BACKGROUND & AIMS: There is evidence that neutrophil functions such as chemotaxis and oxygen radical formation are disturbed in rheumatoid arthritis (RA). Medication might also influence these functions. Cyclic formation and depolymerisation of actin microfilaments is crucial in cell motility, but this phenomenon has not been studied in RA. The aim of this study was to investigate basal and dynamic (formyl-methionyl-leucyl-phenylalanine (fMLP)-induced) neutrophil actin polymerisation in ten RA patients (a) during therapy with non-steroidal anti-inflammatory drugs (NSAIDS) and (b) after stopping NSAIDS> The results were compared with those of ten age-matched controls. Basal F-actin content in RA patients with NSAIDS was significantly lower than in RA patients without NSAIDS and controls35.5 (25.0-49.0), 50.5 (27.0-75.0) and 52.5 (32.0-85.0), respectively. Conversely, upon stimulation with fMLP, the actin polymerisation curve of RA patients with NSAIDS was higher than for RA patients without NSAIDS and controls. These results suggest that, in RA, the effects orf NSAIDS on neutrophil functions might be related to changes in the actin polymerisation-depolymerisation cycle.

背景与目标： 有证据表明，类风湿关节炎（RA）中性粒细胞功能（如趋化性和氧自由基的形成）受到干扰。药物治疗也可能影响这些功能。肌动蛋白微丝的循环形成和解聚对于细胞运动至关重要，但尚未在RA中研究此现象。本研究的目的是研究10名RA患者的基础和动态（甲酰基-甲硫酰基-亮氨酰-苯丙氨酸（fMLP）诱导）嗜中性白细胞肌动蛋白聚合反应（a）在使用非甾体抗炎药（NSAIDS）治疗期间和（ b）停止NSAIDS后>将结果与十个年龄匹配的对照组的结果进行比较。 NSAIDS的RA患者的基础F-肌动蛋白含量显着低于非NSAIDS的RA患者和对照组，分别为35.5（25.0-49.0），50.5（27.0-75.0）和52.5（32.0-85.0）。相反，在用fMLP刺激后，患有NSAIDS的RA患者的肌动蛋白聚合曲线高于没有NSAIDS和对照的RA患者。这些结果表明，在RA中，NSAIDS对中性粒细胞功能的影响可能与肌动蛋白聚合-解聚循环的变化有关。

BACKGROUND & AIMS: :Staphylococcus epidermidis infections are usually nosocomial and involve colonization of biomaterials. The immune defense system cannot efficiently control the bacteria during these infections, which often results in protracted chronic inflammation, in which a key event is disturbed removal of neutrophils by tissue macrophages. While ingesting uninfected apoptotic neutrophils, macrophages release anti-inflammatory cytokines that lead to resolution of inflammation. In clinical studies, we have previously found elevated levels of the proinflammatory cytokines tumor necrosis factor-alpha (TNF-alpha) and interleukin-6 in synovial fluid from prostheses infected with coagulase negative staphylococci. We show that macrophages phagocytosing apoptotic neutrophils containing S. epidermidis released TNF-alpha and interleukin-6, whereas macrophages phagocytosing spontaneously apoptotic neutrophils did not. This difference was not due to dissimilar phagocytic capacities, because macrophages ingested both types of neutrophils to the same extent. The activation was induced mainly by the apoptotic neutrophils themselves, not by the few remaining extracellular bacteria. Macrophages were not activated by apoptotic neutrophils that contained paraformaldehyde-killed S. epidermidis. Proinflammatory reactions induced by clearance of apoptotic neutrophils containing S. epidermidis might represent an important mechanism to combat the infective agent. This activation of macrophages may contribute to the development of chronic inflammation instead of inflammation resolution.

背景与目标： ：表皮葡萄球菌感染通常在医院内发生，涉及生物材料的定殖。在这些感染过程中，免疫防御系统无法有效控制细菌，这通常会导致长期的慢性炎症，其中关键事件是组织巨噬细胞对嗜中性白细胞的去除受到干扰。在摄取未感染的凋亡中性粒细胞时，巨噬细胞释放抗炎细胞因子，导致炎症消退。在临床研究中，我们以前发现来自被凝固酶阴性葡萄球菌感染的假体的滑液中促炎细胞因子肿瘤坏死因子-α（TNF-alpha）和白介素6的水平升高。我们显示巨噬细胞吞噬含有表皮葡萄球菌的凋亡嗜中性粒细胞释放TNF-α和白介素6，而巨噬细胞吞噬自发性凋亡的嗜中性粒细胞则没有。这种差异不是由于吞噬能力不同而引起的，因为巨噬细胞会以相同程度摄入两种类型的中性粒细胞。活化主要是由凋亡的中性粒细胞本身诱导的，而不是由少数剩余的细胞外细菌诱导的。巨噬细胞没有被凋亡的嗜中性粒细胞激活，所述嗜中性粒细胞含有多聚甲醛杀死的表皮葡萄球菌。由含有表皮葡萄球菌的凋亡中性粒细胞清除引起的促炎反应可能代表了与传染病作斗争的重要机制。巨噬细胞的这种活化可能有助于慢性炎症的发展，而不是炎症的消退。

