• Journal of Microbiology
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• 제39권3호
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• pp.186-194
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• 2001

Scrub typhus, caused by Orientia tsutsugamushi infection, is clinically and histopathologically characterized by local as well as systemic inflammatory reactions, indicating that orientiae induce mechanisms that amplify the inflammatory response. To reveal underlying mechanisms of chemoattraction and activation of responding leukocytes, expression of chemokine and tumor necrosis factor alpha (TNF-$\alpha$) genes in murine peritoneal macrophages after infection with the obligate intracellular bacterium Ο.tsutsugamushi was investigated. The genes that were unregulated included macrophage inflammatory proteins l$\alpha$/$\beta$(MIP-l$\alpha$/$\beta$), MIP-2, monocyte chemoattractant protein 1(MCP-1), RANTES (regulated upon activation, normal T-cell expressed and secreted), gamma-interferon-inducible protein 10(IP-10) and TNF-$\alpha$. Peak expression of these chemokines and TNF-$\alpha$ was observed between 1 and 3 h after infection. These responses returned to or approached baseline preinfection levels 6 h after challenge. Semiquantitative reverse transcription (RT)-PCR analysis revealed dramatic Increases during infection in the steady-state levels of mRNA ceding for the inhibitory subunit of NF-kB (IkB$\alpha$), whose transcription is enhanced by binding of NF-kB within the IkB$\alpha$promoter region. Thus, Ο. tsutsugamushi appears to be a stung inducer of chemokines and TNF-$\alpha$ which may significantly contribute to inflammation and tissue damage observed in scrub typhus by attracting and activating phagocytic leukocytes.

• Bulletin of the Korean Chemical Society
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• 제33권9호
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• pp.2878-2882
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• 2012

Amentoflavone is naturally occurring bioflavonoid that is found in a number of plants. In this paper, the anti-inflammatory activity of amentoflavone in LPS-stimulated macrophages and its mode of action were examined. Using LPS-stimulated RAW264.7 macrophage cells, we found that amentoflavone exerted anti-inflammatory activities through inhibition of nitric oxide (NO) production and tumor necrosis factor (TNF)-${\alpha}$ and macrophage inflammatory protein (MIP)-2 secretion. Amentoflavone (1.0-20 ${\mu}M$) gradually inhibited nitrite production without cytotoxicity. Amentoflavone (1.0 and 10 ${\mu}M$) effectively suppressed both TNF-${\alpha}$ and MIP-2 cytokine release from LPS-stimulated RAW264.7 cells. The expression of mIL-$1{\beta}$ and mMIP-2 cytokine mRNAs was completely inhibited while expression of mMIP-1 was effectively suppressed and mTNF-${\alpha}$ expression was slightly inhibited by 10 ${\mu}M$ amentoflavone. We also demonstrated that the innate immune response to amentoflavone involves the toll-like receptor (TLR) and mitogen-activated protein kinase (MAPK) pathways. LPS-induced upregulation of p38 MAPK phosphorylation was significantly reduced by 10 ${\mu}M$ amentoflavone. These results suggest that amentoflavone exhibits effective anti-inflammatory activities through regulation of TLR4 and phosphorylation of p38 MAPKs.

• Tuberculosis and Respiratory Diseases
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• 제62권4호
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• pp.299-307
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• 2007

