Effect of priming in subpopulations of peripheral neutrophils from patients with chronic periodontitis

Background: Patients with periodontal disease are reported to generate more reactive oxygen species (ROS) than matched controls, suggesting increased inflammatory defense activity. The purpose of this study is to determine whether there are subpopulations of peripheral neutrophils in patients with chronic periodontitis (CP) that generate different levels of intracellular ROS when primed with tumor necrosis factor‐α (TNF‐α) or the chemokine interleukin‐8 (IL‐8, CXCL8) compared to controls. Methods: Venous blood was collected from 13 patients with CP despite careful maintenance over 2 to 8 years and from 13 healthy age‐ and sex‐matched controls. Neutrophils were separated from whole blood over a Percoll gradient and then activated via the Fcγ receptor with opsonized Staphylococcus aureus after priming with TNF‐α or IL‐8. The samples were analyzed by flow cytometry using the fluorescent probe dihydrorhodamine 123. Generation of ROS was measured as the intensity of fluorescence (IFL). Results: Two subpopulations were found in both patients and controls: one with low and one with high generation of IFL. The subpopulation with high generation of IFL in patients with CP was more responsive to IL‐8 (P <0.05) than the same subpopulation in healthy controls. No other differences in generation of ROS or priming effects were found between patients with CP and controls. Generation of ROS was dependent on nicotinamide adenine dinucleotide phosphate oxidase, and the intracellular ROS was primarily the oxygen anion. Conclusion: Patients with CP had a subpopulation of peripheral neutrophils that were more responsive to IL‐8 priming than controls.     

Infections and neutrophils in the pathogenesis of bronchiolitis obliterans syndrome in children after allogeneic stem cell transplantation

It is plausible that infections post‐hematopoietic SCT play a role in the pathogenesis of BOS. A prospective study for children with history, questionnaire, examination, PFTs, and blood counts at one, three, six, nine, 12, 18, and 24 months post‐SCT was conducted. Between September 2009 and September 2011 (n = 39), six developed BOS at 200 days (range 94–282), three patients had probable clinical respiratory infection, and all six had higher neutrophil count compared to non‐BOS patients (4.7 vs. 2.4 at three months and 6.3 vs. 2.9 at six months ×10⁹/L, p = 0.03). Contribution of clinical and subclinical infection needs to be considered in the pathogenesis of BOS.     

Mechanisms of neutrophil death in human immunodeficiency virus-infected patients: role of reactive oxygen species, caspases and map kinase pathways

Neutrophils from human immunodeficiency virus-positive (HIV+) patients have an increased susceptibility to undergo programmed cell death (PCD), which could explain neutropenia during advanced disease. In this work, key steps of PCD have been evaluated in neutrophils from HIV+ patients. The role of caspase-3, caspase-8, mitogen activated protein kinase (MAPK) and reactive oxygen species (ROS) was analysed. Spontaneous neutrophil death is dependent upon caspase-3 but independent of caspase-8, suggesting that the intrinsic pathway is involved as a pathogenic mechanism of PCD. Inhibition of ROS decreased spontaneous PCD and caspase-3 hydrolysis, connecting oxidative stress and caspase-3 activation with neutrophil PCD in HIV-infected patients. Additionally, an increased neutrophil death was observed in HIV+ patients, following inhibition of p38 MAPK, suggesting a role for p38 MAPK in cell survival during the disease. We conclude that oxidative stress secondary to HIV infection can accelerate neutrophil death.     

The intriguing normal acute inflammatory response in mice lacking vimentin

Neutrophils express only two intermediate filament proteins, vimentin and, to a lesser extent, lamin B. Lamin B mutant mice die shortly after birth; however, mice lacking vimentin (vim(-/-)) develop and reproduce normally. Herein, we investigate for the first time the role of vimentin in general inflammation in vivo and in neutrophil functions ex vivo. Using the murine air pouch model, we show that the inflammatory response induced by lipopolysaccharide, interleukin-21 or carageenan is, intriguingly, uncompromised in vim(-/-) mice and that neutrophil functions are not altered ex vivo. Our results suggest that vimentin is dispensable for the establishment of an acute inflammatory response in vivo. In addition, based on several criteria presented in this study, one has to accept the existence of a very complex compensatory mechanism to explain the intriguing normal inflammatory response in absence of vimentin.     

