Activation of acid-sensing ion channels （ASICs） plays an important role in neuroinflammation. Macrophage recruitment to the sites of inflammation is an essential step in host defense. ASIC1 and ASIC3 have been repor...Activation of acid-sensing ion channels （ASICs） plays an important role in neuroinflammation. Macrophage recruitment to the sites of inflammation is an essential step in host defense. ASIC1 and ASIC3 have been reported to mediate the endocytosis and maturation of bone marrow derived macrophages. However, the expression and inflammation-related functions of ASICs in RAW 264.7 cells, another common macrophage, are still elusive. In the present study, we first demonstrated the presence of ASIC 1, ASIC2a and ASIC3 in RAW 264.7 macrophage cell line by using reverse transcriptase polymerase chain reaction （RT-PCR）, Western blotting and immunofluorescence experiments. The non-specific ASICs inhibitor amiloride and specific homomeric ASICla blocker PcTxl reduced the production of iNOS and COX-2 by LPS-induced activating RAW 264.7 cells. Furthermore, not only amiloride but also PcTxl inhibited the migration and LPS-induced apoptosis of RAW 264.7 cells. Taken together, our findings suggest that ASICs promote the inflammatory response and apoptosis of RAW 264.7 cells, and ASICs may serve as a potential novel target for immunological disease therapy.展开更多
Objective:To investigate whether the ethanol extract of Chondracanthus tenellus(Harvey)Hommersand,a type of red algae,could exhibit anti-inflammatory potential in lipopolysaccharide(LPS)-stimulated macrophages.Methods...Objective:To investigate whether the ethanol extract of Chondracanthus tenellus(Harvey)Hommersand,a type of red algae,could exhibit anti-inflammatory potential in lipopolysaccharide(LPS)-stimulated macrophages.Methods:The ethanol extract of Chondracanthus tenellus was applied to 100 ng/mL LPS-stimulated RAW 264.7 cells,and cell viability,phagocytic ability,levels of pro-inflammatory factors,and the production of reactive oxygen species were measured.To identify the underlying mechanism of the ethanol extract of Chondracanthus tenellus,the expression of inflammation-regulated genes was estimated.Results:The ethanol extract of Chondracanthus tenellus had no cytotoxic effect at concentrations below 300μg/mL,and reduced the LPS-induced production of inflammatory mediators including nitric oxide(NO)and prostaglandin E2.Furthermore,the extract markedly suppressed the expression of inducible NO synthase and cyclooxygenase-2,as well as the production of reactive oxygen species.The LPS-induced up-regulation of pro-inflammatory cytokines was attenuated by treatment with the ethanol extract of Chondracanthus tenellus,reducing their extracellular secretion.The Chondracanthus tenellus extract also inhibited LPS-mediated activation of nuclear factor-kappa B(NF-κB).In addition,the phosphorylation of mitogen activated protein kinases(MAPKs)and phosphatidylinositol 3 kinase(PI3K)/Akt was markedly increased by LPS,which was significantly abolished by the Chondracanthus tenellus extract.Conclusions:Our findings indicate that the ethanol extract of Chondracanthus tenellus exhibited potential anti-inflammatory and antioxidant effects through downregulating the NF-κB,MAPKs,and PI3K/Akt signaling pathways in LPS stimulated RAW 264.7 macrophages.tenellus extract.Conclusions:Our findings indicate that the ethanol extract of Chondracanthus tenellus exhibited potential anti-inflammatory and antioxidant effects through downregulating the NF-κB,MAPKs,and PI3K/Akt signaling pathways in LPS stimulated RAW 264.7 macrophages.展开更多
