IL-33敲除促进弓形虫感染小鼠腹腔巨噬细胞的M1偏移

目的 研究IL-33敲除(IL-33^-/-)对小鼠急性弓形虫感染腹腔巨噬细胞(pMφ)极化的影响,探讨IL-33^-/-在弓形虫感染免疫应答中的作用。方法 收集C57BL/6 IL-33^-/-小鼠和野生型(WT)小鼠pMφ,各分为弓形虫感染组和未感染组,比较各组pMφ 感染率、细胞因子及表面分子表达水平的变化。结果 IL-33^-/-小鼠pMφ 弓形虫感染30 min后的感染率低于WT小鼠(t=-2.49,P<0.05);IL-33^-/-小鼠感染组pMφ M1型标志物NO(t=29.71,P<0.05)、MHCⅡ(t=19.05,P<0.05)、TLR4(t=8.34,P<0.05)表达高于WT小鼠感染组,而M2型标志物CD206表达低于WT小鼠感染组(t=-3.34,P<0.05);感染组小鼠pMφ NO、IL-10、TNF-α、MHCⅡ类分子及CD86的表达高于各自未感染对照组;IL-33敲除和弓形虫感染对IL-33^-/-与WT小鼠pMφ MHCⅡ(F=5.25,P<0.05)、TLR2(F=14.88,P<0.05)分子的表达有交互作用。结论 在急性弓形虫感染早期,IL-33敲除驱动小鼠pMφ 向M1方向极化,促进pMφ 抗感染。     

弓形虫RH株ROP16I/III基因缺陷虫株感染BALB/c鼠的致病性研究

目的 旨在深入研究弓形虫棒状体效应分子ROP16与虫株毒力及致病性的关系。方法 采用弓形虫ROP16_I/III基因敲除RH株(RH_ΔROP16)攻击感染BALB/c小鼠,在感染后不同时间点与野生株感染鼠比较,动态观察感染动物发病症状、存活时间; HE染色观察脑组织、肺组织病理学差异,qRT-PCR检测脾细胞炎性及抑炎细胞因子表达水平。结果 两组小鼠的发病症状无明显差异;经HE染色显微镜下观察小鼠肺部及脑部病理学改变亦未见无明显差异;qRT-PCR检测并用Graph pad分析两组虫株感染小鼠的脾细胞Arg-1、IL-10、IL-12、TNF -α及IFN-γ等细胞因子表达水平。数据表明在感染相同时间ROP16缺陷株感染小鼠Arg-1的表达量明显低于野生型虫株(P< 0.05);而IL-10、IL-12、TNF-α和IFN -γ的表达水平无统计学差异(P> 0.05)。结论 I型弓形虫RH株的ROP16_I/III并非是决定急性感染期弓形虫毒力的唯一效应分子;ROP16_I/III可诱导宿主Arg-1高表达,提示与巨噬细胞内虫体的增殖有关。     

弓形虫RH株调节JAK1-STAT6信号通路诱导人原代肺泡巨噬细胞极化的研究

目的探讨弓形虫感染人原代肺泡巨噬细胞的特点以及调节JAK1-STAT6信号通路对人肺泡巨噬细胞极化的影响。方法收集肺癌患者肺泡灌洗液,分离纯化肺泡巨噬细胞并作细胞爬片处理,体外选择1∶1的比例感染弓形虫并设置不同感染时间,Diff染色分析巨噬细胞内弓形虫的增殖情况。分别在不同感染时间收集细胞,提取全蛋白,Western blot检测信号通路蛋白的表达;Trizol法提取mRNA,反转录为cDNA后采用Real-time PCR检测相关mRNA的表达。结果成功分离并纯化得到肺泡巨噬细胞并感染弓形虫。Diff染色显示弓形虫可在肺泡巨噬细胞生存繁殖,从24 h开始弓形虫繁殖力较为明显增强,36 h时弓形虫数量显著增多并且从细胞内释放到细胞外。Western blot检测M1型巨噬细胞信号通路蛋白STAT1、P-STAT1和P-NF-κB呈先增高后降低的趋势,以P-NF-κB先升高后增加的趋势尤为明显;M2型巨噬细胞信号通路JAK1、STAT6和P-STAT6信号通路蛋白也呈现逐渐升高的趋势,iNOS的表达呈显著降低的趋势,Arg-1的表达量在6 h时较高(t=9.35,P0.05)。Real-time PCR检测TNF mRNA相对表达量逐渐降低,iNOS mRNA相对表达量在6 h时升高,之后降低;Arg-1 mRNA相对表达量的相对表达量逐渐升高,48 h时表达量升高尤为显著(t=4.27,P0.05)。结论弓形虫感染能诱导巨噬细胞表达M2型细胞炎性因子,并抑制M1型细胞因子的表达,可通过JAK1-STAT6信号通路调节人原代肺泡巨噬细胞向M2型巨噬细胞偏移。     

