壮药三七姜醇提物及含药血清对脂多糖诱导RAW264.7细胞炎症的抗炎作用及机制

目的 研究壮药三七姜醇提物及含药血清对脂多糖（LPS）诱导RAW264.7细胞炎症的抗炎作用及机制。方法 将三七姜醇提物（75.35 g/kg）或纯净水灌胃大鼠以制备含药血清或空白血清。以RAW264.7细胞为研究对象,将细胞分为正常对照组,LPS组（1μg/mL）,三七姜醇提物高、中、低剂量组（50、25、12.5μg/mL）,4%或15%空白血清组,4%或15%空白血清+LPS组,4%或15%含药血清组,4%或15%含药血清+LPS组,按相应条件培养24 h后,检测各组细胞活力和细胞中一氧化氮（NO）、肿瘤坏死因子α(TNF-α)、白细胞介素1β(IL-1β)、IL-6含量,以及Toll样受体4(TLR4)、核因子κB(NF-κB)mRNA表达水平,一氧化氮合酶（NOS）、环氧化酶2(COX-2)蛋白表达水平。结果 各组细胞培养24 h后,细胞活力差异无统计学意义。与正常对照组比较,LPS组细胞中NO、TNF-α、IL-1β、IL-6含量,TLR4、NF-κB mRNA表达水平和NOS、COX-2蛋白表达水平均显著升高（P<0.05）。与4%或15%空白血清组比较,4%或15...     

三七总皂苷调节iNOS-NO-NF-κB信号通路抑制LPS诱导的RAW246.7细胞炎症研究

目的 探讨三七总皂苷（PNS）对脂多糖（LPS）诱导的RAW264.7细胞iNOS-NO-NF-κB信号通路相关分子表达及活性的影响。方法 MTT比色法检测PNS对RAW264.7细胞增殖的抑制作用;不同浓度PNS（25、50、100 μg/mL）干预体外培养LPS诱导的RAW264.7细胞24、48 h后,Griess法检测NO变化;PNS干预24 h后,Western blotting检测iNOS、NF-κB、p-NF-κB、IKKα、p-IKKα、p-ΙκΒα蛋白表达量的变化;PNS干预4 h后,激光共聚焦显微镜检测NF-κB转位入核情况。结果 PNS浓度大于150 μg/mL时,才表现出显著抑制细胞增殖的作用。25、50、100 μg/mL PNS作用于RAW264.7细胞24和48 h后,与模型组比较,NO生成量均显著降低（P<0.001）。50、100 μg/mL PNS作用于RAW264.7 24 h后,iNOS、NF-κB蛋白表达量显著降低,磷酸化蛋白p-NF-κB、p-IKKα、p-ΙκΒα表达水平也显著降低（P<0.01、0.001）。与模型组比较,PNS 25、50、100 μg/mL组核因子p65入核荧光强度显著降低（P<0.01、0.001）。结论 PNS能够显著抑制LPS诱导的RAW264.7细胞iNOS-NO-NF-κB信号通路的活性,降低细胞炎症反应。     

海洋共生菌中脑苷脂对RAW264.7细胞的抗炎作用研究

海洋微生物因具有生长迅速、代谢可调控、菌种易选育、可持续利用等特点成为寻找药理活性物质的新源泉。使用从海洋共附生菌中提取的脑苷脂,采用CCK8法检测其对小鼠单核巨噬细胞（RAW264.7）细胞的毒性,建立RAW264.7细胞炎症模型,检测脑苷脂对炎症因子IL-6和IL-1β的含量变化,运用实时荧光定量PCR技术检测IL-6、IL-1β、TNF-α、MCP-1、MIP-2和INOS mRNA表达量。通过Western Blot技术检测炎症相关信号通路p-ERK1/2、MyD88、NF-κB、NLRP3、p62、p-IKB和p-JNK蛋白的表达量。研究结果显示,脑苷脂可以降低IL-6、IL-1β、TNF-α、MCP-1、MIP-2和INOS炎症因子的含量和mRNA表达量,抑制p-ERK1/2、MyD88、NF-κB、NLRP3、p-IKB和p-JNK蛋白的表达量。这些结果表明,脑苷脂可以通过抑制MyD88/NF-κB/MAPK/NLRP3信号通路实现对脂多糖诱导的RAW264.7细胞的抗炎作用。这为脑苷脂类化合物用于治疗炎症相关疾病提供数据参考,为开发新型海洋药物奠定基础。     