BACKGROUND & AIMS: :The tuberculosis agent Mycobacterium tuberculosis is primarily transmitted through air, but little is known about the tenacity of mycobacterium-containing aerosols derived from either suspensions or infected neutrophils. Analysis of mycobacterial aerosol particles generated from bacterial suspensions revealed an average aerodynamic diameter and mass density that may allow distant airborne transmission. The volume and mass of mycobacterial aerosol particles increased with elevated relative humidity. To more closely mimic aerosol formation that occurs in active TB patients, aerosols from mycobacterium-infected neutrophils were analysed. Mycobacterium-infected intact neutrophils showed a smaller particle size distribution and lower viability than free mycobacteria. In contrast, mycobacterium-infected necrotic neutrophils, predominant in M. tuberculosis infection, revealed particle sizes and viability rates similar to those found for free mycobacteria, but in addition, larger aggregates of viable mycobacteria were observed. Therefore, mycobacteria are shielded from environmental stresses in multibacillary aggregates generated from necrotic neutrophils, which allows improved tenacity but emphasizes short distance transmission between close contacts.

背景与目标： ：结核病菌结核分枝杆菌主要通过空气传播，但对于悬浮液或感染的中性粒细胞衍生的含分枝杆菌的气溶胶的韧性知之甚少。由细菌悬浮液产生的分枝杆菌气溶胶颗粒的分析显示，平均空气动力学直径和质量密度可允许远距离的空气传播。分枝杆菌气溶胶颗粒的体积和质量随着相对湿度的增加而增加。为了更精确地模拟活动性结核病患者中发生的气溶胶形成，分析了分枝杆菌感染的中性粒细胞的气溶胶。与游离分枝杆菌相比，分枝杆菌感染的完整中性粒细胞显示出更小的粒径分布和更低的生存力。相反，主要感染结核分枝杆菌的分枝杆菌感染坏死性中性粒细胞的粒径和存活率与游离分枝杆菌相似，但此外，观察到更大的存活分枝杆菌聚集体。因此，分枝杆菌不受坏死性嗜中性粒细胞产生的多杆菌聚集物中环境压力​​的影响，这可以提高强度，但强调紧密接触之间的短距离传播。

BACKGROUND & AIMS: OBJECTIVES:Intracellular components of transfusion-related acute lung injury (TRALI) were investigated by transmission electron microscopy. METHODS:The lungs from 2 fatal TRALI cases and 2 controls, previously studied by scanning electron microscopy, were studied by transmission electron microscopy. Morphologic data by light and phase microscopy, along with scanning and transmission electron microscopic observations, were collated. RESULTS:The 2 fatal TRALI cases exhibited dense laminated material within capillaries and postcapillary venules, similar to material identified within their neutrophils when viewed by transmission electron microscopy. This material polarized light and is presumed to be cholesterol crystals. CONCLUSIONS:The damage to the pulmonary vascular endothelium in TRALI is related to formation of cholesterol crystals originating within neutrophils.

背景与目标： 目的：通过透射电镜观察与输血相关的急性肺损伤（TRALI）的细胞内成分。
方法：采用透射电子显微镜研究了2例致命的TRALI病例和2例对照的肺，这些患者先前已通过扫描电子显微镜进行了研究。通过光学和相显微镜的形态学数据，以及扫描和透射电子显微镜观察，进行了整理。
结果：2例致命的TRALI病例在毛细血管和毛细血管后小静脉内均表现出致密的层状物质，类似于通过透射电镜观察时在其嗜中性粒细胞中鉴定出的物质。该物质偏振光，推测是胆固醇晶体。
结论：TRALI对肺血管内皮的损害与中性粒细胞内胆固醇晶体的形成有关。

BACKGROUND & AIMS: :TLRs are key elements of the pathogen recognition mechanism used by the host immune system. Neutrophils express almost all TLRs, and activation of TLRs, such as TLR2 and TLR4, has been shown to induce the production of proinflammatory cytokines and chemokines, potentially linking innate and adaptive immunity. In the present study, we investigated whether activation of TLRs induces neutrophil production of MCP-1/CCL2, a key mediator involved in the development of adaptive immunity. Activation of neutrophils with LPS, lipoteichoic acid, or N-palmitoyl-S-[2,3-bis(palmitoyloxy)-(2RS)-propyl]-[R]-Cys-[S]-Ser-[S]-Lys did not induce significant MCP-1 production and release; however, the Th1 cytokine IFN-gamma dramatically up-regulated MCP-1 production in cells activated with each TLR ligand. The majority of MCP-1 was released between 24 and 48 h of culture, indicating that this is a late event. The effect of IFN-gamma appeared to be due to its antiapoptotic effect, but not priming effect, revealing a biological consequence of IFN-gamma-induced neutrophil survival. Although IFN-gamma failed to protect neutrophils from cell death at a higher dose of LPS, the p38 MAPK inhibitor SB203580 dramatically increased MCP-1 release and neutrophil survival at this LPS concentration. Thus, p38 MAPK plays a previously uncharacterized role in neutrophil function. Taken together, our results indicate that human neutrophils produce MCP-1 in a Th1 microenvironment and this neutrophil-derived MCP-1 potentially amplifies the development of Th1 adaptive responses.

