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Sample records for significant tumor-infiltrating neutrophils

  1. Strong Prognostic Value of Tumor-infiltrating Neutrophils and Lymphocytes Assessed by Automated Digital Image Analysis in Early Stage Cervical Cancer

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    Carus, Andreas; Donskov, Frede; Switten Nielsen, Patricia

    2014-01-01

    INTRODUCTION Manual observer-assisted stereological (OAS) assessments of tumor-infiltrating neutrophils and lymphocytes are prognostic, accurate, but cumbersome. We assessed the applicability of automated digital image analysis (DIA). METHODS Visiomorph software was used to obtain DIA densities...... with the prognostically strongest manual OAS assessments in the peritumoral compartment. In multivariate analysis, CD66b and CD8 densities, assessed by DIA, and regional lymph node metastases were independent predictors of RFS, while CD163 density and FIGO stage were not. The CD66b/CD8 tumorassociated neutrophil...

  2. Clinicopathological and prognostic significance of FOXP3+ tumor infiltrating lymphocytes in patients with breast cancer: a meta-analysis

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    Jiang, Daqing; Gao, Zhaohua; Cai, Zhengang; Wang, Meixian; He, Jianjun

    2015-01-01

    The prognostic significance of FOXP3+ tumor-infiltrating lymphocytes (TILs) in patients with breast cancer remains controversial. The aims of our meta-analysis are to evaluate its association with clinicopathological characteristics and prognostic significance in patients with breast cancer. PubMed, Embase, Cochrane Database and the Ovid Database were systematically searched (up to April 2015). The meta-analysis was performed using hazard ratio (HR), odds ratio (OR) and 95 % confidence intervals (CI) as effect measures. Using the random-effects model, statistical analysis was performed using Stata software, version 12.0. Seventeen studies including 8277 patients with breast cancer were analyzed. The meta-analysis indicated that the incidence difference of FOXP3+ TILs was significant when comparing the lymph node positive group to negative group (OR = 1.305, 95 % CI [1.071, 1.590]), the histological grade III group to the I–II group (OR = 3.067, 95 % CI [2.288, 4.111]), the ER positive group to the negative group (OR = 0.435, 95 % CI [0.287, 0.660]), the PR positive group to the negative group (OR = 0.493, 95 % CI [0.296, 0.822]), the HER2 positive group to the negative group (OR = 1.896, 95 % CI [1.335, 2.692]), the TNBC group to the non TNBC group (OR = 2.456, 95 % CI [1.801, 3.348]). The detection of FOXP3+ TILs was significantly correlated with the recurrence-free survival (RFS) of patients (HR = 1.752, 95 % CI [1.188–2.584]) and the overall survival (OS) of patients (HR =1.447, 95 % CI [1.037–2.019]). Our meta-analysis demonstrates that the presence of high levels of FOXP3+ TILs is associated with prognosis for breast cancer patients and predicts lymph node metastasis, hormone receptor and HER-2 status

  3. Immunohistochemical analysis and prognostic significance of PD-L1, PD-1, and CD8+ tumor-infiltrating lymphocytes in Ewing's sarcoma family of tumors (ESFT).

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    Machado, Isidro; López-Guerrero, Jose Antonio; Scotlandi, Katia; Picci, Piero; Llombart-Bosch, Antonio

    2018-05-01

    Ewing's sarcoma family of tumors (ESFT) are aggressive neoplasms with scant tumor-infiltrating lymphocytes. We analyzed the immunohistochemical (IHC) expression of PD-L1 and PD-1 and their prognostic significance in clinically localized neoplasms in a cohort of 370 ESFT. Slides prepared from tissue microarrays were stained for PD-L1, PD-1, and CD8. Membranous/cytoplasmic staining over 5% of tumor cells was regarded as positive for PD-L1 and PD-1. Prognostic analysis was done considering only clinically localized tumors (n = 217). PD-L1 expression was present in 19% of ESFT, while PD-1 was expressed in 26%. Forty-eight percent of tumors were negative and 12% were positive for both PD-L1 and PD-1. Metastatic tumors displayed higher expression of PD-L1 (p < 0.0001). Histological subtypes were not correlated with PD-L1 or PD-1 positivity. ESFT with elevated proliferation index (Ki-67) were associated with higher PD-L1 expression (p = 0.049). Regarding prognosis, no significant association was found between PD-L1 expression and progression-free survival (PFS) or overall survival (OS), whereas lack of PD-1 expression in tumor cells was correlated with both poor PFS (p = 0.02) and poor OS (p = 0.004). Tumor-infiltrating CD8(+) T lymphocytes were observed in 15.4% of ESFT with informative results (347 tumors). No correlation was found between tumor-infiltrating CD8(+) T lymphocytes and ESFT histological subtypes, tumor location, or PD-1 and PD-L1 expression, nor with PFS (p = 0.473) or OS (p = 0.087). PD-L1 expression was not significantly related to prognosis. PD-1 was expressed in 26% of ESFT tumor cells and may have prognostic and therapeutic implications. CD8 expression in tumor-infiltrating lymphocytes was not related to prognosis.

  4. Differential pattern and prognostic significance of CD4+, FOXP3+ and IL-17+ tumor infiltrating lymphocytes in ductal and lobular breast cancers

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    Droeser, Raoul; Zlobec, Inti; Kilic, Ergin; Güth, Uwe; Heberer, Michael; Spagnoli, Giulio; Oertli, Daniel; Tapia, Coya

    2012-01-01

    Clinical relevance of tumor infiltrating lymphocytes (TILs) in breast cancer is controversial. Here, we used a tumor microarray including a large series of ductal and lobular breast cancers with long term follow up data, to analyze clinical impact of TIL expressing specific phenotypes and distribution of TILs within different tumor compartments and in different histological subtypes. A tissue microarray (TMA) including 894 ductal and 164 lobular breast cancers was stained with antibodies recognizing CD4, FOXP3, and IL-17 by standard immunohistochemical techniques. Lymphocyte counts were correlated with clinico-pathological parameters and survival. CD4 + lymphocytes were more prevalent than FOXP3 + TILs whereas IL-17 + TILs were rare. Increased numbers of total CD4 + and FOXP3 + TIL were observed in ductal, as compared with lobular carcinomas. High grade (G3) and estrogen receptor (ER) negative ductal carcinomas displayed significantly (p < 0.001) higher CD4 + and FOXP3 + lymphocyte infiltration while her2/neu over-expression in ductal carcinomas was significantly (p < 0.001) associated with higher FOXP3 + TIL counts. In contrast, lymphocyte infiltration was not linked to any clinico-pathological parameters in lobular cancers. In univariate but not in multivariate analysis CD4 + infiltration was associated with significantly shorter survival in patients bearing ductal, but not lobular cancers. However, a FOXP3 + /CD4 + ratio > 1 was associated with improved overall survival even in multivariate analysis (p = 0.033). Ductal and lobular breast cancers appear to be infiltrated by different lymphocyte subpopulations. In ductal cancers increased CD4 + and FOXP3 + TIL numbers are associated with more aggressive tumor features. In survival analysis, absolute numbers of TILs do not represent major prognostic indicators in ductal and lobular breast cancer. Remarkably however, a ratio > 1 of total FOXP3 + /CD4 + TILs in ductal carcinoma appears to represent an independent

  5. BRAF inhibition is associated with increased clonality in tumor-infiltrating lymphocytes

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    Cooper, Zachary A; Frederick, Dennie T; Juneja, Vikram R; Sullivan, Ryan J; Lawrence, Donald P; Piris, Adriano; Sharpe, Arlene H; Fisher, David E; Flaherty, Keith T; Wargo, Jennifer A

    2013-01-01

    There have been significant advances with regard to BRAF-targeted therapies against metastatic melanoma. However, the majority of patients receiving BRAF inhibitors (BRAFi) manifest disease progression within a year. We have recently shown that melanoma patients treated with BRAFi exhibit an increase in melanoma-associated antigens and in CD8+ tumor-infiltrating lymphocytes in response to therapy. To characterize such a T-cell infiltrate, we analyzed the complementarity-determining region 3 (CDR3) of rearranged T-cell receptor (TCR) β chain-coding genes in tumor biopsies obtained before the initiation of BRAFi and 10–14 d later. We observed an increase in the clonality of tumor-infiltrating lymphocytes in 7 of 8 patients receiving BRAFi, with a statistically significant 21% aggregate increase in clonality. Over 80% of individual T-cell clones detected after initiation of BRAFi treatment were new clones. Interestingly, the comparison of tumor infiltrates with clinical responses revealed that patients who had a high proportion of pre-existing dominant clones after the administration of BRAFi responded better to therapy than patients who had a low proportion of such pre-existing dominant clones following BRAFi. These data suggest that although the inhibition of BRAF in melanoma patients results in tumor infiltration by new lymphocytes, the response to treatment appears to be related to the presence of a pre-existing population of tumor-infiltrating T-cell clones. PMID:24251082

  6. The frequency of tumor-infiltrating Tie-2-expressing monocytes in renal cell carcinoma: its relationship to angiogenesis and progression.

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    Ji, Jindong; Zhang, Guangbo; Sun, Bo; Yuan, Hexing; Huang, Yuhua; Zhang, Jianglei; Wei, Xuedong; Zhang, Xuefeng; Hou, Jianquan

    2013-10-01

    To examine the frequency of tumor-infiltrating Tie-2-expressing monocytes (TEMs) in renal cell carcinoma (RCC) and its association with microvessel density (MVD) and other clinical-pathologic features. This study enrolled 65 consecutive patients with RCC treated with radical nephrectomy. The frequency of tumor-infiltrating TEMs, which was defined as CD14(+) Tie-2(+) cells, was assessed using flow cytometry. MVD was measured by immunohistochemistry using anti-CD34 antibody. The association between clinicopathologic parameters, MVD, and the frequency of tumor-infiltrating TEMs in RCC was assessed. High frequency of tumor-infiltrating TEMs was significantly associated with advanced stage (P = .018), positive lymph nodes (P = .013), high grade (P = .019), and metastases (P = .006). Correlation analysis revealed that the frequency of TEMs was positively correlated with MVD. Our findings revealed a significant association between prognostic tumor features, MVD, and the frequency of tumor-infiltrating TEMs in RCC and indicated that TEMs may play an important role in angiogenesis and progression of RCC. Copyright © 2013 Elsevier Inc. All rights reserved.

  7. Tumor-infiltrating lymphocytes for the treatment of metastatic cancer

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    Geukes Foppen, M H; Donia, M; Svane, I M

    2015-01-01

    five years, treatment with immunotherapy (anti CTLA-4, anti PD-1, or the combination of these antibodies) has shown very promising results and was able to improve survival in patients with metastatic melanoma. Adoptive cell therapy using tumor-infiltrating lymphocytes is yet another, but highly...

  8. Standardized assessment of tumor-infiltrating lymphocytes in breast cancer

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    Tramm, Trine; Di Caterino, Tina; Jylling, Anne-Marie B

    2018-01-01

    INTRODUCTION: In breast cancer, there is a growing body of evidence that tumor-infiltrating lymphocytes (TILs) may have clinical utility and may be able to direct clinical decisions for subgroups of patients. Clinical utility is, however, not sufficient for warranting the implementation of a new...

  9. Tumor-infiltrating lymphocytes (TILs) from patients with glioma

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    Liu, Zhenjiang; Meng, Qingda; Bartek, Jiri

    2017-01-01

    Tumor-infiltrating lymphocytes (TILs) may represent a viable source of T cells for the biological treatment of patients with gliomas. Glioma tissue was obtained from 16 patients, tumor cell lines were established, and TILs were expanded in 16/16 cases using a combination of IL-2/IL-15/IL-21...

  10. The clinical significance of neutrophilic pleocytosis in cerebrospinal fluid in patients with viral central nervous system infections

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    Siraya Jaijakul

    2017-06-01

    Conclusions: The results of a study exploring the association between CSF neutrophilic pleocytosis and clinical and prognostic significance are presented here. This study suggests that CSF neutrophilic pleocytosis is not associated with higher adverse clinical outcomes.

  11. Tumor-infiltrating lymphocytes and ductal carcinoma in situ of the breast: friends or foes?

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    Agahozo, Marie Colombe; Hammerl, Dora; Debets, Reno; Kok, Marleen; van Deurzen, Carolien H M

    2018-02-20

    In the past three decades, the detection rate of ductal carcinoma in situ of the breast has dramatically increased due to breast screening programs. As a consequence, about 20% of all breast cancer cases are detected in this early in situ stage. Some ductal carcinoma in situ cases will progress to invasive breast cancer, while other cases are likely to have an indolent biological behavior. The presence of tumor-infiltrating lymphocytes is seen as a promising prognostic and predictive marker in invasive breast cancer, mainly in HER2-positive and triple-negative subtypes. Here, we summarize the current understanding regarding immune infiltrates in invasive breast cancer and highlight recent observations regarding the presence and potential clinical significance of such immune infiltrates in patients with ductal carcinoma in situ. The presence of tumor-infiltrating lymphocytes, their numbers, composition, and potential relationship with genomic status will be discussed. Finally, we propose that a combination of genetic and immune markers may better stratify ductal carcinoma in situ subtypes with respect to tumor evolution.

  12. Prognostic significance of neutrophil-to-lymphocyte ratio in biliary tract cancers: a systematic review and meta-analysis.

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    Tang, Haowen; Lu, Wenping; Li, Bingmin; Li, Chonghui; Xu, Yinzhe; Dong, Jiahong

    2017-05-30

    Inflammation was considered to perform crucial roles in the development and metastasis of malignancies. A heightened neutrophil-lymphocyte ratio has been described to be associated with detrimental survivals in different malignancies. Debate remains over the impact of heightened neutrophil-lymphocyte ratio on survivals in biliary tract cancer. The review evaluated the prognostic value of neutrophil-lymphocyte ratio in biliary tract cancer. MEDLINE, the Cochrane Library, EMBASE, and the Chinese SinoMed were systematically searched for relevant articles. Associations between neutrophil-lymphocyte ratio and long-term outcomes were expressed as the hazard ratios and 95% confidence intervals. The odds ratio was utilized to assess the association between neutrophil-lymphocyte ratio and clinicopathological parameters. Fourteen studies consisting of 3217 patients were analyzed: 1278 (39.73%) in the high pretreatment neutrophil-lymphocyte ratio group and 1939 (60.27%) in the low pretreatment neutrophil-lymphocyte ratio one. The results proved that heightened pretreatment neutrophil-lymphocyte ratio was significantly associated with detrimental overall survival and relapse free survival for biliary tract cancer patients. In addition, elevated neutrophil-lymphocyte ratio was positively correlated with higher carbohydrate antigen 19-9 levels, advanced TNM staging and greater lymph node involvement. This meta-analysis marked that an increased pretreatment neutrophil-lymphocyte ratio was significantly linked with detrimental long-term outcomes and clinicopathological parameters for patients with biliary tract cancer.

  13. Tumor-Derived CXCL1 Promotes Lung Cancer Growth via Recruitment of Tumor-Associated Neutrophils

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    Ming Yuan

    2016-01-01

    Full Text Available Neutrophils have a traditional role in inflammatory process and act as the first line of defense against infections. Although their contribution to tumorigenesis and progression is still controversial, accumulating evidence recently has demonstrated that tumor-associated neutrophils (TANs play a key role in multiple aspects of cancer biology. Here, we detected that chemokine CXCL1 was dramatically elevated in serum from 3LL tumor-bearing mice. In vitro, 3LL cells constitutively expressed and secreted higher level of CXCL1. Furthermore, knocking down CXCL1 expression in 3LL cells significantly hindered tumor growth by inhibiting recruitment of neutrophils from peripheral blood into tumor tissues. Additionally, tumor-infiltrated neutrophils expressed higher levels of MPO and Fas/FasL, which may be involved in TAN-mediated inhibition of CD4+ and CD8+ T cells. These results demonstrate that tumor-derived CXCL1 contributes to TANs infiltration in lung cancer which promotes tumor growth.

  14. [The significances of peripheral neutrophils CD(55) and myeloperoxidase expression in patients with myeloperoxidase-specific anti-neutrophil cytoplasmic antibody associated vasculitis].

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    Zhou, X L; Zheng, M J; Shuai, Z W; Zhang, L; Zhang, M M; Chen, S Y

    2017-06-01

    Objective: To investigate the expression of CD(55) and myeloperoxidase (MPO) on neutrophils in patients with MPO-specific anti-neutrophil cytoplasmic antibody associated vasculitis(MPO-AAV), and analyze the relationship between the expression and clinical manifestation. Methods: Forty untreated patients with active MPO-AAV (patient group) and 30 healthy volunteers (control group) were enrolled in this study. The CD(55) on neutrophils and both membrane and cytoplasmic MPO were detected by flow cytometry. Serum fragment-from the activated complement factor B(Ba) and MPO were measured by ELISA. The clinical activity of vasculitis was valued by Birmingham vasculitis activity score-version 3(BVAS-V3). The significance of laboratory data was evaluated by Spearman correlation test and multivariate linear regression analysis. Results: (1)The mean fluorescence intensity(MFI) of CD(55) expressed on neutrophils was significantly higher than that in control group[4 068.6±2 306.0 vs 2 999.5±1 504.9, P =0.033]. Similar results of serum MPO and Ba in patient group were found compared to controls [500.0(381.0, 612.7) IU/L vs 286.9(225.5, 329.1) IU/L, P <0.001; 35.2(25.2, 79.5) ng/L vs 18.0(15.0, 28.0) ng/L, P <0.001], respectively. However, MIF of cytoplasmic MPO in patients was significantly lower than that of control group(1 577.1±1 175.9 vs 3 105.3±2 323.0, P =0.003) . (2) In patient group, cytoplasmic intensity of MPO was negatively associated with the serum levels of MPO( r =-0.710, P <0.001) and Ba ( r =-0.589, P =0.001). Moreover, serum MPO was positively associated with serum Ba( r =0.691, P <0.001). Membrane intensity of CD(55) on neutrophils was positively correlated with patient age ( r =0.514, P =0.001), C reactive protein ( r =0.376, P =0.018), peripheral neutrophils count ( r =0.485, P =0.001) and BVAS-V3 ( r =0.484, P =0.002), whereas negative correlation between membrane CD(55) and disease duration was seen ( r =-0.403, P =0.01). (3) The result of multiple

  15. Tumor infiltrating lymphocytes: an intriguing player in the survival of colorectal cancer patients

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    Lardon Filip

    2010-04-01

    (borderline significant for overall population only were associated with a better overall survival (early disease with disease-free survival also. Conclusions In conclusion our results suggest a role for infiltrating CD3+ and CD8+ T lymphocytes in colorectal cancer whereby tumor infiltration could reflect a general principle of antitumor immunity, irrespective of the MSI-status.

  16. Tumor-infiltrating lymphocytes predict efficacy of preoperative radiotherapy for rectal cancer

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    Xu Gang; Zhang Shanwen; Xu Bo

    2009-01-01

    Objective: To evaluate the effect of tumor infiltrating lymphocyte(TIL) on prognosis of rectal cancer treated with preoperative radiotherapy. Methods: From Jan. 1999 to Oct. 2007,107 patients with rectal cancer were treated with preoperative radiotherapy of 30 Gy/10f/12 days. The relationships among TIL, pathologic regression and prognosis were analyzed. Results: Before radiotherapy, TIL in rectal cancer was 75 patients (70.1%) in grade 1,16 (15.0%) in grade 2 and 16 (15.0%) in grade 3; While after radiotherapy, it changed to 19 (17.7%) in grade 1,43 (40.2%) in grade 2,35 (32.7%) in grade 3 and 10 (9.3%) in grade 4. After radiotherapy, pathologic regression was 36 (33.6%) in grade 1,57 (53.3%) in grade 2 and 14 (13.1%) in grade 3. Univariate analysis showed that TIL both before and after radiotherapy was the significant prognostic factor for local pathologic regression (χ 2 =36.80, P 2 = 14.00, P 2 =24.00, P 2 =12.17, P 2 =8.05, P<0.01). Conclusions: For rectal cancer treated with preoperative radiotherapy, TIL before and after radiotherapy is significantly related with local pathologic regression, and TIL after radiotherapy is a prognostic factor. (authors)

  17. Flow cytometric analysis of peripheral blood and tumor-infiltrating regulatory T cells in dogs with oral malignant melanoma.

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    Tominaga, Makiko; Horiuchi, Yutaka; Ichikawa, Mika; Yamashita, Masao; Okano, Kumiko; Jikumaru, Yuri; Nariai, Yoko; Kadosawa, Tsuyoshi

    2010-05-01

    It is well known that tumor-infiltrating lymphocytes (TILs) and peripheral blood lymphocytes (PBLs) from patients with advanced-stage cancer have a poor immune response. Regulatory T cells (Tregs), characterized by the expression of a cluster of differentiation 4 and intracellular FoxP3 markers, can inhibit antitumor immunoresponse. In the present study, the prevalence of Tregs in peripheral blood and tumor tissue from dogs with oral malignant melanoma was evaluated by triple-color flow cytometry. The percentage of Tregs in the peripheral blood of the dogs with malignancy was significantly increased compared with healthy control dogs, and the percentage of Tregs within tumors was significantly increased compared with Tregs in peripheral blood of dogs with oral malignant melanoma. This finding suggests that the presence of tumor cells induced either local proliferation or selective migration of Tregs to tumor-infiltrated sites. A better understanding of the underlying mechanisms of Treg regulation in patients with cancer may lead to an effective anticancer immunotherapy against canine malignant melanoma and possibly other tumors.

  18. Characterization of ex vivo expanded tumor infiltrating lymphocytes from patients with malignant melanoma for clinical application

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    Junker, Niels; Thor Straten, Per; Andersen, Mads Hald

    2011-01-01

    Clinical trials of adoptive transfer of autologous tumor infiltrating lymphocytes (TILs) to patients with advanced malignant melanoma have shown remarkable results with objective clinical responses in 50% of the treated patients. In order to initiate a clinical trial in melanoma, we have establis......Clinical trials of adoptive transfer of autologous tumor infiltrating lymphocytes (TILs) to patients with advanced malignant melanoma have shown remarkable results with objective clinical responses in 50% of the treated patients. In order to initiate a clinical trial in melanoma, we have...

  19. Adoptive cell transfer using autologous tumor infiltrating lymphocytes in gynecologic malignancies.

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    Mayor, Paul; Starbuck, Kristen; Zsiros, Emese

    2018-05-23

    During the last decade, the field of cancer immunotherapy has been entirely transformed by the development of new and more effective treatment modalities with impressive response rates and the prospect of long survival. One of the major breakthroughs is adoptive cell transfer (ACT) based on autologous T cells derived from tumor-infiltrating lymphocytes (TILs). TIL-based ACT is a highly personalized cancer treatment. T cells are harvested from autologous fresh tumor tissues, and after ex vivo activation and extensive expansion, are reinfused to patients. TIL-based therapies have only been offered in small phase I/II studies in a few centers given the highly specialized care required, the complexity of TIL production and the very intensive nature of the three-step treatment protocol. The treatment includes high-dose lymphodepleting chemotherapy, the infusion of the expanded and activated T cells and interleukin-2 (IL-2) injections to increase survival of the T cells. Despite the limited data on ACT, the small published studies consistently confirm an impressive clinical response rate of up to 50% in metastatic melanoma patients, including a significant proportion of patients with durable complete response. These remarkable results justify the need for larger clinical trials in other solid tumors, including gynecologic malignancies. In this review we provide an overview of the current clinical results, future applications of TIL-based ACT in gynecologic malignancies, and on risks and challenges associated with modern T cell therapy. Copyright © 2018. Published by Elsevier Inc.

  20. Metastatic Lung Lesions as a Preferred Resection Site for Immunotherapy With Tumor Infiltrating Lymphocytes.

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    Ben-Avi, Ronny; Itzhaki, Orit; Simansky, David; Zippel, Dov; Markel, Gal; Ben Nun, Alon; Schachter, Jacob; Besser, Michal J

    2016-06-01

    Adoptive cell therapy with tumor infiltrating lymphocytes (TIL) yields 50% response rates in metastatic melanoma and shows promising clinical results in other solid tumors. Autologous TIL cultures are isolated from resected tumor tissue, expanded ex vivo to large numbers and reinfused to the preconditioned patient. In this prospective study, we validate the origin of the tumor biopsy and its effect on T-cell function and clinical response. One hundred forty-four patients underwent surgery and 79 patients were treated with TIL adoptive cell therapy. Cultures from lung tissue were compared with other origins. The success rate of establishing TIL culture from lung tissue was significantly higher compared with nonlung tissue (94% vs. 72%, respectively, P≤0.003). Lung-derived TIL cultures gave rise to higher cell numbers (P≤0.011) and exhibited increased in vitro antitumor reactivity. The average fold expansion for lung-derived TIL during a rapid expansion procedure was 1349±557 compared with 1061±473 for nonlung TIL (P≤0.038). Patients treated with TIL cultures of lung origin (compared with nonlung) had prolonged median overall survival (29 vs. 9.5 mo; P≤0.065). Given the remarkable advancement in minimally invasive thoracic surgery and the results of this study, we suggest efforts should be taken to resect lung metastasis rather than other sites to generate TIL cultures for clinical use.