感染旋毛虫小鼠免疫能力的性别差异研究

目的 研究感染旋毛虫早期雌雄小鼠免疫能力的性别差异。方法 不同剂量的旋毛虫感染雌雄小鼠7 d后,测定脏器指数和白细胞组成比例,脾脏和胸腺组织进行HE染色分析,ELISA法测定血清中IL-2、IL-4、IL-10的含量,流式细胞仪检测脾脏免疫调节性T细胞比例。结果 与感染旋毛虫的雌性小鼠相比,感染旋毛虫雄性小鼠的胸腺指数明显增大(t=4.595, P<0.05);淋巴细胞百分比明显下降而单核细胞和粒细胞百分比明显上升(t=2.604, P<0.05);胸腺组织变化不明显而脾脏组织变化明显;免疫调节性T细胞比例上升,但无统计学意义。血清中细胞因子的变化也存在雌雄差异,与正常小鼠相比,感染旋毛虫的雌性小鼠IL-2含量上升而雄性小鼠下降(F=5.664,P<0.05);IL-4、IL-10含量均上升(F=10.461,P<0.05;F=1.170,P>0.05),且雄性高于雌性。结论 感染旋毛虫7 d后,雄性小鼠的免疫应答强于雌性小鼠。     

金丝桃素对小鼠巨噬细胞内刚地弓形虫速殖子的增殖抑制作用

目的 观察金丝桃素(Hypericin,Hy)对小鼠巨噬细胞内弓形虫的增殖抑制作用,为抗弓形虫天然药物的筛选提供线索。方法 分离小鼠腹腔巨噬细胞(Macrophage,Mφ),建立体外Mφ感染稳定表达绿色荧光蛋白的弓形虫(RH-green fluorescent protein,RH-GFP)速殖子模型,观察不同剂量Hy的最佳作用效果。实验分4组:①A组(对照组),不加Hy;②B组(Mφ感染RH-GFP+100 μg/mL Hy);③C组(Mφ感染RH-GFP+200 μg/mL Hy);④D组(Mφ感染RH-GFP+400 μg/mL Hy),分别在培养1、2和3 h后利用荧光显微镜和流式细胞术观察计数感染的Mφ和游离速殖子在各药物浓度及各时间点的变化;透射电镜观察虫体超微结构的变化。结果 与对照组相比,随着Hy浓度的增加和用药时间的延长,Mφ内速殖子数量明显减少,游离速殖子数量也显著减少,荧光强度逐渐降低;经药物组不同浓度Hy作用后,游离/Mφ内弓形虫的比值逐渐升高,至400 μg/mL剂量作用3 h后开始下降,游离与胞内速殖子比例的差异在同一时间段内均有统计学意义 (P<0.05),但药物作用1 h或2 h各实验组之间无统计学差异 (P>0.05),作用3 h各浓度组之间差异有统计学意义(P<0.05),各实验组同一药物浓度下在孵育1、2和3 h后效果增强(P<0.05)。透射电镜观察显示,随Hy作用时间延长,虫体逐渐出现肿胀,胞膜与基质之间空隙明显,空泡形成且增多、变大,胞膜断裂,内部结构溶解溢出。结论 体外实验初步显示Hy呈剂量和时间依赖性地增强对Mφ内弓形虫速殖子增殖的抑制作用,可降低宿主细胞感染率,且对胞外速殖子有一定杀伤作用。     