Skin aging and most age-related diseases are associated with a low-grade chronic inflammation. The nucleoside adenosine, a potent endogenous anti-inflammatory agent, is deeply involved in inflammatory diseases and, by...Skin aging and most age-related diseases are associated with a low-grade chronic inflammation. The nucleoside adenosine, a potent endogenous anti-inflammatory agent, is deeply involved in inflammatory diseases and, by interaction with the adenosine A2 receptor (A2AR) it immediately promotes a mechanism of defence against the inflammatory damage. The aim of our study was to investigate whether polydeoxyribonucleotide (PDRN), a mixture of deoxyribonucleotides polymers of different lengths that like adenosine, binds the A2A receptor, can reduce the inflammatory state in the macrophage cell line. RAW264.7, murine macrophage cells, were incubated with PDRN in the presence and in the absence of lipopolysaccaride (LPS), which was the major component of the outer membrane of gram-negative bacteria and which acted as a strong macrophage activator. We assessed the production of nitric oxide and the secretion of inflammatory mediators (i.e., TNF-α, IL-10, IL-12 and VEGF-A). Our data showed that PDRN produced a significant decrease of inflammation in macrophages pre-stimulated with LPS, assessed in terms of the nitric oxide content (p 2A receptor, contributed to a great extent towards reducing inflammation.展开更多
Objective:To demonstrate the anti-inflammatory activity of Brassica napus L.hydrosols(BNH)in lipopolysaccharide(LPS)-stimulated RAW 264.7 cells.Methods:Composition analysis of BNH was conducted via gas chromatography-...Objective:To demonstrate the anti-inflammatory activity of Brassica napus L.hydrosols(BNH)in lipopolysaccharide(LPS)-stimulated RAW 264.7 cells.Methods:Composition analysis of BNH was conducted via gas chromatography-mass spectrometry after BNH were extracted.The nitric oxide(NO)production was measured using the Griess assay.Prostaglandin E2(PGE2)production was evaluated with enzyme-linked immunosorbent assay.The effects of BNH on LPS-induced pro-inflammatory enzymes including inducible nitric oxide synthase(iNOS)and cyclooxygenase-2(COX-2)were evaluated using Western blot analysis.Furthermore,phosphorylation of nuclear factor-kappa B(N F-k B)and nuclear translocation of N F-k B p65 were evaluated with Western blot analysis and immunofluorescence staining,respectively.Results:Compared with LPS-stimulated cells,BNH markedly decreased the generation of NO and PGE_(2)in LPS-stimulated RAW 264.7 cells(P<0.01 or P<0.05).Moreover,BNH inhibited protein levels of iNOS and COX-2(P<0.01).Phosphorylation of NF-k B and nuclear translocation of NF-k B p65 was significantly inhibited by BNH(P<0.01 or P<0.05).Conclusion:The anti-inflammatory activities of BNH were mediated via blockage of the N F-k B signaling pathways in LPS-stimulated RAW 264.7 cells.展开更多
The saponin ginsenoside Rk1 is a major compound isolated from ginseng.Ginsenoside Rk1 has been reported to have anti-inflammatory and anti-tumor properties and to be involved in the regulation of metabolism.However,th...The saponin ginsenoside Rk1 is a major compound isolated from ginseng.Ginsenoside Rk1 has been reported to have anti-inflammatory and anti-tumor properties and to be involved in the regulation of metabolism.However,the effect and mechanism of anti-inflammatory action of ginsenoside Rk1 has not been fully clarified.We investigated whether ginsenoside Rk1 could suppress the inflammatory response in lipopolysaccharide-stimulated RAW264.7 macrophages and to explore its mechanism of the action.RAW264.7 cells were treated with LPS(1 μg×mL^(–1))in the absence or the presence of Ginsenoside Rk1(10,20,and 40 μmol×L^(–1)).Then the