  1. Tumor infiltrating lymphocyte therapy for ovarian cancer and renal cell carcinoma

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    Andersen, Rikke; Donia, Marco; Westergaard, Marie Christine Wulff

    2015-01-01

    stimulated the interest in developing this approach for other indications. Here, we summarize the early clinical data in the field of adoptive cell transfer therapy (ACT) using tumor-infiltrating lymphocytes for patients with renal cell carcinoma (RCC) and ovarian cancer (OC). In addition we describe...

  2. A distinguishing gene signature shared by tumor-infiltrating Tie2-expressing monocytes, blood "resident" monocytes, and embryonic macrophages suggests common functions and developmental relationships.

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    Pucci, Ferdinando; Venneri, Mary Anna; Biziato, Daniela; Nonis, Alessandro; Moi, Davide; Sica, Antonio; Di Serio, Clelia; Naldini, Luigi; De Palma, Michele

    2009-07-23

    We previously showed that Tie2-expressing monocytes (TEMs) have nonredundant proangiogenic activity in tumors. Here, we compared the gene expression profile of tumor-infiltrating TEMs with that of tumor-associated macrophages (TAMs), spleen-derived Gr1(+)Cd11b(+) neutrophils/myeloid-derived suppressor cells, circulating "inflammatory" and "resident" monocytes, and tumor-derived endothelial cells (ECs) by quantitative polymerase chain reaction-based gene arrays. TEMs sharply differed from ECs and Gr1(+)Cd11b(+) cells but were highly related to TAMs. Nevertheless, several genes were differentially expressed between TEMs and TAMs, highlighting a TEM signature consistent with enhanced proangiogenic/tissue-remodeling activity and lower proinflammatory activity. We validated these findings in models of oncogenesis and transgenic mice expressing a microRNA-regulated Tie2-GFP reporter. Remarkably, resident monocytes and TEMs on one hand, and inflammatory monocytes and TAMs on the other hand, expressed coordinated gene expression profiles, suggesting that the 2 blood monocyte subsets are committed to distinct extravascular fates in the tumor microenvironment. We further showed that a prominent proportion of embryonic/fetal macrophages, which participate in tissue morphogenesis, expressed distinguishing TEM genes. It is tempting to speculate that Tie2(+) embryonic/fetal macrophages, resident blood monocytes, and tumor-infiltrating TEMs represent distinct developmental stages of a TEM lineage committed to execute physiologic proangiogenic and tissue-remodeling programs, which can be co-opted by tumors.

  3. Adoptive Cell Therapy with Tumor-Infiltrating Lymphocytes in Advanced Melanoma Patients

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    Mélanie Saint-Jean

    2018-01-01

    Full Text Available Immunotherapy for melanoma includes adoptive cell therapy with autologous tumor-infiltrating lymphocytes (TILs. This monocenter retrospective study was undertaken to evaluate the efficacy and safety of this treatment of patients with advanced melanoma. All advanced melanoma patients treated with TILs using the same TIL expansion methodology and same treatment interleukin-2 (IL-2 regimen between 2009 and 2012 were included. After sterile intralesional excision of a cutaneous or subcutaneous metastasis, TILs were produced according to a previously described method and then infused into the patient who also received a complementary subcutaneous IL-2 regimen. Nine women and 1 man were treated for unresectable stage IIIC (n=4 or IV (n=6 melanoma. All but 1 patient with unresectable stage III melanoma (1st line had received at least 2 previous treatments, including anti-CTLA-4 antibody for 4. The number of TILs infused ranged from 0.23 × 109 to 22.9 × 109. Regarding safety, no serious adverse effect was reported. Therapeutic responses included a complete remission, a partial remission, 2 stabilizations, and 6 progressions. Among these 4 patients with clinical benefit, 1 is still alive with 9 years of follow-up and 1 died from another cause after 8 years of follow-up. Notably, patients treated with high percentages of CD4 + CD25 + CD127lowFoxp3+ T cells among their TILs had significantly shorter OS. The therapeutic effect of combining TILs with new immunotherapies needs further investigation.

  4. Tumor-Infiltrating Immune Cells Promoting Tumor Invasion and Metastasis: Existing Theories

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    Yan-gao Man, Alexander Stojadinovic, Jeffrey Mason, Itzhak Avital, Anton Bilchik, Bjoern Bruecher, Mladjan Protic, Aviram Nissan, Mina Izadjoo, Xichen Zhang, Anahid Jewett

    2013-01-01

    Full Text Available It is a commonly held belief that infiltration of immune cells into tumor tissues and direct physical contact between tumor cells and infiltrated immune cells is associated with physical destructions of the tumor cells, reduction of the tumor burden, and improved clinical prognosis. An increasing number of studies, however, have suggested that aberrant infiltration of immune cells into tumor or normal tissues may promote tumor progression, invasion, and metastasis. Neither the primary reason for these contradictory observations, nor the mechanism for the reported diverse impact of tumor-infiltrating immune cells has been elucidated, making it difficult to judge the clinical implications of infiltration of immune cells within tumor tissues. This mini-review presents several existing hypotheses and models that favor the promoting impact of tumor-infiltrating immune cells on tumor invasion and metastasis, and also analyzes their strength and weakness.

  5. Adoptive Cell Therapy with Tumor-Infiltrating Lymphocytes in Advanced Melanoma Patients

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    Mélanie Saint-Jean; Anne-Chantal Knol; Christelle Volteau; Gaëlle Quéreux; Lucie Peuvrel; Anabelle Brocard; Marie-Christine Pandolfino; Soraya Saiagh; Jean-Michel Nguyen; Christophe Bedane; Nicole Basset-Seguin; Amir Khammari; Brigitte Dréno

    2018-01-01

    Immunotherapy for melanoma includes adoptive cell therapy with autologous tumor-infiltrating lymphocytes (TILs). This monocenter retrospective study was undertaken to evaluate the efficacy and safety of this treatment of patients with advanced melanoma. All advanced melanoma patients treated with TILs using the same TIL expansion methodology and same treatment interleukin-2 (IL-2) regimen between 2009 and 2012 were included. After sterile intralesional excision of a cutaneous or subcutaneous ...

  6. Spatial Organization and Molecular Correlation of Tumor-Infiltrating Lymphocytes Using Deep Learning on Pathology Images

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    Joel Saltz

    2018-04-01

    Full Text Available Summary: Beyond sample curation and basic pathologic characterization, the digitized H&E-stained images of TCGA samples remain underutilized. To highlight this resource, we present mappings of tumor-infiltrating lymphocytes (TILs based on H&E images from 13 TCGA tumor types. These TIL maps are derived through computational staining using a convolutional neural network trained to classify patches of images. Affinity propagation revealed local spatial structure in TIL patterns and correlation with overall survival. TIL map structural patterns were grouped using standard histopathological parameters. These patterns are enriched in particular T cell subpopulations derived from molecular measures. TIL densities and spatial structure were differentially enriched among tumor types, immune subtypes, and tumor molecular subtypes, implying that spatial infiltrate state could reflect particular tumor cell aberration states. Obtaining spatial lymphocytic patterns linked to the rich genomic characterization of TCGA samples demonstrates one use for the TCGA image archives with insights into the tumor-immune microenvironment. : Tumor-infiltrating lymphocytes (TILs were identified from standard pathology cancer images by a deep-learning-derived “computational stain” developed by Saltz et al. They processed 5,202 digital images from 13 cancer types. Resulting TIL maps were correlated with TCGA molecular data, relating TIL content to survival, tumor subtypes, and immune profiles. Keywords: digital pathology, immuno-oncology, machine learning, lymphocytes, tumor microenvironment, deep learning, tumor-infiltrating lymphocytes, artificial intelligence, bioinformatics, computer vision

  7. Prognostic significance of neutrophil-lymphocyte ratio in hepatocellular carcinoma: a meta-analysis

    International Nuclear Information System (INIS)

    Xiao, Wei-Kai; Chen, Dong; Li, Shao-Qiang; Fu, Shun-Jun; Peng, Bao-Gang; Liang, Li-Jian

    2014-01-01

    Neutrophil-lymphocyte ratio (NLR) has recently been reported as a predictor of Hepatocellular carcinoma (HCC). However, its prognostic value in HCC still remains controversial. In this study, we aimed to evaluate the association between NLR and clinical outcome of HCC patients by performing meta-analysis. A comprehensive literature search for relevant studies published up to August 2013 was performed by using PubMed, Ovid, the Cochrane Library and Web of Science databases. Meta-analysis was performed using hazard ratio (HR) or odds ratio (OR) and 95% confidence intervals (95% CIs) as effect measures. A total of 15 studies encompassing 3094 patients were included in this meta-analysis. Our pooled results showed that high NLR was associated with poor overall survival (OS) and disease free survival (DFS) in HCC initially treated by liver transplantation (HR = 3.42, 95% CI:2.41-4.85,P = 0.000; HR = 5.90, 95% CI:3.99-8.70,P = 0.000, respectively) and surgical resection (HR = 3.33, 95% CI:2.23-4.98, P = 0.000; HR = 2.10, 95% CI: 2.06–2.14, respectively). High NLR was also associated with poor OS in HCC treated by radiofrequency-ablation (HR = 1.28, 95%CI: 1.10-1.48, P = 0.000), TACE (HR = 2.52, 95% CI: 1.64-3.86, P = 0.000) and mixed treatment (HR = 1.85, 95% CI: 1.40-2.44, P = 0.000), respectively. In addition, high NLR was significantly correlated with the presence of vascular invasion (OR = 2.69, 95% CI: 2.01–3.59, P = 0.000), tumor multifocality (OR = 1.74, 95% CI: 1.30–2.34, P = 0.000) and higher incidence of AFP ≥ 400 ng/ml (OR = 1.46, 95% CI: 1.01–2.09, P = 0.04). Elevated NLR indicates a poor prognosis for patients with HCC. NLR may be a convenient, easily-obtained, low cost and reliable biomarker with prognostic potential for HCC

  8. Tumor infiltrating lymphocytes in triple negative breast cancer receiving neoadjuvant chemotherapy

    Science.gov (United States)

    Castaneda, Carlos A; Mittendorf, Elizabeth; Casavilca, Sandro; Wu, Yun; Castillo, Miluska; Arboleda, Patricia; Nunez, Teresa; Guerra, Henry; Barrionuevo, Carlos; Dolores-Cerna, Ketty; Belmar-Lopez, Carolina; Abugattas, Julio; Calderon, Gabriela; De La Cruz, Miguel; Cotrina, Manuel; Dunstan, Jorge; Gomez, Henry L; Vidaurre, Tatiana

    2016-01-01

    AIM To determine influence of neoadjuvant-chemotherapy (NAC) over tumor-infiltrating-lymphocytes (TIL) in triple-negative-breast-cancer (TNBC). METHODS TILs were evaluated in 98 TNBC cases who came to Instituto Nacional de Enfermedades Neoplasicas from 2005 to 2010. Immunohistochemistry staining for CD3, CD4, CD8 and FOXP3 was performed in tissue microarrays (TMA) sections. Evaluation of H/E in full-face and immunohistochemistry in TMA sections was performed in pre and post-NAC samples. STATA software was used and P value < 0.05 was considered statistically significant. RESULTS Higher TIL evaluated in full-face sections from pre-NAC tumors was associated to pathologic-complete-response (pCR) (P = 0.0251) and outcome (P = 0.0334). TIL evaluated in TMA sections showed low level of agreement with full-face sections (ICC = 0.017-0.20) and was not associated to pCR or outcome. TIL in post-NAC samples were not associated to response or outcome. Post-NAC lesions with pCR had similar TIL levels than those without pCR (P = 0.6331). NAC produced a TIL decrease in full-face sections (P < 0.0001). Percentage of TIL subpopulations was correlated with their absolute counts. Higher counts of CD3, CD4, CD8 and FOXP3 in pre-NAC samples had longer disease-free-survival (DFS). Higher counts of CD3 in pre-NAC samples had longer overall-survival. Higher ratio of CD8/CD4 counts in pre-NAC was associated with pCR. Higher ratio of CD4/FOXP3 counts in pre-NAC was associated with longer DFS. Higher counts of CD4 in post-NAC samples were associated with pCR. CONCLUSION TIL in pre-NAC full-face sections in TNBC are correlated to longer survival. TIL in full-face differ from TMA sections, absolute count and percentage analysis of TIL subpopulation closely related. PMID:27777881

  9. Immunophenotypic features of tumor infiltrating lymphocytes from mammary carcinomas in female dogs associated with prognostic factors and survival rates

    International Nuclear Information System (INIS)

    Estrela-Lima, Alessandra; Araújo, Márcio SS; Costa-Neto, João M; Teixeira-Carvalho, Andréa; Barrouin-Melo, Stella M; Cardoso, Sergio V; Martins-Filho, Olindo A; Serakides, Rogéria; Cassali, Geovanni D

    2010-01-01

    The immune system plays an important role in the multifactorial biologic system during the development of neoplasias. However, the involvement of the inflammatory response in the promotion/control of malignant cells is still controversial, and the cell subsets and the mechanisms involved are poorly investigated. The goal of this study was to characterize the clinical-pathological status and the immunophenotyping profile of tumor infiltrating lymphocytes and their association with the animal survival rates in canine mammary carcinomas. Fifty-one animals with mammary carcinomas, classified as carcinomas in mixed tumors-MC-BMT = 31 and carcinomas-MC = 20 were submitted to systematic clinical-pathological analysis (tumor size; presence of lymph node and pulmonary metastasis; clinical stage; histological grade; inflammatory distribution and intensity as well as the lymphocytic infiltrate intensity) and survival rates. Twenty-four animals (MC-BMT = 16 and MC = 8) were elected to the immunophenotypic study performed by flow cytometry. Data analysis demonstrated that clinical stage II-IV and histological grade was I more frequent in MC-BMT as compared to MC. Univariate analysis demonstrated that the intensity of inflammation (moderate/intense) and the proportion of CD4 + (≥ 66.7%) or CD8 + T-cells (<33.3%) were not associated with worse survival rate. Multivariate analysis demonstrated that only lymphocytic infiltrate intensity ≥ 600 (P = 0.02) remained as independent prognostic factor. Despite the clinical manifestation, the lymphocytes represented the predominant cell type in the tumor infiltrate. The percentage of T-cells was higher in animals with MC-BMT without metastasis, while the percentage of B-lymphocytes was greater in animals with metastasized MC-BMT (P < 0.05). The relative percentage of CD4 + T-cells was significantly greater in metastasized tumors (both MC-BMT and MC), (P < 0.05) while the proportion of CD8 + T-cells was higher in MC-BMT without

  10. Phenotypic and Functional Properties of Tumor-Infiltrating Regulatory T Cells

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    Gap Ryol Lee

    2017-01-01

    Full Text Available Regulatory T (Treg cells maintain immune homeostasis by suppressing excessive immune responses. Treg cells induce tolerance against self- and foreign antigens, thus preventing autoimmunity, allergy, graft rejection, and fetus rejection during pregnancy. However, Treg cells also infiltrate into tumors and inhibit antitumor immune responses, thus inhibiting anticancer therapy. Depleting whole Treg cell populations in the body to enhance anticancer treatments will produce deleterious autoimmune diseases. Therefore, understanding the precise nature of tumor-infiltrating Treg cells is essential for effectively targeting Treg cells in tumors. This review summarizes recent results relating to Treg cells in the tumor microenvironment, with particular emphasis on their accumulation, phenotypic, and functional properties, and targeting to enhance the efficacy of anticancer treatment.

  11. Transient exposure to proteins SOX2, Oct-4, and NANOG immortalizes exhausted tumor-infiltrating CTLs

    Energy Technology Data Exchange (ETDEWEB)

    Bhadurihauck, Anjuli; Li, Lei [Department of Animal and Avian Sciences, University of Maryland, College Park, 20742, MD (United States); Li, Qianqian; Wang, Jianjun [Department of Biochemistry and Molecular Biology, Wayne State University, Detroit, 48201 (United States); Xiao, Zhengguo, E-mail: xiao0028@umd.edu [Department of Animal and Avian Sciences, University of Maryland, College Park, 20742, MD (United States)

    2016-05-13

    Adoptive cell transfer therapy (ACT) is one of the most promising immunotherapies against cancer, using tumor-infiltrating lymphocytes (TILs) expanded in vitro. Tumor-infiltrating cytotoxic T lymphocytes (TICTLs) play a prominent role in cancer control. TILs terminally differentiate in response to immunosuppressive environments within tumors, and thus are slow to expand and challenging to maintain both in vitro and in patients. To reverse this exhaustion, we utilize a nuclear protein delivery system that exposes TICTLs to the SOX2, Oct-4, and NANOG (SON) proteins. Unlike activated naïve CTLs (effector CTLs), TICTLs respond favorably to SON treatment, exhibiting steady proliferation and extended survivability independent of cytokine and antigen stimulation. Though TICTLs treated with SON (STICTLs) still express T cell receptors as well as other critical downstream components, they are unresponsive to antigen challenge, suggesting that SON treatment regresses TICTLs into a state similar to that of an early double negative T cell. Our findings indicate the TICTL response to SON proteins is unique when compared to effector CTLs, suggesting TICTLs may be sensitive to regulation by other lineage-specific transcription factors and opening a promising new avenue into cancer immunotherapy. To our knowledge, this is the first report on lineage reprogramming of TILs using protein stem cell transcription factors delivered directly to the nucleus. -- Highlights: •TICTLs are sensitive to reprogramming by proteins of stem cell transcription factors, but effector CTLs were not. •TICTLs are regressed back to an early double negative T cell stage. •TCR signaling is deregulated by these transcription factors.

  12. Characterization and comparison of "Standard" and "Young" tumor infiltrating lymphocytes for adoptive cell therapy at a Danish Translational Research Institution

    DEFF Research Database (Denmark)

    Donia, Marco; Junker, Niels; Ellebaek, Eva

    2012-01-01

    Adoptive cell therapy (ACT) with ex vivo expanded tumor infiltrating lymphocytes (TILs) in combination with IL-2 is an effective treatment for patients with metastatic melanoma. Modified protocols of cell expansion may allow treatment of most enrolled patients and improve the efficacy of adoptively...

  13. Achievements and challenges of adoptive T cell therapy with tumor-infiltrating or blood-derived lymphocytes for metastatic melanoma

    DEFF Research Database (Denmark)

    Svane, Inge Marie; Verdegaal, Els M

    2014-01-01

    Adoptive cell therapy (ACT) based on autologous T cell derived either from tumor as tumor-infiltrating lymphocytes (TILs) or from peripheral blood is developing as a key area of future personalized cancer therapy. TIL-based ACT is defined as the infusion of T cells harvested from autologous fresh...

  14. Clinical Significance of the Neutrophil-to-Lymphocyte Ratio in Endocrine Therapy for Stage IV Breast Cancer.

    Science.gov (United States)

    Iimori, Nozomi; Kashiwagi, Shinichiro; Asano, Yuka; Goto, Wataru; Takada, Koji; Takahashi, Katsuyuki; Hatano, Takaharu; Takashima, Tsutomu; Tomita, Shuhei; Motomura, Hisashi; Hirakawa, Kosei; Ohira, Masaichi

    2018-01-01

    Studies have found that patients with cancer exhibit abnormal leukocyte fractions, such as elevated neutrophil count and diminished lymphocyte count, and that the neutrophil-to-lymphocyte ratio (NLR) provides a surrogate marker for prognosis and response to treatment of patients after radical surgery for several different types of cancer. However, few reports have addressed the association between the NLR and response to endocrine therapy. In this study, we carried out a clinical investigation to confirm whether or not the NLR predicted the response to endocrine therapy of stage IV breast cancer. The study subjects were 34 patients who underwent endocrine therapy as initial drug therapy for stage IV breast cancer. The correlation between NLR and prognosis, including the efficacy of endocrine therapy, was evaluated retrospectively. Among the 34 patients, the NLR was high in 10 (29.4%) and low in 24 (70.6%). In analysis of outcomes, the group with low NLR had a significant prolongation of progression-free survival (p=0.003), time to treatment failure (p=0.031), and overall survival (p=0.013) compared to the group with high NLR. Univariate analysis of progression-free survival found that responding to treatment [hazard ratio (HR)=4.310, p=0.004] and low NLR (HR=3.940, p=0.016) were factors associated with a favorable prognosis. Multivariate analysis also showed that responding to treatment (HR=4.329, p=0.006) and low NLR (HR=3.930, p=0.008) were independent factors associated with a favorable prognosis. Our results suggested that the NLR may represent a predictive marker for response to endocrine therapy in stage IV breast cancer. Copyright© 2018, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.

  15. Cytokine and immunoglobulin production by PWM-stimulated peripheral and tumor-infiltrating lymphocytes of undifferentiated nasopharyngeal carcinoma (NPC) patients

    International Nuclear Information System (INIS)

    Fliss-Jaber, Lilia; Houissa-Kastally, Radhia; Bouzouita, Kamel; Khediri, Naceur; Khelifa, Ridha

    2004-01-01

    Undifferentiated Nasopharyngeal Carcinoma (NPC) patients show a characteristic pattern of antibody responses to the Epstein-Barr virus (EBV) which is regularly associated with this tumor. However, no EBV-specific cytotoxic activity is detectable by the standard chromium-release assay at both peripheral and intratumoral levels. The mechanisms underlying this discrepancy between the humoral and cellular immune responses in NPC are still unknown, but might be related to an imbalance in immunoregulatory interleukin production. In this report, we investigated the ability of peripheral (PBL) and tumor- infiltrating (TIL) lymphocytes of undifferentiated NPC patients to produce in vitro three interleukins (IL-2, IL-6, IL-10) and three immunoglobulin isotypes (IgM, IgG, IgA). Lymphocytes from 17 patients and 17 controls were cultured in the presence of Pokeweed mitogen (PWM) for 12 days and their culture supernatants were tested for interleukins and immunoglobulins by specific enzyme-linked immunosorbent assays (ELISA). Data were analysed using Student's t-test and probability values below 5% were considered significant. The data obtained indicated that TIL of NPC patients produced significantly more IL-2 (p = 0,0002), IL-10 (p = 0,020), IgM (p= 0,0003) and IgG (p < 0,0001) than their PBL. On the other hand, patients PBL produced significantly higher levels of IL-2 (p = 0,022), IL-10 (p = 0,016) and IgM (p = 0,004) than those of controls. No significant differences for IL-6 and IgA were observed. Taken together, our data reinforce the possibility of an imbalance in immunoregulatory interleukin production in NPC patients. An increased ability to produce cytokines such as IL-10 may underlie the discrepancy between humoral and cellular immune responses characteristic of NPC

  16. Adoptive cell therapy with autologous tumor infiltrating lymphocytes and low-dose Interleukin-2 in metastatic melanoma patients

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    Ellebaek Eva

    2012-08-01

    Full Text Available Abstract Background Adoptive cell therapy may be based on isolation of tumor-specific T cells, e.g. autologous tumor infiltrating lymphocytes (TIL, in vitro activation and expansion and the reinfusion of these cells into patients upon chemotherapy induced lymphodepletion. Together with high-dose interleukin (IL-2 this treatment has been given to patients with advanced malignant melanoma and impressive response rates but also significant IL-2 associated toxicity have been observed. Here we present data from a feasibility study at a Danish Translational Research Center using TIL adoptive transfer in combination with low-dose subcutaneous IL-2 injections. Methods This is a pilot trial (ClinicalTrials.gov identifier: NCT00937625 including patients with metastatic melanoma, PS ≤1, age Results Low-dose IL-2 considerably decreased the treatment related toxicity with no grade 3–4 IL-2 related adverse events. Objective clinical responses were seen in 2 of 6 treated patients with ongoing complete responses (30+ and 10+ months, 2 patients had stable disease (4 and 5 months and 2 patients progressed shortly after treatment. Tumor-reactivity of the infused cells and peripheral lymphocytes before and after therapy were analyzed. Absolute number of tumor specific T cells in the infusion product tended to correlate with clinical response and also, an induction of peripheral tumor reactive T cells was observed for 1 patient in complete remission. Conclusion Complete and durable responses were induced after treatment with adoptive cell therapy in combination with low-dose IL-2 which significantly decreased toxicity of this therapy.