배경: HMGB1은 염증반응의 후기에 분비되는 중요한 염증유발물질 중 하나이다. 본 연구에서는 기존에 염증유발물질로 잘 알려진 LPS와 새롭게 염증유발물질로 관심을 받고 있는 HMGB1의 염증유발작용을 생체 외 및 생체 내 실험을 통해 비교하고자 하였다. 또한 HMGB1의 자극에 의한 IL-8 promoter region의 활성화에 중요한 역할을 수행하는 전사인자들을 확인하고자 하였다. 방법: RAW264.7 세포에 LPS(100 ng/ml) 또는 HMGB1(500 ng/ml)을 투여하고 각각 0, 2, 4, 8, 12 그리고 24시간 뒤에 세포상층액의 $TNF-{\alpha}$, MIP-2 그리고 $IL-1{\beta}$의 농도를 ELISA법으로 측정하였다. 생쥐의 복강에 LPS(5 mg/kg) 또는 HMGB1(2.5 mg/kg)을 주입하여 급성폐손상을 유발한 후에 폐의 사이토카인의 발현과 MPO 활성도를 측정하였다(LPS는 4시간 뒤, HMGB1은 24 시간 뒤). IL-8 promoter 부위에 있는 NF-IL6, $NF-{\kappa}B$ 그리고 AP-1에 대한 결합부위에 대해 돌연변이를 일으킨 후에 각각의 돌연변이체를 pIL-6luc에 결합시킨 뒤 RAW264.7 세포에 삽입하였다. 이 세포들을 36시간 배양한 후에 HMGB1(500 ng/ml)으로 자극하고, 한 시간 뒤에 세포를 녹인 후 luciferase 활성도를 측정하였다. 결과: LPS 투여 후에 RAW264.7 세포 배양상층액의 $TNF-{\alpha}$농도는 24시간 뒤에, MIP-2 농도는 8시간 뒤에 최고치를 보였다. 한편 HMGB1 투여 후에는 $TNF-{\alpha}$와 MIP-2 농도 모두 24시간 뒤에 최고치를 나타내었다. LPS 복강 내 투여 후 4시간 뒤에 생쥐의 폐의 $TNF-{\alpha}$, MIP-2 그리고 $IL-1{\beta}$의 농도는 대조군에 비해 현저히 증가하였으나, HMGB1 복강 내 투여 후 24시간 뒤에 생쥐의 폐에서는 $IL-1{\beta}$의 농도만 약간 증가하였다. MPO 활성도는 LPS와 HMGB1 투여 후에 모두 증가하였으며, LPS 투여 후가 더 의미있게 증가하였다. $NF-{\kappa}B$ 돌연변이체와 AP-1 돌연변이체에서 luciferase 활성도가 의미있게 감소하였다. 결론: 이상의 결과를 살펴볼 때 HMGB1은 염증유발효과는 LPS에 비해 강도가 떨어지나 지속시간은 오래 계속되는 것으로 보이며, HMGB1에 의한 IL-8의 활성화에 $NF-{\kappa}B$ 뿐만 아니라 AP-1도 중요한 역할을 수행하는 것으로 판단된다.

• 대한한방내과학회지
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• 제25권4호
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• pp.289-299
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• 2004

Background : Although the pathophysiology of asthma has been reported, its mechanism has not been fully elucidated. The mast cell is an effector cells in allergic inflammation and secretes a number of chemokines. Chemokines are important for the recruitment of leukocytes to sites of infection, which is essential in host defense. Chemokines also contribute to the pathogenesis of several disorders such as asthma, chronic bronchitis, atopic dermatitis, allergic rhinitis, and rheumatoid arthritis. Objective : In this study, the aim was to identify the effect of Baekryunchihyo-tang(白蓮治哮湯) on expression of chemokines. This was examined by RT-PCR using the human mast cell line (HMC-l) Materials and Methods : HMC-l cells were used, which is known to secrete and express chemokines. In order to investigate the protective effect of Baekryunchihyo-tang(白蓮治哮湯), HMC-l cells were incubated with pretreatment of Baekryunchihyo-tang(白蓮治哮湯) for 24 hrs. RT-PCR analyses of chemokine genes of cells pretreated with Baekryunchihyo-tang(白蓮治哮湯) showed that expressions of IL-8, $MIP-l{\beta}$, and RANTES genes in these cells were lower and $MIP-l{\alpha}$ showed a similar pattern compared to the calcium ionophore-treated group. In addition, cell cytotoxicity concentration measurements were performed by MTT assay method. Results : After stimulation with 1 uM calcium ionophore A23178 for 2 hrs, IL-8, major one of CXC chemokines, was highly expressed, and expression of $MIP-l{\beta}$ and RANTES (CC chemokines) increased, while expression of $MIP-l{\alpha}$ did not change. The cell cytotoxicity of Baekryunchihyo-tang(白蓮治哮湯) with treatments at various concentrations and times was not observed, respectively. Conclusion : This study suggests that Baekryunchihyo-tang(白蓮治哮湯) has dose-dependent effects on mRNA expression of IL-8(CXC chemokines), $MIP-l{\beta}$ and RANTES(CC chemokines) in human mast cellline(HMC-l). So these herbal medicines may inhibit the inflammatory process of asthma. Advanced studies are required to investigate the mechanism of inhibition by herbal medicine in the asthma model. This study provides basic data on the possibility of the clinical treatment of Baekryunchihyo-tang(白蓮治哮湯) for allergic disorders.