青蒿乙素对脂多糖诱导小鼠单核巨噬细胞RAW264.7炎症反应的抑制及机制

目的 探讨青蒿乙素的抗炎活性及其作用机制.方法 采用细菌脂多糖(LPS)诱导小鼠单核巨噬细胞RAW264.7建立炎症反应模型,经青蒿乙素处理后,采用Griess试剂检测NO的含量;采用酶联免疫法(ELISA)检测促炎症因子的水平;采用Western Blot检测炎症相关蛋白及激酶的表达水平.结果 青蒿乙素能明显抑制RA...     

参附注射液对LPS诱导的RAW264.7细胞HMGB1核转位的干预作用

目的:探讨参附注射液(SFI)对脂多糖(LPS)诱导的巨噬细胞中高迁移率族蛋白B1(HMGB1)核转位的干预作用。方法:以经LPS诱导的小鼠单核巨嗜细胞RAW264.7为对象,以组蛋白去乙酰化酶抑制剂RGFP966为阳性对照,在CCK-8法筛选给药剂量的基础上,采用免疫荧光法观察低、中、高剂量(3、6、12μL/m L)SFI对细胞中HMGB1核转位的影响,采用实时荧光聚合酶链式反应法检测细胞中HMGB1 mRNA的表达情况;采用Western blotting法检测细胞中HMGB1、Toll样受体4(TLR4)的表达情况,并比较细胞胞核、胞浆中HMGB1的表达情况;采用酶联免疫吸附测定法检测细胞上清液中HMGB1、白细胞介素1β(IL-1β)、肿瘤坏死因子α(TNF-α)水平。结果:在空白对照组中,HMGB1主要定位于细胞核;经LPS诱导后,HMGB1由细胞核向胞浆迁移。与空白对照组比较,LPS组细胞中HMGB1 m RNA及其蛋白、TLR4的蛋白表达量以及上清液中HMGB1、IL-1β、TNF-α水平均显著升高(P<0.01);细胞胞核中HMGB1的蛋白表达量显著降低,而胞浆...     

MYR通过NF-κκB信号通路改善LPS诱导小鼠纹状体炎症反应

目的研究MYR对LPS诱导小鼠纹状体内神经炎症的作用及机制。方法雄性BALB/c小鼠随机分为正常对照组、LPS模型组、MYR 20和50 mg·kg~(-1)给药组。连续给予MYR 7 d,于末次给药后,模型组和给药组小鼠采用腹腔注射LPS5 mg·kg~(-1)诱导小鼠急性神经炎症的发生,LPS注射6 h后,ELISA法检测小鼠纹状体中IL-1β,IL-6,TNF-α,MCP-1以及ICAM-1等炎症因子的变化;Western印迹法检测小鼠纹状体中NF-κB信号通路相关蛋白的表达变化。结果与正常对照组比较,LPS诱导组小鼠纹状体中炎症因子IL-1β,IL-6,TNF-α,MCP-1以及ICAM-1显著增加(所有P<0.01),NF-κB信号通路相关蛋白表达显著增加。而给予MYR 20和50 mg·kg~(-1)可显著抑制LPS诱导的小鼠纹状体炎症因子的释放,抑制NF-κB,IκB蛋白的磷酸化,抑制胞浆内NF-κB的核转位。结论 MYR可有效抑制LPS诱导的神经炎症,保护小鼠纹状体中多巴胺神经元,其抗炎保护作用与抑制NF-κB信号通路密切相关。     

