Human Neutrophil의 수명연장과 Superoxide 생산에 관여하는 미지의 Monocyte 생성물질

허억,배진우 大韓免疫學會 1993 大韓免疫學會誌 Vol.15 No.-

It has long been known that neutrophils are quickly infilterated and recruited to infected sites and then kill invaders by phagocytic action. Unfortunatly it is not yet revealed which molecule or cytokine is involved in the phagocytic action and viability sustaining activity of neutrophils. The aim of this study was whether lipopolysaccharide (LPS)-stimulated monocyte may control those neutrophil actions. Human peripheral blood monocytes and neutrophils were isolated by Ficollpaque density sedimentation from heparin anti-coagulated blood of healthy adult donors. After preparation of these cells, the purity of both was more than 90%. Monocytes stimulated in various dose(0.1-10pug/ml) of LPS for various times of incubation(0-3 days). and then LPS-stimulated monocyte conditioned medium was collected in order to find an optimal dose and incubation time for the neutrophil viability. It was found out that 3,ug/ml of LPS in 24 hours incubation was maximal effective condition for the activity of neutrophil sustaining viability. Monocyte conditioned medium (MCM) under this condition was used for the comparison with LPS-nonstimulated monocyte conditioned medium or enriched medium alone. When neutrophils were stimulated with each medium for 1-3 days, the activity of neutrophil sustaining viability with MCM was significantly higher than the activity with other medium (in 1 day of culture, 72-1:8 vs 4311:7 vs 17 ±10; p <O. 01). The superoxide production of neutrophil stimulated with MCM for 24 hours incubation was significantly higher than that with other medium under fMLP doses of 0.1-100,uM (p <0.01). Under fMLP l,uM, the superoxide production is predominantly different between them(23.8±2 vs 10.3±3 vs 7.8±1.6). The maximal effective dose of GM-CSF(granulocyte/macrophage colony stimulating factor ; 10pM) enhanced the neutrophil viability in 1 day of culture (50 ± 4%) . In the study to assess whether MCM contains GM-CSF, anti-GM-CSF antibody slightly blocked the MCM-dependent neutrophil viability(73±9 vs 50±12; p <0.07), indicating that MCM might not contain GM-CSF. These data indicate that LPS-stimulated monocyte surely product a factor for the neutrophil sustaining viability and the enhancement of superoxide production, suggesting that a factor is not GMCSF. If more than one factor were producted form LPS-stimulated monocyte, one minor factor might be GM-CSF.

5-Lipoxygenase in monocytes emerges as a therapeutic target for intimal hyperplasia in a murine wire-injured femoral artery

Baek, S.E.,Jang, M.A.,Lee, S.J.,Park, S.Y.,Bae, S.S.,Kim, C.D. Elsevier/North Holland [etc.] 2017 Biochimica et biophysica acta Vol.1863 No.9

Given the importance of leukotrienes in vascular inflammation induced by local tissue injury, this study investigated the role for 5-lipoxygenase (5-LO) in monocytes in the development of intimal hyperplasia. As a mechanistic study, the importance of monocyte 5-LO in monocyte-macrophage differentiation with subsequent infiltration in neointima was evaluated. In a mouse model of wire-injured femoral artery, intimal hyperplasia started as early as 2wks after injury, and luminal area and blood flow were reduced due to increased neointima formation. Time-dependent increases in macrophage infiltration were observed in neointima and showed a positive relationship with neointima volume. In 5-LO-deficient (KO) mice or wild-type (WT) mice treated with an inhibitor of 5-LO activating protein (MK886, 1 and 10mg/kg), intimal hyperplasia and macrophage infiltration into neointima were reduced, but monocyte adhesion to injured luminal surface was not inhibited, which suggested 5-LO participates in monocyte-macrophage differentiation. In an in vitro study, monocyte-macrophage differentiation was found to be increased by high mobility group box 1 protein (HMGB1), but this effect was attenuated in cells isolated from 5-LO-KO mice. Furthermore, macrophage infiltration and intimal hyperplasia were more prominent in 5-LO-KO mice transplanted with monocytes from WT mice than in 5-LO-KO mice transplanted with monocytes from 5-LO-KO mice. Taken together, it was suggested that 5-LO in monocytes played a pivotal role in monocyte-macrophage differentiation and subsequent infiltration of macrophage in neointima, leading to vascular remodeling after vascular injury.