背景与目标： ：TLRs是宿主免疫系统使用的病原体识别机制的关键要素。中性粒细胞几乎表达所有TLR，TLR的激活（如TLR2和TLR4）已显示出诱导促炎性细胞因子和趋化因子的产生，并可能将先天免疫和适应性免疫联系在一起。在本研究中，我们调查了TLR的激活是否诱导中性粒细胞产生MCP-1 / CCL2，MCP-1 / CCL2是参与适应性免疫发展的关键介体。用LPS，脂蛋白酸或N-棕榈酰-S- [2,3-双（棕榈酰氧基）-（2RS）-丙基]-[R] -Cys- [S] -Ser- [S] -Lys活化嗜中性粒细胞没有引起明显的MCP-1产生和释放；然而，Th1细胞因子IFN-γ显着上调了每种TLR配体激活的细胞中MCP-1的产生。 MCP-1的大多数在培养24至48小时之间释放，表明这是一个晚期事件。 IFN-γ的作用似乎是由于其抗凋亡作用，而不是引发作用，这揭示了IFN-γ诱导的中性粒细胞存活的生物学结果。尽管在较高的LPS剂量下，IFN-γ不能保护嗜中性粒细胞免于细胞死亡，但在此LPS浓度下，p38 MAPK抑制剂SB203580显着增加了MCP-1的释放和嗜中性粒细胞的存活。因此，p38 MAPK在嗜中性粒细胞功能中发挥了以前未表征的作用。两者合计，我们的结果表明，人类嗜中性粒细胞在Th1微环境中产生MCP-1，而这种嗜中性粒细胞衍生的MCP-1可能会放大Th1适应性反应的发展。

BACKGROUND & AIMS: :TNFalpha-induced expression of CD83 in leukocytes is mediated by NF-kappab. The aim of our present study was to investigate the underlying mechanism of a unique functional antagonism between GM-CSF and TNFalpha-induced up-regulation of CD83 in human neutrophils. CD83 was down-regulated by co-stimulation of neutrophils with TNFalpha and GM-CSF compared to TNFalpha alone both at the level of mRNA and protein. In marked contrast, the expression of IL-1RA was up-regulated under the same conditions. The down-regulation of CD83 was not mediated by modulation of the NF-kappab signaling pathway. Neither was it mediated by a decrease in mRNA stability of CD83. NF-kappab was modulated under these conditions as both the expression of the target gene IL-1RA as well as the phosphorylation of IkBalpha were up-regulated. Our results show that co-stimulation with pro-inflammatory cytokines such as TNFalpha and GM-CSF can have differential effects on inflammatory pathways initiated in the same target cell. GM-CSF can both synergize with TNFalpha in the case of expression of IL1-RA and antagonize in the case of CD83. Therefore, expression of CD83 as read out for activation of neutrophils in patients with inflammatory diseases is complicated by the presence of cross-modulating cytokines such as GM-CSF.

背景与目标： ：TNFalpha诱导的白细胞CD83表达是由NF-κB介导的。我们本研究的目的是研究人类嗜中性粒细胞中GM-CSF与TNFalpha诱导的CD83上调之间独特的功能拮抗的潜在机制。与单独的TNFα相比，通过与TNFα和GM-CSF共同刺激嗜中性粒细胞CD83被下调。与之形成鲜明对比的是，在相同条件下，IL-1RA的表达上调。 CD83的下调不是由NF-κB信号通路的调节介导的。它也不是由CD83的mRNA稳定性的降低介导的。在这些条件下调节了NF-kappab，因为上调了靶基因IL-1RA的表达以及IkBalpha的磷酸化。我们的结果表明，与促炎细胞因子（如TNFalpha和GM-CSF）共同刺激可对同一靶细胞中起始的炎症途径产生不同的影响。 GM-CSF可以在IL1-RA表达的情况下与TNFalpha协同作用，而在CD83情况下则可以与TNFα拮抗作用。因此，由于存在交叉调节性细胞因子（例如GM-CSF），在炎症性疾病患者中读出CD83以激活嗜中性粒细胞的表达变得复杂。

BACKGROUND & AIMS: :CD28, described as a T cell costimulatory molecule so far, is expressed on human peripheral blood neutrophils, as shown by cell surface staining and immunoprecipitation with anti-CD28 monoclonal antibody, and by reverse transcription PCR. The phorbol 12-myristate 13-acetate-augmented expression of CD28 on these cells can be blocked by actinomycin D, an RNA transcription inhibitor, and staurosporin, a protein kinase inhibitor. Cross-linking of CD28 results in an early increase in IL-8 receptor A (IL-8RA or CXCR-1) expression and a concurrent increase in IL-8-induced chemotaxis. The expression of CXCR-1 is down-regulated by receptor internalization 3 h after CD28 cross-linking with concurrent decrease in IL-8-induced chemotactic migration. Thus, our results demonstrate for the first time that CD28 is expressed on human peripheral blood neutrophils and that CD28 may play an important role in the regulation of IL-8RA expression and migration of neutrophils in response to IL-8.

