• Title/Summary/Keyword: cytokine ELISA

Search Result 398, Processing Time 0.028 seconds

Inhibition of Lipopolysaccharide-stimulated Inflammatory Cytokine Production by LY303511 in Human Macrophagic THP-1 Cells

  • Kim, So-Hee;Paek, Yun-Woong;Kang, In-Chol
    • International Journal of Oral Biology
    • /
    • v.42 no.3
    • /
    • pp.117-121
    • /
    • 2017
  • We have previously shown that the specific phosphatidylinositol 3-kinase inhibitor LY294002 (LY29), and its inactive analog LY303511 (LY30), inhibit a monocyte chemoattractant protein-1 (MCP-1) expression in human umbilical vein endothelial cells; these results suggest the potential of LY30 as an anti-inflammatory drug. In this study, we determined the effects of LY30 on the production of various inflammatory cytokines in human macrophagic THP-1 cells which were stimulated with lipopolysaccharide (LPS). LY30 selectively suppressed the mRNA expression of IL-12 p40, $TNF-{\alpha}$, and MCP-1 without affecting the expression of $IL-1{\alpha}$, IL-6, and IL-8. Inhibition of the production of IL-12 and $TNF-{\alpha}$ by LY30 was also demonstrated using ELISA assays. In order to elucidate the mechanisms of the action of LY30, we examined the role played by the mitogen-activated protein kinases and the key transcription factors, AP-1 and $NF-{\kappa}B$ in LPS-stimulated THP-1 cells. The results revealed that LY30 inhibited LPS-induced activation of ERK, but not p38 or JNK. Furthermore, the AP-1 DNA binding activity was suppressed by LY30 based upon the dosage, whereas $NF-{\kappa}B$ DNA binding was not affected. These results suggest that LY30 selectively inhibits cytokine production in the LPS-stimulated macrophagic THP-1 cells by down-regulating the activation of ERK and AP-1.

Nuclear Factor-${\kappa}B$ Dependent Induction of TNF-${\alpha}$ and IL-$1{\beta}$ by the Aggregatibacter actinomycetemcomitans Lipopolysaccharide in RAW 264.7 Cells

  • Na, Hee Sam;Jeong, So Yeon;Park, Mi Hee;Kim, Seyeon;Chung, Jin
    • International Journal of Oral Biology
    • /
    • v.39 no.1
    • /
    • pp.15-22
    • /
    • 2014
  • Aggregatibacter actinomycetemcomitans is an important pathogen in the development of localized aggressive periodontitis. Lipopolysaccharide (LPS) is a virulent factor of periodontal pathogens that contributes to alveolar bone loss and connective tissue degradation in periodontal disease. Our present study was designed to investigate the cytokine expression and signaling pathways regulated by A. actinomycetemcomitans LPS (Aa LPS). Cytokine gene expression profiling in RAW 264.7 cells was performed by microarray analyses. The cytokine mRNA and protein levels and related signaling pathways induced by Aa LPS were measured by RT-PCR, ELISA and western blotting. Microarray results showed that Aa LPS strongly induced the expression of NF-${\kappa}B$, NF-${\kappa}B$-related genes, inflammatory cytokines, TNF-${\alpha}$ and IL-$1{\beta}$ in RAW 264.7 cells. NF-${\kappa}B$ inhibitor pretreatment significantly reduced the levels of TNF-${\alpha}$ and IL-$1{\beta}$ mRNA and protein. In addition, the Aa LPS-induced TNF-${\alpha}$ and IL-$1{\beta}$ expression was inhibited by p38/JNK MAP kinase inhibitor pretreatment. These results show that Aa LPS stimulates TNF-${\alpha}$ and IL-$1{\beta}$ expression through NF-${\kappa}B$ and p38/JNK activation in RAW 264.7 cells, suggesting the essential role of this pathway in the pathogenesis of localized aggressive periodontitis.