5-Lipoxygenase plays a pivotal role in endothelial adhesion of monocytes via an increased expression of Mac-1

Lee, Seung Jin,Choi, Eun Kyoung,Seo, Kyo Won,Bae, Jin Ung,Kim, Yun Hak,Park, So Youn,Oh, Sae Ock,Kim, Chi Dae Oxford University Press 2013 Cardiovascular research Vol.99 No.4

<P><B>Aims</B></P><P>5-Lipoxygenase (5-LO) is known to participate in the pathogenesis of atherosclerosis; however, the underlying mechanisms are unclear. Thus, this study investigated the molecular mechanisms responsible for 5-LO expression in monocytes as well as the role of 5-LO in monocyte adhesion to the vascular endothelium, which is a key early event in macrophage foam cell formation.</P><P><B>Methods and results</B></P><P>An <I>en face</I> immunohistochemistry of endothelial surfaces revealed a marked increase in monocyte adhesion to the aortic endothelium in wild-type (WT) mice treated with lipopolysaccharide (LPS), which was significantly attenuated in 5-LO<SUP>(−/−)</SUP> mice. Likewise, the adhesion capacity of primary monocytes isolated from LPS-treated WT mice was higher than those of monocytes from 5-LO<SUP>(−/−)</SUP> mice. In <I>in vitro</I> study, LPS increased monocyte adhesion to endothelial cells with an enhanced Mac-1 expression. These were attenuated by a 5-LO inhibitor, MK886, as well as by molecular depletion of 5-LO in monocytes. Furthermore, LPS-induced Mac-1 expression on monocytes was significantly inhibited by pre-treatment with U-75302, a BLT<SUB>1</SUB>-receptor antagonist, suggesting a pivotal role of 5-LO-derived leukotrienes. In promoter activity analysis and chromatin immunoprecipitation assays to identify transcription factors involved in 5-LO expression, both NF-κB and Sp1 played central roles to increase 5-LO expression in LPS-treated monocytes.</P><P><B>Conclusion</B></P><P>5-LO expression in monocytes is modulated via NF-κB and Sp1 signalling pathways, and 5-LO plays a pivotal role in LPS-mediated monocyte adhesion to the vascular endothelium through an increased expression of Mac-1 on monocytes.</P>

한국산 겨우살이 추출물 M11C(렉틴 구성물질)가단구세포의 TNF-α 유전자 발현유도 및 분비에 비치는 효과

허억 한국약용작물학회 2007 韓國藥用作物學會誌 Vol.15 No.1

It is well-known that Korean mistletoe (Viscum album) extract has an immune activity and anticancer effect. In this study, Korean mistletoe extract, M11C (non-lectin components), was used to examine whether this extract might activate human peripheral monocyte to produce tumor necrosis factor-α (TNF-α). To examine the effect of M11C on the production of TNF-α from monocyte, the monocyte were stimulated by the M11C, and then collected the supernatant (M11C stimulated monocyte-conditioned media; MCM). MCM was treated to the TNF-α sensitive L929 cells, and then L929 cytotoxicity was measured by means of MTT. MCM had cytotoxic effect on L929. And the cytotoxic effect of MCM on L929 was almost abolished by anti-TNF-α antibody. These data indicated that MCM contained TNF-α, suggesting the TNF-α generation from M11C-stimulated monocyte. This suggestion was confirmed from the data that TNF-α was highly detected in MCM by immunoblotting technique. M11C effect on TNF-α production from monocyte was in the dose and stimulating time dependent manners. Also the effect of M11C on the expression of TNF-α mRNA from monocyte was shown in the dose and stimulating time dependent manners. As a result, Korean mistletoe extract, M11C, could be used for an immunostimulator.