香青兰总黄酮对RAW264.7巨噬细胞泡沫化及炎症反应的调控作用北大核心CSCD

目的研究香青兰总黄酮(total flavonoids of Dracocephalum moldavica L.,TFDM)对氧化低密度脂蛋白(oxidized low density lipoprotein,ox-LDL)诱导的小鼠单核巨噬细胞白血病细胞(RAW264.7)泡沫化及炎症的影响,进一步阐明TFDM抗动脉粥样硬化(atherosclerosis,AS)的作用机制。方法体外培养RAW264.7巨噬细胞,采用ox-LDL刺激诱导使其成为泡沫细胞,TFDM(25、50、100 mg·L^(-1))及辛伐他汀(10μmol·L^(-1))进行干预,油红O染色法观察胞内脂滴的聚集情况,CCK-8法检测细胞活力,活性氧试剂盒测定ROS的生成,实时荧光定量PCR测定细胞中NF-κB、NLRP3、caspase-1、IL-18和IL-1βmRNA的表达,免疫蛋白印迹法检测巨噬细胞中IκBα、NF-κB p65、NLRP3、pro-caspase-1、caspase-1、IL-1β以及IL-18蛋白的表达,ELISA法检测TNF-α和IL-10的表达。结果TFDM可以减少泡沫巨噬细胞的形成,降低炎症因子IL-1β、IL-18和TNF-α的表达,增加抑炎因子IL-10的表达;并且下调NF-κB p65、NLRP3、pro-caspase-1、caspase-1蛋白的表达,上调IκBα的蛋白表达。结论TFDM能够减轻巨噬细胞的泡沫化,抑制炎症因子的表达,从而可能延缓动脉粥样硬化的发展进程。其作用机制可能是通过抑制NF-κB途径,减少ox-LDL诱导的RAW264.7细胞中炎症介质的产生。     

化合物4h对脂多糖诱导RAW264.7细胞炎症的抑制作用及机制研究

摘要：目的:探讨化合物4h对脂多糖（LPS）诱导小鼠腹腔巨噬细胞炎症的抑制作用。方法:小鼠腹腔巨噬细胞RAW264.7经LPS诱导建立炎症模型,实验分为空白组、对照组、阳性罗格列酮组(5,10μmol·L-1)、阳性吲哚美辛组(5,10μmol·L-1)、4h组(5,10μmol·L-1),采用酶联免疫吸附实验测定目标化合物4h对于炎症因子肿瘤坏死因子α（TNF-α）分泌的影响;实验分为空白组、对照组、4h组(5,10μmol·L-1),采用蛋白免疫印迹实验测定目标化合物4h对于信号通路Toll样受体4/核因子κB（TLR4/NF-κB）蛋白的表达水平。结果:与空白组比较,LPS作用后,对照组TNF-α含量显著升高（P<0.01）,并且随着LPS作用时间的延长,TNF-α含量随之升高。加入化合物处理后,各实验组TNF-α含量呈下降趋势,与对照组比较,化合物4h各时间点的TNF-α含量差异均有统计学意义（P<0.05或P<0.01）,随着化合物浓度的升高,炎症因子的含量逐渐下降。化合物4h对TNF-α的抑制作用明显优于两个阳性对照药罗格列酮和吲哚美辛（P<0.05或P<0.01）。与空白组比较,LPS作用后对照组RAW 264.7细胞中IκBα蛋白表达水平显著降低,TLR4、p-NF-κB p65和p-IκBα蛋白表达水平显著升高（P<0.01）。与对照组比较,4h作用后实验组RAW 264.7细胞中IκBα蛋白表达水平显著升高,TLR4、p-NF-κB p65和p-IκBα蛋白表达水平显著降低（P<0.05或P<0.01）。结论:4h通过抑制信号通路TLR4/NF-κB蛋白的表达,进而抑制炎症因子TNF-α的产生,从而发挥抗炎作用。     

牛膝-杜仲药对对小鼠巨噬细胞RAW264.7的抗炎作用

目的 研究牛膝-杜仲药对的抗炎作用.方法 将小鼠巨噬细胞RAW264.7分为空白组、模型组、牛膝组(800μg/mL)、杜仲组(800μg/mL)和牛膝-杜仲药对低、中、高浓度组(400、800、1600μg/mL),除空白组和模型组外,其余各组加入相应药物培养6 h后,空白组继续加入培养基,模型组加入10μg/mL脂...     