Mechanism of Differential Ag-specific Immune Induction by Different Tumor Cell Lysate Pulsed DC (종양 세포 용해액에 따른 수지상세포 유도 항원 특이 면역반응 차이의 기전 연구)

  • Lee, Kang-Eun;Shon, Hye-Jin;Kim, Myung-Joo;Baek, So-Young;Lee, Hyun-Ah
    • IMMUNE NETWORK
    • /
    • v.6 no.3
    • /
    • pp.145-153
    • /
    • 2006
  • Background: Tumor cell lysate has been considered as a preferential antigen source for the therapeutic dendritic cell pulsing. Our experiences with in vivo study with animal tumor model indicate the tumor cell lysate dependent differential effect of DC therapy. Our previous data show that MC38 lysate pulsed-DC induced stronger ag-specific immunity than CT26 lysate pulsed-DC in vitro. In this study we tried to reveal the mechanism for differential induction of ag-specific immunity of different colon cancer cell lysate pulsed-DCs. Methods: MC38 and CT26 cell lines were prepared as lysate by freezing-thawing procedure. Tumor cell antigenicity was confirmed by detecting the surface expression of MHC I/II & B7.1/2 molecules. IL-10, IL-12 and TGF-beta in the tumor cell lysate were detected by ELISA and the presence of heat shock proteins were analysed by western blotting. Results: The secretion of IL-10, a immune-inhibitory cytokine was about 470% higher in CT26 lysate than in MC38. Hsp 70 was detected only in the MC38 lysate but not in the CT26. On the other hand, Hsp 60 and 90 expression were not different in two colon cancer cell lysates. Conclusion: In two different colon cancer cell lysate, immune inhibitory IL-10 (higher in CT26) and Hsp70 (MC38 superiority) were differentially expressed. These data indicate that higher agspecific immunity induction by MC38 lysate pulsed-DC may due to the expression of hsp70 and lower secretion of IL-10, a immune-inhibitory cytokine than CT26 lysate. The significance of other cytokine and the surface marker expression will be discussed.

Xylitol Mitigate Neutrophil Inflammatory Response Against Porphyromonas gingivalis Infection

  • Na, Hee Sam;Song, YuRi;Choi, Yoon Hee;Chung, Jin
    • International Journal of Oral Biology
    • /
    • v.43 no.3
    • /
    • pp.141-146
    • /
    • 2018
  • Periodontitis is generally a chronic disorder characterized by breakdown of tooth-supporting tissues, producing dentition loss. Porphyromonas gingivalis (P. gingivalis), a Gramnegative anaerobic rod, is one of the major pathogens associated with periodontitis. Neutrophils are first line defense cells in the oral cavity that play a significant role in inflammatory response. Xylitol is a known anti-caries agent and has anti-inflammatory effects. In this study, we conducted experiments to evaluate anti-inflammatory effects of xylitol on P. gingivalis infected neutrophils for possible usage in prevention and treatment of periodontal infections. P. gingivalis was intraperitoneally injected and peritoneal lavage was collected for cytokine determination. For in vitro study, neutrophils were collected from mouse peritoneal cells after zymosan injection or bone marrow cells. Neutrophils were stimulated with live P. gingivalis and ELISA was used to determine the effect of xylitol on P. gingivalis induced cytokine production. $IL-1{\beta}$, IL-6, $TNF-{\alpha}$ concentration and neutrophil population in the peritoneal lavage was increased in P. gingivalis-infected mouse. Peritoneal cells infected with live P. gingivalis revealed significantly increased production of $IL-1{\beta}$, IL-6 and $TNF-{\alpha}$ at multiplicity of infection of 10. Neutrophils from bone marrow and peritoneal lavage revealed increased production of $IL-1{\beta}$, IL-6 and $TNF-{\alpha}$. Xylitol significantly mitigated P. gingivalis induced cytokine production in neutrophils. Findings indicate that xylitol is an anti-inflammatory agent in neutrophils infected with live P. gingivalis, that suggests its use in periodontitis management.