• Journal of Yeungnam Medical Science
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• 제20권2호
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• pp.187-196
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• 2003

The cytokines are the hormone-like proteins, which are produced in the mononuclear cells. They have many roles, such as immune mediators, cell differentiations, angiogenesis. The chemokines have chemotactic effects which control the host immune response. There were few reports about the cytokines associated with musculoskeletal tumors. From late 1980s, the cytokine studies of bone tumors such as osteosarcoma were started, but most studies for benign and malignant musculoskeletal tumors were left to be explored. To evaluate the characteristics of the cytokines in variable musculoskeletal tumors, tissues were obtained from the seven patients who visited the Yeungnam University hospital from February to July 2000. They were lipoma (1 case), parosteal osteoma (1 case), enchondroma (2 cases), pigmented villonodular synovitis (1 case), ganglion (1 case), and metastaic squamous cell carcinoma (1 case). The gene experession of the cytokines were analyzed by RNase protection assay (RPA) and reverse transcription-polymerase chain reaction (RT-PCR). The lipoma and parosteal osteoma expressed MIP-$1{\beta}$, and IP-10 genes. The two enchondromas showed different results, one expressed all of MIP-$1{\alpha}$, MIP-$1{\beta}$ and IP-10 genes but the other expressed none of above. The pigmented villonodular synovitis strongly expressed MIP-$1{\alpha}$ and IP-10 when compared with the other cases. The ganglion did not express all of the chemokines mentioned above. And the metastatic squamous cell carcinoma expressed all of the chemokines and especially IP-10 was highly expressed. Even though this study has only a few cases, these results provide a basis for the cytokine mediating network study in musculoskeletal tumors.

• Journal of Microbiology
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• 제41권3호
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• pp.266-270
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• 2003

Orientia tsutsugamushi, an obligate intracellular bacterium, is the causative agent of scrub typhus which is histopathologically characterized by inflammatory manifestations. To understand the pathogenesis of scrub typhus, chemokine gene expression in mice after infection with O. tsutsugamushi was investigated. The mRNAs that were upregulated included macrophage inflammatory proteins 1${\alpha}$/${\beta}$ (MIP-1${\alpha}$/${\beta}$), MIP-2, monocyte chemoattractant protein 1, RANTES (regulated upon activation, normal T-cell expressed and secreted), and gamma-interferon-inducible protein 10. Peak expression of these chemokines was observed six days after infection. These responses returned to or approached baseline preinfection levels by eight days after infection. Chemokine profiles in infected mice were well correlated with the kinetics of inflammatory cell infiltration. Thus, O. tsutsugamushi appears to be a strong inducer of chemokines which may significantly contribute to the inflammation observed in scrub typhus by attracting and activating phagocytic leukocytes.