MCP-1에 의해 유도된 THP-1 유주에 미치는 Zerumbone의 영향

김사현,김시현,유성률,이평재,문철,Kim, Sa Hyun,Kim, Si Hyun,Ryu, Sung Ryul,Lee, Pyeongjae,Moon, Cheol 대한임상검사과학회 2018 대한임상검사과학회지(KJCLS) Vol.50 No.2

본 연구는 zerumbone이 단구의 유주에 어떠한 영향을 미치는지 알아보고자 진행되었다. 단구는 다양한 염증 질환의 중요한 매개자로 인식되고 있으며, 활성, 유주 등 단구의 기능 조절을 통해 염증 질환을 조절하는 가능성이 보고 되고 있다. 염증 발생 시 증가하는 케모카인인 MCP-1에 의해 단핵구 세포주 THP-1의 유주가 유발되는 것을 확인하였다. 10 ng/mL의 농도에서 유주가 발생하였으며, 100 ng/mL과 200 ng/mL의 농도에서 가장 높은 유주 현상이 나타났다. MCP-1에 의해 유발된 THP-1 유주는 zerumbone 존재 시 50% 이상 감소하였다. MCP-1 수용체인 CCR2 신호전달 과정의 중요 2차 전달자인 cAMP의 배양액 내 농도는 zerumbone 단독 처리 시 세포 단독 배양 조건에 비해 증가하였으며, MCP-1 단독 처리 시에는 의미있게 감소하였다. 그러나, zerumbone과 MCP-1을 동시에 처리했을 때에는 다시 cAMP의 증가가 관찰되었다. MCP-1 처리에 의해 일어나는 Erk 인산화도 zerumbone과 동시 처리 시 감소하는 결과를 확인했다. 본 연구는 염증성 질환에 중요한 매개자로 인식되고 있는 단구의 유주 현상을 조절하는 zerumbone의 가능성을 보여준다. This study examined the effects of zerumbone on monocyte migration. Monocytes are recognized as important mediators of various inflammatory diseases, and the possibility of controlling inflammatory diseases by regulating the monocyte functions, such as activity and mobility, has been reported. MCP-1, which is a chemokine with levels that increase upon inflammation, causes the migration of the monocyte cell line, THP-1. Migration occurred at a concentration of 10 ng/mL MCP-1, and the highest migration occurred at 100 ng/mL and 200 ng/mL. MCP-1-induced THP-1 migration decreased by more than 40% in the presence of zerumbone. The concentration of cAMP, an important secondary messenger of the CCR2 signaling pathway, the MCP-1 receptor, was increased in the culture medium after a zerumbone treatment. The concentrations of cAMP decreased significantly under the MCP-1 treatment condition only. On the other hand, an increase in cAMP was observed when zerumbone and MCP-1 were treated simultaneously. Erk phosphorylation induced by an MCP-1 treatment was also found to decrease with the zerumbone treatment. This study introduces the possibility of controlling inflammatory diseases through the function of zerumbone, which regulates the migration of monocytes.

Ulmus davidiana ethanol extract inhibits monocyte adhesion to tumor necrosis factor-alpha-stimulated endothelial cells

이기모,주희경,이유란,박명수,강건,최성아,전병화 한국한의학연구원 2016 Integrative Medicine Research Vol.5 No.2

Background Ulmus davidiana var. japonica Rehder (UD) has long been used in traditional folk medicine in Asia. This study is designed to investigate the antiadhesive activity of the ethanol extract of UD (UDE) and its underlying mechanisms in cultured endothelial cells. Methods The dried root bark of UD was extracted with 80% (v/v) ethanol. The antiadhesive activity of the UDE was investigated in cultured human umbilical vein endothelial cells and human embryonic kidney epithelial 293T (HEK 293T) cells stably transfected with pGL3-vascular cell adhesion molecule (VCAM)-1-luc. Monocyte adhesion in endothelial cells was induced by tumor necrosis factor-alpha (TNF-α), and the protective effects of UDE on monocyte–endothelial cell adhesion, VCAM-1 expression, reactive oxygen species production, and nuclear factor-κB activity were determined. Results Exposure to UDE at a concentration of 3–30 μg/mL for 24 hours produced no detectable cytotoxicity in human umbilical vein endothelial cells, but it significantly inhibited TNF-α-induced monocyte adhesion and VCAM-1 expression. TNF-α treatment of HEK 293T/VCAM-1-luc cells resulted in increased luciferase activity of the VCAM-1 promoter, which was inhibited by treatment with UDE. Additionally, TNF-α-induced reactive oxygen species generation, nuclear translocation of nuclear factor-κB, and IκBα degradation in human umbilical vein endothelial cells were effectively reduced by treatment with 30 μg/mL of UDE. Conclusion Our results indicated that UDE treatment inhibited TNF-α-induced monocyte adhesion in endothelial cells, suggesting that UD may reduce vascular endothelial inflammation.