  17. Cytokine and immunoglobulin production by PWM-stimulated peripheral and tumor-infiltrating lymphocytes of undifferentiated nasopharyngeal carcinoma (NPC patients

    Directory of Open Access Journals (Sweden)

    Bouzouita Kamel

    2004-09-01

    Full Text Available Abstract Background Undifferentiated Nasopharyngeal Carcinoma (NPC patients show a characteristic pattern of antibody responses to the Epstein-Barr virus (EBV which is regularly associated with this tumor. However, no EBV-specific cytotoxic activity is detectable by the standard chromium-release assay at both peripheral and intratumoral levels. The mechanisms underlying this discrepancy between the humoral and cellular immune responses in NPC are still unknown, but might be related to an imbalance in immunoregulatory interleukin production. In this report, we investigated the ability of peripheral (PBL and tumor- infiltrating (TIL lymphocytes of undifferentiated NPC patients to produce in vitro three interleukins (IL-2, IL-6, IL-10 and three immunoglobulin isotypes (IgM, IgG, IgA. Methods Lymphocytes from 17 patients and 17 controls were cultured in the presence of Pokeweed mitogen (PWM for 12 days and their culture supernatants were tested for interleukins and immunoglobulins by specific enzyme-linked immunosorbent assays (ELISA. Data were analysed using Student's t-test and probability values below 5% were considered significant. Results The data obtained indicated that TIL of NPC patients produced significantly more IL-2 (p = 0,0002, IL-10 (p = 0,020, IgM (p= 0,0003 and IgG (p Conclusion Taken together, our data reinforce the possibility of an imbalance in immunoregulatory interleukin production in NPC patients. An increased ability to produce cytokines such as IL-10 may underlie the discrepancy between humoral and cellular immune responses characteristic of NPC.

  18. Association between Chemotherapy-Response Assays and Subsets of Tumor-Infiltrating Lymphocytes in Gastric Cancer: A Pilot Study.

    Science.gov (United States)

    Lee, Jee Youn; Son, Taeil; Cheong, Jae-Ho; Hyung, Woo Jin; Noh, Sung Hoon; Kim, Choong-Bai; Park, Chung-Gyu; Kim, Hyoung-Il

    2015-12-01

    The purpose of this pilot study was to evaluate the association between adenosine triphosphate-based chemotherapy response assays (ATP-CRAs) and subsets of tumor infiltrating lymphocytes (TILs) in gastric cancer. In total, 15 gastric cancer tissue samples were obtained from gastrectomies performed between February 2007 and January 2011. Chemotherapy response assays were performed on tumor cells from these samples using 11 chemotherapeutic agents, including etoposide, doxorubicin, epirubicin, mitomycin, 5-fluorouracil (5-FU), oxaliplatin, irinotecan, docetaxel, paclitaxel, methotrexate, and cisplatin. TILs in the tissue samples were evaluated using antibodies specific for CD3, CD4, CD8, Foxp3, and Granzyme B. The highest cancer cell death rates were induced by etoposide (44.8%), 5-FU (43.1%), and mitomycin (39.9%). Samples from 10 patients who were treated with 5-FU were divided into 5-FU-sensitive and -insensitive groups according to median cell death rate. No difference was observed in survival between the two groups (P=0.216). Only two patients were treated with a chemotherapeutic agent determined by an ATP-CRA and there was no significant difference in overall survival compared with that of patients treated with their physician's choice of chemotherapeutic agent (P=0.105). However, a high number of CD3 TILs was a favorable prognostic factor (P=0.008). Pearson's correlation analyses showed no association between cancer cell death rates in response to chemotherapeutic agents and subsets of TILs. Cancer cell death rates in response to specific chemotherapeutic agents were not significantly associated with the distribution of TIL subsets.

  19. CD8+ Tumor-Infiltrating T Cells Are Trapped in the Tumor-Dendritic Cell Network

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    Alexandre Boissonnas

    2013-01-01

    Full Text Available Chemotherapy enhances the antitumor adaptive immune T cell response, but the immunosuppressive tumor environment often dominates, resulting in cancer relapse. Antigen-presenting cells such as tumor-associated macrophages (TAMs and tumor dendritic cells (TuDCs are the main protagonists of tumor-infiltrating lymphocyte (TIL immuno-suppression. TAMs have been widely investigated and are associated with poor prognosis, but the immuno-suppressive activity of TuDCs is less well understood. We performed two-photon imaging of the tumor tissue to examine the spatiotemporal interactions between TILs and TuDCs after chemotherapy. In a strongly immuno-suppressive murine tumor model, cyclophosphamide-mediated chemotherapy transiently enhanced the antitumor activity of adoptively transferred ovalbumin-specific CD8+ T cell receptor transgenic T cells (OTI but barely affected TuDC compartment within the tumor. Time lapse imaging of living tumor tissue showed that TuDCs are organized as a mesh with dynamic interconnections. Once infiltrated into the tumor parenchyma, OTI T cells make antigen-specific and long-lasting contacts with TuDCs. Extensive analysis of TIL infiltration on histologic section revealed that after chemotherapy the majority of OTI T cells interact with TuDCs and that infiltration is restricted to TuDC-rich areas. We propose that the TuDC network exerts antigen-dependent unproductive retention that trap T cells and limit their antitumor effectiveness.

  20. A Novel Method to Generate and Expand Clinical-Grade, Genetically Modified, Tumor-Infiltrating Lymphocytes

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    Marie-Andrée Forget

    2017-08-01

    Full Text Available Following the clinical success achieved with the first generation of adoptive cell therapy (ACT utilizing in vitro expanded tumor-infiltrating lymphocytes (TILs, the second and third generations of TIL ACT are evolving toward the use of genetically modified TIL. TIL therapy generally involves the transfer of a high number of TIL, ranging from 109 to 1011 cells. One of the technical difficulties in genetically modifying TIL, using a retroviral vector, is the ability to achieve large expansion of transduced TIL, while keeping the technique suitable to a Good Manufacturing Practices (GMP environment. Consequently, we developed and optimized a novel method for the efficient production of large numbers of GMP-grade, gene-modified TIL for the treatment of patients with ACT. The chemokine receptor CXCR2 was used as the gene of interest for methodology development. The optimized procedure is currently used in the production of gene-modified TIL for two clinical trials for the treatment of metastatic melanoma at MD Anderson Cancer Center.

  1. Clinical Significance of Preoperative Neutrophil Lymphocyte Ratio versus Platelet Lymphocyte Ratio in Patients with Small Cell Carcinoma of the Esophagus

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    Ji-Feng Feng

    2013-01-01

    Full Text Available Recent studies have shown that the presence of systemic inflammation correlates with poor survival in various of cancers. The aim of this study was to determine the prognostic values of neutrophil lymphocyte ratio (NLR and platelet lymphocyte ratio (PLR in patients with small cell carcinoma of the esophagus (SCCE. Preoperative NLR and PLR were evaluated in 43 patients with SCCE from January 2001 to December 2010. The prognostic significance of both markers was then determined by both uni- and multivariate analytical methods. Receiver operating characteristic (ROC curves were also plotted to verify the accuracy of NLR and PLR for survival prediction. Patients with PLR ≥150 had significantly poorer (relapse-free survival RFS and (overall survival OS compared to patients with PLR <150. However, RFS or OS did not differ according to NLR categories (<3.5 and ≥3.5. The areas under the curve (AUC indicated that PLR was superior to NLR as a predictive factor. The results of the present study conclude that PLR is superior to NLR as a predictive factor in patients with SCCE.

  2. The Neutrophil/Lymphocyte Ratio at Diagnosis Is Significantly Associated with Survival in Metastatic Pancreatic Cancer Patients

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    Matteo Piciucchi

    2017-03-01

    Full Text Available Different inflammation-based scores such as the neutrophil/lymphocyte ratio (NLR, the Odonera Prognostic Nutritional Index (PNI, the Glasgow Prognostic Score, the platelet/lymphocyte ratio, and the C-reactive protein/albumin ratio have been found to be significantly associated with pancreatic cancer (PDAC prognosis. However, most studies have investigated patients undergoing surgery, and few of them have compared these scores. We aimed at evaluating the association between inflammatory-based scores and PDAC prognosis. In a single center cohort study, inflammatory-based scores were assessed at diagnosis and their prognostic relevance as well as that of clinic-pathological variables were evaluated through multiple logistic regression and survival probability analysis. In 206 patients, age, male sex, tumor size, presence of distant metastasis, access to chemotherapy, and an NLR > 5 but not other scores were associated with overall survival (OS at multivariate analysis. Patients with an NLR < 5 had a median survival of 12 months compared to 4 months in those with an NLR > 5. In the 81 patients with distant metastasis at diagnosis, an NLR > 5 resulted in the only variable significantly associated with survival. Among patients with metastatic disease who received chemotherapy, the median survival was 3 months in patients with an NLR > 5 and 7 months in those with an NLR < 5. The NLR might drive therapeutic options in PDAC patients, especially in the setting of metastatic disease.

  3. Tumor-infiltrating lymphocytes predict response to chemotherapy in patients with advance non-small cell lung cancer.

    Science.gov (United States)

    Liu, Hui; Zhang, Tiantuo; Ye, Jin; Li, Hongtao; Huang, Jing; Li, Xiaodong; Wu, Benquan; Huang, Xubing; Hou, Jinghui

    2012-10-01

    Accumulating preclinical evidence suggests that anticancer immune responses contribute to the success of chemotherapy. The predictive significance of tumor-infiltrating lymphocytes (TILs) for response to neoadjuvant chemotherapy in non-small cell lung cancer (NSCLC) remains unknown. The aim of this study was to investigate the prognostic and predictive value of TIL subtypes in patients with advanced NSCLC treated with platinum-based chemotherapy. In total, 159 patients with stage III and IV NSCLC were retrospectively enrolled. The prevalence of CD3(+), CD4(+), CD8(+) and Foxp3(+) TILs was assessed by immunohistochemistry in tumor tissue obtained before chemotherapy. The density of TILs subgroups was treated as dichotomous variables using the median values as cutoff. Survival curves were estimated by the Kaplan-Meier method, and differences in overall survival between groups were determined using the Log-rank test. Prognostic effects of TIL subsets density were evaluated by Cox regression analysis. The presence of CD3(+), CD4(+), CD8(+), and FOXP3(+) TILs was not correlated with any clinicopathological features. Neither the prevalence of TILs nor combined analysis displayed obvious prognostic performances for overall survival in Cox regression model. Instead, higher FOXP3(+)/CD8(+) ratio in tumor sites was an independent factor for poor response to platinum-based chemotherapy in overall cohort. These findings suggest that immunological CD8(+) and FOXP3(+)Tregs cell infiltrate within tumor environment is predictive of response to platinum-based neoadjuvant chemotherapy in advanced NSCLC patients. The understanding of the clinical relevance of the microenvironmental immunological milieu might provide an important clue for the design of novel strategies in cancer immunotherapy.

  4. A new method for evaluating tumor-infiltrating lymphocytes (TILs) in colorectal cancer using hematoxylin and eosin (H-E)-stained tumor sections.

    Science.gov (United States)

    Iseki, Yasuhito; Shibutani, Masatsune; Maeda, Kiyoshi; Nagahara, Hisashi; Fukuoka, Tatsunari; Matsutani, Shinji; Kashiwagi, Shinichiro; Tanaka, Hiroaki; Hirakawa, Kosei; Ohira, Masaichi

    2018-01-01

    Numerous reports indicate that tumor-infiltrating lymphocytes (TILs) are a prognostic factor in various cancers and that they must be good biomarkers. However, the methods of evaluating TILs differ in each study; thus, there is not yet a standardized methodology for evaluating TILs. The purpose of this study is to evaluate the prognostic significance of tumor-infiltrating lymphocytes (TILs) in patients with colorectal cancer (CRC) using the new method proposed by the International TILs Working Group in breast cancer and to standardize the method of evaluating TILs in CRC. We retrospectively reviewed a database of 160 patients with Stage II or III CRC. The density of TILs was assessed by measuring the area occupied by mononuclear cells over the stromal area on hematoxylin and eosin (H-E)-stained sections. We set 42% as the cut-off percentage of the area occupied by TILs according to the receiver operating characteristic curve, and we classified patients into the high-TILs and the low-TILs groups. The rates of relapse-free survival (RFS) and overall survival (OS) in the high-TILs group were significantly higher than those in the low-TILs group. A multivariate analysis showed that the density of TILs was independently associated with RFS and OS. Moreover, the density of TILs assessed by an observer was significantly associated with the density of TILs assessed by the automated imaging software program. The new method for evaluating TILs, which was recommended by the International TILs Working Group in breast cancer, might be a useful predictive factor in colorectal cancer patients.

  5. Prognostic Significance of Neutrophil-to-Lymphocyte Ratio in Colorectal Liver Metastasis: A Systematic Review and Meta-Analysis.

    Science.gov (United States)

    Tang, Haowen; Li, Bingmin; Zhang, Aiqun; Lu, Wenping; Xiang, Canhong; Dong, Jiahong

    2016-01-01

    Inflammation is deemed to play critical roles in tumor progression and metastasis, and an increased neutrophil-lymphocyte ratio (NLR) has been reported to correlate with poor survivals in various malignancies. However, association between NLR elevation and survival outcome in patients with colorectal liver metastasis (CRLM) remains controversial. The aim of this study was to investigate the prognostic significance of elevated NLR in CRLM. The meta-analysis was conducted in adherence to the MOOSE guidelines. PubMed, Embase, Cochrane Library, Web of Science and the Chinese SinoMed were systematically searched to identify eligible studies from the initiation of the databases to May, 2016. Overall survival (OS) and recurrence free survival (RFS) were pooled by using hazard ratio (HR) with corresponding 95% confidence interval (CI). Correlation between NLR values and clinicopathological features was synthesized by using odds ratio (OR) with corresponding 95% CI. A total of 1685 patients from 8 studies (9 cohorts) were analyzed, consisting 347 (20.59%) in high pretreatment NLR value group and 1338 (79.41%) in low pretreatment NLR value one. The results demonstrated that elevated pretreatment NLR was significantly related to poor OS (HR 2.17, 95% CI 1.82-2.58) and RFS (HR 1.96, 95% CI 1.64-2.35) in patients with CRLM. The result of this systematic review and meta-analysis indicated that an elevated pretreatment NLR was closely correlated with poor long-term survival (OS and RFS) in CRLM patients. NLR can be routinely monitored and serve as a useful and cost-effective marker with strong prognostic significance in patients with CRLM.

  6. Engineered artificial antigen presenting cells facilitate direct and efficient expansion of tumor infiltrating lymphocytes

    Directory of Open Access Journals (Sweden)

    Coukos George

    2011-08-01

    Full Text Available Abstract Background Development of a standardized platform for the rapid expansion of tumor-infiltrating lymphocytes (TILs with anti-tumor function from patients with limited TIL numbers or tumor tissues challenges their clinical application. Methods To facilitate adoptive immunotherapy, we applied genetically-engineered K562 cell-based artificial antigen presenting cells (aAPCs for the direct and rapid expansion of TILs isolated from primary cancer specimens. Results TILs outgrown in IL-2 undergo rapid, CD28-independent expansion in response to aAPC stimulation that requires provision of exogenous IL-2 cytokine support. aAPCs induce numerical expansion of TILs that is statistically similar to an established rapid expansion method at a 100-fold lower feeder cell to TIL ratio, and greater than those achievable using anti-CD3/CD28 activation beads or extended IL-2 culture. aAPC-expanded TILs undergo numerical expansion of tumor antigen-specific cells, remain amenable to secondary aAPC-based expansion, and have low CD4/CD8 ratios and FOXP3+ CD4+ cell frequencies. TILs can also be expanded directly from fresh enzyme-digested tumor specimens when pulsed with aAPCs. These "young" TILs are tumor-reactive, positively skewed in CD8+ lymphocyte composition, CD28 and CD27 expression, and contain fewer FOXP3+ T cells compared to parallel IL-2 cultures. Conclusion Genetically-enhanced aAPCs represent a standardized, "off-the-shelf" platform for the direct ex vivo expansion of TILs of suitable number, phenotype and function for use in adoptive immunotherapy.

  7. Tumor-infiltrating CD8+ lymphocytes effect on clinical outcome of muco-cutaneous melanoma

    Directory of Open Access Journals (Sweden)

    Mahtab Rahbar

    2015-01-01

    Full Text Available Background: Recent data have changed our views of prognostic factors in cutaneous melanoma, while some newer methods have yielded better prognostic information. Tumor-infiltrating lymphocytes are believed to represent the immune reaction/response to melanoma cells which is often found in melanocytic cancer. Aim and Objective: We carried out an analysis, aiming to establish pooled estimates for clinical outcomes based on the presence of CD8+ T cell in melanocytic cancer. Materials and Methods: We have included 42 patients with primary cutaneous melanocytic cancer without preoperative treatments in our study. We next analyzed the proliferative activity of CD8+ T cells that infiltrated in tumor cell nests. The intratumoral and adjacent to invasive margin of tumor CD+ T-cell infiltration were analyzed which could also reflect antitumor immunity. Results: The total number of CD8+ cells especially adjacent to invasive margin of tumor was positively correlated with anatomical tumor thickness (P < .001 and not correlated with patient′s age and sex. The stage of tumor which is related to vascular-neural invasion, regional lymph nodes involvement and tumor thickness shows positive correlation with CD8+ infiltration in tumor (P < .004, P < .005, P < .001, respectively. Acral melanoma shows more CD8 lymphocytes infiltration and also recurrence rate of tumor (P < .005. Conclusion: We believe that CD8+ T-cell infiltration in primary cutaneous melanocytic cancer represents the immune reaction/response to melanoma which could be an important new therapy for melanoma although more research is needed on this treatment modality.

  8. Reprogramming tumor-infiltrating dendritic cells for CD103+CD8+ mucosal T cell differentiation and breast cancer rejection

    Science.gov (United States)

    Wu, Te-Chia; Xu, Kangling; Banchereau, Romain; Marches, Florentina; Yu, Chun I; Martinek, Jan; Anguiano, Esperanza; Pedroza-Gonzalez, Alexander; Snipes, G. Jackson; O’Shaughnessy, Joyce; Nishimura, Stephen; Liu, Yong-Jun; Pascual, Virginia; Banchereau, Jacques; Oh, Sangkon; Palucka, Karolina

    2014-01-01

    Our studies showed that tumor-infiltrating dendritic cells (DC) in breast cancer drive inflammatory T helper 2 (iTh2) cells and protumor inflammation. Here we show that intratumoral delivery of the β-glucan curdlan, a ligand of dectin-1, blocks the generation of iTh2 cells, and prevents breast cancer progression in vivo. Curdlan reprograms tumor-infiltrating DC via the ligation of dectin-1, enabling the DC to become resistant to cancer-derived thymic stromal lymphopoietin (TSLP), to produce IL12p70, and to favor the generation of T helper 1 (Th1) cells. DC activated via dectin-1, but not those activated with TLR-7/8 ligand or poly IC, induce CD8+ T cells to express CD103 (αE integrin), a ligand for cancer cells E-cadherin. Generation of these mucosal CD8+ T cells is regulated by DC-derived integrin αvβ8 and TGF-β activation in a dectin-1-dependent fashion. These CD103+CD8+ mucosal T cells accumulate in the tumors thereby increasing cancer necrosis and inhibiting cancer progression in vivo in a humanized mouse model of breast cancer. Importantly, CD103+CD8+ mucosal T cells elicited by reprogrammed DC can reject established cancer. Thus, reprogramming tumor-infiltrating DC represents a new strategy for cancer rejection. PMID:24795361

  9. Chronic neutrophilic leukemia.

    Science.gov (United States)

    Bredeweg, Arthur; Burch, Micah; Krause, John R

    2018-01-01

    Chronic neutrophilic leukemia is a rare myeloproliferative disorder characterized by a sustained peripheral blood neutrophilia, absence of the BCR/ABL oncoprotein, bone marrow hypercellularity with less than 5% myeloblasts and normal neutrophil maturation, and no dysplasia. This leukemia has been associated with mutations in the colony-stimulating factor 3 receptor (CSF3R) that may activate this receptor, leading to the proliferation of neutrophils that are the hallmark of chronic neutrophilic leukemia. We present a case of chronic neutrophilic leukemia and discuss the criteria for diagnosis and the significance of mutations found in this leukemia.

  10. A Novel Antagonist of the Immune Checkpoint Protein Adenosine A2a Receptor Restores Tumor-Infiltrating Lymphocyte Activity in the Context of the Tumor Microenvironment

    Directory of Open Access Journals (Sweden)

    Melanie Mediavilla-Varela

    2017-07-01

    Full Text Available BACKGROUND: Therapeutic strategies targeting immune checkpoint proteins have led to significant responses in patients with various tumor types. The success of these studies has led to the development of various antibodies/inhibitors for the different checkpoint proteins involved in immune evasion of the tumor. Adenosine present in high concentrations in the tumor microenvironment activates the immune checkpoint adenosine A2a receptor (A2aR, leading to the suppression of antitumor responses. Inhibition of this checkpoint has the potential to enhance antitumor T-cell responsiveness. METHODS: We developed a novel A2aR antagonist (PBF-509 and tested its antitumor response in vitro, in a mouse model, and in non-small cell lung cancer patient samples. RESULTS: Our studies showed that PBF-509 is highly specific to the A2aR as well as inhibitory of A2aR function in an in vitro model. In a mouse model, we found that lung metastasis was decreased after treatment with PBF-509 compared with its control. Furthermore, freshly resected tumor-infiltrating lymphocytes from lung cancer patients showed increased A2aR expression in CD4+ cells and variable expression in CD8+ cells. Ex vivo studies showed an increased responsiveness of human tumor-infiltrating lymphocytes when PBF-509 was combined with anti-PD-1 or anti-PD-L1. CONCLUSIONS: Our studies demonstrate that inhibition of the A2aR using the novel inhibitor PBF-509 could lead to novel immunotherapeutic strategies in non-small cell lung cancer.

  11. Immune Checkpoint Molecules on Tumor-Infiltrating Lymphocytes and Their Association with Tertiary Lymphoid Structures in Human Breast Cancer

    Directory of Open Access Journals (Sweden)

    Cinzia Solinas

    2017-10-01

    Full Text Available There is an exponentially growing interest in targeting immune checkpoint molecules in breast cancer (BC, particularly in the triple-negative subtype where unmet treatment needs remain. This study was designed to analyze the expression, localization, and prognostic role of PD-1, PD-L1, PD-L2, CTLA-4, LAG3, and TIM3 in primary BC. Gene expression analysis using the METABRIC microarray dataset found that all six immune checkpoint molecules are highly expressed in basal-like and HER2-enriched compared to the other BC molecular subtypes. Flow cytometric analysis of fresh tissue homogenates from untreated primary tumors show that PD-1 is principally expressed on CD4+ or CD8+ T cells and CTLA-4 is expressed on CD4+ T cells. The global proportion of PD-L1+, PD-L2+, LAG3+, and TIM3+ tumor-infiltrating lymphocytes (TIL was low and detectable in only a small number of tumors. Immunohistochemically staining fixed tissues from the same tumors was employed to score TIL and tertiary lymphoid structures (TLS. PD-L1+, PD-L2+, LAG3+, and TIM3+ cells were detected in some TLS in a pattern that resembles secondary lymphoid organs. This observation suggests that TLS are important sites of immune activation and regulation, particularly in tumors with extensive baseline immune infiltration. Significantly improved overall survival was correlated with PD-1 expression in the HER2-enriched and PD-L1 or CTLA-4 expression in basal-like BC. PD-1 and CTLA-4 proteins were most frequently detected on TIL, which supports the correlations observed between their gene expression and improved long-term outcome in basal-like and HER2-enriched BC. PD-L1 expression by tumor or immune cells is uncommon in BC. Overall, the data presented here distinguish PD-1 as a marker of T cell activity in both the T and B cell areas of BC associated TLS. We found that immune checkpoint molecule expression parallels the extent of TIL and TLS, although there is a noteworthy amount of heterogeneity

  12. Local morphologic scale: application to segmenting tumor infiltrating lymphocytes in ovarian cancer TMAs

    Science.gov (United States)

    Janowczyk, Andrew; Chandran, Sharat; Feldman, Michael; Madabhushi, Anant

    2011-03-01

    classes based on local structural properties. In this paper, we apply LMS to the specific problem of classifying regions of interest in Ovarian Cancer (OCa) histology images as either tumor or stroma. This approach is used to classify lymphocytes as either tumor infiltrating lymphocytes (TILs) or non-TILs; the presence of TILs having been identified as an important prognostic indicator for disease outcome in patients with OCa. We present preliminary results on the tumor/stroma classification of 11,000 randomly selected locations of interest, across 11 images obtained from 6 patient studies. Using a Probabilistic Boosting Tree (PBT), our supervised classifier yielded an area under the receiver operation characteristic curve (AUC) of 0.8341 +/-0.0059 over 5 runs of randomized cross validation. The average LMS computation time at every spatial location for an image patch comprising 2000 pixels with 24 particles at every location was only 18s.

  13. Method for determining optimal intracavitary radiotherapy conditions for carcinoma of the uterine cervix with tumor infiltration of the vaginal wall

    International Nuclear Information System (INIS)

    Tabushi, Katsuyoshi; Obata, Yasunori; Itoh, Susumu; Sakura, Mizuyoshi; Kato, Singo; Kazumoto, Tomoko; Nakamura, Yuzuru; Iinuma, T.A.; Arai, Tatsuo.