背景与目标： ：迄今为止，被描述为T细胞共刺激分子的CD28在人外周血中性粒细胞中表达，如细胞表面染色和用抗CD28单克隆抗体进行的免疫沉淀以及逆转录PCR所示。这些细胞上CD28的佛波12-肉豆蔻酸酯13-醋酸酯增强表达可以被放线菌素D（一种RNA转录抑制剂）和星形孢菌素（一种蛋白激酶抑制剂）阻断。 CD28的交联导致IL-8受体A（IL-8RA或CXCR-1）表达的早期增加，并同时引起IL-8诱导的趋化性增加。 CD28交联后3小时，受体内化作用下调了CXCR-1的表达，并同时降低了IL-8诱导的趋化性迁移。因此，我们的结果首次证明了CD28在人外周血中性粒细胞上表达，并且CD28可能在调节IL-8RA表达和中性粒细胞对IL-8的迁移中起重要作用。

BACKGROUND & AIMS: :In this study we investigated the effect of adrenomedullin (AM) on fMLP-mediated activation of human neutrophils. AM partially, but significantly, suppressed fMLP-induced upregulation of CD11b expression. The inhibitory effects of AM upon fMLP-induced upregulation of CD11b expression were completely blocked by CGRP [8-37], a CGRP receptor antagonist. AM significantly increased cAMP content in neutrophils and SQ-22,536, an adenylate cyclase inhibitor, and KT-5720, a PKA inhibitor, significantly blocked the inhibitory effects of AM upon fMLP-induced upregulation of CD11b expression. This study indicates that binding of AM to the CGRP receptor suppresses fMLP-induced upregulation of CD11b expression of human neutrophils by increasing intracellular cAMP levels. AM may play an important role in the regulation of inflammatory processes, especially in the binding of neutrophils to vascular endothelial cells and subsequent neutrophil emigration evident in acute pulmonary inflammation.

背景与目标： ：在这项研究中，我们研究了肾上腺髓质素（AM）对fMLP介导的人类嗜中性粒细胞活化的影响。 AM部分但显着抑制fMLP诱导的CD11b表达上调。 CGRP [8-37]是一种CGRP受体拮抗剂，完全抑制了AM对fMLP诱导的CD11b表达上调的抑制作用。 AM显着增加了中性粒细胞和腺苷酸环化酶抑制剂SQ-22,536和PKA抑制剂KT-5720中的cAMP含量，从而显着阻断了AM对fMLP诱导的CD11b表达上调的抑制作用。这项研究表明，AM与CGRP受体的结合可通过增加细胞内cAMP水平来抑制fMLP诱导的人类嗜中性粒细胞CD11b表达的上调。 AM在炎症过程的调节中可能起重要作用，特别是在嗜中性粒细胞与血管内皮细胞的结合以及随后在急性肺部炎症中明显的嗜中性粒细胞迁移中。

BACKGROUND & AIMS: :Receptors involved in the phagocytosis of microorganisms under nonopsonic conditions have been little studied in neutrophils. Complement receptor type 3 (CR3) is a pattern recognition receptor able to internalize zymosan and C3bi-coated particles. We report that Abs directed against CR3 strongly inhibited nonopsonic phagocytosis of Mycobacterium kansasii in human neutrophils. In these cells CR3 has been found associated with several GPI-anchored proteins localized in cholesterol-rich microdomains (rafts) of the plasma membrane. Cholesterol sequestration by nystatin, filipin, or beta-cyclodextrin as well as treatment of neutrophils with phosphatidylinositol phospholipase C to remove GPI-anchored proteins from the cell surface markedly inhibited phagocytosis of M. kansasii, without affecting phagocytosis of zymosan or serum-opsonized M. kansasii. Abs directed against several GPI-anchored proteins inhibited phagocytosis of M. kansasii, but not of zymosan. N:-acetyl-D-glucosamine, which is known to disrupt interactions between CR3 and GPI proteins, also strongly diminished phagocytosis of these mycobacteria. In conclusion, phagocytosis of M. kansasii involved CR3, GPI-anchored receptors, and cholesterol. In contrast, phagocytosis of zymosan or opsonized particles involved CR3, but not cholesterol or GPI proteins. We propose that CR3, when associated with a GPI protein, relocates in cholesterol-rich domains where M. kansasii are internalized. When CR3 is not associated with a GPI protein, it remains outside of these domains and mediates phagocytosis of zymosan and opsonized particles, but not of M. kansasii.