结核分枝杆菌CFP10-ESAT6融合蛋白免疫小鼠效果初步评价

目的 融合表达和纯化结核分枝杆菌(MTB)CFP10-ESAT6(CE),初步评价免疫效果。方法 构建pET32a(+)-CE重组质粒,E.coli BL21(DE3)中融合表达,Ni-NTA层析纯化。用CE加铝佐剂,50 μg CE/只/次,分3次免疫BALB/c小鼠。ELISA测定血清IgG抗体;ELISpot与Luminex检测细胞因子,进行MTB体外生长抑制试验。卡介苗(BCG)为阳性对照。结果 CE免疫小鼠,能诱导产生高效价的IgG、IgG1和IgG2a,且IgG1(t=19.1,P<0.000 1)和IgG2a(t=8.7,P<0.000 1)抗体水平均高于BCG组。CE组与BCG组诱导产生的分泌IFN-γ的斑点形成细胞(SFC)数量差异无统计学意义(t=0.4, P>0.05),但诱导分泌IL-4的SFC数量高于BCG组(t=8.0, P<0.000 1)。CE组诱导分泌GM-CSF(t=8.4,P<0.05)、IL-6(t=8.3,P<0.05)、IL-10(t=2.5,P<0.05)和IL-4(t=7.0,P<0.05)均高于BCG组,而CE组诱导分泌IFN-γ(t=1.4,P>0.05)、TNF-α(t=1.8,P>0.05)、IL-2(t=2.0,P>0.05)、IL-12(t=0.9,P>0.05)和IL-17(t=1.3,P>0.05)均与BCG组相似,差异无统计学意义。CE组小鼠脾细胞的MTB体外生长抑制结-果与BCG组相似(t=0.8,P>0.05)。结论 CE免疫小鼠可诱导较强的Th1与Th2混合型免疫反应,且有较强的体外抑制MTB生长的能力,表明CE具有良好的结核疫苗研制应用价值。     

葡萄糖调节蛋白78对单核巨噬细胞分化的影响

目的:探讨葡萄糖调节蛋白78(GRP78)对单核巨噬细胞分化的影响。方法:利用佛波酯(PMA)诱导形成THP-1单核细胞来源巨噬细胞模型。实验分为4组,分别为对照组(PMA浓度为50 ng/ml)、GRP78 10 ng/ml组(GRP78浓度10 ng/ml+PMA 50 ng/ml)、GRP78 20 ng/ml组(GRP78浓度20 ng/ml+PMA 50 ng/ml)、GRP78 40 ng/ml组(GRP78浓度40 ng/ml+PMA 50 ng/ml),培养72 h。应用流式细胞术检测巨噬细胞M1型特异表面标志物CD86、M2型特异表面标志物CD163的表达;实时荧光定量PCR检测巨噬细胞M1型相关因子iNOS、M2型相关因子FIZZ-1、TGF-β、ARG-1及IL-10的mRNA表达。结果:与对照组比较,其他3组的CD163阳性巨噬细胞百分比显著增加(P0.01);而各实验组CD86阳性巨噬细胞百分比没有明显改变。与对照组相比,GRP78实验组中各组FIZZ-1、TGF-β的mRNA水平均有增加,其中GRP78 40 ng/ml组FIZZ-1、TGF-β的mRNA水平明显增高(P0.01);各组其他因子iNOS、IL-10和ARG-1的mRNA水平则没有明显变化。结论:GRP78可诱导THP-1单核巨噬细胞极化成M2型巨噬细胞。     