2-Nonadecanone Alleviates Depression through Inflammation Relief in SD Rat

  • Lee, Gil-Hyun;Hyun, Kyung-Yae
    • Biomedical Science Letters
    • /
    • v.24 no.3
    • /
    • pp.206-212
    • /
    • 2018
  • Depression is a type of mood disorder characterized by hypochondriasis, decreased appetite, and insomnia. Depression is a disease that affects more than 100 million people worldwide. 2-Nonadecanone (NAC) is a bioactive substance that constitutes Fomes fomentarius, and NAC is expected to have an antidepressant effect. By using the forced swimming test (FST), we investigated the effects of treatment with NAC on immobility subacutely in rats after oral dosing once a day for 2 days. Serum levels of cytokine interleukin-1 beta (IL-1beta) and tumor necrosis factor-alpha (TNF-alpha) were determined by enzyme-linked immunosorbent assay (ELISA). Nuclear factor-2 (Nrf-2) and inducible nitric oxide synthases (iNOS) were analyzed by western blot method. NAC dose-dependently decreased immobility in the FST. NAC dosedependently decreased FST-induced increase of cytokine levels, as manifested by significantly stronger effects on $IL-1{\beta}$ and $TNF-{\alpha}$ levels at higher doses than the lowest dose of NAC. Western blot analysis showed that Nrf-2 was significantly lower in the NAC-treated group than in the disease-induced group. The iNOS results were also significantly lower in the NAC-treated group than in the other groups. Considering FST results, the antidepressant effect of NAC is effective. Considering the results of cytokine and protein expression, this anti-depressant effect may be related to the anti-inflammatory effect. Therefore, it can be said that the anti-inflammatory effect of NAC increases the antidepressant effect in the FST experiment.

Inhibitory Effects of Scrophulariae Radix on β-hexosaminidase release and cytokine production in RBL-2H3 cells (현삼(玄蔘) 추출물이 RBL-2H3 비만세포에서 β-hexosaminidase 및 cytokine 분비에 미치는 효과)

  • Kim, Se-Gie
    • The Korea Journal of Herbology
    • /
    • v.32 no.6
    • /
    • pp.9-15
    • /
    • 2017
  • Objectives : Traditional medicines isolated from natural products often have positive effects in the prevention and healing of various immune disorders, such as allergy and atopic inflammation. Scrophulariae Radix (SR) been used in oriental medicine used for treatment of acute and chronic inflammatory diseases. Mast cells are known to play important roles in the initiation of allergic reactions. In this study, we investigated the effects of SR ethanol extract on inflammatory responses in IgE-stimulated RBL-2H3 mast cells. Methods : Rat basophilic leukemia RBL-2H3 cells were purchased from Korean Cell Line Bank (KCLB No. 22256). Cell viability was measured by MTT assay. Assays for ${\beta}-Hexosaminidase$ Secretion : RBL-2H3 cells were sensitized with dinitrophenyl-ImmunoglobulinE (DNP IgE). The next antigen DNP-BSA ($25ng/m{\ell}$) was added for 10 minutes and the reaction was terminated after 5 minutes in the ice bath. To determine ${\beta}-Hexosaminidase$ release, supernatants were aliquoted into 96-well plates. Samples were mixed with substrate solution and incubated for 1 h at $37^{\circ}C$. Absorbance was measured with a spectrophotometer at 405 nm. IL-4 and tumor necrosis $factor-{\alpha}$($TNF-{\alpha}$) concentrations in cell culture supernatants were measured using enzyme-linked immunosorbent assay (ELISA) kits. Results : The cytotoxicity of SRE in RBL-2H3 cells was less than 5%. SRE inhibited DNP-IgE-imduced degranulation of mast cells in RBL-2H3 cells. Also significantly decreased the levels of inflammatory cytokine, IL-4 and TNF-alpha. In this study, the SRE showed potential anti-allergic and antiinflammatory. Conclusions : These results indicate that SRE could be inhibit the allergic response through suppressing the mast cell activation.