자궁내막증 환자의 복강액으로 처리한 단핵구가 T-세포 반응에 미치는 영향에 관한 연구

이규섭 부산대학교 병원 암연구소 2006 부산대병원학술지 Vol.- No.20

Purpose: This study examined whether or not the PF from endometriosis patients (EPF)-treated monocytes affect the T cell response. Subject and Methods: Peritoneal fluid was obtained from endometriosis patient(n=20) and healthy control group(n=10) respectively. Peripheral blood monocytes were obtained from healthy donors by density-gradient centrifgation and THP-l cells derived from a monocyte/macrophage cell line. We examined the effects of heat-treatment and IL-10 neutralizing antibody on the down-regulation of MHC class IT expression level by EPF. The monocytes pre-treated with 10% of EPF mixture were mixed with T-cells, cultured with or without Staphylococcus enterotoxin B(SEB), and then pulsed with 1μCi of [^(5)H] thymidine. The radioactivity incorporated was determined. EPF-treated or CPF-treated monocytes were mixed with T-cells and cultured. The expression of MHC class Ⅱ was evaluated by confocal microscopic analysis, and the levels of IL-10, TGF-β and VEGF were measured by standard enzyme-linked immunosorbent assay kits(R and D system). Results: Heat-pretreatment of EPF and neutralizing IL-10 antibody abrogated MHC class Ⅱ down-regulation on monocytes, which had been induced by EPF. However, in a functional assay, monocytes treated with EPF failed to stimulate T cell in mixed leucocyte reaction, although T cell proliferation was increased with EPF-treated monocytes and SEB. Conclusion: These results suggest that the cell stimulatory ability of monocytes dose not coincide with MHC class Ⅱ expression level.

실리콘에 의한 말초혈액 단핵구의 아세포 형성능 및 단핵세포의 interleukin-1β와 tumor necrosis factor-α의 분비효과

한만욱,장익수,안상태 大韓成形外科學會 1998 Archives of Plastic Surgery Vol.25 No.7

Although there are numerous reviews of clinical and epidemiologic data, there has been no critical analysis of silicone immunology. The purpose of this study is to identify human celluar immune reaction to silicone through mononuclear cell blastogenesis as well as by measuring IL-1βand TNF-α released from human monocyte/macrophage incubated with silicone in vitro. In the study, total 14 healthy volunteers participated in the experiment as blood donors. in the peripheral blood mononuclear cell blastogenesis assay, one control group and three experimental groups were designed. The three experimental groups were composed of a silicone treated group, a silicone/phytohemagglutinin treated group, and a phytohemagglutinin treated group. The peripheral blood mononuclear cells were isolated with the Ficoll-Hypaque gradient method, and they were incubated for 72 hours. The proliferation of the peripheral blood mononuclear cells was measured with the [³H]-thymidme uptake. In the cytokine assay of monocyte stimulated by silicone, human monocyte was isolated from the peripheral blood mononuclear cells through the magnetic cell sorting(MACS) method. One control group and three experimental groups were designed also in the experiment. The experimental groups were composed of a silicone treated group, a silicone/lipopolysaccharide treated group, and a lipopolysaccharide treated group. The monocytes of each group were incubated for 1,3,6,24 and 48 hours. The supernatants were preserved at -20℃ to measure IL-1βand TNF-α with ELISA. Statistical analysis was done by two-way ANOVA and Scheffe test(p < 0.05). In the peripheral blood mononuclear cell blastogenesis study, no statistical difference was found either between the silicon treated group and the control group, or between the silicone/phytohemagglutinin treated group and the phytohemagglutinin treated group. The silicone treated group had higher IL-1βlevel than the control group 1 hour and 3 horus(p < 0.05), and the silicone/LPS treated group and had higher IL-1βlevel than the LPS treated group at 1 hour(p < 0.05). However the TNF-αlevel had shown statistical difference neither between the silicone treated group and control group nor between the silicone/LPS treated group and the LPS treated group at any time. In conclusion, IL-1β was released from the human monocyte/macrophage induced by silicione, and silicone did not induce peripheral blood mononuclear cell blastogenesis. It has been known that IL-1βcan induce the chemotaxis of T lymphocyte-subpopulation and T cell infiltration in tissue. Therefore the results of this experiments are regarded as an evidence of silicone-induced inflammation and tissue destruction such as rheumatic disease.