葡萄糖调节蛋白78对单核巨噬细胞分化的影响

目的:探讨葡萄糖调节蛋白78(GRP78)对单核巨噬细胞分化的影响。方法:利用佛波酯(PMA)诱导形成THP-1单核细胞来源巨噬细胞模型。实验分为4组,分别为对照组(PMA浓度为50 ng/ml)、GRP78 10 ng/ml组(GRP78浓度10 ng/ml+PMA 50 ng/ml)、GRP78 20 ng/ml组(GRP78浓度20 ng/ml+PMA 50 ng/ml)、GRP78 40 ng/ml组(GRP78浓度40 ng/ml+PMA 50 ng/ml),培养72 h。应用流式细胞术检测巨噬细胞M1型特异表面标志物CD86、M2型特异表面标志物CD163的表达;实时荧光定量PCR检测巨噬细胞M1型相关因子iNOS、M2型相关因子FIZZ-1、TGF-β、ARG-1及IL-10的mRNA表达。结果:与对照组比较,其他3组的CD163阳性巨噬细胞百分比显著增加(P0.01);而各实验组CD86阳性巨噬细胞百分比没有明显改变。与对照组相比,GRP78实验组中各组FIZZ-1、TGF-β的mRNA水平均有增加,其中GRP78 40 ng/ml组FIZZ-1、TGF-β的mRNA水平明显增高(P0.01);各组其他因子iNOS、IL-10和ARG-1的mRNA水平则没有明显变化。结论:GRP78可诱导THP-1单核巨噬细胞极化成M2型巨噬细胞。     

TIPE2通过TLR4/IκBα/NF-κB通路调节脂肪组织巨噬细胞表型转化

目的探讨肿瘤坏死因子-α诱导蛋白8样分子-2(TIPE2)对脂多糖或白细胞介素4(IL-4)诱导的脂肪组织巨噬细胞(ATM)表型转化的作用和分子机制。方法应用免疫组化、Western印迹法和实时定量PCR(RT-qPCR)检测肥胖小鼠和TIPE2敲除小鼠(KO)及其各自对照小鼠内脏脂肪组织中TIPE2、诱导型一氧化氮合酶(iNOS)、单核细胞趋化蛋白1(MCP-1)、CD206和精氨酸酶1(Arg-1)的表达水平。分离培养TIPE2敲除小鼠(KO)和野生型小鼠(WT)的腹腔巨噬细胞及RAW 264.7小鼠巨噬细胞系, 给予脂多糖(100 ng/mL)或IL-4(20 ng/mL)刺激6 h, Western印迹法和RT-qPCR检测TIPE2、iNOS、MCP-1、CD206和Arg-1的表达水平。结果在肥胖小鼠中, TIPE2表达下调, 促炎因子iNOS和MCP-1表达升高, 抑炎因子CD206和Arg-1表达降低。脂多糖降低RAW 264.7细胞和小鼠腹腔巨噬细胞TIPE2的表达, 诱导经典活化的巨噬细胞(M1表型)标志物iNOS和MCP-1表达升高, 降低替代活化的巨噬细胞(M2...     

结核分枝杆菌Rv3621c原核表达及免疫功能研究

目的 以结核分枝杆菌(Mycobacterium tuberculosis,M.tb)H37Rv基因组为模板,构建、纯化及鉴定原核表达质粒pPROEX-Rv3621c,通过人群、小鼠试验进行免疫原性评价。方法 构建重组质粒pPROEX-Rv3621c,并以全血干扰素释放分析技术(Whole-blood IFN-γ release assay,WBIA)检测其能否被安徽省淮南市M.tb感染者T细胞特异性识别。rRv3621c混合佐剂MTM[母牛分枝杆菌(M.vaccae),人工合成海藻糖-6'6,二分枝菌酸(TDB),单磷酰脂质A(MPLA)]免疫小鼠后,检测血清中特异性抗体分泌水平、脾细胞中抗原特异性Th1型细胞因子分泌水平及肺脏细胞因子mRNA表达水平。结果 成功构建重组质粒pPROEX-Rv3621c,并使之诱导表达、纯化和鉴定。在rRv3621c蛋白诱导下,活动性结核(Active tuberculosis, ATB)患者外周血淋巴细胞释放的IFN-γ水平明显较高(t=4.813, P<0.01),且ATB患者产生的IFN-γ水平高于潜伏性结核(Latent tuberculosis infection, LTBI)人群(t=4.442, P<0.01)。BCG+Rv3621c/MTM组小鼠产生的特异性抗体滴度水平明显高于Rv3621c/MTM组(P<0.01)和BCG组(P<0.01),Rv3621c/MTM组和BCG+Rv3621c/MTM组小鼠的IgG2a/IgG1比值大于1,明显高于MTM组和BCG组。BCG+Rv3621c/MTM组小鼠均分泌高水平IFN-γ、TNF-α和IL-2。Rv3621c/MTM组小鼠肺脏组织中IFN-γ、TNF-α及iNOS表达水平较高。结论 M.tb感染者外周血T细胞可特异性识别rRv3621c蛋白,rRv3621c混合佐剂MTM可以诱导较强烈的抗原特异性Th1型免疫应答。     