Activation of granulocytes by direct interaction with dendritic cells

Granulocytes from human peripheral blood were co-cultured with conventional dendritic cells (cDC) or plasmacytoid DCs (pDC) to examine the effects of DCs on the activation or function of granulocytes. After co-culture of granulocytes with DCs, expression of the activation markers of granulocytes (CD63 and CD64) was up-regulated, and increased expression of CD50, the activation marker and ligand for CD209 (DC-SIGN) was also observed. The interaction of granulocytes with DCs was visualized as the cluster where DCs, especially cDCs, were surrounded by granulocytes to form a 'rosette'. After co-culture of granulocytes with cDCs, the secretion of elastase from granulocytes was enhanced significantly when examined cytohistochemically and by enzyme-linked immunosorbent assay. An increase in myeloperoxidase (another activation index of granulocytes) was also observed after co-culture with DCs. These findings suggest the functional and phenotypical activation of granulocytes by interaction with DCs. Furthermore, we examined the involvement of adhesion molecules in the granulocyte-DC interaction, and found that CD209 participates to some extent in this interaction.     

中性粒细胞与淋巴细胞比值和急性冠状动脉综合征的相关性及其对急性心肌梗死的诊断价值

目的 探讨中性粒细胞与淋巴细胞比值（NLR）与急性冠状动脉综合征（ACS）患者冠脉严重程度之间的关 系,并分析NLR对急性心肌梗死的诊断价值。方法 回顾性分析2016年11月—2017年3月于我院行冠状动脉造影 检查的259例ACS患者的临床资料,其中不稳定型心绞痛组（UA组）148例、急性非ST段抬高型心肌梗死组（NSTEMI组）46例、急性ST段抬高型心肌梗死组（STEMI组）65例。比较3组的白细胞计数（WBC）、中性粒细胞计数（N）、淋巴细胞计数（L）、NLR、高敏C反应蛋白（hs-CRP）、心肌酶谱、血脂、血糖、肝功能等指标的差异。根据冠状动脉造影结果将患者按照冠状动脉病变支数分为单支病变组（70例）、双支病变组（70例）和三支病变组（119例）,比较不同病变支数组间炎性细胞的差异。同时绘制NLR的受试者工作特征（ROC）曲线,评价NLR对急性心肌梗死的预测价值。结果 UA组、NSTEMI组及STEMI组间WBC、N、NLR、hs-CRP、cTnT、CK、CK-MB水平依次升高,STEMI组L值低于UA组,差异有统计学意义（P＜0.05）;WBC、NLR随冠状动脉病变支数的增加而呈上升趋势,但仅三支病变组与单支病变组间差异有统计学意义;NLR诊断急性心肌梗死的ROC曲线下面积（AUC）为0.865（95%CI：0.814~0.916）。当NLR=4.22时,其对急性心肌梗死的诊断效能最高,敏感度为72.1％,特异度为95.3％。结论 NLR水平与ACS患者病情严重程度呈正相关,并对诊断急性心肌梗死具有较高的诊断价值,可作为ACS患者病情的预测因素。     

Stimulatory response of neutrophils from periodontitis patients with periodontal pathogens