inflammatory factors were tested with Griess reagents,ELISA,and RT-PCR.The proteins were analyzed by Western blotting.Ginsenoside Rk1 inhibited lipopolysaccharide-induced expression of nitric oxide(NO),interleukin(IL)-6,IL-1β,tumor necrosis factor(TNF)-α,and monocyte chemotactic protein(MCP)-1.Ginsenoside Rk1 inhibited the lipopolysaccharide-stimulated phosphorylation of NF-κB and janus kinase(Jak)2 and signal transducer and activator of transcription(Stat)3 at Ser727 and Tyr705.These data suggested that ginsenoside Rk1 could inhibit expression of inflammatory mediators and suppress inflammation further by blocking activation of NF-κB and the Jak2/Stat3 pathway in LPS-stimulated RAW264.7 cells.展开更多
OBJECTIVE: To investigate the effect of Young Yum pill (YYP) on inflammatory mediators in cultured RAW 264.7 cells and elucidate the nuclear factor- kappa B (NF-κB)-related mechanism behind the action. METHODS: YYP w...OBJECTIVE: To investigate the effect of Young Yum pill (YYP) on inflammatory mediators in cultured RAW 264.7 cells and elucidate the nuclear factor- kappa B (NF-κB)-related mechanism behind the action. METHODS: YYP was extracted with 95% ethanol Lipopolysaccharide (LPS)-stimulated RAW 264.7 macrophages were used to evaluate the effect of YYP on inflammatory mediators. Production of nitric oxide (NO) and prostaglandin E2 (PGE2) were measured by Griess test and enzyme-linked immunosorbent assay, respectively. The levels of genes and proteins involved in the generation of inflammatory mediators were examined using real-time polymerase chain reaction and Western blotting, respectively. RESULTS: YYP dose-dependently suppressed LPS-induced production of NO, PGE2 and tumor necrosis factor-α(TNF-α), and elevation of mRNA and protein levels of inducible NO synthase and cyclooxygenase- 2 in RAW 264.7 macrophages. These observations were associated with decreased NF-κB p65 phosphorylation and nuclear localization, enhanced Akt (protein kinase B) phosphorylation, as well as reduced inhibitor of κB (IκB)α degradation and IκB kinase α/β phosphorylation. CONCLUSION: The present study demonstrated an inhibitory effect of YYP on the NF-κB-regulated inflammatory mediators NO, PGE2 and TNF-α in LPS-stimulated RAW 264.7 macrophages, providing a pharmacological basis for the use of YYP in treating inflammatory disorders.展开更多
基金This work was supported by grants from the National Natural science Foundation of China （No. 81473199）, and the Fundamental Research Funds for the Central Universities （No, 015TS 125）.
文摘Activation of acid-sensing ion channels （ASICs） plays an important role in neuroinflammation. Macrophage recruitment to the sites of inflammation is an essential step in host defense. ASIC1 and ASIC3 have been reported to mediate the endocytosis and maturation of bone marrow derived macrophages. However, the expression and inflammation-related functions of ASICs in RAW 264.7 cells, another common macrophage, are still elusive. In the present study, we first demonstrated the presence of ASIC 1, ASIC2a and ASIC3 in RAW 264.7 macrophage cell line by using reverse transcriptase polymerase chain reaction （RT-PCR）, Western blotting and immunofluorescence experiments. The non-specific ASICs inhibitor amiloride and specific homomeric ASICla blocker PcTxl reduced the production of iNOS and COX-2 by LPS-induced activating RAW 264.7 cells. Furthermore, not only amiloride but also PcTxl inhibited the migration and LPS-induced apoptosis of RAW 264.7 cells. Taken together, our findings suggest that ASICs promote the inflammatory response and apoptosis of RAW 264.7 cells, and ASICs may serve as a potential novel target for immunological disease therapy.