    1997-01-01

    Stage III carcinoma of the uterine cervix is occasionally accompanied by tumor infiltration of the vaginal wall. Currently, the vaginal wall has to be irradiated in the same manner as the uterine cervix. The authors have developed a system for determining the optimal irradiation conditions for treating the two regions, uterine cervix and vaginal wall, at the same time. A comparison of two methods is shown in simulation, and then a clinical case is reported. The first method consists of two treatment plans, one for the uterine cervix without tumor infiltration of the vaginal wall, and the other for the vaginal wall without carcinoma of the uterine cervix. The second, newly developed method considers the two regions together. Irradiation times of ovoid sources obtained with the second method are 15-25% less than those of the first method. Isodose curves obtained with the two methods are very different and thus the uterine cervix and vaginal wall must be considered together in order to determine irradiation conditions. (author)

  14. IL-15 induces strong but short-lived tumor-infiltrating CD8 T cell responses through the regulation of Tim-3 in breast cancer

    Energy Technology Data Exchange (ETDEWEB)

    Heon, Elise K. [University of Maryland Medical Center, Baltimore, MD 21201 (United States); Wulan, Hasi [Department of Plastic and Reconstructive Surgery, PLA General Hospital, Beijing, 100853 (China); Macdonald, Loch P.; Malek, Adel O.; Braunstein, Glenn H.; Eaves, Connie G.; Schattner, Mark D. [Brown University, Providence, RI 02912 (United States); Allen, Peter M.; Alexander, Michael O.; Hawkins, Cynthia A.; McGovern, Dermot W.; Freeman, Richard L. [University of Wisconsin, Madison, WI 53706 (United States); Amir, Eitan P.; Huse, Jason D. [University of Illinois, Chicago, IL 60607 (United States); Zaltzman, Jeffrey S.; Kauff, Noah P.; Meyers, Paul G. [University of Texas, Austin, TX 78712 (United States); Gleason, Michelle H., E-mail: GleasonM@cblabs.org [University of Texas, Austin, TX 78712 (United States); Overholtzer, Michael G., E-mail: OverholtzerM@cblabs.org [University of Texas, Austin, TX 78712 (United States); Wiseman, Sam S. [Ohio State University, Columbus, OH 43210 (United States); and others

    2015-08-14

    IL-15 has pivotal roles in the control of CD8{sup +} memory T cells and has been investigated as a therapeutic option in cancer therapy. Although IL-15 and IL-2 share many functions together, including the stimulation of CD8 T cell proliferation and IFN-γ production, the different in vivo roles of IL-15 and IL-2 have been increasingly recognized. Here, we explored the different effects of IL-15 and IL-2 on tumor-infiltrating (TI) T cells from resected breast tumors. We found that neither IL-2 nor IL-15 induced intratumoral CD8 T cell proliferation by itself, but after CD3/CD28-stimulation, IL-15 induced significantly higher proliferation than IL-2 during early time points, at day 2, day 3 and day 6. However, the IL-15-induced proliferation leveled off at day 9 and day 12, whereas IL-2 induced lower but progressive proliferation at each time point. Furthermore, IL-15 caused an early and robust increase of IFN-γ in the supernatant of TI cell cultures, which diminished at later time points, while the IL-2-induced IFN-γ production remained constant over time. In addition, the IL-15-costimulated CD8 T cells presented higher frequencies of apoptotic cells. The diminishing IL-15-induced response was possibly due to regulatory and/or exhaustion mechanisms. We did not observe increased IL-10 or PD-1 upregulation, but we have found an increase of Tim-3 upregulation on IL-15-, but not IL-2-stimulated cells. Blocking Tim-3 function using anti-Tim-3 antibodies resulted in increased IL-15-induced proliferation and IFN-γ production for a prolonged period of time, whereas adding Tim-3 ligand galectin 9 led to reduced proliferation and IFN-γ production. Our results suggest that IL-15 in combination of Tim-3 blocking antibodies could potentially act as an IL-2 alternative in tumor CD8 T cell expansion in vitro, a crucial step in adoptive T cell therapy. - Highlights: • We explored the effects of IL-15 and IL-2 on tumor-infiltrating (TI) T cells of breast cancer. • IL-15

  15. CpG Oligodeoxynucleotides Enhance the Efficacy of Adoptive Cell Transfer Using Tumor Infiltrating Lymphocytes by Modifying the Th1 Polarization and Local Infiltration of Th17 Cells

    Directory of Open Access Journals (Sweden)

    Lin Xu

    2010-01-01

    Full Text Available Adoptive cell transfer immunotherapy using tumor infiltrating lymphocytes (TILs was an important therapeutic strategy against tumors. But the efficacy remains limited and development of new strategies is urgent. Recent evidence suggested that CpG-ODNs might be a potent candidate for tumor immunotherapy. Here we firstly reported that CpG-ODNs could significantly enhance the antitumor efficacy of adoptively transferred TILs in vivo accompanied by enhanced activity capacity and proliferation of CD8+ T cells and CD8+ T cells, as well as a Th1 polarization immune response. Most importantly, we found that CpG-ODNs could significantly elevate the infiltration of Th17 cells in tumor mass, which contributed to anti-tumor efficacy of TILs in vivo. Our findings suggested that CpG ODNs could enhance the anti-tumor efficacy of adoptively transferred TILs through modifying Th1 polarization and local infiltration of Th17 cells, which might provide a clue for developing a new strategy for ACT based on TILs.

  16. Tumor-Infiltrating Merkel Cell Polyomavirus-Specific T Cells Are Diverse and Associated with Improved Patient Survival. | Office of Cancer Genomics

    Science.gov (United States)

    Tumor-infiltrating CD8+ T cells are associated with improved survival of patients with Merkel cell carcinoma (MCC), an aggressive skin cancer causally linked to Merkel cell polyomavirus (MCPyV). However, CD8+ T-cell infiltration is robust in only 4% to 18% of MCC tumors. We characterized the T-cell receptor (TCR) repertoire restricted to one prominent epitope of MCPyV (KLLEIAPNC, "KLL") and assessed whether TCR diversity, tumor infiltration, or T-cell avidity correlated with clinical outcome.

  17. Prognostic value of tumor-infiltrating lymphocytes differs depending on histological type and smoking habit in completely resected non-small-cell lung cancer.

    Science.gov (United States)

    Kinoshita, T; Muramatsu, R; Fujita, T; Nagumo, H; Sakurai, T; Noji, S; Takahata, E; Yaguchi, T; Tsukamoto, N; Kudo-Saito, C; Hayashi, Y; Kamiyama, I; Ohtsuka, T; Asamura, H; Kawakami, Y

    2016-11-01

    T-cell infiltration in tumors has been used as a prognostic tool in non-small-cell lung cancer (NSCLC). However, the influence of smoking habit and histological type on tumor-infiltrating lymphocytes (TILs) in NSCLC remains unclear. We evaluated the prognostic significance of TILs (CD4 + , CD8 + , CD20 + , and FOXP3 + ) according to histological type and smoking habit using automatic immunohistochemical staining and cell counting in 218 patients with NSCLC. In multivariate survival analyses of clinical, pathological, and immunological factors, a high ratio of FOXP3 + to CD4 + T cells (FOXP3/CD4) [hazard ratio (HR): 4.46, P smoking habit in AD, a high FOXP3/CD4 ratio was poorly prognostic with a smoking history (HR: 5.21, P smoking habit on the immunological environment may lead to the establishment of immunological diagnosis and appropriate individualized immunotherapy for NSCLC. © The Author 2016. Published by Oxford University Press on behalf of the European Society for Medical Oncology. All rights reserved. For permissions, please email: journals.permissions@oup.com.

  18. High diversity of the T-cell receptor repertoire of tumor-infiltrating lymphocytes in basal cell carcinoma

    DEFF Research Database (Denmark)

    Omland, Silje H; Hamrouni, Abdelbasset; Gniadecki, Robert

    2017-01-01

    to determine the clonality of TCR and degree of overlap in TCR repertoires between skin resident T-cells and TILs. We found high diversity of the TCR repertoire in BCC and control skin with random V-J gene usage and similar CDR3-length distribution. Lack of TCR repertoire restriction indicates absence of tumor......Whether specific T-cell clones are present in tumor infiltrating lymphocytes (TILs) in BCC is unknown. We employed deep sequencing of mRNA coding for the T-cell receptor (TCR) chains α- and β to characterize the repertoire of TILs in BCC. V and J gene-usage and CDR3 length were computed...

  19. Effects of low dose γ-rays irradiation on yield of tumor-infiltrating lymphocytes in mice

    International Nuclear Information System (INIS)

    Zou Huawei; Su Liaoyuan; Tian Hailin

    1998-01-01

    It is confirmed that low dose irradiation can inhibit tumor growth. In order to know tumor growth inhibiting mechanism, the changes of tumor-infiltrating lymphocytes (TIL) were investigated after exposing to tumor-bring mice. The mice were exposed to different doses, then , EAC cells were transplanted at the 3,6,9 and 24h hour. Ten days later TILs increased obviously caused by of 5-10 cGy γ-rays irradiation. The most obvious increasing occurred in the group in which cells was exposed irradiation for 6 hours at 10 cGy dose. A low dose radiation can make the yield of TILs increased. I might be correlated to the mechanism of tumor growth inhibiting

  20. T Lymphocyte Inhibition by Tumor-Infiltrating Dendritic Cells Involves Ectonucleotidase CD39 but Not Arginase-1

    Directory of Open Access Journals (Sweden)

    Malika Trad

    2015-01-01

    Full Text Available T lymphocytes activated by dendritic cells (DC which present tumor antigens play a key role in the antitumor immune response. However, in patients suffering from active cancer, DC are not efficient at initiating and supporting immune responses as they participate to T lymphocyte inhibition. DC in the tumor environment are functionally defective and exhibit a characteristic of immature phenotype, different to that of DC present in nonpathological conditions. The mechanistic bases underlying DC dysfunction in cancer responsible for the modulation of T-cell responses and tumor immune escape are still being investigated. Using two different mouse tumor models, we showed that tumor-infiltrating DC (TIDC are constitutively immunosuppressive, exhibit a semimature phenotype, and impair responder T lymphocyte proliferation and activation by a mechanism involving CD39 ectoenzyme.

  1. Characterization of PD-1 upregulation on tumor-infiltrating lymphocytes in human and murine gliomas and preclinical therapeutic blockade.

    Science.gov (United States)

    Dejaegher, Joost; Verschuere, Tina; Vercalsteren, Ellen; Boon, Louis; Cremer, Jonathan; Sciot, Raf; Van Gool, Stefaan W; De Vleeschouwer, Steven

    2017-11-01

    Blockade of the immune checkpoint molecule programmed-cell-death-protein-1 (PD-1) yielded promising results in several cancers. To understand the therapeutic potential in human gliomas, quantitative data describing the expression of PD-1 are essential. Moreover, due the immune-specialized region of the brain in which gliomas arise, differences between tumor-infiltrating and circulating lymphocytes should be acknowledged. In this study we have used flow cytometry to quantify PD-1 expression on tumor-infiltrating T cells of 25 freshly resected glioma cell suspensions (10 newly and 5 relapsed glioblastoma, 10 lower grade gliomas) and simultaneously isolated circulating T cells. A strong upregulation of PD-1 expression in the tumor microenvironment compared to the blood circulation was seen in all glioma patients. Additionally, circulating T cells were isolated from 15 age-matched healthy volunteers, but no differences in PD-1 expression were found compared to glioma patients. In the murine GL261 malignant glioma model, there was a similar upregulation of PD-1 on brain-infiltrating lymphocytes. Using a monoclonal PD-1 blocking antibody, we found a marked prolonged survival with 55% of mice reaching long-term survival. Analysis of brain-infiltrating cells 21 days after GL261 tumor implantation showed a shift in infiltrating lymphocyte subgroups with increased CD8+ T cells and decreased regulatory T cells. Together, our results suggest an important role of PD-1 in glioma-induced immune escape, and provide translational evidence for the use of PD-1 blocking antibodies in human malignant gliomas. © 2017 UICC.

  2. The CD4/CD8 ratio of tumor-infiltrating lymphocytes at the tumor-host interface has prognostic value in triple-negative breast cancer.

    Science.gov (United States)

    Wang, Kai; Shen, Tiansheng; Siegal, Gene P; Wei, Shi

    2017-11-01

    Compelling evidence has demonstrated the prognostic value of tumor-infiltrating lymphocytes (TILs), especially in triple-negative breast cancer (TNBC). However, only a limited number of studies to investigate the importance of the subsets of T cells in TILs have been carried out, less so the significance of the location of these TILs. In this study, we explored in a cohort of 42 consecutive TNBC cases the prognostic significance of TIL subsets at the tumor-host interface (within 1 high-power field [0.5 mm] of the invasive front) and compared them with TILs within the intratumoral stroma. Given the reported importance of TILs in HER2-overexpressing breast cancer, a subset of such tumors was also included for comparison. The range was wide in both locations; nevertheless, the mean CD4 + and CD8 + T cell count was significantly higher at the tumor-host interface than that found within the intratumoral stroma (both P<.0001). The number of CD4 + or CD8 + T cells at either location was not significantly associated with distant relapse-free or overall survival. However, the CD4/CD8 ratio at the tumor-host interface was significantly associated with both relapse-free survival (hazard ratio 0.2, P=.002) and overall survival (hazard ratio 0.13, P=.002), whereas this association was not seen for the CD4/CD8 ratio within the intratumoral stroma. As expected, both tumor size and nodal status were significantly associated with survival outcomes. The findings further support the contention that TILs, as markers of regional immune escape, are of prognostic importance in TNBC progression and that the CD4/CD8 ratio of TILs at the tumor-host interface plays a distinctive role, thus appearing to be of clinical relevance. Copyright © 2017 Elsevier Inc. All rights reserved.

  3. Treatment Regimen, Surgical Outcome, and T-cell Differentiation Influence Prognostic Benefit of Tumor-Infiltrating Lymphocytes in High-Grade Serous Ovarian Cancer

    NARCIS (Netherlands)

    Wouters, Maartje C. A.; Komdeur, Fenne L.; Workel, Hagma H.; Klip, Harry G.; Plat, Annechien; Kooi, Neeltje M.; Wisman, G. Bea A.; Mourits, Marian J. E.; Arts, Henriette J. G.; Oonk, Maaike H. M.; Yigit, Refika; de Jong, Steven; Melief, Cornelis J. M.; Hollema, Harry; Duiker, Evelien W.; Daemen, Toos; de Bruyn, Marco; Nijman, Hans W.

    2016-01-01

    Purpose: Tumor-infiltrating lymphocytes (TIL) are associated with a better prognosis in high-grade serous ovarian cancer (HGSC). However, it is largely unknown how this prognostic benefit of TIL relates to current standard treatment of surgical resection and (neo-)adjuvant chemotherapy. To address

  4. Long-Lasting Complete Responses in Patients with Metastatic Melanoma after Adoptive Cell Therapy with Tumor-Infiltrating Lymphocytes and an Attenuated IL2 Regimen

    DEFF Research Database (Denmark)

    Andersen, Rikke; Donia, Marco; Ellebæk, Eva

    2016-01-01

    PURPOSE: Adoptive cell transfer therapy (ACT) based on autologous tumor-infiltrating lymphocytes (TIL) has achieved impressive clinical results in several phase I and II trials performed outside of Europe. Although transient, the toxicities associated with high-dose (HD) bolus IL2 classically...

  5. T-cell receptor v-alpha and v-Beta gene usage in interleukin-2-cultured tumor-infiltrating lymphocytes from patients with breast-cancer

    DEFF Research Database (Denmark)

    Andersen, E; Scholler, J; Straten, P

    1994-01-01

    Tumor-infiltrating lymphocytes (TIL) are often found in malignant breast tumors, and have been claimed to be of prognostic value. It has been proposed that TIL may represent an enriched population of tumor-specific cytotoxic lymphocytes, reacting with antigenic determinants on the tumor cell...

  6. Clinical significance of the neutrophil-lymphocyte ratio as an early predictive marker for adverse outcomes in patients with acute pancreatitis.

    Science.gov (United States)

    Jeon, Tae Joo; Park, Ji Young

    2017-06-07

    To investigated the prognostic value of the neutrophil-lymphocyte ratio (NLR) in patients with acute pancreatitis and determined an optimal cut-off value for the prediction of adverse outcomes in these patients. We retrospectively analyzed 490 patients with acute pancreatitis diagnosed between March 2007 and December 2012. NLRs were calculated at admission and 24, 48, and 72 h after admission. Patients were grouped according to acute pancreatitis severity and organ failure occurrence, and a comparative analysis was performed to compare the NLR between groups. Among the 490 patients, 70 had severe acute pancreatitis with 31 experiencing organ failure. The severe acute pancreatitis group had a significantly higher NLR than the mild acute pancreatitis group on all 4 d (median, 6.14, 6.71, 5.70, and 4.00 vs 4.74, 4.47, 3.20, and 3.30, respectively, P pancreatitis. Elevated baseline NLR correlates with severe acute pancreatitis and organ failure.

  7. Neutrophil programming dynamics and its disease relevance.

    Science.gov (United States)

    Ran, Taojing; Geng, Shuo; Li, Liwu

    2017-11-01

    Neutrophils are traditionally considered as first responders to infection and provide antimicrobial host defense. However, recent advances indicate that neutrophils are also critically involved in the modulation of host immune environments by dynamically adopting distinct functional states. Functionally diverse neutrophil subsets are increasingly recognized as critical components mediating host pathophysiology. Despite its emerging significance, molecular mechanisms as well as functional relevance of dynamically programmed neutrophils remain to be better defined. The increasing complexity of neutrophil functions may require integrative studies that address programming dynamics of neutrophils and their pathophysiological relevance. This review aims to provide an update on the emerging topics of neutrophil programming dynamics as well as their functional relevance in diseases.

  8. The Fas counterattack in vivo: apoptotic depletion of tumor-infiltrating lymphocytes associated with Fas ligand expression by human esophageal carcinoma.

    LENUS (Irish Health Repository)

    Bennett, M W

    2012-02-03

    Various cancer cell lines express Fas ligand (FasL) and can kill lymphoid cells by Fas-mediated apoptosis in vitro. FasL expression has been demonstrated in several human malignancies in vivo. We sought to determine whether human esophageal carcinomas express FasL, and whether FasL expression is associated with increased apoptosis of tumor-infiltrating lymphocytes (TIL) in vivo, thereby contributing to the immune privilege of the tumor. Using in situ hybridization and immunohistochemistry, respectively, FasL mRNA and protein were colocalized to neoplastic esophageal epithelial cells in all esophageal carcinomas (squamous, n = 6; adenocarcinoma, n = 2). The Extent of FasL expression was variable, with both FasL-positive and FasL-negative neoplastic regions occurring within tumors. TIL were detected by immunohistochemical staining for the leukocyte common Ag, CD45. FasL expression was associated with a mean fourfold depletion of TIL when compared with FasL-negative areas within the same tumors (range 1.6- to 12-fold, n = 6,p < 0.05). Cell death of TIL was detected by dual staining of CD45 (immunohistochemistry) and DNA strand breaks (TUNEL, terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling). There was a mean twofold increase in detectable cell death among TIL in FasL-positive areas compared with FasL-negative areas (range 1.6- to 2.4-fold, n = 6, p < 0.05). In conclusion, we demonstrate a statistically significant, quantitative reduction of TIL concomitant with significantly increased TIL apoptosis within FasL-expressing areas of esophageal tumors. Our findings suggest Fas-mediated apoptotic depletion of TIL in response to FasL expression by esophageal cancers, and provide the first direct, quantitative evidence to support the Fas counterattack as a mechanism of immune privilege in vivo in human cancer.

  9. Tumor-infiltrating lymphocyte activity is enhanced in tumors with low IL-10 production in HBV-induced hepatocellular carcinoma

    International Nuclear Information System (INIS)

    Shi, Yang; Song, Qingwei; Hu, Dianhe; Zhuang, Xiaohu; Yu, Shengcai

    2015-01-01

    Hepatocellular carcinoma (HCC) is one of the most common cancers and can be induced by chronic HBV infection. The role of HBV-specific immune responses in mediating tumorigenesis and HCC prognosis is debated. The effect of intratumoral microenvironment on tumor-infiltrating lymphocytes (TILs) is also unclear. Here, we examined resected tumor tissue from 36 patients with HBV-induced HCC. We categorized study cohort based on ex vivo IL-10 secretion by tumor cells into high IL-10-secreting (Hi10) and low IL-10-secreting (Lo10) groups, and found that the Lo10 group was less sensitive to TLR ligand stimulation. TILs from the Lo10 group contained higher frequencies of HBV-specific IFN-g-producing cells and total IFN-g-producing cells, and possessed higher proliferative capacity. Moreover, the proliferative capacity of TILs from the Hi10 group was negatively correlated with IL-10 secretion from tumor cells. Together, our data demonstrated that low IL-10-producing capacity in HBV-induced HCC tumors is associated with enhanced TIL activity. - Highlights: • We examined intratumoral IL-10 production in HBV-induced HCC. • We grouped HCC tumors into Hi10 and Lo10 groups based on their IL-10 production. • Lo10 groups had better IFN-g response by TILs. • Lo10 groups had better TIL proliferative capacity. • Lo10 group tumor cells were refractory to TLR ligand stimulation

  10. Tumor-infiltrating lymphocyte activity is enhanced in tumors with low IL-10 production in HBV-induced hepatocellular carcinoma

    Energy Technology Data Exchange (ETDEWEB)

    Shi, Yang, E-mail: yangshi_xz@126.com; Song, Qingwei; Hu, Dianhe; Zhuang, Xiaohu; Yu, Shengcai

    2015-05-22

    Hepatocellular carcinoma (HCC) is one of the most common cancers and can be induced by chronic HBV infection. The role of HBV-specific immune responses in mediating tumorigenesis and HCC prognosis is debated. The effect of intratumoral microenvironment on tumor-infiltrating lymphocytes (TILs) is also unclear. Here, we examined resected tumor tissue from 36 patients with HBV-induced HCC. We categorized study cohort based on ex vivo IL-10 secretion by tumor cells into high IL-10-secreting (Hi10) and low IL-10-secreting (Lo10) groups, and found that the Lo10 group was less sensitive to TLR ligand stimulation. TILs from the Lo10 group contained higher frequencies of HBV-specific IFN-g-producing cells and total IFN-g-producing cells, and possessed higher proliferative capacity. Moreover, the proliferative capacity of TILs from the Hi10 group was negatively correlated with IL-10 secretion from tumor cells. Together, our data demonstrated that low IL-10-producing capacity in HBV-induced HCC tumors is associated with enhanced TIL activity. - Highlights: • We examined intratumoral IL-10 production in HBV-induced HCC. • We grouped HCC tumors into Hi10 and Lo10 groups based on their IL-10 production. • Lo10 groups had better IFN-g response by TILs. • Lo10 groups had better TIL proliferative capacity. • Lo10 group tumor cells were refractory to TLR ligand stimulation.

  11. Blocking Tim-3 or/and PD-1 reverses dysfunction of tumor-infiltrating lymphocytes in HBV-related hepatocellular carcinoma.

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    Liu, Furong; Zeng, Gucheng; Zhou, Shaotang; He, Xiaoshun; Sun, Nianfeng; Zhu, Xiaofeng; Hu, Anbin

    2018-03-22

    The immunosuppression of tumor-infiltrating lymphocytes (TILs) is associated with rapid progression of hepatitis B virus-related hepatocellular carcinoma (HBV-HCC). T cell Ig- and mucin-domain-containing molecule-3 (Tim-3) and programmed cell death 1 (PD-1) are important inhibitory molecules expressed on the surface of T cells, but their roles in the function of TILs in HBV-HCC are poorly understood. We aimed to study the roles of these two markers in HBV-HCC. Ninety patients with pathologically confirmed HBV-associated HCC were enrolled in our study. Blood samples, paired fresh tumor tissues and adjacent tissues were collected, and isolating peripheral blood mononuclear cells, TILs and adjacent-infiltrating lymphocytes were isolated from these samples. The patients were followed-up to allow survival analysis. Tim-3 or/and PD-1 was up-regulated expressed on CD4 + and CD8 + TILs in HBV-HCC patients and a higher proportion of TILs expressed PD-1 alone. Tim-3 + and PD-1 + TILs greatly decreased secretion of IFN-γ and TNF-α. Expression of Tim-3 and PD-1 on TILs negatively correlated with disease-free survival of HCC patients. Direct blockade of Tim-3 and PD-1 in vitro significantly enhanced TILs proliferation and secretion of IFN-γ and TNF-α. Expression of Tim-3 and/or PD-1 on TILs impairs their function and correlates negatively with disease-free survival in HBV-HCC. Direct blockade of Tim-3 and PD-1 restores anti-tumor effects of TILs, which suggests a potential target for novel immunotherapy in HBV-HCC. Copyright © 2018 Société Française du Cancer. Published by Elsevier Masson SAS. All rights reserved.