背景与目标： ：在中性粒细胞中很少研究非调理条件下参与微生物吞噬作用的受体。 3型补体受体（CR3）是一种模式识别受体，能够内化酵母聚糖和C3bi包被的颗粒。我们报道，针对CR3的Abs强烈抑制人嗜中性粒细胞中堪萨斯分枝杆菌的非调理吞噬作用。在这些细胞中，已发现CR3与定位在质膜富含胆固醇的微区（筏）中的几种GPI锚定蛋白有关。制霉菌素，菲利普林或β-环糊精的胆固醇隔离以及用磷脂酰肌醇磷脂酶C处理嗜中性白细胞从细胞表面去除GPI锚定的蛋白显着抑制了堪萨斯支原体的吞噬作用，而又不影响酵母聚糖的吞噬作用或血清调理的M.堪萨斯州。针对几种GPI锚定蛋白的Abs抑制了堪萨斯分枝杆菌的吞噬作用，但抑制了酵母聚糖的吞噬作用。众所周知，N：-乙酰基-D-葡萄糖胺会破坏CR3和GPI蛋白之间的相互作用，也大大降低了这些分枝杆菌的吞噬作用。总之，堪萨斯分枝杆菌的吞噬作用涉及CR3，GPI锚定受体和胆固醇。相反，酵母聚糖或调理过的颗粒的吞噬作用涉及CR3，但不涉及胆固醇或GPI蛋白。我们建议CR3，当与GPI蛋白相关时，重新定位在坎萨西分枝杆菌被内化的富含胆固醇的域中。当CR3与GPI蛋白不相关时，它将保留在这些结构域之外，并介导酵母聚糖和调理过的颗粒的吞噬作用，而不是堪萨斯分枝杆菌的吞噬作用。

BACKGROUND & AIMS: :Asbestos exposure causes diffuse interstitial pulmonary fibrosis. Since alveolar epithelial cell injury is hypothesized to precede the fibrotic response in asbestosis, we investigated whether asbestos, either alone or in conjunction with neutrophils (PMNs), injured cultured human pulmonary epithelial cells (HPECs). HPEC cytotoxicity was assessed with a standard 51chromium release assay after a 16-hour incubation with asbestos and PMNs. Negligible HPEC cytotoxicity was observed after incubation with either amosite asbestos (500 micrograms/ml) or PMNs alone in serum-free media. However, incubation with both asbestos and PMNs caused significant HPEC injury, which was asbestos dose-dependent; causing 25% +/- 4% detachment and 52% +/- 8% 51chromium release with 500 micrograms/ml asbestos. The cytotoxic effects of asbestos plus PMNs were nearly completely attenuated with serum (20%) or catalase (100 micrograms/ml) but were not prevented with scavengers of superoxide anion, hydroxyl radical, or hypochlorous acid. A role for hydrogen peroxide (H2O2) in mediating HPEC injury was also suggested by the demonstration of asbestos-induced generation of H2O2 by PMNs. Furthermore, H2O2 alone (10(-4)mol/L) caused significant HPEC damage. Intimate contact between asbestos-activated PMNs and HPECs was a necessary requirement for PMN-mediated HPEC cytotoxicity. These data suggest that pulmonary epithelial cell injury is mediated in part by H2O2 release from asbestos-activated PMNs as well as intimate contact between the epithelial cell, PMNs, and asbestos.

背景与目标： ：石棉暴露引起弥漫性间质性肺纤维化。由于假设肺泡上皮细胞损伤先于石棉沉滞中的纤维化反应，所以我们调查了石棉（单独还是与中性粒细胞（PMN）联合使用）是否损伤了培养的人肺上皮细胞（HPEC）。与石棉和PMN孵育16小时后，使用标准的51铬释放分析法评估HPEC的细胞毒性。在无血清培养基中与铁石棉（500微克/毫升）或单独的PMN孵育后，观察到的HPEC细胞毒性可忽略不计。但是，与石棉和PMNs一起孵育会引起严重的HPEC损伤，这是石棉剂量依赖性的；导致25％/-4％的脱离和52％/-8％的51铬释放，含500微克/毫升的石棉。血清（20％）或过氧化氢酶（100微克/毫升）几乎完全减弱了石棉和PMN的细胞毒性作用，但用超氧阴离子，羟基自由基或次氯酸清除剂却无法阻止。 PMN在石棉诱导的H2O2生成中的作用也暗示了过氧化氢（H2O2）在介导HPEC损伤中的作用。此外，仅H2O2（10（-4）mol / L）会对HPEC造成严重损害。石棉激活的PMN和HPEC之间的紧密接触是PMN介导的HPEC细胞毒性的必要条件。这些数据表明，肺部上皮细胞损伤部分是由石棉激活的PMN释放的H2O2以及上皮细胞，PMN和石棉之间的紧密接触所介导的。

BACKGROUND & AIMS: :Peripheral blood polymorphonuclear neutrophils (PMN) suppressed the induction of PBL lymphokine-activated killer (LAK) function by rIL-2 in vitro. The suppression depended on the concentration of PMN in the IL-2 culture, and required intact PMN. However, PMN did not require treatment with immunoregulators such as IL-2, LPS, or TNF to express the suppressive activity, and no direct contact with PBL was needed for the suppression. Addition of anti-TNF antibodies had no effect on the suppression, suggesting that no endogenous TNF in the culture was involved in the suppression. PMN did not inhibit LAK function by preventing utilization of IL-2 by PBL or by selective depletion of NKH-1+ cells which constitute the majority of LAK precursors in PBL. The suppression was reversed by superoxide dismutase but not by catalase, suggesting that superoxide anion, not hydrogen peroxide, was involved in the suppression. No other suppressive factor was detectable in PMN culture supernates. Our results of PMN regulating LAK induction in vitro suggest that PMN may have a role in determining the outcome of immunotherapy with IL-2 in vivo.