基金This research was a part of the project titled‘Omics based on fishery disease control technology development and industrialization(20150242)’,funded by the Ministry of Oceans and Fisheries,Republic of Korea。
文摘Objective:To investigate whether the ethanol extract of Chondracanthus tenellus(Harvey)Hommersand,a type of red algae,could exhibit anti-inflammatory potential in lipopolysaccharide(LPS)-stimulated macrophages.Methods:The ethanol extract of Chondracanthus tenellus was applied to 100 ng/mL LPS-stimulated RAW 264.7 cells,and cell viability,phagocytic ability,levels of pro-inflammatory factors,and the production of reactive oxygen species were measured.To identify the underlying mechanism of the ethanol extract of Chondracanthus tenellus,the expression of inflammation-regulated genes was estimated.Results:The ethanol extract of Chondracanthus tenellus had no cytotoxic effect at concentrations below 300μg/mL,and reduced the LPS-induced production of inflammatory mediators including nitric oxide(NO)and prostaglandin E2.Furthermore,the extract markedly suppressed the expression of inducible NO synthase and cyclooxygenase-2,as well as the production of reactive oxygen species.The LPS-induced up-regulation of pro-inflammatory cytokines was attenuated by treatment with the ethanol extract of Chondracanthus tenellus,reducing their extracellular secretion.The Chondracanthus tenellus extract also inhibited LPS-mediated activation of nuclear factor-kappa B(NF-κB).In addition,the phosphorylation of mitogen activated protein kinases(MAPKs)and phosphatidylinositol 3 kinase(PI3K)/Akt was markedly increased by LPS,which was significantly abolished by the Chondracanthus tenellus extract.Conclusions:Our findings indicate that the ethanol extract of Chondracanthus tenellus exhibited potential anti-inflammatory and antioxidant effects through downregulating the NF-κB,MAPKs,and PI3K/Akt signaling pathways in LPS stimulated RAW 264.7 macrophages.tenellus extract.Conclusions:Our findings indicate that the ethanol extract of Chondracanthus tenellus exhibited potential anti-inflammatory and antioxidant effects through downregulating the NF-κB,MAPKs,and PI3K/Akt signaling pathways in LPS stimulated RAW 264.7 macrophages.
文摘Skin aging and most age-related diseases are associated with a low-grade chronic inflammation. The nucleoside adenosine, a potent endogenous anti-inflammatory agent, is deeply involved in inflammatory diseases and, by interaction with the adenosine A2 receptor (A2AR) it immediately promotes a mechanism of defence against the inflammatory damage. The aim of our study was to investigate whether polydeoxyribonucleotide (PDRN), a mixture of deoxyribonucleotides polymers of different lengths that like adenosine, binds the A2A receptor, can reduce the inflammatory state in the macrophage cell line. RAW264.7, murine macrophage cells, were incubated with PDRN in the presence and in the absence of lipopolysaccaride (LPS), which was the major component of the outer membrane of gram-negative bacteria and which acted as a strong macrophage activator. We assessed the production of nitric oxide and the secretion of inflammatory mediators (i.e., TNF-α, IL-10, IL-12 and VEGF-A). Our data showed that PDRN produced a significant decrease of inflammation in macrophages pre-stimulated with LPS, assessed in terms of the nitric oxide content (p 2A receptor, contributed to a great extent towards reducing inflammation.
基金Supported by the Korea Basic Science Institute(No.C38915 and C030360)。
文摘Objective:To demonstrate the anti-inflammatory activity of Brassica napus L.hydrosols(BNH)in lipopolysaccharide(LPS)-stimulated RAW 264.7 cells.Methods:Composition analysis of BNH was conducted via gas chromatography-mass spectrometry after BNH were extracted.The nitric oxide(NO)production was measured using the Griess assay.Prostaglandin E2(PGE2)production was evaluated with enzyme-linked immunosorbent assay.The effects of BNH on LPS-induced pro-inflammatory enzymes including inducible nitric oxide synthase(iNOS)and cyclooxygenase-2(COX-2)were evaluated using Western blot analysis.Furthermore,phosphorylation of nuclear factor-kappa B(N F-k B)and nuclear translocation of N F-k B p65 were evaluated with Western blot analysis and immunofluorescence staining,respectively.Results:Compared with LPS-stimulated cells,BNH markedly decreased the generation of NO and PGE_(2)in LPS-stimulated RAW 264.7 cells(P<0.01 or P<0.05).Moreover,BNH inhibited protein levels of iNOS and COX-2(P<0.01).Phosphorylation of NF-k B and nuclear translocation of NF-k B p65 was significantly inhibited by BNH(P<0.01 or P<0.05).Conclusion:The anti-inflammatory activities of BNH were mediated via blockage of the N F-k B signaling pathways in LPS-stimulated RAW 264.7 cells.