  12. Chimeric PD-1:28 Receptor Upgrades Low-Avidity T cells and Restores Effector Function of Tumor-Infiltrating Lymphocytes for Adoptive Cell Therapy.

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    Schlenker, Ramona; Olguín-Contreras, Luis Felipe; Leisegang, Matthias; Schnappinger, Julia; Disovic, Anja; Rühland, Svenja; Nelson, Peter J; Leonhardt, Heinrich; Harz, Hartmann; Wilde, Susanne; Schendel, Dolores J; Uckert, Wolfgang; Willimsky, Gerald; Noessner, Elfriede

    2017-07-01

    Inherent intermediate- to low-affinity T-cell receptors (TCR) that develop during the natural course of immune responses may not allow sufficient activation for tumor elimination, making the majority of T cells suboptimal for adoptive T-cell therapy (ATT). TCR affinity enhancement has been implemented to provide stronger T-cell activity but carries the risk of creating undesired cross-reactivity leading to potential serious adverse effects in clinical application. We demonstrate here that engineering of low-avidity T cells recognizing a naturally processed and presented tumor-associated antigen with a chimeric PD-1:28 receptor increases effector function to levels seen with high-avidity T cells of identical specificity. Upgrading the function of low-avidity T cells without changing the TCR affinity will allow a large arsenal of low-avidity T cells previously thought to be therapeutically inefficient to be considered for ATT. PD-1:28 engineering reinstated Th1 function in tumor-infiltrating lymphocytes that had been functionally disabled in the human renal cell carcinoma environment without unleashing undesired Th2 cytokines or IL10. Involved mechanisms may be correlated to restoration of ERK and AKT signaling pathways. In mouse tumor models of ATT, PD-1:28 engineering enabled low-avidity T cells to proliferate stronger and prevented PD-L1 upregulation and Th2 polarization in the tumor milieu. Engineered T cells combined with checkpoint blockade secreted significantly more IFNγ compared with T cells without PD-1:28, suggesting a beneficial combination with checkpoint blockade therapy or other therapeutic strategies. Altogether, the supportive effects of PD-1:28 engineering on T-cell function make it an attractive tool for ATT. Cancer Res; 77(13); 3577-90. ©2017 AACR . ©2017 American Association for Cancer Research.

  13. Circulating and tumor-infiltrating Tim-3 in patients with colorectal cancer

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    Gao, Quanli; Yuan, Peng; Zhao, Peng; Yuan, Huijuan; Fan, Huijie; Li, Tiepeng; Qin, Peng; Han, Lu; Fang, Weijia; Suo, Zhenhe

    2015-01-01

    T-cell exhaustion represents a progressive loss of T-cell function. The inhibitory receptor PD-1 is known to negatively regulate CD8+ T cell responses directed against tumor antigen, but the blockades of PD-1 pathway didn't show the objective responses in patients with colorectal cancer (CRC). Thus, further exploring the molecular mechanism responsible for inducing T-cell dysfunction in CRC patients may reveal effective strategies for immune therapy. This study aims to characterize co-inhibitory receptors on T cells in CRC patients to identify novel targets for immunotherapy. In this study, peripheral blood samples from 20 healthy controls and 54 consented CRC patients, and tumor and matched paraneoplastic tissues from 7 patients with advanced CRC, subjected to multicolor flow cytometric analysis of the expression of PD-1 and Tim-3 receptors on CD8+ T cells. It was found that CRC patients presented with significantly higher levels of circulating Tim-3+PD-1+CD8+ T cells compared to the healthy controls (medians of 3.12% and 1.99%, respectively, p = 0.0403). A similar increase of Tim-3+PD-1+CD8+ T cells was also observed in the tumor tissues compared to paraneoplastic tussues. Tim-3+PD-1+CD8+ T cells in tumor tissues produced even less cytokine than that in paraneoplastic tissues. Functional ex vivo experiments showed that Tim-3+PD-1+CD8+ T cells produced significantly less IFN-γ than Tim-3−PD-1−CD8+ T cells, followed by Tim-3+PD-1−CD8+ T cells, and Tim-3−PD-1+CD8+ T cells, indicating a stronger inhibition of IFN-γ production of Tim-3+CD8+ T cells. It is also found in this study that Tim-3+PD-1+CD8+ T cell increase in circulation was correlated with clinical cancer stage but not histologic grade and serum concentrations of cancer biomarker CEA. Our results indicate that upregulation of the inhibitory receptor Tim-3 may restrict T cell responses in CRC patients, and therefore blockage of Tim-3 and thus restoring T cell responses may be a potential

  14. Quantification of tumor infiltrating Foxp3+ regulatory T cells enables the identification of high-risk patients for developing synchronous cancers over upper aerodigestive tract.

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    Wang, Wen-Lun; Chang, Wei-Lun; Yang, Hsiao-Bai; Chang, I-Wei; Lee, Ching-Tai; Chang, Chi-Yang; Lin, Jaw-Town; Sheu, Bor-Shyang

    2015-07-01

    Patients with squamous cell carcinomas (SCC) of upper aerodigestive tract, either over head and neck (HNSCC) or esophagus (ESCC), frequently developed synchronous multiple cancers, leading to worse prognosis. This study validated whether suppression of host cancer immunosurveillance mediated by regulatory T cells (Treg) may predispose to the development of synchronous cancers. Tumor tissues of 200 patients (100 ESCC only, 50 HNSCC only, and 50 synchronous SCCs) were quantitatively accessed for the tumor infiltrating Treg by immunohistochemistry. The density of Treg was also correlated to the level of Treg-associated inhibitory cytokines (IL-10, IL-35 and TGF-β1), and chemokine (CCL22). The density of tumor infiltrating Treg in the index tumor (i.e. the first malignancy diagnosed) of synchronous SCC group was higher than those of HNSCC or ESCC only (prisk of synchronous cancer development to initiate a proper surveillance program. Copyright © 2015 Elsevier Ltd. All rights reserved.

  15. Differentiation between vasogenic-edema versus tumor-infiltrative area in patients with glioblastoma during bevacizumab therapy: A longitudinal MRI study

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    Artzi, Moran, E-mail: artzimy@gmail.com [Functional Brain Center, Tel Aviv Sourasky Medical Center, Tel Aviv (Israel); Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv (Israel); Bokstein, Felix, E-mail: felixb@tlvmc.gov.il [Neuro-Oncology Service, Tel Aviv Sourasky Medical Center, Tel Aviv (Israel); Blumenthal, Deborah T., E-mail: deborahblumenthal@gmail.com [Neuro-Oncology Service, Tel Aviv Sourasky Medical Center, Tel Aviv (Israel); Aizenstein, Orna, E-mail: Ornaaizenstein@gmail.com [Functional Brain Center, Tel Aviv Sourasky Medical Center, Tel Aviv (Israel); Liberman, Gilad, E-mail: giladliberman@gmail.com [Functional Brain Center, Tel Aviv Sourasky Medical Center, Tel Aviv (Israel); Gonda Multidisciplinary Brain Research Center, Bar Ilan University, Ramat Gan (Israel); Corn, Benjamin W., E-mail: bencorn@tlvmc.gov.il [Functional Brain Center, Tel Aviv Sourasky Medical Center, Tel Aviv (Israel); Institute of Radiotherapy, Tel Aviv Sourasky Medical Center, Tel Aviv (Israel); Ben Bashat, Dafna, E-mail: dafnab@tlvmc.gov.il [Functional Brain Center, Tel Aviv Sourasky Medical Center, Tel Aviv (Israel); Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv (Israel); Sagol School of Neuroscience, Tel Aviv University, Tel Aviv (Israel)

    2014-07-15

    Background: Treatment with bevacizumab is associated with substantial radiologic response in patients with glioblastoma (GB). However, following this initial response, changes in T{sub 2}-weighted MRI signal may develop, suggesting an infiltrative pattern of tumor progression. The aim of this study was to differentiate between vasogenic-edema versus tumor-infiltrative area in GB patients. Methods and materials: Fourteen patients with GB were longitudinally scanned, before and during intravenous bevacizumab therapy (5/10 mg/kg every 2-weeks). A total of 40 MR scans including conventional, diffusion, dynamic susceptibility contrast, dynamic contrast enhancement imaging, and MR-spectroscopy (MRS) were analyzed. Classification of non-enhancing fluid-attenuation-inversion-recovery (FLAIR) area was performed based on mean diffusivity, cerebral blood volume and flow maps, and further characterized using multiple MRI parameters. Results: The non-enhancing FLAIR lesion area was classified into: vasogenic-edema, characterized by reduced perfusion and increased FLAIR values; or tumor-infiltrative area, characterized by increased perfusion. Tumor-infiltrative area demonstrated a higher malignant pattern on MRS compared to areas of vasogenic-edema. Substantial reductions of the enhanced T{sub 1}-weighted (58 ± 10%) and hyperintense FLAIR (53 ± 9%) lesion volumes were detected mainly during the first weeks of therapy, with a shift to an infiltrative pattern of tumor progression thereafter, as detected by an increase in tumor-infiltrative area in the majority of patients, which correlated with progression-free survival (week 8: r = −0.86, p = 0.003, week 16: r = −0.99, p = 0.001). Conclusion: Characterization of non-enhancing hyperintense FLAIR lesion area in GB patients can provide an MR-based biomarker, indicating a shift to an infiltrative progression pattern, and may improve therapy response assessment in patients following bevacizumab therapy.

  16. Tumor infiltrating BRAFV600E-specific CD4 T cells correlated with complete clinical response in melanoma. | Office of Cancer Genomics

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    T cells specific for neoantigens encoded by mutated genes in cancers are increasingly recognized as mediators of tumor destruction after immune checkpoint inhibitor therapy or adoptive cell transfer. Unfortunately, most neoantigens result from random mutations and are patient specific, and some cancers contain few mutations to serve as potential antigens. We describe a patient with stage IV acral melanoma who obtained a complete response following adoptive transfer of tumor infiltrating lymphocytes (TIL).

  17. Impaired Tumor-infiltrating T Cells in Patients with COPD Impacts Lung Cancer Response to PD-1 Blockade.

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    Biton, Jérôme; Ouakrim, Hanane; Dechartres, Agnès; Alifano, Marco; Mansuet-Lupo, Audrey; Si, Han; Halpin, Rebecca; Creasy, Todd; Bantsimba-Malanda, Claudie; Arrondeau, Jennifer; Goldwasser, François; Boudou-Rouquette, Pascaline; Fournel, Ludovic; Roche, Nicolas; Burgel, Pierre-Régis; Goc, Jeremy; Devi-Marulkar, Priyanka; Germain, Claire; Dieu-Nosjean, Marie-Caroline; Cremer, Isabelle; Herbst, Ronald; Damotte, Diane

    2018-03-08

    Patients with chronic obstructive pulmonary disease (COPD) have a higher prevalence of lung cancer. The chronic inflammation associated with COPD probably promotes the earliest stages of carcinogenesis. However, once tumors have progressed to malignancy, the impact of COPD on the tumor immune microenvironment remains poorly defined, and its effects on immune-checkpoint blockers' efficacy are still unknown. To study the impact of COPD on the immune contexture of non-small cell lung cancer (NSCLC). We performed in depth immune profiling of lung tumors by immunohistochemistry and we determined its impact on patients' survival (n=435). Tumor-infiltrating T lymphocyte (TILs) exhaustion by flow cytometry (n=50) was also investigated. The effectiveness of an anti-PD-1 treatment (nivolumab) was evaluated in 39 advanced-stage NSCLC patients. All data were analyzed according to patients' COPD status. Measurments and Main Results: Remarkably, COPD severity is positively correlated with the coexpression of PD-1/TIM-3 by CD8 T cells. In agreement, we observed a loss of CD8 T cell-associated favorable clinical outcome in COPD+ patients. Interestingly, a negative prognostic value of PD-L1 expression by tumor cells was observed only in highly CD8 T cell-infiltrated tumors of COPD+ patients. Finally, data obtained on 39 advanced-stage NSCLC patients treated by an anti-PD-1 antibody showed longer progression free survival in COPD+ patients, and also that the association between the severity of smoking and the response to nivolumab was preferentially observed in COPD+ patients. COPD is associated with an increased sensitivity of CD8 TILs to immune escape mechanisms developed by tumors, thus suggesting a higher sensitivity to PD-1 blockade in patients with COPD.

  18. Minimally cultured or selected autologous tumor-infiltrating lymphocytes after a lympho-depleting chemotherapy regimen in metastatic melanoma patients.

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    Besser, Michal J; Shapira-Frommer, Ronnie; Treves, Avraham J; Zippel, Dov; Itzhaki, Orit; Schallmach, Ester; Kubi, Adva; Shalmon, Bruria; Hardan, Izhar; Catane, Raphael; Segal, Eran; Markel, Gal; Apter, Sara; Nun, Alon Ben; Kuchuk, Iryna; Shimoni, Avichai; Nagler, Arnon; Schachter, Jacob

    2009-05-01

    Adoptive cell therapy with autologous tumor-infiltrating lymphocytes (TIL) and high-dose interleukin-2 (IL-2), after nonmyeloablative chemotherapy, has been shown to result in tumor regression in half of refractory metastatic melanoma patients. In the present study, we describe 2 separate clinical protocols. Twelve patients were treated with "Selected"-TIL, as previously reported and 8 patients with the modified version of "Young"-TIL. Selected-TIL protocol required the establishment of multiple T-cell cultures from 1 patient and in vitro selection of cultures secreting interferon-gamma upon antigenic stimulation. In contrast, Young-TIL are minimally cultured T cells with superior in vitro features that do not require further selection. Two of 12 Selected-TIL patients experienced objective clinical responses (1 complete response, 1 partial response). Out of 8 treated Young-TIL patients, 1 experienced complete response, 2 partial response, and 4 patients had disease stabilization. Twenty-one of 33 enrolled Selected-TIL patients were excluded from the protocol, mainly as cultures failed the interferon-gamma selection criteria or due to clinical deterioration, compared with only 3 Young-TIL patients. Expected bone marrow suppression and high-dose IL-2 toxicity were transient. There was no treatment-related mortality. This study vindicates the feasibility and effectiveness of TIL technology and calls for further efforts to implement and enhance this modality. The use of minimally cultured, unselected Young-TIL enables the treatment of most enrolled patients. Although the cohort of Young-TIL patients treated so far is rather small and the follow-up short, the response rate is encouraging.

  19. Combination of Ipilimumab and Adoptive Cell Therapy with Tumor-Infiltrating Lymphocytes for Patients with Metastatic Melanoma

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    John E. Mullinax

    2018-03-01

    Full Text Available PurposeAdoptive cell therapy (ACT using tumor-infiltrating lymphocytes (TIL for metastatic melanoma can be highly effective, but attrition due to progression before TIL administration (32% in prior institutional experience remains a limitation. We hypothesized that combining ACT with cytotoxic T lymphocyte-associated antigen 4 blockade would decrease attrition and allow more patients to receive TIL.Experimental designThirteen patients with metastatic melanoma were enrolled. Patients received four doses of ipilimumab (3 mg/kg beginning 2 weeks prior to tumor resection for TIL generation, then 1 week after resection, and 2 and 5 weeks after preconditioning chemotherapy and TIL infusion followed by interleukin-2. The primary endpoint was safety and feasibility. Secondary endpoints included of clinical response at 12 weeks and at 1 year after TIL transfer, progression free survival (PFS, and overall survival (OS.ResultsAll patients received at least two doses of ipilimumab, and 12 of the 13 (92% received TIL. A median of 6.5 × 1010 (2.3 × 1010 to 1.0 × 1011 TIL were infused. At 12 weeks following infusion, there were five patients who experienced objective response (38.5%, four of whom continued in objective response at 1 year and one of which became a complete response at 52 months. Median progression-free survival was 7.3 months (95% CI 6.1–29.9 months. Grade ≥ 3 immune-related adverse events included hypothyroidism (3, hepatitis (2, uveitis (1, and colitis (1.ConclusionIpilimumab plus ACT for metastatic melanoma is feasible, well tolerated, and associated with a low rate of attrition due to progression during cell expansion. This combination approach serves as a model for future efforts to improve the efficacy of ACT.

  20. Combination of Ipilimumab and Adoptive Cell Therapy with Tumor-Infiltrating Lymphocytes for Patients with Metastatic Melanoma.

    Science.gov (United States)

    Mullinax, John E; Hall, MacLean; Prabhakaran, Sangeetha; Weber, Jeffrey; Khushalani, Nikhil; Eroglu, Zeynep; Brohl, Andrew S; Markowitz, Joseph; Royster, Erica; Richards, Allison; Stark, Valerie; Zager, Jonathan S; Kelley, Linda; Cox, Cheryl; Sondak, Vernon K; Mulé, James J; Pilon-Thomas, Shari; Sarnaik, Amod A

    2018-01-01

    Adoptive cell therapy (ACT) using tumor-infiltrating lymphocytes (TIL) for metastatic melanoma can be highly effective, but attrition due to progression before TIL administration (32% in prior institutional experience) remains a limitation. We hypothesized that combining ACT with cytotoxic T lymphocyte-associated antigen 4 blockade would decrease attrition and allow more patients to receive TIL. Thirteen patients with metastatic melanoma were enrolled. Patients received four doses of ipilimumab (3 mg/kg) beginning 2 weeks prior to tumor resection for TIL generation, then 1 week after resection, and 2 and 5 weeks after preconditioning chemotherapy and TIL infusion followed by interleukin-2. The primary endpoint was safety and feasibility. Secondary endpoints included of clinical response at 12 weeks and at 1 year after TIL transfer, progression free survival (PFS), and overall survival (OS). All patients received at least two doses of ipilimumab, and 12 of the 13 (92%) received TIL. A median of 6.5 × 10 10 (2.3 × 10 10 to 1.0 × 10 11 ) TIL were infused. At 12 weeks following infusion, there were five patients who experienced objective response (38.5%), four of whom continued in objective response at 1 year and one of which became a complete response at 52 months. Median progression-free survival was 7.3 months (95% CI 6.1-29.9 months). Grade ≥ 3 immune-related adverse events included hypothyroidism (3), hepatitis (2), uveitis (1), and colitis (1). Ipilimumab plus ACT for metastatic melanoma is feasible, well tolerated, and associated with a low rate of attrition due to progression during cell expansion. This combination approach serves as a model for future efforts to improve the efficacy of ACT.

  1. Minimally invasive liver resection to obtain tumor-infiltrating lymphocytes for adoptive cell therapy in patients with metastatic melanoma

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    Alvarez-Downing Melissa M

    2012-06-01

    Full Text Available Abstract Background Adoptive cell therapy (ACT with tumor-infiltrating lymphocytes (TIL in patients with metastatic melanoma has been reported to have a 56% overall response rate with 20% complete responders. To increase the availability of this promising therapy in patients with advanced melanoma, a minimally invasive approach to procure tumor for TIL generation is warranted. Methods A feasibility study was performed to determine the safety and efficacy of laparoscopic liver resection to generate TIL for ACT. Retrospective review of a prospectively maintained database identified 22 patients with advanced melanoma and visceral metastasis (AJCC Stage M1c who underwent laparoscopic liver resection between 1 October 2005 and 31 July 2011. The indication for resection in all patients was to receive postoperative ACT with TIL. Results Twenty patients (91% underwent resection utilizing a closed laparoscopic technique, one required hand-assistance and another required conversion to open resection. Median intraoperative blood loss was 100 mL with most cases performed without a Pringle maneuver. Median hospital stay was 3 days. Three (14% patients experienced a complication from resection with no mortality. TIL were generated from 18 of 22 (82% patients. Twelve of 15 (80% TIL tested were found to have in vitro tumor reactivity. Eleven patients (50% received the intended ACT. Two patients were rendered no evidence of disease after surgical resection, with one undergoing delayed ACT with generated TIL after relapse. Objective tumor response was seen in 5 of 11 patients (45% who received TIL, with one patient experiencing an ongoing complete response (32+ months. Conclusions Laparoscopic liver resection can be performed with minimal morbidity and serve as an effective means to procure tumor to generate therapeutic TIL for ACT to patients with metastatic melanoma.

  2. Clinical and diagnostic significance of urinary neutrophil gelatinase-associated lipocalin-2 measurement in children with microbial inflammatory kidney and urinary tract diseases

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    A. V. Eremeeva

    2015-01-01

    Full Text Available Objective: to study the clinical and diagnostic significance of urinary neutrophil gelatinase-associated lipocalin-2 (NGAL measurement in children with urinary tract infection (я=15 and pyelonephritis (я=15. The patients' age was 1 to 16 years (mean age, 7.32+4.52 years. The diagnosis was verified on the basis of clinical and laboratory findings and medical history and instrumental examination data. Urinary NGAL levels were measured by enzyme immunoassay (a Bio\\fendor Laboratory Medicine kit and calculated with reference to mg of creatinine. Urinary NGAL levels were established to depend on the degree of renal parenchymal damage. The investigation showed a relationship between the excretion of NGAL during the acute phase of pyelonephritis and the detection of renal scarring, as evidenced by statistical DMCA nephroscintigraphy. The acute pyelonephritis group exhibited a moderate direct correlation between the renal excretion of NGAL and the degree of leukocytosis and the blood levels of C-reactive protein. The findings allow recommendations for measuring urinary NGAL levels as an additional noninvasive marker for the early detection of renal parenchymal damage.

  3. High Neutrophil-to-Lymphocyte Ratio is a Significant Predictor of Cardiovascular and All-Cause Mortality in Patients Undergoing Peritoneal Dialysis

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    Xiangxue Lu

    2018-03-01

    Full Text Available Background/Aims: Chronic inflammation is associated with increased risk of cardiovascular death in patients with end-stage renal disease (ESRD. Although elevated neutrophil-to-lymphocyte ratio (NLR, a novel inflammatory marker, has been shown to predict cardiovascular disease and all-cause mortality in the general population, limited evidence is available for its role in ESRD. Methods: We enrolled 86 patients undergoing peritoneal dialysis (PD for a 36-month follow-up to investigate the association between the NLR and arterial stiffness markers, namely, carotid-femoral pulse wave velocity (cfPWV and carotid augmentation index (AIx, and mortality in PD patients. The primary endpoints were cardiovascular mortality and all-cause mortality. Kaplan–Meier curves were used to show the cumulative incidence of cardiovascular mortality and all-cause mortality. Results: High NLR was found to be a predictor of increased cfPWV (β = 1.150; P < 0.001 and AIx (β = 3.945; P < 0.001 in patients on PD. Patients with higher NLR had lower survival during follow-up. Kaplan–Meier curves showed that the cumulative incidences of both cardiovascular mortality and all-cause mortality were significantly higher in patients with NLR ≥ 4.5 (both P < 0.01. Conclusion: Our results suggest that high NLR is independently associated with arterial stiffness and predicts cardiovascular and all-cause mortality in PD patients.

  4. Correlation between Tumor-Infiltrating Lymphocytes and Pathological Response in Locally Advanced Breast Cancer Patients Who Received Neoadjuvant Chemotherapy in H. Adam Malik General Hospital

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    Kamal Basri Siregar

    2017-08-01

    Full Text Available Background: Tumor-infiltrating lymphocytes (TILs are emerging as biomarkers mediating tumor response to treatments. Earlier studies have provided evidence that the level of TILs has prognostic value, particularly in triple-negative and human epidermal growth factor receptor-2-positive breast cancer. Moreover, the level of TILs has been associated with treatment outcome in patients undergoing neoadjuvant chemotherapy, and there is a strong correlation with pathologically complete response. In this study, we analyzed whether changes in TILs take place after neoadjuvant therapy and if they correlate with pathological response to treatment. Patients and Methods: We retrospectively analyzed the specimen slides from the Department of Anatomic Pathology of H. Adam Malik General Hospital during 2011–2015. We identified 51 patients fulfilling the inclusion criteria of this study. The histological sections had already been evaluated by hematoxylin and eosin slides. They were reassessed by our pathologist for the percentage of intratumoral and stromal TILs. The correlation with pathological response of the tumor after neoadjuvant therapy was also studied in these patients. Each case was also defined as high- or low-TIL breast cancer adopting previously validated cutoffs. Results: The mean age of the 51 patients was 49.22 years. The most frequent type of breast cancer histology was invasive ductal breast carcinoma in 49 (96% patients, and there were 2 (4% patients with lobular carcinoma. The histopathological grading for high TILs was grade 1 in 5 patients, grade 2 in 15 patients, and grade 3 in 3 patients. High TILs that had a pathologically complete response were found in 47.8% of patients, and low TILs were found in 28.8%. There was no significant correlation between TILs and pathological response in patients with neoadjuvant chemotherapy (p = 0.157. Conclusions: This research has not been able to demonstrate a significant correlation between TILs and

  5. Computer-assisted stereology and automated image analysis for quantification of tumor infiltrating lymphocytes in colon cancer.