背景与目标： ：外周血多形核中性粒细胞（PMN）在体外抑制了rIL-2对PBL淋巴因子激活的杀手（LAK）功能的诱导。抑制取决于IL-2培养物中PMN的浓度，并需要完整的PMN。但是，PMN不需要用免疫调节剂（例如IL-2，LPS或TNF）进行治疗即可表达抑制活性，并且不需要直接与PBL接触即可进行抑制。添加抗TNF抗体对抑制没有影响，表明培养物中没有内源性TNF参与抑制。 PMN不能通过阻止PBL或通过选择性消耗NKH-1细胞（构成PBL中大部分LAK前体）来利用IL-2来抑制LAK功能。抑制作用被超氧化物歧化酶逆转，但不被过氧化氢酶逆转，表明抑制作用涉及超氧阴离子而不是过氧化氢。在PMN培养上清液中未检测到其他抑制因子。我们的PMN在体外调节LAK诱导的结果表明，PMN可能在体内确定IL-2免疫疗法的结果中具有作用。

BACKGROUND & AIMS: :Nonopsonic interaction of host immune cells with pathogens is an important first line of defense. We hypothesized that nonopsonic recognition between type III group B streptococcus and human neutrophils would occur and that the interaction would be sufficient to trigger neutrophil activation. By using a serum-free system, it was found that heat-killed type III group B streptococci bound to neutrophils in a rapid, stable, and inoculum-dependent manner that did not result in ingestion. Transposon-derived type III strain COH1-13, which lacks capsular polysaccharide, and strain COH1-11 with capsular polysaccharide lacking terminal sialic acid demonstrated increased neutrophil binding, suggesting that capsular polysaccharide masks an underlying binding site. Experiments using monoclonal antibodies to complement receptor 1 and to the I domain or lectin site of complement receptor 3 did not inhibit binding, indicating that the complement receptors used for ingestion of opsonized group B streptococci were not required for nonopsonic binding. Nonopsonic binding resulted in rapid activation of cellular p38 and p44/42 mitogen-activated protein kinases. This interaction was not an effective trigger for superoxide production but did promote release of the proinflammatory cytokine interleukin-8. The release of interleukin-8 was markedly suppressed by the p38 mitogen-activated protein kinase inhibitor SB203580 but was only minimally suppressed by the mitogen-activated protein/extracellular signal-regulated kinase inhibitor PD98059. Thus, nonopsonic binding of type III group B streptococci to neutrophils is sufficient to initiate intracellular signaling pathways and could serve as an arm of innate immunity of particular importance to the immature host.

背景与目标： ：宿主免疫细胞与病原体的正典相互作用是重要的第一道防线。我们假设III型B组链球菌与人类嗜中性粒细胞之间会发生非调理性识别，并且这种相互作用足以触发嗜中性粒细胞的活化。通过使用无血清系统，发现热灭活的III型B组链球菌以快速，稳定和接种物依赖性的方式与嗜中性粒细胞结合，而不会导致摄入。缺乏荚膜多糖的转座子衍生的III型菌株COH1-13，以及缺少末端唾液酸的荚膜多糖的COH1-11菌株显示嗜中性粒细胞结合增加，表明荚膜多糖掩盖了潜在的结合位点。使用针对补体受体1以及补体受体3的I结构域或凝集素位点的单克隆抗体进行的实验不会抑制结合，这表明用于非调理结合不需要摄入调理B组链球菌所使用的补体受体。非光子结合导致细胞p38和p44 / 42丝裂原活化蛋白激酶的快速活化。这种相互作用不是产生超氧化物的有效触发器，但确实促进了促炎细胞因子白细胞介素8的释放。 p38丝裂原活化的蛋白激酶抑制剂SB203580明显抑制了白细胞介素8的释放，但丝裂原活化的蛋白/细胞外信号调节的激酶抑制剂PD98059仅最小程度地抑制了白介素8的释放。因此，III型B组链球菌与嗜中性粒细胞的非调理结合足以启动细胞内信号传导途径，并可作为对未成熟宿主特别重要的先天免疫的一部​​分。