雷公藤次碱通过调控TLR4/MyD88/TRAF6信号通路抑制LPS诱导的RAW264.7细胞炎症反应

目的 探讨雷公藤次碱抗炎活性及其作用机制。方法 用细胞计数盒-8 (CCK-8)法考察雷公藤次碱对小鼠单核巨噬细胞白血病细胞RAW 264.7 增殖活性的影响,用酶联免疫吸附法（ELISA）检测雷公藤次碱对脂多糖（LPS）诱导的RAW264.7细胞分泌细胞因子一氧化氮（NO）、白细胞介素-1β（IL-1β）、 肿瘤坏死因子α（TNF-α）和白细胞介素6（IL-6）的影响,用免疫印迹法考察雷公藤次碱对白介素受体相关激酶（IRAK）、肿瘤坏死因子受体相关蛋白6（TRAF6）、核因子κB抑制因子α（IκBα）、核因子κB（NF-κB p65）、分裂原激活的蛋白激酶p38（p38）、c-Jun氨基末端激酶（JNK）、细胞外调节蛋白激酶（ERK）在LPS刺激的RAW264.7细胞中的表达及其磷酸化的影响。结果 雷公藤次碱在25、50、100 μmol/L浓度下对RAW264.7细胞无显著毒性,并可显著抑制细胞因子NO、IL-1β、TNF-α和IL-6含量。免疫印迹法检测结果显示雷公藤次碱可显著抑制IRAK及TRAF6的表达;显著抑制ERK、P38和JNK的磷酸化;抑制IκBα的降解,降低NF-κB p65的核转运水平。结论 雷公藤次碱具有体外抗炎活性,其作用机制可能介导TLR4/MyD88/TRAF6信号通路。     

Anti-inflammatory effect of pomegranate flower in lipopolysaccharide (LPS)-stimulated RAW264.7 macrophages

Context: Punica granatum L (Punicaceae) flower is an important diabetes treatment in oriental herbal medicine.

Objective: This study investigates the inflammation effects of pomegranate flower (PFE) ethanol extract in LPS-induced RAW264.7 cells.

Materials and methods: PFE (10, 25, 50, 100?μg/mL) was applied to 1?μg/mL LPS-induced RAW 264.7 macrophages in vitro. Levels of nitric oxide (NO), prostaglandin E₂ (PGE₂) and pro-inflammatory cytokines interleukin (IL)-1β (IL-1β), interleukin (IL)-6 (IL-6) and tumor necrosis factor (TNF-α) in the supernatant fraction were determined using enzyme-linked immunosorbent assay (ELISA). Expression of cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS), phosphorylation of mitogen-activated protein kinase (MAPK) subgroups extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK) and P38, as well as nuclear factor-κB (NF-κB) activation in extracts were detected via Western blot.

Results: 10–100?μg/mL PFE decreased the production of NO (IC₅₀ value?=?31.8?μg/mL), PGE₂ (IC₅₀ value?=?54.5?μg/mL), IL-6 (IC₅₀ value?=?48.7?μg/mL), IL-1β (IC₅₀ value?=?71.3?μg/mL) and TNF-α (IC₅₀ value?=?62.5?μg/mL) in LPS-stimulated RAW 264.7 cells significantly. A mechanism-based study showed that phosphorylation of ERK1/2, p38, JNK and translocation of the NF-B p65 subunit into nuclei were inhibited by the PFE treatment.

Discussion and conclusion: These results show that PFE produced potential anti-inflammatory effect through modulating the synthesis of several mediators and cytokines involved in the inflammatory process.     

金骨莲胶囊对炎症模型大鼠的抗炎作用及机制研究

目的:研究金骨莲胶囊对炎症模型大鼠的抗炎作用及机制.方法:将48只大鼠随机分为空白对照组、模型组、金骨莲胶囊低、中、高剂量组(0.66、1.32、2.64 g/kg)和地塞米松组(阳性对照,0.945 mg/kg),每组8只.空白对照组和模型组大鼠灌胃等体积水,其余各组大鼠灌胃相应药物,每天给药2次,连续3天.末次给药...     