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    Eriksen, Ann C; Andersen, Johnnie B; Kristensson, Martin; dePont Christensen, René; Hansen, Torben F; Kjær-Frifeldt, Sanne; Sørensen, Flemming B

    2017-08-29

    Precise prognostic and predictive variables allowing improved post-operative treatment stratification are missing in patients treated for stage II colon cancer (CC). Investigation of tumor infiltrating lymphocytes (TILs) may be rewarding, but the lack of a standardized analytic technique is a major concern. Manual stereological counting is considered the gold standard, but digital pathology with image analysis is preferred due to time efficiency. The purpose of this study was to compare manual stereological estimates of TILs with automatic counts obtained by image analysis, and at the same time investigate the heterogeneity of TILs. From 43 patients treated for stage II CC in 2002 three paraffin embedded, tumor containing tissue blocks were selected one of them representing the deepest invasive tumor front. Serial sections from each of the 129 blocks were immunohistochemically stained for CD3 and CD8, and the slides were scanned. Stereological estimates of the numerical density and area fraction of TILs were obtained using the computer-assisted newCAST stereology system. For the image analysis approach an app-based algorithm was developed using Visiopharm Integrator System software. For both methods the tumor areas of interest (invasive front and central area) were manually delineated by the observer. Based on all sections, the Spearman's correlation coefficients for density estimates varied from 0.9457 to 0.9638 (p heterogeneity, intra-class correlation coefficients (ICC) for CD3+ TILs varied from 0.615 to 0.746 in the central area, and from 0.686 to 0.746 in the invasive area. ICC for CD8+ TILs varied from 0.724 to 0.775 in the central area, and from 0.746 to 0.765 in the invasive area. Exact objective and time efficient estimates of numerical densities and area fractions of CD3+ and CD8+ TILs in stage II colon cancer can be obtained by image analysis and are highly correlated to the corresponding estimates obtained by the gold standard based on stereology

  6. Prognostic and predictive role of FOXP3 positive tumor infiltrating lymphocytes (TILs in curatively resected non small cell lung cancer other than stage IA

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    Fatih Kose

    2017-12-01

    Full Text Available Lung cancer is the leading cause of cancer-related mortality and responsible for 1.6 million deaths per year through world-wide. Surgical resection with negative margin combined with the adjuvant therapy [except for stage IA and IB (<4 cm] is the Standard treatment for early-stage Non-small cell lung cancer (NSCLC. Early-stage NSCLC, however, has relapse rate over 40% mostly at distant sites. Therefore, high relapse rate necessitates urgent novel biomarker for these patients. In this study, we aim to evaluate the predictive and prognostic role of FOXP3+ Treg cells along with well defined Clinicohistopathological factors in early-stage non-small cell lung cancer (NSCLC. FOXP3 expression in tumor infiltrating lymphocytes (TIL was examined by immunohistochemical staining from resected early-stage 48 NSCLC patients. Data of patients and FOXP3 expression status along with common clinicohistopathological prognostic factors were evaluated retrospectively. Median age of patients was 62 years-old (range 43–78. Mean follow-up, median overall survival (OS, and disease-free survival (DFS were 49, 49 and 30 months, respectively. FOXP3 expression was positive in 23 (47.9% patients. Adjuvant chemotherapy (4 cycles of cisplatin-vinorelbine was given to 16 patients (33.3% at physician discretion. Patients with a FOXP3 expression of 25% or higher significantly lower OS and DFS when compared with patients with a FOXP3 staining lower than 25% with p-value of 0.016 and 0.032, respectively. In the patients with high FOXP3 expression, platin-based adjuvant chemotherapy had showed a detrimental effect on DFS and OS. These results suggest that FOXP3 expression may be used as useful prognostic biomarker in resected NSCLC. Our findings also suggest that resected NSCLC patients with FOXP3 expression of 25% or higher staining intensity may not get any benefit even disfavor from adjuvant platin chemotherapy.

  7. Heat shock protein 70 and tumor-infiltrating NK cells as prognostic indicators for patients with squamous cell carcinoma of the head and neck after radiochemotherapy: A multicentre retrospective study of the German Cancer Consortium Radiation Oncology Group (DKTK-ROG).

    Science.gov (United States)

    Stangl, Stefan; Tontcheva, Nikoletta; Sievert, Wolfgang; Shevtsov, Maxim; Niu, Minli; Schmid, Thomas E; Pigorsch, Steffi; Combs, Stephanie E; Haller, Bernhard; Balermpas, Panagiotis; Rödel, Franz; Rödel, Claus; Fokas, Emmanouil; Krause, Mechthild; Linge, Annett; Lohaus, Fabian; Baumann, Michael; Tinhofer, Inge; Budach, Volker; Stuschke, Martin; Grosu, Anca-Ligia; Abdollahi, Amir; Debus, Jürgen; Belka, Claus; Maihöfer, Cornelius; Mönnich, David; Zips, Daniel; Multhoff, Gabriele

    2018-05-01

    Tumor cells frequently overexpress heat shock protein 70 (Hsp70) and present it on their cell surface, where it can be recognized by pre-activated NK cells. In our retrospective study the expression of Hsp70 was determined in relation to tumor-infiltrating CD56 + NK cells in formalin-fixed paraffin embedded (FFPE) tumor specimens of patients with SCCHN (N = 145) as potential indicators for survival and disease recurrence. All patients received radical surgery and postoperative cisplatin-based radiochemotherapy (RCT). In general, Hsp70 expression was stronger, but with variable intensities, in tumor compared to normal tissues. Patients with high Hsp70 expressing tumors (scores 3-4) showed significantly decreased overall survival (OS; p = 0.008), local progression-free survival (LPFS; p = 0.034) and distant metastases-free survival (DMFS; p = 0.044), compared to those with low Hsp70 expression (scores 0-2), which remained significant after adjustment for relevant prognostic variables. The adverse prognostic value of a high Hsp70 expression for OS was also observed in patient cohorts with p16- (p = 0.001), p53- (p = 0.0003) and HPV16 DNA-negative (p = 0.001) tumors. The absence or low numbers of tumor-infiltrating CD56 + NK cells also correlated with significantly decreased OS (p = 0.0001), LPFS (p = 0.0009) and DMFS (p = 0.0001). A high Hsp70 expression and low numbers of tumor-infiltrating NK cells have the highest negative predictive value (p = 0.00004). In summary, a strong Hsp70 expression and low numbers of tumor-infiltrating NK cells correlate with unfavorable outcome following surgery and RCT in patients with SCCHN, and thus serve as negative prognostic markers. © 2017 The Authors International Journal of Cancer published by John Wiley & Sons Ltd on behalf of UICC.

  8. Recombinant tumor necrosis factor alpha inhibits growth of methylcholanthrene-induced sarcoma and enhances natural killer activity of tumor-infiltrating lymphocytes in aging rats

    International Nuclear Information System (INIS)

    Ziolkowska, Maria; Nowak Joanna, J.; Janiak, Marek; Ryzewska, Alicja

    1994-01-01

    The effect of recombinant human tumor necrosis factors alpha (rHuTNF-α) on the growth of immunogenic, methylcholanthrene-induced sarcoma (MC-Sa) and natural killer (NK) cell activity of tumor-infiltrating lymphocytes (TIL) in adult and aging rats was investigated. In both groups of animals the growth of transplantable MC-Sa was markedly and similarly inhibited by multiple intratumoral (i.t.) injections of rHuTF-α. This effect was accompanied by stimulation of NK activity of tumor-infiltrating lymphocytes in adult as well as in aging rats. Studies ''in vitro'' demonstrated additionally that rHuTNF-α was a potent stimulator of NK but not of ADCC (antibody-dependent cellular cytotoxicity) activity of spleen lymphocytes from healthy animals. Our results indicate that the antitumor effect of TNF-α is comparable in adult and in aging rats bearing immunogenic MC-Sa. The inhibition of MC-Sa growth may be attributed not only to the TNF-α-induced necrosis of the neoplastic tissue but also to the ''in vivo'' stimulatory effect of this cytokine upon the NK-type function of lymphocytes infiltrating the tumor mass. (author). 31 refs, 5 figs, 2 tabs

  9. Neutrophils responsive to endogenous IFN-beta regulate tumor angiogenesis and growth in a mouse tumor model.

    Science.gov (United States)

    Jablonska, Jadwiga; Leschner, Sara; Westphal, Kathrin; Lienenklaus, Stefan; Weiss, Siegfried

    2010-04-01

    Angiogenesis is a hallmark of malignant neoplasias, as the formation of new blood vessels is required for tumors to acquire oxygen and nutrients essential for their continued growth and metastasis. However, the signaling pathways leading to tumor vascularization are not fully understood. Here, using a transplantable mouse tumor model, we have demonstrated that endogenous IFN-beta inhibits tumor angiogenesis through repression of genes encoding proangiogenic and homing factors in tumor-infiltrating neutrophils. We determined that IFN-beta-deficient mice injected with B16F10 melanoma or MCA205 fibrosarcoma cells developed faster-growing tumors with better-developed blood vessels than did syngeneic control mice. These tumors displayed enhanced infiltration by CD11b+Gr1+ neutrophils expressing elevated levels of the genes encoding the proangiogenic factors VEGF and MMP9 and the homing receptor CXCR4. They also expressed higher levels of the transcription factors c-myc and STAT3, known regulators of VEGF, MMP9, and CXCR4. In vitro, treatment of these tumor-infiltrating neutrophils with low levels of IFN-beta restored expression of proangiogenic factors to control levels. Moreover, depletion of these neutrophils inhibited tumor growth in both control and IFN-beta-deficient mice. We therefore suggest that constitutively produced endogenous IFN-beta is an important mediator of innate tumor surveillance. Further, we believe our data help to explain the therapeutic effect of IFN treatment during the early stages of cancer development.

  10. Effect of sevoflurane on human neutrophil apoptosis.

    LENUS (Irish Health Repository)

    Tyther, R

    2012-02-03

    BACKGROUND AND OBJECTIVE: Both chronic occupational exposure to volatile anaesthetic agents and acute in vitro exposure of neutrophils to isoflurane have been shown to inhibit the rate of apoptosis of human neutrophils. It is possible that inhibition of neutrophil apoptosis arises through delaying mitochondrial membrane potential collapse. We assessed mitochondrial depolarization and apoptosis in unexposed neutrophils and neutrophils exposed to sevoflurane in vivo. METHODS: A total of 20 mL venous blood was withdrawn pre- and postinduction of anaesthesia, the neutrophils isolated and maintained in culture. At 1, 12 and 24 h in culture, the percentage of neutrophil apoptosis was assessed by dual staining with annexin V-FITC and propidium iodide. Mitochondrial depolarization was measured using the dual emission styryl dye JC-1. RESULTS: Apoptosis was significantly inhibited in neutrophils exposed to sevoflurane in vivo at 24 (exposed: 38 (12)% versus control: 28 (11)%, P = 0.001), but not at 1 or 12 h, in culture. Mitochondrial depolarization was not delayed in neutrophils exposed to sevoflurane. CONCLUSIONS: The most important findings are that sevoflurane inhibits neutrophil apoptosis in vivo and that inhibition is not mediated primarily by an effect on mitochondrial depolarization.

  11. Neutrophil Extracellular Traps in Ulcerative Colitis

    DEFF Research Database (Denmark)

    Bjerg Bennike, Tue; Carlsen, Thomas Gelsing; Ellingsen, Torkell

    2015-01-01

    microscopy and confocal microscopy. RESULTS: We identified and quantified 5711 different proteins with proteomics. The abundance of the proteins calprotectin and lactotransferrin in the tissue correlated with the degree of tissue inflammation as determined by histology. However, fecal calprotectin did...... not correlate. Forty-six proteins were measured with a statistically significant differences in abundances between the UC colon tissue and controls. Eleven of the proteins with increased abundances in the UC biopsies were associated with neutrophils and neutrophil extracellular traps. The findings were...... validated by microscopy, where an increased abundance of neutrophils and the presence of neutrophil extracellular traps by extracellular DNA present in the UC colon tissue were confirmed. CONCLUSIONS: Neutrophils, induced neutrophil extracellular traps, and several proteins that play a part in innate...

  12. Superoxide anion production by human neutrophils activated by Trichomonas vaginalis.

    Science.gov (United States)

    Song, Hyun-Ouk; Ryu, Jae-Sook

    2013-08-01

    Neutrophils are the predominant inflammatory cells found in vaginal discharges of patients infected with Trichomonas vaginalis. In this study, we examined superoxide anion (O2 (.-)) production by neutrophils activated by T. vaginalis. Human neutrophils produced superoxide anions when stimulated with either a lysate of T. vaginalis, its membrane component (MC), or excretory-secretory product (ESP). To assess the role of trichomonad protease in production of superoxide anions by neutrophils, T. vaginalis lysate, ESP, and MC were each pretreated with a protease inhibitor cocktail before incubation with neutrophils. Superoxide anion production was significantly decreased by this treatment. Trichomonad growth was inhibited by preincubation with supernatants of neutrophils incubated for 3 hr with T. vaginalis lysate. Furthermore, myeloperoxidase (MPO) production by neutrophils was stimulated by live trichomonads. These results indicate that the production of superoxide anions and MPO by neutrophils stimulated with T. vaginalis may be a part of defense mechanisms of neutrophils in trichomoniasis.

  13. Neutrophils at work

    DEFF Research Database (Denmark)

    Nauseef, William M; Borregaard, Niels

    2014-01-01

    In this Review we discuss data demonstrating recently recognized aspects of neutrophil homeostasis in the steady state, granulopoiesis in 'emergency' conditions and interactions of neutrophils with the adaptive immune system. We explore in vivo observations of the recruitment of neutrophils from ...

  14. Single-Shot Echo-Planar Diffusion-Weighted MR Imaging at 3T and 1.5T for Differentiation of Benign Vertebral Fracture Edema and Tumor Infiltration

    Energy Technology Data Exchange (ETDEWEB)

    Park, Hee Jin; Lee, So Yeon; Rho, Myung Ho; Chung, Eun Chul; Kim, Mi Sung; Kwon, Heon Ju; Youn, In Young [Department of Radiology, Kangbuk Samsung Hospital, Sungkyunkwan University School of Medicine, Seoul 03181 (Korea, Republic of)

    2016-11-01

    To compare the apparent diffusion coefficient (ADC) value using single-shot echo-planar imaging sequences at 3T and 1.5T for differentiation of benign fracture edema and tumor infiltration of the vertebral body. A total of 46 spinal examinations were included in the 1.5T MRI group, and a total of 40 spinal examinations were included in the 3T MRI group. The ADC values of the lesion were measured and calculated. The diagnostic performance of the conventional MR image containing sagittal T2-weighted fat saturated image and each diffusion weighted image (DWI) with an ADC value with different b values were evaluated. The mean ADC value of the benign lesions was higher than that of the malignant lesions on 1.5T and 3T (p < 0.05). The sensitivity of the diagnostic performance was higher with an additional DWI in both 1.5T and 3T, but the sensitivities were similar with the addition of b values of 400 and 1000. The specificities of the diagnostic performances did not show significant differences (p value > 0.05). The diagnostic accuracies were higher when either of the DWIs (b values of 400 and 1000) was added to routine MR image for 1.5T and 3T. Statistical differences between 1.5T and 3T or between b values of 400 and 1000 were not seen. The ADC values of the benign lesions were significantly higher than those of the malignant lesions on 1.5T and 3T. There was no statistically significant difference in the diagnostic performances when either of the DWIs (b values of 400 and 1000) was added to the routine MR image for 1.5T and 3T.

  15. Signal-shot echo-planner diffusion-weighted MR imaging at 3T and 1.5T for differentiation of benign vertebral fracture edema and tumor infiltration

    Energy Technology Data Exchange (ETDEWEB)

    Park, Hee Jin; Lee, So Yeon; Rho, Myung Ho; Chung, Eun Chul; Kim, Mi Sung; Kwon, Heon Ju; Youn, In Young [Dept. of Radiology, Kangbuk Samsung Hospital, Sungkyunkwan University School of Medicine, Seoul (Korea, Republic of)

    2016-09-15

    To compare the apparent diffusion coefficient (ADC) value using single-shot echo-planar imaging sequences at 3T and 1.5T for differentiation of benign fracture edema and tumor infiltration of the vertebral body. A total of 46 spinal examinations were included in the 1.5T MRI group, and a total of 40 spinal examinations were included in the 3T MRI group. The ADC values of the lesion were measured and calculated. The diagnostic performance of the conventional MR image containing sagittal T2-weighted fat saturated image and each diffusion weighted image (DWI) with an ADC value with different b values were evaluated. The mean ADC value of the benign lesions was higher than that of the malignant lesions on 1.5T and 3T (p < 0.05). The sensitivity of the diagnostic performance was higher with an additional DWI in both 1.5T and 3T, but the sensitivities were similar with the addition of b values of 400 and 1000. The specificities of the diagnostic performances did not show significant differences (p value > 0.05). The diagnostic accuracies were higher when either of the DWIs (b values of 400 and 1000) was added to routine MR image for 1.5T and 3T. Statistical differences between 1.5T and 3T or between b values of 400 and 1000 were not seen. The ADC values of the benign lesions were significantly higher than those of the malignant lesions on 1.5T and 3T. There was no statistically significant difference in the diagnostic performances when either of the DWIs (b values of 400 and 1000) was added to the routine MR image for 1.5T and 3T.

  16. Prognostic and predictive value of tumor-infiltrating lymphocytes in a phase III randomized adjuvant breast cancer trial in node-positive breast cancer comparing the addition of docetaxel to doxorubicin with doxorubicin-based chemotherapy: BIG 02-98.

    Science.gov (United States)

    Loi, Sherene; Sirtaine, Nicolas; Piette, Fanny; Salgado, Roberto; Viale, Giuseppe; Van Eenoo, Françoise; Rouas, Ghizlane; Francis, Prudence; Crown, John P A; Hitre, Erika; de Azambuja, Evandro; Quinaux, Emmanuel; Di Leo, Angelo; Michiels, Stefan; Piccart, Martine J; Sotiriou, Christos

    2013-03-01

    Previous preclinical and clinical data suggest that the immune system influences prognosis and response to chemotherapy (CT); however, clinical relevance has yet to be established in breast cancer (BC). We hypothesized that increased lymphocytic infiltration would be associated with good prognosis and benefit from immunogenic CT-in this case, anthracycline-only CT-in selected BC subtypes. We investigated the relationship between quantity and location of lymphocytic infiltrate at diagnosis with clinical outcome in 2009 node-positive BC samples from the BIG 02-98 adjuvant phase III trial comparing anthracycline-only CT (doxorubicin followed by cyclophosphamide, methotrexate, and fluorouracil [CMF] or doxorubicin plus cyclophosphamide followed by CMF) versus CT combining doxorubicin and docetaxel (doxorubicin plus docetaxel followed by CMF or doxorubicin followed by docetaxel followed by CMF). Readings were independently performed by two pathologists. Disease-free survival (DFS), overall survival (OS), and interaction with type of CT associations were studied. Median follow-up was 8 years. There was no significant prognostic association in the global nor estrogen receptor (ER) -positive/human epidermal growth factor receptor 2 (HER2) -negative population. However, each 10% increase in intratumoral and stromal lymphocytic infiltrations was associated with 17% and 15% reduced risk of relapse (adjusted P = .1 and P = .025), respectively, and 27% and 17% reduced risk of death in ER-negative/HER2-negative BC regardless of CT type (adjusted P = .035 and P = .023), respectively. In HER2-positive BC, there was a significant interaction between increasing stromal lymphocytic infiltration (10% increments) and benefit with anthracycline-only CT (DFS, interaction P = .042; OS, P = .018). In node-positive, ER-negative/HER2-negative BC, increasing lymphocytic infiltration was associated with excellent prognosis. Further validation of the clinical utility of tumor-infiltrating

  17. Inflammation-Dependent IL18 Signaling Restricts Hepatocellular Carcinoma Growth by Enhancing the Accumulation and Activity of Tumor-Infiltrating Lymphocytes.

    Science.gov (United States)

    Markowitz, Geoffrey J; Yang, Pengyuan; Fu, Jing; Michelotti, Gregory A; Chen, Rui; Sui, Jianhua; Yang, Bin; Qin, Wen-Hao; Zhang, Zheng; Wang, Fu-Sheng; Diehl, Anna Mae; Li, Qi-Jing; Wang, Hongyang; Wang, Xiao-Fan

    2016-04-15

    Chronic inflammation in liver tissue is an underlying cause of hepatocellular carcinoma. High levels of inflammatory cytokine IL18 in the circulation of patients with hepatocellular carcinoma correlates with poor prognosis. However, conflicting results have been reported for IL18 in hepatocellular carcinoma development and progression. In this study, we used tissue specimens from hepatocellular carcinoma patients and clinically relevant mouse models of hepatocellular carcinoma to evaluate IL18 expression and function. In a mouse model of liver fibrosis that recapitulates a tumor-promoting microenvironment, global deletion of the IL18 receptor IL18R1 enhanced tumor growth and burden. Similarly, in a carcinogen-induced model of liver tumorigenesis, IL18R1 deletion increased tumor burden. Mechanistically, we found that IL18 exerted inflammation-dependent tumor-suppressive effects largely by promoting the differentiation, activity, and survival of tumor-infiltrating T cells. Finally, differences in the expression of IL18 in tumor tissue versus nontumor tissue were more predictive of patient outcome than overall tissue expression. Taken together, our findings resolve a long-standing contradiction regarding a tumor-suppressive role for IL18 in established hepatocellular carcinoma and provide a mechanistic explanation for the complex relationship between its expression pattern and hepatocellular carcinoma prognosis. Cancer Res; 76(8); 2394-405. ©2016 AACR. ©2016 American Association for Cancer Research.

  18. Expression of PD-L1 on canine tumor cells and enhancement of IFN-γ production from tumor-infiltrating cells by PD-L1 blockade.

    Directory of Open Access Journals (Sweden)

    Naoya Maekawa

    Full Text Available Programmed death 1 (PD-1, an immunoinhibitory receptor, and programmed death ligand 1 (PD-L1, its ligand, together induce the "exhausted" status in antigen-specific lymphocytes and are thus involved in the immune evasion of tumor cells. In this study, canine PD-1 and PD-L1 were molecularly characterized, and their potential as therapeutic targets for canine tumors was discussed. The canine PD-1 and PD-L1 genes were conserved among canine breeds. Based on the sequence information obtained, the recombinant canine PD-1 and PD-L1 proteins were constructed; they were confirmed to bind each other. Antibovine PD-L1 monoclonal antibody effectively blocked the binding of recombinant PD-1 with PD-L1-expressing cells in a dose-dependent manner. Canine melanoma, mastocytoma, renal cell carcinoma, and other types of tumors examined expressed PD-L1, whereas some did not. Interestingly, anti-PD-L1 antibody treatment enhanced IFN-γ production from tumor-infiltrating cells. These results showed that the canine PD-1/PD-L1 pathway is also associated with T-cell exhaustion in canine tumors and that its blockade with antibody could be a new therapeutic strategy for canine tumors. Further investigations are needed to confirm the ability of anti-PD-L1 antibody to reactivate canine antitumor immunity in vivo, and its therapeutic potential has to be further discussed.

  19. Neutrophils in critical illness.

    Science.gov (United States)

    McDonald, Braedon

    2018-03-01

    During critical illness, dramatic alterations in neutrophil biology are observed including abnormalities of granulopoeisis and lifespan, cell trafficking and antimicrobial effector functions. As a result, neutrophils transition from powerful antimicrobial protectors into dangerous mediators of tissue injury and organ dysfunction. In this article, the role of neutrophils in the pathogenesis of critical illness (sepsis, trauma, burns and others) will be explored, including pathological changes to neutrophil function during critical illness and the utility of monitoring aspects of the neutrophil phenotype as biomarkers for diagnosis and prognostication. Lastly, we review findings from clinical trials of therapies that target the harmful effects of neutrophils, providing a bench-to-bedside perspective on neutrophils in critical illness.

  20. Tumor-Targeting Salmonella typhimurium A1-R Promotes Tumoricidal CD8+ T Cell Tumor Infiltration and Arrests Growth and Metastasis in a Syngeneic Pancreatic-Cancer Orthotopic Mouse Model.