Saponin attenuates diesel exhaust particle (DEP)-induced MUC5AC expression and pro-inflammatory cytokine upregulation via TLR4/TRIF/NF-𝛋B signaling pathway in airway epithelium and ovalbumin (OVA)-sensitized mice

  • Jo, Sooyeon;Na, Hyung Gyun;Choi, Yoon Seok;Bae, Chang Hoon;Song, Si-Youn;Kim, Yong-Dae
    • Journal of Ginseng Research
    • /
    • v.46 no.6
    • /
    • pp.801-808
    • /
    • 2022
  • Background: Diesel exhaust particle (DEP) is a harmful kind of particulate matter known to exacerbate pre-existing respiratory diseases. Although their adverse effects on airway pathologies have been widely studied, the mechanistic analysis of signaling pathways and potential targets in reducing DEP-induced mucin secretion and pro-inflammatory cytokine production remain elusive. We, for the first time, investigated the effects of Korean Red Ginseng (KRG) extracts on mucin overproduction and airway inflammation induced by DEP. Methods: The effects of KRG and saponin on DEP-induced expression of MUC5AC and interleukin (IL)-6/8 were examined by real-time polymerase chain reaction (PCR) and enzyme-linked immunosorbent assay (ELISA) in human airway epithelial NCI-H292 cells. We conducted Western blotting analysis to analyze the associated signaling pathways. To evaluate the effects of saponin treatment on DEP-induced MUC5AC expression and inflammatory cell infiltrations in ovalbumin (OVA)-sensitized mice, immunohistochemical (IHC) staining and real-time PCR were implemented. Results: The KRG extracts markedly attenuated DEP-induced MUC5AC expression in vitro by inhibiting the TLR4/TRIF/NF-𝛋B pathway. Furthermore, KRG and saponin inhibited DEP-induced pro-inflammatory cytokine IL-6/8 production. The in vivo study revealed that saponin blocked DEP-induced inflammation, mucin production and MUC5AC expression. Conclusion: Our study revealed that KRG extracts have inhibitory effects on DEP-induced expression of MUC5AC and the production of pro-inflammatory cytokines. This finding provides novel insights into the mechanism by which saponin alleviates diesel-susceptible airway inflammation, elucidating its potential as a phytotherapeutic agent for inflammatory pathologies of airway.

The Effect of Sonicated Extracts of Treponema Denticola and Treponema Lecithinolyticum on the Cytokine Secretion and Matrix Metalloproteinase Activation of Gingival Fibroblast (Treponema denticola와 Treponema lecithinolyticum의 분쇄액이 치은섬유아세포의 Cytokine 분비 및 Matrix metalloproteinase 활성에 미치는 영향)

  • Suh, Hye-Yuhn;Choi, Bong-Kyu;Choi, Seong-Ho;Cho, Kyoo-Sung;Kim, Chong-Kwan;Chai, Jung-Kiu
    • Journal of Periodontal and Implant Science
    • /
    • v.29 no.4
    • /
    • pp.979-995
    • /
    • 1999
  • This study was investigated to observe the effect of Treponema denticola cell sonicates(TDC) and Treponema lecithinolyticum cell sonicates(TLC) on cytokine secretion and matix metalloproteinase-2(MMP-2) activation of cultured human gingival fibroblast. Several experiments were performed including $IL-1{\beta}$, IL-6 ELISA for the effect on the $IL-1{\beta}$, IL-6 secretion of human gingival fibroblast. Also gelatinase zymography and gelatin dissolubility test for the activation of MMP-2 secreted by gingival fibroblast. The results were as follows. 1. The effect of TDC and TLC on IL-6 secretion of human gingival fibroblast showed statistically significant increase of IL-6 secretion in the TDC and TLC treated group compared to no treatment group(p<0.05) . 2. The amount of $IL-1{\beta}$ secretion was below the lower limit and there was no difference in the $IL-1{\beta}$ secretion of gingival fibroblast between TDC, TLC treated group and no treatment group. 3. The active form of pro MMP-2 with 72 kDa molecular weight was activated in both TDC and TLC treated group and clear band was appeared at 62kDa site on the zymography. 4. Gelatin dissolubility of MMP-2 secreted by gingival fibroblast was higher in TDC and TLC treated group compared to no treatment group(p<0.05). 5. In the TDC treated group, serine protease of T. denticola affect gelatin dissolubility. But in the TLC treated group gelatin was degraded by only MMP secreted by gingival fibroblast. Regarding to the above results, TDC and TLC have an effect on the IL-6 secretion increase of human gingival fibroblast and appears to activate pro MMP-2 which degrades collagen.