• Journal of Periodontal and Implant Science
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• 제27권2호
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• pp.425-441
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• 1997

The neuropeptide substance P(SP) has been implicated in the mediation of inflammation and immune-mediated disease such as arthritis. Recently, it was reported that SP was markedly increased around the blood vessels in inflamed gingiva as well as in close association with the inflammatory cell infiltrate. These results support that SP may contribute to the pathophysiology of neuronal inflammation in human periodontal tissues. SP may regulate inflammatory/immune responses by stimulating the proliferation of human T cells, differentiation and antibody-secreting potential of B cells, macrophage respiratory burst, connective tissue proliferation, and the secretion of cytokines from monocytes and T cells. Here, I studied potential role of SP as a costimulatory chemical signal in inflammatory/immune responses, by determining the released proinflammatory cytokines such as $MIP-1{\alpha}$, $IL-1{\beta}$, and IL-6 from culture supernatants of homogeneous immune cell lines. Serum free cell supernatants were concentrated with TCA precipitation, fractionated with SDS-PAGE, and subjected into western blot analysis. Among 15 cell lines tested, macrophage/monocyte cell line RAW264.7 and WRl9m.1 showed the highest level of induction of $MIP-1{\alpha}$ when stimulated with LPS. Discrete IL-6 bands with multiple forms of molecular mass were detected from supernatants of B cell lines A20(32kDa), Daudi(32, 35kDa), and SKW6.4(29kDa), which were expressed constitutively. $IL-1{\beta}$ could not be detected by the method of western blot analysis from supernatants of all cell lines tested except RAW264.7, WRl9m.1, and erythroid cell line K562 which showed the least amount of $IL-{\beta}$ secretion. SP $10^{-9}M$ with suboptimal dose of LPS treatment showed synergistic induction of $MIP-1{\alpha}$ release from RAW264.7 or WR19m.1, and also IL-6 release from A20, but this synergism is not the case in costimulation of RAW264.7 or WRl9m.1 with SP $10^{-9}M$ and TPA. Although treatment of T cell line CTLL-R8 with SP $10^{-7}M$ or PHA+TPA induced modest level of $MIP-1{\alpha}$ secretion, synergism was not observed when they are applied together. These findings all together suggest the possibility of a regulatory role of SP in inflammatory/immune reaction through differential modulation of bioactivities of other chemical cosignals.

• IMMUNE NETWORK
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• 제2권1호
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• pp.53-59
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• 2002

Background: Clinical observations and laboratory studies have supported an immune basis for most acquired aplastic anemias, with the majority of patients responding to immunosuppressive therapy. Fas, a member of the tumor necrosis factor (TNF) receptor superfamily is a critical downregulator of cellular immune responses. Proinflammatory cytokines like interferon gamma (IFN-${\gamma}$) and TNF-${\alpha}$ can induce Fas expression and render hematopoietic progenitor cells susceptible to Fas-induced growth suppression and apoptosis. Methods: In order to investigate the involvement of apoptosis in the pathogenesis of aplastic anemia (AA), we measured the expression of Fas antigen and caspase-3 on bone marrow (BM) mononuclear cells (MNCs) of AA in the presence or absence of IFN-${\gamma}$, TNF-${\alpha}$, or macrophage inflammatory protein 1-${\alpha}$ (MIP-$1{\alpha}$). Results: We confirmed that AA BM MNCs were more apoptotic and highly expressed Fas antigen than normal donors. Stimulation by IFN-${\gamma}$, TNF-${\alpha}$, or MIP-$1{\alpha}$ increased Fas antigen and caspase-3 expression in AA BM MNCs than BM MNCs of normal donors. Anti-Fas monoclonal antibody enhanced IFN-${\gamma}$, TNF-${\alpha}$, or MIP$1{\alpha}$ mediated caspase-3 expression in BM MNCs of normal donors. Among these three cytokines, IFN-${\gamma}$ enhanced apoptosis most strongly via Fas-caspase-3 pathway. Conclusion: These results suggest that Fas signal pathway may play a role in the pathophysiology of aplastic anemia and negative hematopoietic regulators like IFN-${\gamma}$ can induce apoptosis of bone marrow progenitors in part by Fas induction.