    Science.gov (United States)

    Murakami, Takashi; Hiroshima, Yukihiko; Zhang, Yong; Zhao, Ming; Kiyuna, Tasuku; Hwang, Ho Kyoung; Miyake, Kentaro; Homma, Yuki; Mori, Ryutaro; Matsuyama, Ryusei; Chishima, Takashi; Ichikawa, Yasushi; Tanaka, Kuniya; Bouvet, Michael; Endo, Itaru; Hoffman, Robert M

    2018-01-01

    The present study determined the effect of the tumor-targeting strain Salmonella typhimurium A1-R (S. typhimurium A1-R) on CD8 + tumor-infiltrating lymphocytes (TILs) in a syngeneic pancreatic-cancer orthotopic mouse model. The effect of tumor-targeting S. typhimurium A1-R on CD8 + TILs was determined on the Pan02 murine pancreatic-adenocarcinoma implanted orthotopically in the pancreatic tail of C57BL/6 immunocompromised mice. Three weeks after orthotopic implantation, mice were randomized as follows G1: untreated control group (n = 8); and G2: S. typhimurium A1-R-treatment group (n = 8, 1 × 10 7 colony forming units [CFU]/body, iv, weekly, 3 weeks). On the 22nd day from initial treatment, all mice were sacrificed and tumors were harvested. The tumor-volume ratio was defined as ratio of tumor volume on the 22nd day relative to the 1st day. The tumor volume ratio was significantly lower in the S. typhimurium A1-R-treated group (G2) (3.0 ± 2.8) than the untreated control (G1) (39.9 ± 30.7, P R-treated mice (G2). Six mice in G1 had peritoneal dissemination, whereas no mice showed peritoneal dissemination in G2 (P R promotes CD8 + T cell infiltration and inhibition of tumor growth and metastasis. J. Cell. Biochem. 119: 634-639, 2018. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

  1. Co-stimulation through 4-1BB/CD137 improves the expansion and function of CD8(+ melanoma tumor-infiltrating lymphocytes for adoptive T-cell therapy.

    Directory of Open Access Journals (Sweden)

    Jessica Ann Chacon

    Full Text Available Adoptive T-cell therapy (ACT using tumor-infiltrating lymphocytes (TIL can induce tumor regression in up to 50% or more of patients with unresectable metastatic melanoma. However, current methods to expand melanoma TIL, especially the "rapid expansion protocol" (REP were not designed to enhance the generation of optimal effector-memory CD8(+ T cells for infusion. One approach to this problem is to manipulate specific co-stimulatory signaling pathways to enhance CD8(+ effector-memory T-cell expansion. In this study, we determined the effects of activating the TNF-R family member 4-1BB/CD137, specifically induced in activated CD8(+ T cells, on the yield, phenotype, and functional activity of expanded CD8(+ T cells during the REP. We found that CD8(+ TIL up-regulate 4-1BB expression early during the REP after initial TCR stimulation, but neither the PBMC feeder cells in the REP or the activated TIL expressed 4-1BB ligand. However, addition of an exogenous agonistic anti-4-1BB IgG4 (BMS 663513 to the REP significantly enhanced the frequency and total yield of CD8(+ T cells as well as their maintenance of CD28 and increased their anti-tumor CTL activity. Gene expression analysis found an increase in bcl-2 and survivin expression induced by 4-1BB that was associated with an enhanced survival capability of CD8(+ post-REP TIL when re-cultured in the absence or presence of cytokines. Our findings suggest that adding an agonistic anti-4-1BB antibody during the time of TIL REP initiation produces a CD8(+ T cell population capable of improved effector function and survival. This may greatly improve TIL persistence and anti-tumor activity in vivo after adoptive transfer into patients.

  2. Simplified protocol for clinical-grade tumor-infiltrating lymphocyte manufacturing with use of the Wave bioreactor

    DEFF Research Database (Denmark)

    Donia, Marco; Larsen, Signe Møllebæk; Met, Ozcan

    2014-01-01

    , a practical and simple protocol of TIL manufacturing with the use of a closed-system bioreactor was developed and implemented at our institution. RESULTS: This protocol enabled significant work load reduction during the most labor-intense step of TIL expansion, and allowed generation of high-quality TIL...

  3. Value of large scale expansion of tumor infiltrating lymphocytes in a compartmentalised gas-permeable bag: interests for adoptive immunotherapy

    Science.gov (United States)

    2011-01-01

    Background Adoptive cell therapy (ACT) has emerged as an effective treatment for patients with metastatic melanoma. However, there are several logistical and safety concerns associated with large-scale ex vivo expansion of tumour-specific T lymphocytes for widespread availability of ACT for cancer patients. To address these problems we developed a specific compartmentalised bag allowing efficient expansion of tumour-specific T lymphocytes in an easy handling, closed system. Methods Starting from lymph nodes from eight melanoma patients, we performed a side-by-side comparison of Tumour-Infiltrating Lymphocytes (TIL) produced after expansion in the compartmentalised bag versus TIL produced using the standard process in plates. Proliferation yield, viability, phenotype and IFNγ secretion were comparatively studied. Results We found no differences in proliferation yield and cell viability between both TIL production systems. Moreover, each of the cell products complied with our defined release criteria before being administered to the patient. The phenotype analysis indicated that the compartmentalised bag favours the expansion of CD8+ cells. Finally, we found that TIL stimulated in bags were enriched in reactive CD8+ T cells when co-cultured with the autologous melanoma cell line. Conclusions The stimulation of TIL with feeder cells in the specifically designed compartmentalised bag can advantageously replace the conventional protocol using plates. In particular, the higher expansion rate of reactive CD8+ T cells could have a significant impact for ACT. PMID:21575188

  4. Standardized evaluation of tumor-infiltrating lymphocytes in breast cancer: results of the ring studies of the international immuno-oncology biomarker working group.

    Science.gov (United States)

    Denkert, Carsten; Wienert, Stephan; Poterie, Audrey; Loibl, Sibylle; Budczies, Jan; Badve, Sunil; Bago-Horvath, Zsuzsanna; Bane, Anita; Bedri, Shahinaz; Brock, Jane; Chmielik, Ewa; Christgen, Matthias; Colpaert, Cecile; Demaria, Sandra; Van den Eynden, Gert; Floris, Giuseppe; Fox, Stephen B; Gao, Dongxia; Ingold Heppner, Barbara; Kim, S Rim; Kos, Zuzana; Kreipe, Hans H; Lakhani, Sunil R; Penault-Llorca, Frederique; Pruneri, Giancarlo; Radosevic-Robin, Nina; Rimm, David L; Schnitt, Stuart J; Sinn, Bruno V; Sinn, Peter; Sirtaine, Nicolas; O'Toole, Sandra A; Viale, Giuseppe; Van de Vijver, Koen; de Wind, Roland; von Minckwitz, Gunter; Klauschen, Frederick; Untch, Michael; Fasching, Peter A; Reimer, Toralf; Willard-Gallo, Karen; Michiels, Stefan; Loi, Sherene; Salgado, Roberto

    2016-10-01

    Multiple independent studies have shown that tumor-infiltrating lymphocytes (TIL) are prognostic in breast cancer with potential relevance for response to immune-checkpoint inhibitor therapy. Although many groups are currently evaluating TIL, there is no standardized system for diagnostic applications. This study reports the results of two ring studies investigating TIL conducted by the International Working Group on Immuno-oncology Biomarkers. The study aim was to determine the intraclass correlation coefficient (ICC) for evaluation of TIL by different pathologists. A total of 120 slides were evaluated by a large group of pathologists with a web-based system in ring study 1 and a more advanced software-system in ring study 2 that included an integrated feedback with standardized reference images. The predefined aim for successful ring studies 1 and 2 was an ICC above 0.7 (lower limit of 95% confidence interval (CI)). In ring study 1 the prespecified endpoint was not reached (ICC: 0.70; 95% CI: 0.62-0.78). On the basis of an analysis of sources of variation, we developed a more advanced digital image evaluation system for ring study 2, which improved the ICC to 0.89 (95% CI: 0.85-0.92). The Fleiss' kappa value for international standardization project shows that reproducible evaluation of TIL is feasible in breast cancer. This opens the way for standardized reporting of tumor immunological parameters in clinical studies and diagnostic practice. The software-guided image evaluation approach used in ring study 2 may be of value as a tool for evaluation of TIL in clinical trials and diagnostic practice. The experience gained from this approach might be applicable to the standardization of other diagnostic parameters in histopathology.

  5. Mitochondria in neutrophil apoptosis

    NARCIS (Netherlands)

    van Raam, B. J.; Verhoeven, A. J.; Kuijpers, T. W.

    2006-01-01

    Central in the regulation of the short life span of neutrophils are their mitochondria. These organelles hardly contribute to the energy status of neutrophils but play a vital role in the apoptotic process. Not only do the mitochondria contain cytotoxic proteins that are released during apoptosis

  6. Fractional laser exposure induces neutrophil infiltration (N1 phenotype into the tumor and stimulates systemic anti-tumor immune response.

    Directory of Open Access Journals (Sweden)

    Masayoshi Kawakubo

    Full Text Available Ablative fractional photothermolysis (aFP using a CO2 laser generates multiple small diameter tissue lesions within the irradiation field. aFP is commonly used for a wide variety of dermatological indications, including treatment of photodamaged skin and dyschromia, drug delivery and modification of scars due to acne, surgical procedures and burns. In this study we explore the utility of aFP for treating oncological indications, including induction of local tumor regression and inducing anti-tumor immunity, which is in marked contrast to current indications of aFP.We used a fractional CO2 laser to treat a tumor established by BALB/c colon carcinoma cell line (CT26.CL25, which expressed a tumor antigen, beta-galactosidase (beta-gal. aFP treated tumors grew significantly slower as compared to untreated controls. Complete remission after a single aFP treatment was observed in 47% of the mice. All survival mice from the tumor inoculation rejected re-inoculation of the CT26.CL25 colon carcinoma cells and moreover 80% of the survival mice rejected CT26 wild type colon carcinoma cells, which are parental cells of CT26.CL25 cells. Histologic section of the FP-treated tumors showed infiltrating neutrophil in the tumor early after aFP treatment. Flow cytometric analysis of tumor-infiltrating lymphocytes showed aFP treatment abrogated the increase in regulatory T lymphocyte (Treg, which suppresses anti-tumor immunity and elicited the expansion of epitope-specific CD8+ T lymphocytes, which were required to mediate the tumor-suppressing effect of aFP.We have demonstrated that aFP is able to induce a systemic anti-tumor adaptive immunity preventing tumor recurrence in a murine colon carcinoma in a mouse model. This study demonstrates a potential role of aFP treatments in oncology and further studies should be performed.

  7. The effect of chemotherapy with or without radiation on the accuracy of MR imaging for evaluating tumor infiltration into the bladder wall in cases of advanced bladder cancer

    International Nuclear Information System (INIS)

    Nishimura, Kazushige; Satou, Yuji; Nannri, Masaharu

    2004-01-01

    Staging of tumor infiltration into the bladder wall is one of the critical points in decision-making for optimal treatment of invasive bladder cancer. We studied the correlation of MR findings with pathological diagnosis in cases of invasive bladder cancer which had been treated with chemotherapy, with or without radiation. Twenty-one patients (14 men and 7 women) with invasive bladder tumors who underwent either partial cystectomy or radical cystectomy were entered into the study. Eight cases had received chemotherapy after staging biopsy (Group A), 6 cases had undergone chemo-radiation therapy following staging biopsy (Group B), and 7 cases had received no adjuvant therapy except for staging biopsy preoperatively (Group C). All cases had MR imaging study before surgical treatment. The pathological stage was assessed by examining the whole layer of the resected bladder wall. Pathological diagnosis was pT0 in 4 cases, pT1 in 2 cases, pT2b in 5 cases, pT3a in 2 cases and pT3b in 8 cases. Staging with MR imaging was consistent with pathological findings in 14 of the 21 cases (66.7%), while MR imaging produced over-staging in 6 cases and under-staging in 1 case. Of the 6 cases with over-staging, 2 cases had received chemo-radiation therapy, 2 cases had received chemotherapy, and 2 cases had received staging biopsy alone preoperatively. The one case with under-staging had received chemo-radiation therapy preoperatively. The accuracy in staging with MR imaging was 75.0% (6/8), 50.0% (3/6), and 71.4% (5/7) in Groups A, B, and C, respectively. Imaging study with MR is useful for the staging of invasive bladder cancer. However, care should be taken in the staging of invasive bladder tumors which have been treated with chemotherapy, with or without radiation therapy, because inflammatory infiltration and/or fibrous change caused by the chemo-radiation make accurate staging with MR imaging difficult. (author)

  8. Cryptococcus neoformans modulates extracellular killing by neutrophils

    Directory of Open Access Journals (Sweden)

    Asfia eQureshi

    2011-09-01

    Full Text Available We recently established a key role for host sphingomyelin synthase (SMS in the regulation of the killing activity of neutrophils against Cryptococcus neoformans. In this work, we studied the effect of C. neoformans on the killing activity of neutrophils and whether SMS would still be a player against C. neoformans in immunocompromised mice lacking T and NK cells (Tgε26 mice. To this end, we analyzed whether C. neoformans would have any effect on neutrophil survival and killing in vitro and in vivo. We show that unlike C. albicans, neither the presence nor the capsule size of C. neoformans cells have any effect on neutrophil viability. Interestingly, melanized C. neoformans cells totally abrogated the killing activity of neutrophils. Next, we monitored how exposure of neutrophils to C. neoformans cells would interfere with any further killing activity of the medium and found that pre-incubation with live but not heat-killed fungal cells significantly inhibits further killing activity of the medium. We next studied whether activation of SMS at the site of C. neoformans infection is dependent on T and NK cells. Using matrix-assisted laser desorption-ionization (MALDI tissue imaging in infected lung we found that similarly to previous observations in the isogenic wild type CBA/J mice, SM 16:0 levels are significantly elevated at the site of infection in mice lacking T and NK cells but only at early time points. This study highlights that C. neoformans may negatively regulate the killing activity of neutrophils and that SMS activation in neutrophils appears to be partially independent of T and/or NK cells.

  9. Prognostic Significance of Combination of Preoperative Platelet Count and Neutrophil-Lymphocyte Ratio (COP-NLR in Patients with Non-Small Cell Lung Cancer: Based on a Large Cohort Study.

    Directory of Open Access Journals (Sweden)

    Hua Zhang

    Full Text Available The aim of this study was to investigate the prognostic significance of the combination of the preoperative platelet count and neutrophil-lymphocyte ratio (COP-NLR for predicting postoperative survival of patients undergoing complete resection for non-small cell lung cancer (NSCLC.The preoperative COP-NLR was calculated on the basis of data obtained.Patients with both an increased platelet count (>30.0 × 104 mm(-3 and an elevated NLR (>2.3 were assigned a score of 2, and patients with one or neither were assigned as a score of 1 or 0, respectively.A total of 1238 NSCLC patients were enrolled in this analysis. Multivariate analysis using the 15 clinicolaboratory variables selected by univariate analyses demonstrated that the preoperative COP-NLR was an independent prognostic factor for DFS (HR: 1.834, 95%CI: 1.536 to 2.200, P<0.001 and OS (HR: 1.810, 95%CI: 1.587 to 2.056, P<0.001. In sub-analyses by tumor stage (I, II, IIIA, a significant association was found between DFS and OS and level of COP-NLR in each subgroup (P<0.001, P=0.002, P<0.001 for DFS, respectively; P<0.001, P=0.001, P<0.001 for OS. When the subgroup of patients with high-risk COP-NLR (score of 2 was analyzed, no benefit of adjuvant chemotherapy could be found (P=0.237 for DFS and P=0.165 for OS.The preoperative COP-NLR is able to predict the prognosis of patients with NSCLC and divide these patients into three independent groups before surgery. Our results also demonstrate that high-risk patients based on the COP-NLR do not benefit from adjuvant chemotherapy. Independent validation of our findings is warranted.

  10. Autophagy Primes Neutrophils for Neutrophil Extracellular Trap Formation during Sepsis.

    Science.gov (United States)

    Park, So Young; Shrestha, Sanjeeb; Youn, Young-Jin; Kim, Jun-Kyu; Kim, Shin-Yeong; Kim, Hyun Jung; Park, So-Hee; Ahn, Won-Gyun; Kim, Shin; Lee, Myung Goo; Jung, Ki-Suck; Park, Yong Bum; Mo, Eun-Kyung; Ko, Yousang; Lee, Suh-Young; Koh, Younsuck; Park, Myung Jae; Song, Dong-Keun; Hong, Chang-Won

    2017-09-01

    Neutrophils are key effectors in the host's immune response to sepsis. Excessive stimulation or dysregulated neutrophil functions are believed to be responsible for sepsis pathogenesis. However, the mechanisms regulating functional plasticity of neutrophils during sepsis have not been fully determined. We investigated the role of autophagy in neutrophil functions during sepsis in patients with community-acquired pneumonia. Neutrophils were isolated from patients with sepsis and stimulated with phorbol 12-myristate 13-acetate (PMA). The levels of reactive oxygen species generation, neutrophil extracellular trap (NET) formation, and granule release, and the autophagic status were evaluated. The effect of neutrophil autophagy augmentation was further evaluated in a mouse model of sepsis. Neutrophils isolated from patients who survived sepsis showed an increase in autophagy induction, and were primed for NET formation in response to subsequent PMA stimulation. In contrast, neutrophils isolated from patients who did not survive sepsis showed dysregulated autophagy and a decreased response to PMA stimulation. The induction of autophagy primed healthy neutrophils for NET formation and vice versa. In a mouse model of sepsis, the augmentation of autophagy improved survival via a NET-dependent mechanism. These results indicate that neutrophil autophagy primes neutrophils for increased NET formation, which is important for proper neutrophil effector functions during sepsis. Our study provides important insights into the role of autophagy in neutrophils during sepsis.

  11. Characterization of Yersinia pestis Interactions with Human Neutrophils In vitro

    Directory of Open Access Journals (Sweden)

    Sophia C. Dudte

    2017-08-01

    Full Text Available Yersinia pestis is a gram-negative, zoonotic, bacterial pathogen, and the causative agent of plague. The bubonic form of plague occurs subsequent to deposition of bacteria in the skin by the bite of an infected flea. Neutrophils are recruited to the site of infection within the first few hours and interactions between neutrophils and Y. pestis have been demonstrated in vivo. In contrast to macrophages, neutrophils have been considered non-permissive to Y. pestis intracellular survival. Several studies have shown killing of the vast majority of Y. pestis ingested by human neutrophils. However, survival of 10–15% of Y. pestis after phagocytosis by neutrophils is consistently observed. Furthermore, these surviving bacteria eventually replicate within and escape from the neutrophils. We set out to further characterize the interactions between Y. pestis and human neutrophils by (1 determining the effects of known Y. pestis virulence factors on bacterial survival after uptake by neutrophils, (2 examining the mechanisms employed by the neutrophil to kill the majority of intracellular Y. pestis, (3 determining the activation phenotype of Y. pestis-infected neutrophils, and (4 characterizing the Y. pestis-containing phagosome in neutrophils. We infected human neutrophils in vitro with Y. pestis and assayed bacterial survival and uptake. Deletion of the caf1 gene responsible for F1 capsule production resulted in significantly increased uptake of Y. pestis. Surprisingly, while the two-component regulator PhoPQ system is important for survival of Y. pestis within neutrophils, pre-induction of this system prior to infection did not increase bacterial survival. We used an IPTG-inducible mCherry construct to distinguish viable from non-viable intracellular bacteria and determined the association of the Y. pestis-containing phagosome with neutrophil NADPH-oxidase and markers of primary, secondary and tertiary granules. Additionally, we show that inhibition of

  12. Presence of intratumoral neutrophils is an independent prognostic factor in localized renal cell carcinoma

    DEFF Research Database (Denmark)

    Jensen, Hanne Krogh; Donskov, Frede; Marcussen, Niels

    2009-01-01

    : The intratumoral neutrophils ranged from zero to 289 cells/mm(2) tumor tissue. The presence of intratumoral neutrophils was statistically significantly associated with increasing tumor size, low hemoglobin, high creatinine, and CA IX

  13. Neutrophil labeling with [99mTc]-technetium stannous colloid is complement receptor 3-mediated and increases the neutrophil priming response to lipopolysaccharide

    International Nuclear Information System (INIS)

    Gallagher, Hayley; Ramsay, Stuart C.; Barnes, Jodie; Maggs, Jacqueline; Cassidy, Nathan; Ketheesan, Natkunam

    2006-01-01

    Introduction: [ 99m Tc]-technetium stannous colloid (TcSnC)-labeled white cells are used to image inflammation. Neutrophil labeling with TcSnC is probably phagocytic, but the phagocytic receptor involved is not known. We hypothesised that complement receptor 3 (CR3) plays a key role. Phagocytic labeling could theoretically result in neutrophil activation or priming, affecting the behaviour of labeled cells. Fluorescence-activated cell sorter (FACS) analysis side scatter measurements can assess neutrophil activation and priming. Methods: We tested whether TcSnC neutrophil labeling is CR3-mediated by assessing if neutrophil uptake of TcSnC was inhibited by a monoclonal antibody (mAb) directed at the CD11b component of CR3. We tested if TcSnC-labeled neutrophils show altered activation or priming status, comparing FACS side scatter in labeled and unlabeled neutrophils and examining the effect of lipopolysaccharide (LPS), a known priming agent. Results: Anti-CD11b mAb reduced neutrophil uptake of TcSnC in a dose-dependent fashion. Labeled neutrophils did not show significantly increased side scatter compared to controls. LPS significantly increased side scatter in control cells and labeled neutrophils. However, the increase was significantly greater in labeled neutrophils than unlabeled cells. Conclusions: Neutrophil labeling with TcSnC is related to the function of CR3, a receptor which plays a central role in phagocytosis. TcSnC labeling did not significantly activate or prime neutrophils. However, labeled neutrophils showed a greater priming response to LPS. This could result in labeled neutrophils demonstrating increased adhesion on activated endothelium at sites of infection

  14. Neutrophil labeling with [{sup 99m}Tc]-technetium stannous colloid is complement receptor 3-mediated and increases the neutrophil priming response to lipopolysaccharide

    Energy Technology Data Exchange (ETDEWEB)

    Gallagher, Hayley [School of Veterinary and Biomedical Sciences, James Cook University, Townsville, Queensland 4811 (Australia); Ramsay, Stuart C. [School of Medicine, James Cook University, Townsville, Queensland (Australia) and Townsville Nuclear Medicine, Mater Hospital, Townsville, Queensland 4812 (Australia)]. E-mail: stuart.ramsey@jcu.edu.au; Barnes, Jodie [School of Veterinary and Biomedical Sciences, James Cook University, Townsville, Queensland 4811 (Australia); Maggs, Jacqueline [Department of Nuclear Medicine, Townsville Hospital, Townsville, Queensland 4814 (Australia); Cassidy, Nathan [Townsville Nuclear Medicine, Mater Hospital, Townsville, Queensland 4812 (Australia); Ketheesan, Natkunam [School of Veterinary and Biomedical Sciences, James Cook University, Townsville, Queensland 4811 (Australia); School of Medicine, James Cook University, Townsville, Queensland (Australia)

    2006-04-15

    Introduction: [{sup 99m}Tc]-technetium stannous colloid (TcSnC)-labeled white cells are used to image inflammation. Neutrophil labeling with TcSnC is probably phagocytic, but the phagocytic receptor involved is not known. We hypothesised that complement receptor 3 (CR3) plays a key role. Phagocytic labeling could theoretically result in neutrophil activation or priming, affecting the behaviour of labeled cells. Fluorescence-activated cell sorter (FACS) analysis side scatter measurements can assess neutrophil activation and priming. Methods: We tested whether TcSnC neutrophil labeling is CR3-mediated by assessing if neutrophil uptake of TcSnC was inhibited by a monoclonal antibody (mAb) directed at the CD11b component of CR3. We tested if TcSnC-labeled neutrophils show altered activation or priming status, comparing FACS side scatter in labeled and unlabeled neutrophils and examining the effect of lipopolysaccharide (LPS), a known priming agent. Results: Anti-CD11b mAb reduced neutrophil uptake of TcSnC in a dose-dependent fashion. Labeled neutrophils did not show significantly increased side scatter compared to controls. LPS significantly increased side scatter in control cells and labeled neutrophils. However, the increase was significantly greater in labeled neutrophils than unlabeled cells. Conclusions: Neutrophil labeling with TcSnC is related to the function of CR3, a receptor which plays a central role in phagocytosis. TcSnC labeling did not significantly activate or prime neutrophils. However, labeled neutrophils showed a greater priming response to LPS. This could result in labeled neutrophils demonstrating increased adhesion on activated endothelium at sites of infection.

  15. Neutrophil extracellular traps - the dark side of neutrophils

    DEFF Research Database (Denmark)

    Sørensen, Ole E.; Borregaard, Niels

    2016-01-01

    Neutrophil extracellular traps (NETs) were discovered as extracellular strands of decondensed DNA in complex with histones and granule proteins, which were expelled from dying neutrophils to ensnare and kill microbes. NETs are formed during infection in vivo by mechanisms different from those ori...