BACKGROUND & AIMS: 1. Inositol hexakisphosphate (InsP6) is a ubiquitous and abundant cytosolic inositol phosphate that has been reported to prime human neutrophils for enhanced agonist-stimulated superoxide anion generation. This led to the proposal that the release of InsP6 from necrotic cells may augment the functional responsiveness of neutrophils at an inflammatory focus. The aim of this study was to examine whether the functional effects of InsP6 in neutrophils are receptor-mediated and establish the magnitude of this priming effect relative to other better characterized priming agents. 2. Analysis of [3H]-InsP6 binding to human neutrophil membranes in 20 mM Tris, 20 mM NaCl, 100 mM KCl, 5 mM EDTA (pH 7.7) buffer using 0.1 mg ml-1 membrane protein and 2.5 nM [3H]-InsP6 (90 min, 4 degrees C), demonstrated specific low affinity [3H]-InsP6 binding that was non-saturable up to a radioligand concentration of 10 nM. 3. [3H]-InsP6 displacement by InsP6 gave a Hill coefficient of 0.55 and best fitted a two-site logistic model (53% KD 150 nM, 47% KD 5 microM). [3H]-InsP6 binding also displayed low (3 fold) selectivity for InsP6 over Ins(1,3,4,5,6)P5. 4. The specific [3H]-InsP6 binding displayed a pH optimum of 8, was abolished by pre-boiling the membranes, and was enhanced by Ca2+, Mg2+ and Na+. 5. In incubations with intact neutrophils, where high levels of specific [3H]-LTB4 binding was observed, no [3H]-InsP6 binding could be identified. 6. Preincubation of neutrophils with 100 microM InsP6 had no effect on resting cell morphology, but caused a minor and transient (maximal at 30 s) enhancement of (0.1 nM) fMLP-induced shape change (% cells shape changedfMLP 53 +/- 3%, fMLP+InsP6 66 +/- 4%).

Similarly, InsP6 (100 microM, 30 s) had no effect on basal superoxide anion generation and, compared to lipopolysaccharide (LPS, 100 ng ml-1, 60 min), tumour necrosis factor-alpha (TNF alpha, 200 u ml-1, 30 min) or platelet-activating factor (PAF, 100 nM, 5 min) caused only a small enhancement of 100 nM fMLP-stimulated superoxide anion generation (fold-increase in superoxide anion generation over fMLP aloneInsP6 1.8 +/- 0.3, LPS 6.8 +/- 0.6, TNF alpha 5.2 +/- 0.7, PAF 5.8 +/- 0.6). 7. While these data support the presence of a specific, albeit low affinity, [3H]-InsP6 binding site in human neutrophil membrane preparations, the lack of binding to intact cells implies that the functional effects of InsP6 (ie. enhanced fMLP-stimulated superoxide anion generation and shape change) are not receptor-mediated.

背景与目标： 1.肌醇六磷酸肌醇（InsP6）是一种普遍存在且富含胞质的肌醇磷酸肌醇，据报道可引发人类嗜中性粒细胞，以增强激动剂刺激的超氧阴离子生成。这导致了一个提议，即从坏死细胞释放InsP6可能会增强炎症中心的嗜中性粒细胞的功能反应性。这项研究的目的是检查InsP6在嗜中性粒细胞中的功能作用是否是受体介导的，并确定这种引发作用相对于其他更好表征的引发剂的强度。 2.在使用0.1 mg ml-1膜蛋白和2.5 nM [3H]-的20 mM Tris，20 mM NaCl，100 mM KCl，5 mM EDTA（pH 7.7）缓冲液中分析[3H] -InsP6与人嗜中性白细胞膜的结合。 InsP6（90分钟，4摄氏度）显示出特定的低亲和力[3H] -InsP6结合，直到10nM的放射性配体浓度，该结合都是不饱和的。 3.用InsP6置换[3H] -InsP6的希尔系数为0.55，最适合两点逻辑模型（53％KD 150 nM，47％KD 5 microM）。与Ins（1,3,4,5,6）P5相比，[3H] -InsP6结合对InsP6的选择性也低（3倍）。 4.特定的[3H] -InsP6结合表现出最适的pH值为8，已通过预煮膜来消除，并被Ca2，Mg2和Na增强。 5.在与完整的中性粒细胞温育中，观察到高水平的特异性[3H] -LTB4结合，无法鉴定到[3H] -InsP6结合。 6.中性粒细胞与100 microM InsP6的预温育对静息细胞形态没有影响，但引起（0.1 nM）fMLP诱导的形状变化的轻微和短暂的增强（最大30 s）（％细胞形状变化fMLP 53 /-3％ ，fMLP InsP6 66 /-4％）。

同样，InsP6（100 microM，30 s）对基础超氧阴离子的生成没有影响，并且与脂多糖（LPS，100 ng ml-1，60分钟） ），肿瘤坏死因子-α（TNFα，200 u ml-1，30分钟）或血小板活化因子（PAF，100 nM，5分钟）仅引起100 nM fMLP刺激的超氧阴离子生成的少量增强（折叠-仅通过fMLP可以增加超氧阴离子的产生-InsP6 1.8 /-0.3，LPS 6.8 /-0.6，TNFα5.2 /-0.7，PAF 5.8 /-0.6）。 7.虽然这些数据支持人嗜中性粒细胞膜制剂中存在特定的，尽管亲和力很低的[3H] -InsP6结合位点，但缺乏与完整细胞结合的能力暗示了InsP6的功能作用（即增强的fMLP刺激超氧阴离子的产生和形状变化）不是受体介导的。