基金supported by the National Natural Science Foundation of China(Nos.81173369,81303253,81530097,and 81222051)the National Key Technology R&D Program “New Drug Innovation” of China(Nos.2012ZX09301002-002-002 and 2012ZX09304-005)+1 种基金the Natural Science Foundation of Beijing,China(No.132210)the Doctoral Scientific Fund Project of the Ministry of Education of China(No.20120001110105)
文摘The saponin ginsenoside Rk1 is a major compound isolated from ginseng.Ginsenoside Rk1 has been reported to have anti-inflammatory and anti-tumor properties and to be involved in the regulation of metabolism.However,the effect and mechanism of anti-inflammatory action of ginsenoside Rk1 has not been fully clarified.We investigated whether ginsenoside Rk1 could suppress the inflammatory response in lipopolysaccharide-stimulated RAW264.7 macrophages and to explore its mechanism of the action.RAW264.7 cells were treated with LPS(1 μg×mL^(–1))in the absence or the presence of Ginsenoside Rk1(10,20,and 40 μmol×L^(–1)).Then the inflammatory factors were tested with Griess reagents,ELISA,and RT-PCR.The proteins were analyzed by Western blotting.Ginsenoside Rk1 inhibited lipopolysaccharide-induced expression of nitric oxide(NO),interleukin(IL)-6,IL-1β,tumor necrosis factor(TNF)-α,and monocyte chemotactic protein(MCP)-1.Ginsenoside Rk1 inhibited the lipopolysaccharide-stimulated phosphorylation of NF-κB and janus kinase(Jak)2 and signal transducer and activator of transcription(Stat)3 at Ser727 and Tyr705.These data suggested that ginsenoside Rk1 could inhibit expression of inflammatory mediators and suppress inflammation further by blocking activation of NF-κB and the Jak2/Stat3 pathway in LPS-stimulated RAW264.7 cells.
基金Supported by Wai Yuen Tong Medicine Company Limited,Innovation and Technology Fund(UIT/315)General Research Fund(GRF:12125116)of Hong KongFRG1/16-17/048 and FRG2/16-17/033 from the Hong Kong Baptist University
文摘OBJECTIVE: To investigate the effect of Young Yum pill (YYP) on inflammatory mediators in cultured RAW 264.7 cells and elucidate the nuclear factor- kappa B (NF-κB)-related mechanism behind the action. METHODS: YYP was extracted with 95% ethanol Lipopolysaccharide (LPS)-stimulated RAW 264.7 macrophages were used to evaluate the effect of YYP on inflammatory mediators. Production of nitric oxide (NO) and prostaglandin E2 (PGE2) were measured by Griess test and enzyme-linked immunosorbent assay, respectively. The levels of genes and proteins involved in the generation of inflammatory mediators were examined using real-time polymerase chain reaction and Western blotting, respectively. RESULTS: YYP dose-dependently suppressed LPS-induced production of NO, PGE2 and tumor necrosis factor-α(TNF-α), and elevation of mRNA and protein levels of inducible NO synthase and cyclooxygenase- 2 in RAW 264.7 macrophages. These observations were associated with decreased NF-κB p65 phosphorylation and nuclear localization, enhanced Akt (protein kinase B) phosphorylation, as well as reduced inhibitor of κB (IκB)α degradation and IκB kinase α/β phosphorylation. CONCLUSION: The present study demonstrated an inhibitory effect of YYP on the NF-κB-regulated inflammatory mediators NO, PGE2 and TNF-α in LPS-stimulated RAW 264.7 macrophages, providing a pharmacological basis for the use of YYP in treating inflammatory disorders.
基金National Natural Science Foundation of China(Grant No.41906033)the Natural Science Foundation of Guangdong Province of China(Grant No.2019A1515012084)。