P038 Elucidation of the expression and clinical significance of tripartite motif-containing proteins in Behcet`s disease

( Yuri Ahn ),( Zhenlong Zheng ),( Yeongjoo Oh ),( Hemin Lee ),( Dongsik Bang ),( Do Young Kim ) 대한피부과학회 2016 대한피부과학회 학술발표대회집 Vol.68 No.2

<div style="display:none">fiogf49gjkf0d</div><div style="display:none">fiogf49gjkf0d</div><div style="display:none">fiogf49gjkf0d</div><div style="display:none">fiogf49gjkf0d</div><div style="display:none">fiogf49gjkf0d</div><div style="display:none">fiogf49gjkf0d</div><div style="display:none">fiogf49gjkf0d</div> Background: Behcet’s disease (BD) is a chronic, multisystem vasculitis and autoinflammatory disorder. Many study suggest that BD is triggered by environmental factor including infectious agents. TRIM21, E3 ligase protein, regulates the production of cytokine by ubiquitination of transcriptional factors. TRIM21 can present antiviral properties Objectives: To elucidate TRIM21 proteins expression in monocyte of BD patient, and to identify its role on immune dysregulation in BD. Methods: Monocytes and T cells were isolated from peripheral blood. Protein levels were measured using western blot, ELISA, and flow cytometry. Co-cultures between monocyte and responder T cells were maintained in media for 7~14 days. Knock-down of TRIM21 was performed using siRNA technique Results: TRIM21 expression was increased in peripheral blood monocytes from BD patients. The expression of IRF8, a representative ubiquitination target of TRIM21, was also decreased. BD monocyte promoted secretion of pro- inflammatory cytokines including Th17 promoting cytokine following NF-kB activation. BD monocytes promoted Th17 polarization after co-culture with responder T cells from healthy donors, and IL-17A production from these T cells was also increased. Knock-down of TRIM21 in monocyte prevents NF-kB activation and decreased Th1,Th17 polarization of responder T cell Conclusion: BD monocytes facilitate Th/1Th17 differentiation of naive T cells and TRIM21 may regulate the secretion of pro-inflammatory cytokine from monocytes.

Inhibitory Effect of Simvastatin on the TNF-α- and Angiotensin II-Induced Monocyte..

Su-Young Park,Jong-Suk Lee,Yu Jin Ko,Ah Ra Kim,Mi Kyoung Choi,Mi-Kyoung Kwak,최한곤,용철순,김정애 대한약학회 2008 Archives of Pharmacal Research Vol.31 No.2

Vascular endothelial cell activation by cytokines and other pro-inflammatory mediators is an initial event in atherosclerosis and in other vascular diseases. Simvastatin, a HMG-CoA reductase inhibitor, suppressed both tumor necrosis factor (TNF)-α- and angiotensin (Ang) IIinduced monocyte adhesion to endothelial cells (an initial step in vascular inflammation) and reactive oxygen species (ROS) production. Diphenyleneiodonium and apocynin, both NADPH oxidase inhibitors, also suppressed TNF-α-induced ROS and monocyte-endothelial cell adhesion, demonstrating that TNF-α-induced monocyte adhesion is mediated through ROS produced by NADPH oxidase activation. Furthermore, exogenously applied mevalonate or geranylgeranylpyrophosphate in combination with simvastatin completely prevented the inhibitory effects of simvastatin on ROS generation and monocyte-endothelial cell adhesion by TNF- α and Ang II. These results suggest that monocyte adhesion to endothelial cells induced by TNF-α or Ang II is mediated via the geranylgeranyl isoprenoid-dependent generation of ROS, and that this is inhibited by simvastatin.