BACKGROUND & AIMS: :The MAPK family member JNK/stress-activated MAPK (SAPK) is involved in extracellular stress and proinflammatory cytokine responses, including production of cytokines such as IL-12. The JNK1 and 2 isoforms are widely expressed, but JNK3 is largely restricted to tissues of the brain, testis, and heart. In this study, we focus on mouse neutrophils, a cell type in which JNK/SAPK expression and activity has been given little study. We used Western blot analysis to examine expression patterns of JNK/SAPK in wild-type and JNK2-/- polymorphonuclear leukocytes (PMN). Surprisingly, neutrophils displayed a major deficiency in JNK1 expression, in contrast to macrophages that expressed high levels of both JNK1 and JNK2 MAPK. JNK1 expression was steadily reduced during the neutrophil maturation in bone marrow. We used PMN infection with the protozoan parasite Toxoplasma gondii to determine whether neutrophil JNK2 was functional. The parasite induced rapid JNK2 phosphorylation and intracellular FACS staining demonstrated preferential activation in infected neutrophils. Use of JNK2-/- neutrophils revealed that this MAPK family member was required for PMN IL-12p40 and CCL2/MCP-1 production. The chemotactic response displayed a minor JNK2 dependence but phagocytosis and oxidative burst activity did not require this MAPK. These findings are important because they demonstrate 1) a previously unrecognized unusual JNK expression pattern in mouse neutrophils, 2) JNK2 in PMN is activated by Toxoplasma invasion, and 3) a requirement for JNK2 in PMN IL-12p40 and CCL2/MCP-1 production in response to a microbial pathogen.

背景与目标： MAPK家族成员JNK /应激激活MAPK（SAPK）与细胞外应激和促炎性细胞因子反应有关，包括细胞因子如IL-12的产生。 JNK1和2亚型广泛表达，但JNK3在很大程度上限于大脑，睾丸和心脏的组织。在这项研究中，我们集中于小鼠嗜中性粒细胞，这是一种细胞类型，其中对JNK / SAPK表达和活性的研究很少。我们使用蛋白质印迹分析来检查JNK / SAPK在野生型和JNK2-/-多形核白细胞（PMN）中的表达模式。出乎意料的是，嗜中性粒细胞在JNK1表达中表现出主要缺陷，这与表达高水平JNK1和JNK2 MAPK的巨噬细胞相反。在骨髓中性粒细胞成熟过程中，JNK1表达稳定降低。我们使用原生动物寄生虫弓形虫PMN感染来确定中性粒细胞JNK2是否起作用。寄生虫诱导的快速JNK2磷酸化和细胞内FACS染色显示在感染的中性粒细胞中优先激活。使用JNK2-/-中性粒细胞表明该MAPK家族成员是PMN IL-12p40和CCL2 / MCP-1生产所必需的。趋化反应显示出较小的JNK2依赖性，但吞噬作用和氧化爆发活性不需要此MAPK。这些发现很重要，因为它们证明了1）小鼠嗜中性粒细胞中以前无法识别的异常JNK表达模式，2）PMN中的JNK2被弓形虫入侵激活，3）PMN IL-12p40和CCL2 / MCP-1生产中对JNK2的要求对微生物病原体的反应。

BACKGROUND & AIMS: :The expression and production of cytokines by cells of the innate immune system, including monocytes/macrophages, dendritic and NK cells, play a critical role not only in defensive and inflammatory but also in immunoregulatory and anti-/pro-tumoral processes. Studies performed in the last years have well ascertained that polymorphonuclear neutrophils can also be induced to express and produce chemokines, proinflammatory, anti-inflammatory, immunoregulatory, angiogenic and fibrogenic cytokines, as well as ligands belonging to the TNF superfamily. Among the latter group of molecules, B-cell-activating factor (BAFF)/B lymphocyte stimulator (BLyS), known to be essential for B lymphocyte homeostasis and related pathologies, has recently been identified as one of the factors potentially expressed by human neutrophils. The addition of this novel TNF superfamily member, and more recently also of the closely related "A Proliferation-Inducing Ligand" (APRIL), to the list of cytokines produced by neutrophils not only testifies to the continuous growth of this area of investigation, but also implies the involvement of neutrophils in B-cell-dependent autoimmune diseases and tumors.

背景与目标： 天然免疫系统细胞（包括单核细胞/巨噬细胞，树突状细胞和NK细胞）表达和产生细胞因子，不仅在防御和炎症中起着关键作用，而且在免疫调节和抗肿瘤/促肿瘤过程中也起着至关重要的作用。近年来进行的研究已经确定，多形核中性粒细胞也可以被诱导表达和产生趋化因子，促炎，抗炎，免疫调节，血管生成和纤维生成细胞因子，以及属于TNF超家族的配体。在后一组分子中，B细胞活化因子（BAFF）/ B淋巴细胞刺激物（BLyS）是已知的B淋巴细胞稳态和相关病理必不可少的，最近已被确定为人类嗜中性粒细胞潜在表达的因子之一。在嗜中性粒细胞产生的细胞因子列表中增加了这种新的TNF超家族成员，以及最近与之密切相关的“增殖诱导配体”（APRIL），不仅证明了这一研究领域的持续发展，而且证明了这一领域的持续发展。还暗示嗜中性粒细胞参与B细胞依赖性自身免疫性疾病和肿瘤。