• Journal of Microbiology and Biotechnology
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• 제14권4호
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• pp.722-729
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• 2004

Leptin is an adipocyte-secreted hormone and its plasma levels correlate with total body fat mass, however, it also plays a regulatory role in immunity, inflammation, and hematopoiesis. Chemokine is known as a chemoattractant cytokine in inflammatory reaction, but its role in leptin reaction has not been well studied. In this study, the direct effect of leptin on the expression of chemokine mRNAs and lipopolysaccharide (LPS)-induced chemokine KC mRNA in mouse peritoneal macrophages was investigated. Leptin did not induce the expression of lymphotactin, RANTES, eotaxin, MIP-1$\beta$, MIP-1$\alpha$, MIP-2, MCP-1, IP-10, TCA-3, and KC mRNA in mouse peritoneal macrophages, and had no direct effect on the expression of these LPS-induced chemokine mRNAs except KC mRNA. The synergistic effect of leptin on the expression of LPS-induced KC mRNA occurred late in the time course of response to LPS. The increased expressions of Ob-Rb mRNA and leptin receptor protein were detected during the LPS treatment. Leptin produced a substantial increase in the stability of the LPS-induced KC mRNA, and the synergistic effect of leptin on LPS-induced KC mRNA expression was further augmented by cycloheximide (CHX). Pyrrolidine dithiocarbamate (PDTC) did not block the synergistic effect of leptin on LPS-induced KC mRNA expression in mouse peritoneal macrophages. These data suggest that although leptin has no direct effect on the expression of lymphotactin, RANTES, eotaxin, MIP-1$\beta$, MIP-1$\alpha$, MIP-2, MCP-1, IP-10, TCA-3, and KC mRNA in mouse peritoneal macrophages, the synergistic effect of leptin on the expression of LPS-induced KC mRNA has the possibility that LPS might induce the expression of the Ob-Rb receptor or an unknown gene(s) that sensitizes macrophages to the synergistic function of leptin. Therefore, further studies are necessary to examine leptin as a regulatory factor of chemokine production.

• Asian Pacific Journal of Cancer Prevention
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• 제15권2호
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• pp.929-935
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• 2014

Two types of nanosized titanium dioxide, anatase ($anTiO_2$) and rutile ($rnTiO_2$), are widely used in industry, commercial products and biosystems. $TiO_2$ has been evaluated as a Group 2B carcinogen. Previous reports indicated that $anTiO_2$ is less toxic than $rnTiO_2$, however, under ultraviolet irradiation $anTiO_2$ is more toxic than $rnTiO_2$ in vitro because of differences in their crystal structures. In the present study, we compared the in vivo and in vitro toxic effects induced by $anTiO_2$ and $rnTiO_2$. Female SD rats were treated with $500{\mu}g/ml$ of $anTiO_2$ or $rnTiO_2$ suspensions by intra-pulmonary spraying 8 times over a two week period. In the lung, treatment with $anTiO_2$ or $rnTiO_2$ increased alveolar macrophage numbers and levels of 8-hydroxydeoxyguanosine (8-OHdG); these increases tended to be lower in the $anTiO_2$ treated group compared to the $rnTiO_2$ treated group. Expression of $MIP1{\alpha}$ mRNA and protein in lung tissues treated with $anTiO_2$ and $rnTiO_2$ was also significantly up-regulated, with $MIP1{\alpha}$ mRNA and protein expression significantly lower in the $anTiO_2$ group than in the $rnTiO_2$ group. In cell culture of primary alveolar macrophages (PAM) treated with $anTiO_2$ and $rnTiO_2$, expression of $MIP1{\alpha}$ mRNA in the PAM and protein in the culture media was significantly higher than in control cultures. Similarly to the in vivo results, $MIP1{\alpha}$ mRNA and protein expression was significantly lower in the $anTiO_2$ treated cultures compared to the $rnTiO_2$ treated cultures. Furthermore, conditioned cell culture media from PAM cultures treated with $anTiO_2$ had less effect on A549 cell proliferation compared to conditioned media from cultures treated with $rnTiO_2$. However, no significant difference was found in the toxicological effects on cell viability of ultra violet irradiated $anTiO_2$ and $rnTiO_2$. In conclusion, our results indicate that $anTiO_2$ is less potent in induction of alveolar macrophage infiltration, 8-OHdG and $MIP1{\alpha}$ expression in the lung, and growth stimulation of A549 cells in vitro than $rnTiO_2$.