The anti-inflammatory effect of Sonchus oleraceus aqueous extract on lipopolysaccharide stimulated RAW 264.7 cells and mice

Context: Sonchus oleraceus L. (Asteraceae) (SO) is a dietary and traditional medicinal plant in China. However, its underlying mechanism of action as an anti-inflammatory agent is not known.

Objective: This study evaluates the anti-inflammatory activity of aqueous extract of SO.

Materials and methods: The extract of SO was used to treat RAW 264.7 cells (in the working concentrations of 500, 250, 125, 62.5, 31.3 and 15.6?μg/mL) for 24?h. Pro-inflammatory cytokines and mediators produced in LPS-stimulated RAW 264.7 cells were assessed. Meanwhile, the expression level of TLR-4, COX-2, pSTATs and NF-κB was tested. Moreover, the anti-inflammatory activity of the extract in vivo was assessed using xylene-induced mouse ear oedema model and the anti-inflammatory compounds in the extracts were analyzed by HPLC-MS.

Results: SO extract significantly inhibited the production of pro-inflammatory cytokines and mediators at gene and protein levels with the concentration of 31.3?μg/mL, and suppressed the expression of TLR-4, COX-2, NF-κB and pSTAT in RAW 264.7 cells. The anti-inflammatory activity of SO in vivo has significant anti-inflammatory effects with the concentration of 250 and 125?mg/kg, and less side effect on the weights of the mice at the concentration of 250?mg/kg. Moreover, HPLC-MS analysis revealed that the anti-inflammatory compounds in the extract were identified as villosol, ferulaic acid, β-sitosterol, ursolic acid and rutin.

Discussion and conclusion: This study indicated that SO extract has anti-inflammatory effects in vitro and in vivo, which will be further developed as novel pharmacological strategies in order to defeat inflammatory diseases.     

Anti-inflammatory effect of essential oil and its constituents from fingered citron (Citrus medica L. var. sarcodactylis) through blocking JNK,ERK and NF-κB signaling pathways in LPS-activated RAW 264.7 cells

We investigated the composition of essential oil from fingered citron (Citrus medica L. var. sarcodactylis) (FCEO) peels by GC–MS and its anti-inflammatory effects on lipopolysaccharide (LPS) – stimulated mouse macrophage (RAW 264.7) cells. Fifteen compounds, representing 98.97% of the essential oil, were tentatively identified; the main constituents were limonene (52.44%) and γ-terpinene (28.41%). FCEO significantly inhibited nitric oxide (NO) and prostaglandin E2 (PGE2) by suppressing the protein expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase (COX)-2, respectively. Additionally, FCEO suppressed the production of tumor necrosis factor-α (TNF-α), interleukin (IL)-1β, and IL-6. FCEO attenuated LPS-induced nuclear factor-κB (NF-κB) activation via inhibition of inhibitor κB-α phosphorylation. Furthermore, FCEO blocked activation of c-Jun N-terminal kinase (JNK) and extracellular signal-regulated kinase (ERK) but not that of p38 mitogen-activated protein kinase. These results indicate that FCEO inhibits LPS-stimulated inflammation by blocking the NF-κB, JNK, and ERK pathways in macrophages, and demonstrate that FCEO possesses anti-inflammatory properties.     

Activation of NLRP3 inflammasome in RAW 264.7 cells by polysaccharides extracted from Grateloupia livida (Harv.) Yamada

Polysaccharides have been proven to be involved in the immune response in both anti-inflammation and pro-inflammation due to their distinct pharmacological properties. Recent studies showed that polysaccharides from Grateloupia livida (Harv.) Yamada possessed several biological activities, including anti-oxidant, anti-angiogenic, anti-cancer and antiviral. Our previous work has elucidated the structural features of the polysaccharides (named WGW) by combining ESI-MS with NMR and infrared (IR) spectroscopic analyses. The polysaccharides were mainly composed of galactose linked with sulfate ester, concluding to be μ-carrageenan and κ-carrageenan. The purpose of this study was to investigate the immunoregulation effects of WGW on macrophage RAW 264.7 cell. The cells were treated with WGW for different times, then Griess reagent was applied to detect the production of NO. The results presented that WGW induced the release of NO in large quantities (ranging from 2 µg/mL to 256 µg/mL) and improved the phagocytosis of macrophage RAW264.7 cells. Furthermore, WGW have a predominant role in the improvement of proinflammatory mediators, such TNF-α, IL-1β, iNOS, MMP-9 and COX2. Moreover, WGW activated NLRP3 inflammasome, in which MAPK/NF-κB signaling pathway played an important role. These results indicated WGW might be a potential immuno-stimulation drug to promote inflammation.     