microRNA?155在弓形虫感染调节巨噬细胞极化中的作用

目的 探讨弓形虫感染对宿主细胞内microRNA?155（miR?155）表达的影响及其对巨噬细胞极化的作用。 方法 利用miRNAs基因芯片检测弓形虫感染宿主细胞内miRNAs表达水平,应用实时定量聚合酶链反应技术（qPCR）检测miR?155表达水平。采用脂质体转染法将pEGFP?miR?155导入人巨噬细胞,利用流式细胞术检测转染效率。采用流式细胞术、qPCR、酶联免疫法检测弓形虫感染组、pEGFP?miR?155过表达组以及miR?155抑制组诱导巨噬细胞表面分子CD86及诱导型一氧化氮合酶（iNOS）和白细胞介素（IL）?12表达水平。结果 基因芯片和qPCR检测结果显示,随着弓形虫感染时间的延长,miR?155表达量上升;pEGFP?miR?155转染宿主细胞的转染效率可达82.6%。弓形虫感染组和pEGFP?miR?155过表达组CD86表达水平显著高于对照组和miR?155抑制组,iNOS和IL?12基因表达量显著升高。结论 弓形虫感染能够通过上调miR?155表达参与驱动人源巨噬细胞向M1型偏移。     

姜黄素对不同亚型THP-1来源巨噬细胞表达炎症因子的抑制作用

目的 探讨姜黄素对M0、M1和M23种亚型巨噬细胞表达炎症因子的影响。方法 构建M0型/M1型/M2型巨噬细胞模型以及不同浓度梯度的姜黄素干预组和对照组，通过实时荧光定量PCR及酶联免疫吸附方法分别测定不同分组中TNFα、IL-6以及IL-123种炎症因子的表达。结果 实时荧光定量PCR结果显示，与对照组相比，姜黄素对3种亚型巨噬细胞TNFα、IL-6以及IL-12 mRNA的表达均具有明显抑制作用(P<0.01)，酶联免疫吸附实验也证实姜黄素能抑制以上3种细胞炎症因子的分泌水平(P<0.01)。结论 姜黄素对M0、M1和M23种亚型的巨噬细胞炎症因子表达水平均有不同程度的抑制作用，且呈剂量依赖关系。     

Toxoplasma gondii infection can regulate the expression of tumour necrosis factor-alpha receptors on human cells in vitro

The in vitro regulation of tumour necrosis factor (TNF)-alpha receptors during Toxoplasma gondii infection of human MRC5 fibroblasts and human myelomonocytic THP-1 cells was investigated. Cells were infected with the virulent RH of T. gondii. TNFR membrane receptors were analysed by flow cytometry with biotinylated TNF-alpha. Shedding of the soluble form of TNFR1 and TNFR2 in cell culture supernatants was measured by enzyme-linked immunosorbent assay, and expression of mRNA production of TNFR1 and TNFR2 was analysed by quantitative real-time polymerase chain reaction, 1 h after infection. In the MRC5 cell line, T. gondii infection did not induce any up- or down-regulation of membrane TNFRs, soluble TNFRs or mRNA of TNFRs. However, THP-1 cell infection with living parasites induced a significant soluble TNFR1 release by THP-1 cells after 1 h. We detected an approximately 50% up-regulation (P < 0.01) of soluble TNFR1 in infected THP-1 cells compared to controls. No change in soluble TNFR2 levels was observed in the same conditions. Moreover, infection decreased the level of TNF membrane receptors, but had no effect on TNFR1 and TNFR2 mRNA levels. TNFR modulation by T. gondii infection, in vitro, depends on the cell type. Furthermore, our data suggest that living parasites control the shedding of the soluble form of TNFR1. This mechanism may influence the role of TNF-alpha in toxoplasmosis.     