  • PDF

Changes of Interleukin-12 and Transforming Growth Factor Beta 1 before and after Antipsychotic Treatments in Schizophrenic Patients (정신분열병 환자에서 Interleukin-12와 Transforming Growth Factor Beta 1의 치료 전후의 변화)

  • Kim, Sung-Jae;Lee, Bun-Hee;Kim, Yong-Ku
    • Korean Journal of Biological Psychiatry
    • /
    • v.12 no.2
    • /
    • pp.143-150
    • /
    • 2005
  • Background:Several reports have suggested that cytokine alterations could be related to the pathophysiology of schizophrenia. In this study, we measured plasma level of interleukin-12(IL-12), a proinflammatory T helper 1(Th1) cytokine and transforming growth factor-${\beta}1$(TGF-${\beta}1$), an anti-inflammatory Th3 cytokine before and after antipsychotic treatment in schizophrenic patients. Methods:The plasma concentrations of IL-12 and TGF-${\beta}1$ were measured by using quantitative ELISA in 23 schizophrenic patients and 31 normal controls at admission and 8 weeks later. The psychopathology was measured by Brief Psychiatric Rating Scale(BPRS). Results:IL-12 and TGF-${\beta}1$ levels were significantly higher in schizophrenic patients than in controls before treatment. At the 8 week of treatment, the TGF-${\beta}1$ levels returned to control values, while IL-12 levels were not significantly changed. There were no significant correlations between the changes of BPRS scores and the changes of IL-12 or TGF-${\beta}1$ levels in schizophrenic patients. Conclusion:Cytokine abnormalities in schizophrenia might be involved in the pathophysiology of the illness. It is possible that TGF-${\beta}1$ plays an important role in the schizophrenia.

  • PDF

Studying of the Effects of Gamichungsangboha-tang water-extract and ethanol-extract on $IFN{\gamma}$, IL-4, T-bet, GATA-3, c-maf in Asthma Model (가미청상보하탕(加味淸上補下湯)의 물과 에탄올 추출법에 따른 항(抗)알레르기 효과(效果)에 대(對)한 연구(硏究))

  • Heo, Tae-Seok;Ko, Eun-Jung;Jung, Hee-Jae;Rhee, Hyung-Koo;Bae, Hyun-Su;Jung, Sung-Ki
    • The Journal of Internal Korean Medicine
    • /
    • v.27 no.1
    • /
    • pp.40-54
    • /
    • 2006
  • Objective: This study aimed to identify the different effects of GMCSBHT water-extract and ethanol-extract on Th1/Th2 differentiation by monitoring Th1/Th2 specific cytokine secretion patterns and the transcriptional activities of T-bet, GATA-3, c-maf, $INF{\gamma}$ and IL-4. Materials and Methods: Spleen cells from eight week-old BALB/c mice were cultured in GMCSBHT extracts containing medium without activation for 24 hours and with activation for 48 hours. CD4+ T cells were isolated and mRNA expression levels of $INF{\gamma}$, IL-4, T-bet, GATA-3, c-maf by RT-PCR and secretion cytokines levels of $IFN{\gamma}$, IL-4 by ELISA were analyzed. Results: GMCSBHT extracts didn't have mitogenic effects on the unstimulated CD4+ T cells. In Th1 skewed condition, GMCSBAHT water extract had no significant effects on mRNA expression levels of $IFN{\gamma}$, T-bet and c-maf, but inhibited mRNA expression levels of IL-4, GATA-3. It showed significantly increased secretion cytokine levels of $IFN{\gamma}$, but had no significant effect on secretion cytokine levels of IL-4. In Th2 skewed condition, GMCSBHT ethanol extract inhibited mRNA expression levels of $INF{\gamma}$, IL-4, GATA-3 and c-maf significantly, but had no significant effects on mRNA expression levels of T-bet. It had no significant effects on secretion cytokine levels of $INF{\gamma}$, but showed remarkable inhibitory effects on secretion cytokine levels of IL-4. Conclusion: Results suggest that on Th1/Th2 deviation, GMCSBHT water extract has both amplifying effects on Th1 differentiation and inhibitory effects on Th2, but GMCSBHT ethanol extract has stronger inhibitory effects on Th2 differentiation than on Th1.

  • PDF