Anti-inflammatory effects of Juncus effusus extract (JEE) on LPS-stimulated RAW 264.7 cells and edema models

Context: Juncus effusus L. var. decipiens BUCHEN. f. leschenaultii GAY has been used in traditional medicine for the treatment of anxiety and insomnia.

Objective: The objective of this study was to evaluate the effects of ethanol extract from the pith of Juncus effusus (JEE) on anti-inflammatory activities in RAW 264.7 cells.

Materials and methods: The production of inflammatory mediators and the underlying mechanisms using 3.1, 6.3, and 12.5?μg/mL concentrations of JEE were investigated. In addition, the topical anti-inflammatory effects of JEE (0.5, 1, and 2?mg/mL) on 12-O-tetradecanoylphorobol-13 acetate (TPA)-induced ear edema and oral administration of JEE (50, 100, and 200?mg/kg) on carrageenan-induced paw-edema were studied in mice.

Results: JEE reduced the release of nitric oxide (NO, IC₅₀ value?=?1.98?μg/mL), prostaglandin E₂ (IC₅₀ value?=?5.5?μg/mL), and pro-inflammatory cytokines, IL-1β (IC₅₀ value?=?4.74?μg/mL) and IL-6 (IC₅₀ value?=?20.48?μg/mL). JEE also suppressed the protein expression of inducible NO synthase and cyclooxygenase-2 in lipopolysaccharide (LPS)-stimulated RAW 264.7 cells. Mechanism studies showed attenuation of LPS-induced activation of NF-κB by JEE via abrogation of IκBα degradation and a subsequent decrease in nuclear p65 level. Phosphorylation of all three MAP kinases (ERK, JNK, and p38) in LPS-stimulated RAW 264.7 cells was also suppressed in a dose-dependent manner. In acute inflammation models of mice, topical application (1 and 2?mg) and oral administration (50, 100, and 200?mg/kg) of JEE ameliorated TPA-induced ear edema and carrageenan-induced paw edema, respectively, in dose-dependent manners.

Discussion and conclusion: These results indicate that JEE exhibited anti-inflammatory activities by suppressing the production of inflammatory mediators in LPS-stimulated RAW 264.7 cells and by attenuating edema in mice.     

Anti-inflammatory effect of sargachromanol G isolated from Sargassum siliquastrum in RAW 264.7 cells

A study on the anti-inflammatory activity of brown alga Sargassum siliquastrum led to the isolation of sargachromanol G (SG). In this study, the anti-inflammatory effect and the action mechanism of SG have been investigated in murine macrophage cell line RAW 264.7. SG dosedependently inhibited the production of inflammatory markers [nitric oxide (NO), inducible nitric oxide synthase (iNOS), prostaglandin E(2) (PGE(2)), and cyclooxygenase-2 (COX-2)] and pro-inflammatory cytokines [tumor necrosis factor-α (TNF-α), interleukin (IL)-1β, and IL-6] induced by LPS treatment. To further elucidate the mechanism of this inhibitory effect of SG, we studied LPS-induced nuclear factor-κB (NF-κB) activation and mitogen-activated protein kinases (MAPKs) phosphorylation. SG inhibited the phosphorylation IκB-α and NF-κB (p65 and p50) and MAPK (ERK1/2, JNK, and p38) in a dose dependent manner. These results suggest that the anti-inflammatory activity of SG results from its modulation of pro-inflammatory cytokines and mediators via the suppression of NF-κB activation and MAPK phosphorylation.