Curcumin induces M2 macrophage polarization by secretion IL-4 and/or IL-13

AimsTo address the underlying mechanisms by which curcumin facilitates M2 phenotype polarization of macrophages and its roles in the protective effects during experimental autoimmune myocarditis (EAM).Methods and resultsThe expression of classic M2 markers, including macrophage mannose receptor (MMR), arginase-1 (Arg-1) and peroxisome proliferator-activated receptor-γ (PPAR-γ) was upregulated in curcumin-treated Raw264.7 macrophages. Curcumin increased interleukin-4 (IL-4) and interleukin-13 (IL-13) mRNA expression and protein secretion. Curcumin notably increased STAT6 phosphorylation. Leflunomide, a STAT6 inhibitor, and IL-4 and/or IL-13 neutralizing antibodies antagonized the induction of MMR, Arg-1 and PPAR-γ by curcumin in Raw264.7 cells. In vivo, 6-week old male Lewis rats were used to induce EAM and orally administrated with curcumin or corn oil for 3 weeks after myosin injection. Cardiac functional parameters, including left ventricular fractional shortening (LVFS), ejection fraction (EF), left ventricular end-systolic diameter (LVEDs) and heart rate (HR) were significantly improved by curcumin treatment. Curcumin also reduced the inflammatory cell infiltration and myocardial mRNA levels of interleukin-1β (IL-1β) and inducible nitric oxide synthase (iNOS). Meanwhile, the myocardial mRNA levels of MMR and Arg-1 were markedly up-regulated by curcumin. Immunofluorescence assay showed that the number of CD68 ⁺ MMR ⁺ and CD68 ⁺ Arg-1 ⁺ double positive macrophages in curcumin-treated myocardial tissue was significantly higher than untreated control. The number of CD68 ⁺ iNOS ⁺ double positive macrophages was increased obviously in EAM group, but decreased markedly by curcumin treatment.ConclusionsTaken together, these results show that curcumin induces macrophage M2 polarization by secretion of IL-4 and/or IL-13. Curcumin ameliorates EAM by reducing infiltration inflammatory macrophages and by polarizing M0 and M1 macrophages to M2 phenotype.     

肺炎衣原体通过上调清道夫受体A1诱导THP-1源性泡沫细胞形成

目的观察清道夫受体A1和CD36在肺炎衣原体诱导THP-1源性泡沫细胞形成中的作用。方法给予不同浓度的肺炎衣原体(1×105～1×106IFU)感染THP-1源性巨噬细胞0～72h。运用油红O染色观察细胞质内脂滴的变化,酶荧光学法检测细胞内胆固醇酯含量的变化。分别运用逆转录聚合酶链反应和Western-Blot检测清道夫受体A1和CD36的mRNA和蛋白表达。结果高浓度的肺炎衣原体(5×105和1×106IFU)感染负荷低密度脂蛋白的THP-1源性巨噬细胞48h后,细胞质内的脂滴明显增多,胆固醇酯占总胆固醇百分比明显增加(>50%)。在负荷低密度脂蛋白的THP-1源性巨噬细胞上,肺炎衣原体感染呈浓度和时间依赖性地上调道夫受体A1 mRNA和蛋白表达,但不影响CD36 mRNA和蛋白表达。结论清道夫受体A1表达上调是肺炎衣原体诱导THP-1源性泡沫细胞形成的机制之一,这可能为进一步阐明肺炎衣原体感染促进动脉粥样硬化发生发展提供一个新的理论依据。     

