (Crlz1) gene, which is mapped 30 kb upstream of the IgJ gene in mice, is shown to be expressed in the pre-B cell stage. These stage-specific expressions of two neighboring genes are found to be regulated by their chromatin accessibility and acetylation. Hypersensitive site 1 on the IgJ promoter is opened in the plasma cells, whereas hypersensitive sites 9/10 on the Crlz1 promoter are opened in the pre-B cells. Furthermore, H3 and H4 histones toward the chromatin of the Crlz1 gene are found to be hyperacetylated, especially on H3, in the pre-B cells, whereas those toward the chromatin of the IgJ gene are found to be hyperacetylated in the plasma cells. Consistently, the hyperacetylation of H3 and H4 toward the chromatin of the IgJ gene but not the Crlz1 gene is induced by an IL-2 treatment of BCL1, which is a model cell line for studying the terminal differentiation of B cells.

Pharmacological inhibition of interleukin-12 (IL-12) production may be a therapeutic strategy for preventing the development and progression of disease in experimental models of autoimmunity. In this study, the effects of myricetin, a naturally occurring flavonoid present in fruits, vegetables and medicinal herbs, on the production of IL-12 were investigated in mouse macrophages stimulated with lipopolysaccharide (LPS). Myricetin significantly inhibited the LPS-induced IL-12 production from both primary macrophages and the RAW264.7 monocytic cell-line in a dose-dependent manner. The effect of myricetin on IL-12 gene promoter activation was analyzed by transfecting RAW264.7 cells with IL-12 gene promoter/luciferase constructs. The repressive effect was mapped to a region in the IL-12 gene promoter containing a binding site for NF-kappaB. Furthermore, activation of macrophages by LPS resulted in markedly enhanced binding activity to the NF-kappaB site, which significantly decreased upon addition of myricetin, indicating that myricetin inhibited IL-12 production in LPS-activated macrophages via the down-regulation of NF-kappaB binding activity.

Pharmacological inhibition of interleukin-12 (IL-12) production may be a therapeutic strategy for preventing development and progression of disease in experimental models of autoimmunity. In this study we investigated the effects of baicalein, a flavonoid present in the root of Scutellaria baicalensis, on the production of IL-12 from mouse macrophages stimulated with lipopolysaccharide (LPS). Baicalein potently inhibited the LPS-induced IL-12 production from both primary macrophages and RAW264.7 monocytic cell-line in a dose-dependent manner (the IC50 values were 43.7 and 17.4 microM, respectively). The effect of baicalein on IL-12 gene promoter activation was analyzed by transfecting RAW264.7 cells with IL-12 gene promoter/luciferase constructs. The repressive effect mapped to a region in the IL-12 gene promoter containing a binding site for NF-kappaB. Furthermore, activation of macrophages by LPS resulted in markedly enhanced binding activity to the NF-kappaB site, which significantly decreased upon addition of baicalein, indicating that baicalein inhibited IL-12 production in LPS-activated macrophages via inhibition of NF-kappaB binding activity.