OBJECTIVE: Neutrophils play a crucial role in the defense of invading bacteria by releasing biologically active molecules. The response of peripheral blood neutrophils was studied in periodontitis-affected patients and in healthy controls towards stimulation to Porphyromonas gingivalis (Pg) and Actinobacillus actinomycetemcomitans (Aa) extracts. MATERIALS AND METHODS: Peripheral venous blood was drawn from 23 adult patients with moderate to advanced chronic periodontitis (probing depth >or=5 mm, attachment loss >or=3 mm), and 30 healthy volunteers. Neutrophil response followed by metalloproteinase-9 (MMP-9) and interleukin-8 (IL-8) secretion was assayed by zymography and enzyme-linked immunosorbent assay, respectively, on both whole blood and purified neutrophils. In addition to periodontal pathogen extracts, known stimulating agents were tested, such as Escherichia coli-lipopolysaccharide (LPS), phytohemagglutinin, and zymosan A. RESULTS: Neutrophil response, expressed as a secretion ratio under stimulated and non-stimulated conditions, measured in whole blood, showed no differences between periodontitis and healthy controls. Instead, in purified neutrophils from patients, MMP-9 exhibited a significantly higher secretion ratio with LPS and Pg (1.5- to 2-fold), whereas IL-8 showed a larger increase in secretion ratio (3- to 7-fold) in the presence of Pg, Aa, LPS, and zymosan A. CONCLUSION: Peripheral neutrophils of periodontitis-affected patients are more reactive as suggested by their significantly higher response toward periodontal pathogen extracts and other stimulating agents.     

馥感啉口服液联合脾氨肽治疗小儿反复呼吸道感染的临床研究

目的探讨馥感啉口服液联合脾氨肽治疗小儿反复呼吸道感染的临床疗效。方法选取2015年9月—2016年3月在郑州市第三人民院治疗的反复呼吸道感染患儿98例,随机分为对照组(49例)和治疗组(49例)。对照组口服脾氨肽口服冻干粉,2 mg/次,1次/d;治疗组在对照组基础上口服馥感啉口服液,10 mL/次,1~3岁患儿3次/d,4~6岁4次/d,7~12岁5次/d。两组患儿均经过3个月治疗。观察两组患儿临床疗效,比较治疗前后两组患儿临床症状改善时间、血清学指标、白细胞分类和白三烯D4(LTD4)水平。结果治疗后,对照组临床总有效率为81.63%,显著低于治疗组的95.92%。两组比较差异具有统计学意义(P0.05)。治疗后,治疗组发热、咳嗽和肺部啰音消失时间均显著短于对照组(P0.05)。治疗后,两组血清IL-1β、IL-6、和肿瘤坏死因子-α(TNF-α)水平均显著降低,胰岛素1号增长因子(IGF-1)、干扰素-γ(IFN-γ)和25-OH维生素D_3[25-(OH)D_3]水平均显著升高,同组比较差异具有统计学意义(P0.05);且治疗组上述血清学指标改善程度明显好于对照组(P0.05)。治疗后,两组鼻咽分泌物中中性粒细胞、嗜酸性粒细胞和LTD4水平均显著降低,单核巨噬细胞显著升高,同组比较差异具有统计学意义(P0.05);且治疗组上述指标改善后水平显著优于对照组(P0.05)。结论馥感啉口服液联合脾氨肽治疗小儿反复呼吸道感染可有效改善临床症状,降低炎症水平,具有一定的临床推广应用价值。     

Proteolysis of the endothelial cell protein C receptor by neutrophil proteinase 3

BACKGROUND: The endothelial cell protein C receptor (EPCR) presents protein C to the thrombin:thrombomodulin complex on the endothelium of large vessels, and enhances the generation of activated protein C (APC) and activation of protease-activated receptor-1. A previous report has demonstrated binding of soluble (s) EPCR to activated neutrophils via surface proteinase 3 (PR3). METHODS: We now report further characterization of this interaction. Activated neutrophils and purified PR3 both decrease endothelial cell (EC) surface EPCR, suggestive of its proteolysis. RESULTS: When added to purified recombinant sEPCR, PR3 produced multiple cleavages, with early products including 20 kDa N-terminal and C-terminal (after Lys(176)) fragments. The binding of active site blocked PR3 to sEPCR was studied by surface plasmon resonance. Estimates of the K(D) of 18.5-102 nM were obtained with heterogeneous binding, suggestive of more than a single interaction site. CONCLUSIONS: This work demonstrates PR3 binding to and proteolysis of EPCR and suggests a mechanism by which anticoagulant and cell protective pathways can be down-regulated during inflammation.