地塞米松对大鼠腹腔巨噬细胞抗弓形虫作用的影响

目的研究地塞米松(Dexamethasone,DXM)对大鼠腹腔巨噬细胞抗弓形虫感染的影响。方法采用瑞-姬染色观察弓形虫在与DXM共孵育大鼠腹腔巨噬细胞内的增殖;以半定量RT-PCR法检测与DXM共孵育大鼠腹腔巨噬细胞IFN-γ、TNF-α和IL-2mRNA的表达,以ELISA法检测细胞培养上清中IFN-γ、TNF-α和IL-2的含量。结果弓形虫在DXM孵育的腹腔巨噬细胞内大量增殖,其弓形虫密度0h为(37±7)个/100个细胞,而24h时为(173±32)个/100个细胞(P<0.01);与DXM共孵育大鼠腹腔巨噬细胞24h时的IL-2、IFN-γ和TNF-αmRNA及其蛋白的表达与对照组相比均明显降低(P<0.01),如:DXM孵育组的TNF-α(187.52±39.41pg/ml)与对照组(115.43±22.46pg/ml)相比差异显著(P<0.01)。结论 DXM可诱发腹腔巨噬细胞易感弓形虫,其机制可能与细胞因子IFN-γ、TNF-α和IL-2表达下调有关。     

Hepcidin Induces M1 Macrophage Polarization in Monocytes or THP-1 Derived Macrophages

Background: Macrophage polarization plays a critical role in determining the inflammatory states. Hepcidin is a key negative regulator of iron homeostasis and functions. Although hepcidin has been shown to affect ferroportin expression in macrophages, whether it affects macrophage polarization is still largely unknown. Objective: To address whether hepcidin induces macrophage polarization. Methods: The expression of iNOS and CD206, and the ratio of IFN-γ vs IL-4 in THP-1 derived macrophages upon hepcidin stimulation were evaluated. Further detected was the percentage of CD16+ M1, CD23+ M1, CD10+ M2 and CCL22+ M2 cells in monocyte derived macrophages. Results: M1 associated molecules were increased in hepcidin-treated cells, yet M2 associated molecules were increased when hepcidin was neutralized. Concomitantly, we observed a significant increase in IRF3 phosphorylation in hepcidin-stimulated cells. However, STAT6 phosphorylation with hepcidin was neutralized. Conclusion: Hepcidin is able to induce macrophage polarization towards M1 type, and might be utilized as a potential M1 macrophage agonist in clinical practice.     

Transient expression of the IL-2 receptor alpha-chain in IL-6-induced myeloid cells is regulated by autocrine production of prostaglandin E2.

The alpha-chain of the interleukin 2 receptor (IL-2R alpha) is expressed on monocytes and macrophages after activation by bacterial lipopolysaccharide (LPS) and interferon-gamma (IFN-gamma). In the present study, we investigated whether the expression of IL-2R alpha is associated with the process of differentiation of myeloid cells to mature macrophages and how this expression is regulated. The murine myeloid M1 cell line, which can be induced by leukemia inhibitory factor (LIF) or interleukin 6 (IL-6) to differentiate from blast cells to mature macrophages, was used as a model system for myeloid differentiation. Bone marrow (BM)-derived macrophages were used as mature myeloid cells. Cytofluorometry revealed that IL-2R alpha is transiently expressed during M1 cell differentiation, with peak levels 24 h after induction by LIF or IL-6, whereas the high affinity receptor for monomeric IgG2a (FcR), a surface marker typical for macrophage differentiation, continues to rise up to 72 h. BM-derived macrophages already express FcR but not IL-2R alpha. IL-2R alpha expression is induced on these cells after treatment by IL-6 for up to 48 h. Treatment of IL-6-induced M1 cells with indomethacin permitted a sustained expression of IL-2R alpha beyond 24 h, and this effect was reversed by the addition of prostaglandin E2 (PGE2). Northern analysis showed that in M1 cells the expression of mRNA for IL-2R alpha, but not for IL-2R beta, is also transient, indicating that cell surface expression of IL-2R alpha is regulated at the mRNA level. These data show that inducers of macrophage differentiation such as LIF and IL-6 can induce a transient expression of the IL-2R alpha-chain in differentiating murine myeloid M1 cells and that autocrine production of PGE2 is involved in the control of the transient expression of this receptor. However, induction of expression of IL-2R alpha by IL-6 appears to be independent of differentiation because it can be induced on fully differentiated BM-derived macrophages as well.