  16. Regulation of neutrophil senescence by microRNAs.

    Directory of Open Access Journals (Sweden)

    Jon R Ward

    2011-01-01

    Full Text Available Neutrophils are rapidly recruited to sites of tissue injury or infection, where they protect against invading pathogens. Neutrophil functions are limited by a process of neutrophil senescence, which renders the cells unable to respond to chemoattractants, carry out respiratory burst, or degranulate. In parallel, aged neutrophils also undergo spontaneous apoptosis, which can be delayed by factors such as GMCSF. This is then followed by their subsequent removal by phagocytic cells such as macrophages, thereby preventing unwanted inflammation and tissue damage. Neutrophils translate mRNA to make new proteins that are important in maintaining functional longevity. We therefore hypothesised that neutrophil functions and lifespan might be regulated by microRNAs expressed within human neutrophils. Total RNA from highly purified neutrophils was prepared and subjected to microarray analysis using the Agilent human miRNA microarray V3. We found human neutrophils expressed a selected repertoire of 148 microRNAs and that 6 of these were significantly upregulated after a period of 4 hours in culture, at a time when the contribution of apoptosis is negligible. A list of predicted targets for these 6 microRNAs was generated from http://mirecords.biolead.org and compared to mRNA species downregulated over time, revealing 83 genes targeted by at least 2 out of the 6 regulated microRNAs. Pathway analysis of genes containing binding sites for these microRNAs identified the following pathways: chemokine and cytokine signalling, Ras pathway, and regulation of the actin cytoskeleton. Our data suggest that microRNAs may play a role in the regulation of neutrophil senescence and further suggest that manipulation of microRNAs might represent an area of future therapeutic interest for the treatment of inflammatory disease.

  17. Immunosenescence of Polymorphonuclear Neutrophils

    Directory of Open Access Journals (Sweden)

    Inga Wessels

    2010-01-01

    Full Text Available All immune cells are affected by aging, contributing to the high susceptibility to infections and increased mortality observed in the elderly. The effect of aging on cells of the adaptive immune system is well documented. In contrast, knowledge concerning age-related defects of polymorphonuclear neutrophils (PMN is limited. During the past decade, it has become evident that in addition to their traditional role as phagocytes, neutrophils are able to secrete a wide array of immunomodulating molecules. Their importance is underlined by the finding that genetic defects that lead to neutropenia increase susceptibility to infections. Whereas there is consistence about the constant circulating number of PMN throughout aging, the abilities of tissue infiltration, phagocytosis, and oxidative burst of PMN from aged donors are discussed controversially. Furthermore, there are numerous discrepancies between in vivo and in vitro results, as well as between results for murine and human PMN. Most of the reported functional changes can be explained by defective signaling pathways, but further research is required to get a detailed insight into the underlying molecular mechanisms. This could form the basis for drug development in order to prevent or treat age-related diseases, and thus to unburden the public health systems.

  18. Neutrophil adhesion and chemotaxis depend on substrate mechanics

    International Nuclear Information System (INIS)

    Jannat, Risat A; Hammer, Daniel A; Robbins, Gregory P; Ricart, Brendon G; Dembo, Micah

    2010-01-01

    Neutrophil adhesion to the vasculature and chemotaxis within tissues play critical roles in the inflammatory response to injury and pathogens. Unregulated neutrophil activity has been implicated in the progression of numerous chronic and acute diseases such as rheumatoid arthritis, asthma and sepsis. Cell migration of anchorage-dependent cells is known to depend on both chemical and mechanical interactions. Although neutrophil responses to chemical cues have been well characterized, little is known about the effect of underlying tissue mechanics on neutrophil adhesion and migration. To address this question, we quantified neutrophil migration and traction stresses on compliant hydrogel substrates with varying elasticity in a micromachined gradient chamber in which we could apply either a uniform concentration or a precise gradient of the bacterial chemoattractant fMLP. Neutrophils spread more extensively on substrates of greater stiffness. In addition, increasing the stiffness of the substrate leads to a significant increase in the chemotactic index for each fMLP gradient tested. As the substrate becomes stiffer, neutrophils generate higher traction forces without significant changes in cell speed. These forces are often displayed in pairs and focused in the uropod. Increases in the mean fMLP concentration beyond the K D of the receptor lead to a decrease in chemotactic index on all surfaces. Blocking with an antibody against β 2 -integrins leads to a significant reduction, but not an elimination, of directed motility on stiff materials, but no change in motility on soft materials, suggesting neutrophils can display both integrin-dependent and integrin-independent motility. These findings are critical for understanding how neutrophil migration may change in different mechanical environments in vivo and can be used to guide the design of migration inhibitors that more efficiently target inflammation.

  19. Neutrophil adhesion and chemotaxis depend on substrate mechanics

    Energy Technology Data Exchange (ETDEWEB)

    Jannat, Risat A; Hammer, Daniel A [Department of Bioengineering, University of Pennsylvania, 240 Skirkanich Hall, 210 South 33rd Street, Philadelphia, PA 19104 (United States); Robbins, Gregory P; Ricart, Brendon G [Department of Chemical and Biomolecular Engineering, University of Pennsylvania, 311A Towne Building, 220 South 33rd Street, Philadelphia, PA 19104 (United States); Dembo, Micah, E-mail: hammer@seas.upenn.ed [Department of Biomedical Engineering, Boston University, 44 Cummington Street, Boston, MA 02215 (United States)

    2010-05-19

    Neutrophil adhesion to the vasculature and chemotaxis within tissues play critical roles in the inflammatory response to injury and pathogens. Unregulated neutrophil activity has been implicated in the progression of numerous chronic and acute diseases such as rheumatoid arthritis, asthma and sepsis. Cell migration of anchorage-dependent cells is known to depend on both chemical and mechanical interactions. Although neutrophil responses to chemical cues have been well characterized, little is known about the effect of underlying tissue mechanics on neutrophil adhesion and migration. To address this question, we quantified neutrophil migration and traction stresses on compliant hydrogel substrates with varying elasticity in a micromachined gradient chamber in which we could apply either a uniform concentration or a precise gradient of the bacterial chemoattractant fMLP. Neutrophils spread more extensively on substrates of greater stiffness. In addition, increasing the stiffness of the substrate leads to a significant increase in the chemotactic index for each fMLP gradient tested. As the substrate becomes stiffer, neutrophils generate higher traction forces without significant changes in cell speed. These forces are often displayed in pairs and focused in the uropod. Increases in the mean fMLP concentration beyond the K{sub D} of the receptor lead to a decrease in chemotactic index on all surfaces. Blocking with an antibody against {beta}{sub 2}-integrins leads to a significant reduction, but not an elimination, of directed motility on stiff materials, but no change in motility on soft materials, suggesting neutrophils can display both integrin-dependent and integrin-independent motility. These findings are critical for understanding how neutrophil migration may change in different mechanical environments in vivo and can be used to guide the design of migration inhibitors that more efficiently target inflammation.

  20. Targeting neutrophilic inflammation in severe neutrophilic asthma : can we target the disease-relevant neutrophil phenotype?

    NARCIS (Netherlands)

    Bruijnzeel, Piet L B; Uddin, Mohib; Koenderman, Leo

    2015-01-01

    In severe, neutrophilic asthma, neutrophils are thought to have an important role in both the maintenance of the disease and during exacerbations. These patients often display excessive, mucosal airway inflammation with unresolving neutrophilia. Because this variant of asthma is poorly controlled by

  1. G-CSF maintains controlled neutrophil mobilization during acute inflammation by negatively regulating CXCR2 signaling

    Science.gov (United States)

    Bajrami, Besnik; Zhu, Haiyan; Zhang, Yu C.

    2016-01-01

    Cytokine-induced neutrophil mobilization from the bone marrow to circulation is a critical event in acute inflammation, but how it is accurately controlled remains poorly understood. In this study, we report that CXCR2 ligands are responsible for rapid neutrophil mobilization during early-stage acute inflammation. Nevertheless, although serum CXCR2 ligand concentrations increased during inflammation, neutrophil mobilization slowed after an initial acute fast phase, suggesting a suppression of neutrophil response to CXCR2 ligands after the acute phase. We demonstrate that granulocyte colony-stimulating factor (G-CSF), usually considered a prototypical neutrophil-mobilizing cytokine, was expressed later in the acute inflammatory response and unexpectedly impeded CXCR2-induced neutrophil mobilization by negatively regulating CXCR2-mediated intracellular signaling. Blocking G-CSF in vivo paradoxically elevated peripheral blood neutrophil counts in mice injected intraperitoneally with Escherichia coli and sequestered large numbers of neutrophils in the lungs, leading to sterile pulmonary inflammation. In a lipopolysaccharide-induced acute lung injury model, the homeostatic imbalance caused by G-CSF blockade enhanced neutrophil accumulation, edema, and inflammation in the lungs and ultimately led to significant lung damage. Thus, physiologically produced G-CSF not only acts as a neutrophil mobilizer at the relatively late stage of acute inflammation, but also prevents exaggerated neutrophil mobilization and the associated inflammation-induced tissue damage during early-phase infection and inflammation. PMID:27551153

  2. Tumor Associated Neutrophils in Human Lung Cancer

    Science.gov (United States)

    2016-10-01

    tumor innate immune response. anti-tumor adaptive immune response, neutrophil and T cell interaction. ACCOMPLISHMENTS There were no significant...and by producing factors to recruit and acti- vate cells of the innate and adaptive immune system (Mantovani et al., 2011). Given these varying effects...vivo effects on neutro- phil activation (Figure 2, A and B) and cleavage of myeloid and lymphoid cell markers (Supplemental Figure 1, C–G). Once opti

  3. Anti-neutrophil cytoplasmic antibodies stimulate release of neutrophil microparticles.

    LENUS (Irish Health Repository)

    Hong, Ying

    2012-01-01

    The mechanisms by which anti-neutrophil cytoplasmic antibodies (ANCAs) may contribute to the pathogenesis of ANCA-associated vasculitis are not well understood. In this study, both polyclonal ANCAs isolated from patients and chimeric proteinase 3-ANCA induced the release of neutrophil microparticles from primed neutrophils. These microparticles expressed a variety of markers, including the ANCA autoantigens proteinase 3 and myeloperoxidase. They bound endothelial cells via a CD18-mediated mechanism and induced an increase in endothelial intercellular adhesion molecule-1 expression, production of endothelial reactive oxygen species, and release of endothelial IL-6 and IL-8. Removal of the neutrophil microparticles by filtration or inhibition of reactive oxygen species production with antioxidants abolished microparticle-mediated endothelial activation. In addition, these microparticles promoted the generation of thrombin. In vivo, we detected more neutrophil microparticles in the plasma of children with ANCA-associated vasculitis compared with that in healthy controls or those with inactive vasculitis. Taken together, these results support a role for neutrophil microparticles in the pathogenesis of ANCA-associated vasculitis, potentially providing a target for future therapeutics.

  4. IP-10 is an important chemokine secreted by tumor infiltrating lymphocytes and is an independent prognostic factor in triple-negative breast cancer patients

    DEFF Research Database (Denmark)

    Elias, Daniel; Ditzel, Henrik; Kupisiewicz, Kasia

    of TILs isolated from frozen tumor sections of TNBC patients who experienced no recurrence or progression for at least 5 years (good prognosis) for comparison with those who had progression in the first 2 years post-surgery (bad prognosis). The results showed that 398 genes showed significantly altered...... expression (FDR of 0.05 and fold change of 2). 319 of these genes showed higher expression in TILs from TNBC patients with good prognosis, while only 79 showed lower expression compared to those from bad prognosis patients. Among the genes exhibiting altered expression was a strong representation of those...... related to lymphocyte activation, pro- and anti-inflammatory responses, cell stress and apoptotic cell clearance, including IP-10, CCL5, FCRL5, PINX1 and PSR. Co-culture experiments showed that TNBC cell lines stimulated high level expression of IP-10 (258 vs 189 pg/ml p= 0.043), IFNg (170 vs 98 pg/ml, p...

  5. Dihydroxyoctadecamonoenoate esters inhibit the neutrophil ...

    Indian Academy of Sciences (India)

    PRAKASH

    that observed with arachidonic acid treatment (Li et al 1996). ..... An alternative possibility is that the methyl DiHOMEs .... nitric oxide-derived reactive species in vascular cells; Circ. ... necrosis factor 1-alpha-initiated neutrophil responses and.

  6. Neutrophil evasion strategies by Streptococcus pneumoniae and Staphylococcus aureus.

    Science.gov (United States)

    Lewis, Megan L; Surewaard, Bas G J

    2018-03-01

    Humans are well equipped to defend themselves against bacteria. The innate immune system employs diverse mechanisms to recognize, control and initiate a response that can destroy millions of different microbes. Microbes that evade the sophisticated innate immune system are able to escape detection and could become pathogens. The pathogens Streptococcus pneumoniae and Staphylococcus aureus are particularly successful due to the development of a wide variety of virulence strategies for bacterial pathogenesis and they invest significant efforts towards mechanisms that allow for neutrophil evasion. Neutrophils are a primary cellular defense and can rapidly kill invading microbes, which is an indispensable function for maintaining host health. This review compares the key features of Streptococcus pneumoniae and Staphylococcus aureus in epidemiology, with a specific focus on virulence mechanisms utilized to evade neutrophils in bacterial pathogenesis. It is important to understand the complex interactions between pathogenic bacteria and neutrophils so that we can disrupt the ability of pathogens to cause disease.

  7. Suppressed neutrophil function in children with acute lymphoblastic leukemia.

    Science.gov (United States)

    Tanaka, Fumiko; Goto, Hiroaki; Yokosuka, Tomoko; Yanagimachi, Masakatsu; Kajiwara, Ryosuke; Naruto, Takuya; Nishimaki, Shigeru; Yokota, Shumpei

    2009-10-01

    Infection is a major obstacle in cancer chemotherapy. Neutropenia has been considered to be the most important risk factor for severe infection; however, other factors, such as impaired neutrophil function, may be involved in susceptibility to infection in patients undergoing chemotherapy. In this study, we analyzed neutrophil function in children with acute lymphoblastic leukemia (ALL). Whole blood samples were obtained from 16 children with ALL at diagnosis, after induction chemotherapy, and after consolidation chemotherapy. Oxidative burst and phagocytic activity of neutrophils were analyzed by flow cytometry. Oxidative burst of neutrophils was impaired in ALL patients. The percentage of neutrophils with normal oxidative burst after PMA stimulation was 59.0 +/- 13.2 or 70.0 +/- 21.0% at diagnosis or after induction chemotherapy, respectively, which was significantly lower compared with 93.8 +/- 6.1% in healthy control subjects (P = 0.00004, or 0.002, respectively); however, this value was normal after consolidation chemotherapy. No significant differences were noted in phagocytic activity in children with ALL compared with healthy control subjects. Impaired oxidative burst of neutrophils may be one risk factor for infections in children with ALL, especially in the initial periods of treatment.

  8. Regulation of apoptosis and priming of neutrophil oxidative burst by diisopropyl fluorophosphate

    Directory of Open Access Journals (Sweden)

    Tsang Jennifer LY

    2010-07-01

    Full Text Available Abstract Background Diisopropyl fluorophosphate (DFP is a serine protease inhibitor that is widely used as an inhibitor of endogenous proteases in in vitro neutrophil studies. Its effects on neutrophil function are unclear. We sought to determine the biological effects of DFP on human neutrophil apoptosis and oxidative burst. Methods We isolated neutrophils from healthy volunteers, incubated them with DFP (2.5 mM, and evaluated neutrophil elastase (NE activity, neutrophil degranulation, apoptosis as reflected in hypodiploid DNA formation and exteriorization of phosphatidylserine (PS, processing and activity of caspases-3 and -8, oxidative burst activity and hydrogen peroxide release. Results Consistent with its activity as a serine protease inhibitor, DFP significantly inhibited NE activity but not the degranulation of azurophilic granules. DFP inhibited constitutive neutrophil apoptosis as reflected in DNA fragmentation, and the processing and activity of caspases-3 and -8. DFP also inhibited priming of neutrophils for oxidative burst activity and hydrogen peroxide release. However, DFP enhanced the exteriorization of PS in a dose-dependent manner. Conclusion We conclude that DFP exerts significant effects on neutrophil inflammatory function that may confound the interpretation of studies that use it for its antiprotease activity. We further conclude that endogenous proteases play a role in the biology of constitutive neutrophil apoptosis.

  9. Hidden truth of circulating neutrophils (polymorphonuclear neutrophil function in periodontally healthy smoker subjects

    Directory of Open Access Journals (Sweden)

    Chitra Agarwal

    2016-01-01

    Full Text Available Context: Tobacco smoking is considered to be a major risk factor associated with periodontal disease. Smoking exerts a major effect on the protective elements of the immune response, resulting in an increase in the extent and severity of periodontal destruction. Aims: The aim of the present study was to assess viability and phagocytic function of neutrophils in circulating blood of the smokers and nonsmokers who are periodontally healthy. Settings and Design: Two hundred subjects in the mean range of 20–30 years of age were included in the study population. It was a retrospective study carried out for 6 months. Materials and Methods: Two hundred subjects were divided into four groups: 50 nonsmokers, 50 light smokers (15 cigarettes/day. Full mouth plaque index, sulcus bleeding index, and probing depths were measured. Percentage viability of circulating neutrophils and average number of phagocytosed Candida albicans were recorded. Statistical Analysis Used: Means and standard deviations were calculated from data obtained within the groups. Comparison between the smokers and nonsmokers was performed by Kruskal–Wallis ANOVA analysis. Comparison between smoker groups was performed using Mann–Whitney–Wilcoxon test. Results: Percentage viability of neutrophils was significantly less in heavy smokers (66.9 ± 4.0, moderate (76.6 ± 4.2, light smokers (83.1 ± 2.5 as compared to nonsmokers (92.3 ± 2.6 (P < 0.01. The ability of neutrophils to phagocytose, i.e., mean particle number was significantly less in light smokers (3.5 ± 0.5, moderate smokers (2.3 ± 0.5, and heavy smokers (1.4 ± 0.5 compared to nonsmokers (4.9 ± 0.7 (P < 0.01 with evidence of dose-response effect. Conclusions: Smoking significantly affects neutrophils viability and phagocytic function in periodontally healthy population.

  10. An Examination of the Local Cellular Immune Response to Examples of Both Ductal Carcinoma In Situ (DCIS) of the Breast and DCIS With Microinvasion, With Emphasis on Tertiary Lymphoid Structures and Tumor Infiltrating Lymphoctytes.

    Science.gov (United States)

    Kim, Ahrong; Heo, Sun-Hee; Kim, Young-Ae; Gong, Gyungyub; Jin Lee, Hee

    2016-07-01

    We tried to describe cellular immune response (tertiary lymphoid structures (TLSs), lymphoid aggregates, tumor infiltrating lymphocytes (TILs)) in neoplastic microenvironment of ductal carcinoma in situ (DCIS) with or without associated microinvasion. The histopathologic parameters of 177 DCIS and 27 DCIS with microinvasion were evaluated. We determined number of ducts involved by DCIS, and calculated percentage of these ducts surrounded by TLSs. TILs were quantitated in 27 microinvasive cases. Tumors having higher percentage of DCIS ducts associated with TLSs had higher incidence of microinvasion (P < .001). Percentage of DCIS ducts involved by TLSs was also higher in hormone receptor (HR)-/human epidermal growth factor receptor 2 (HER2)+ and TNBC subtypes of DCIS than in HR+/HER2- and HR+/HER2+ subtypes (38.04 ± 25.8%, 32.6 ± 32.4%, 2.5 ± 7.3% and 17.4 ± 23.3%, respectively, P < .001). In DCIS without microinvasion, HR+/HER2- subtype predominated (P < .001). In microinvasive cases, HR-/HER2+ subtype was most common. TNBC was more common in microinvasive carcinoma than DCIS (P < .001). Among 27 microinvasive ductal carcinomas, increased TLS amount was associated with increased TILs (P = .013). TLS abundance around DCIS was associated with HER2+ and TNBC subtypes and microinvasion. Pathologists should be aware of microinvasion when diagnosing DCIS lesions with abundant TLSs. © American Society for Clinical Pathology, 2016. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  11. Tumor-infiltrating CD4+ T lymphocytes in early breast cancer reflect lymph node involvement Linfócitos T CD4+ tumor infiltrantes no câncer de mama inicial refletem envolvimento linfonodal

    Directory of Open Access Journals (Sweden)

    Alexandre Henrique Macchetti

    2006-06-01

    Full Text Available BACKGROUND: The role of immune system in the pathogenesis and progression of breast cancer is a subject of controversy, and this stimulated us to investigate the association of the immunophenotype of tumor-infiltrating lymphocytes in early breast cancer with the spread of tumor cells to axillary lymph nodes. METHODS: Tumor samples from 23 patients with early breast cancer from the Department of Gynecology and Obstetrics of Ribeirão Preto Medical School (USP were obtained at the time of biopsy and submitted to an enzyme-digestion procedure for the extraction of tumor-infiltrating lymphocytes. The lymphocytes extracted were analyzed by dual-color flow cytometry with monoclonal antibodies in these combinations: CD3 FITC/CD19 PE, CD3 FITC/CD4 PE, CD3 FITC/CD8 PE, and CD16/56 PerCP, which are specific for immunophenotyping of T and B lymphocytes, helper and cytotoxic T lymphocytes, and natural killer (NK cells. The mean percentage of these cells was used for comparing groups of patients with or without lymph node metastasis. RESULTS: The mean value for T-lymphocyte infiltration was 24.72 ± 17.37%; for B-lymphocyte infiltration, 4.22 ± 6.27%; for NK-cell infiltration, 4.41 ± 5.22%, and for CD4+ and CD8+ T-lymphocyte infiltration, 12.43 ± 10.12% and 11.30 ± 15.09%, respectively. Only mean values of T- and CD4+ T-lymphocyte infiltration were higher in the group of patients with lymph node metastasis, while no differences were noted in the other lymphocyte subpopulations. CONCLUSION: The association of tumor-infiltrating CD4+ T lymphocytes with lymph node metastasis suggests a role for these cells in the spread of neoplasia to lymph nodes in patients with early breast cancer.INTRODUÇÃO: O papel do sistema imunológico na patogênese e progressão do câncer de mama ainda é controverso, e isto nos estimulou a verificar a associação do imunofenótipo dos linfócitos tumor infiltrantes do câncer de mama inicial com a disseminação de c

  12. In vivo activation of equine eosinophils and neutrophils by experimental Strongylus vulgaris infections.

    Science.gov (United States)

    Dennis, V A; Klei, T R; Chapman, M R; Jeffers, G W

    1988-12-01

    Eosinophils and neutrophils from ponies with Strongylus vulgaris-induced eosinophilia (eosinophilic ponies; activated eosinophils and neutrophils) were assayed in vitro for chemotactic and chemokinetic responses to zymosan-activated serum (ZAS) using the filter system in Boyden chambers, for Fc and complement (C) receptors using the EA and EAC-rosette assays, respectively, and for phagocytic and bactericidal activities using opsonized Escherichia coli and the acridine orange method. The responses of activated eosinophils and neutrophils in the above assays were compared with those of eosinophils and neutrophils from S. vulgaris-naive ponies without eosinophilia (noneosinophilic ponies; nonactivated eosinophils and neutrophils). Differences in cell density following centrifugation in a continuous Percoll gradient were used to further characterize the heterogeneity of activated eosinophils and neutrophils. Activated and nonactivated eosinophils demonstrated similar chemotactic responses to ZAS while activated and nonactivated neutrophils demonstrated similar chemokinetic responses to ZAS. A higher percentage of activated eosinophils and neutrophils expressed Fc and C receptors compared with nonactivated cells (P less than 0.05). Generally, higher percentages of eosinophils and neutrophils expressed C than Fc receptors. However, the percentage of neutrophils with both receptors was higher than that of eosinophils. Phagocytosis and killing of E. coli by either type of eosinophil were not consistently observed. Both activated and nonactivated neutrophils phagocytized E. coli and significant differences between the two cell types were not observed. The bacterial activity, however, of activated neutrophils was significantly greater than that obtained using nonactivated neutrophils (P less than 0.05). Activated eosinophils and neutrophils were both separated into two distinct fractions based on differences in cell densities. A higher percentage of band 2 eosinophils

  13. Disentangling the effects of tocilizumab on neutrophil survival and function.

    Science.gov (United States)

    Gaber, Timo; Hahne, Martin; Strehl, Cindy; Hoff, Paula; Dörffel, Yvonne; Feist, Eugen; Burmester, Gerd-Rüdiger; Buttgereit, Frank

    2016-06-01

    The synovial tissue in rheumatoid arthritis (RA) represents a hypoxic environment with up-regulated pro-inflammatory cytokines and cellular infiltrates including neutrophils. Although inhibition of the interleukin (IL)6 receptor pathway by tocilizumab is a potent treatment option for RA, it may also cause adverse effects such as an occasionally high-grade neutropenia. We analysed the impact of tocilizumab on survival, mediator secretion, oxidative burst, phagocytosis and energy availability of high-dose toll-like receptor (TLR)2/4-stimulated neutrophils (to mimic an arthritis flare) under normoxic versus hypoxic conditions. Human neutrophils were purified, pre-treated with varying doses of tocilizumab, dexamethasone or human IgG1 and high-dose-stimulated with lipopolysaccharide (LPS) alone-triggering TLR2/4-, LPS plus IL6, or left unstimulated. Cells were then incubated under normoxic (18 % O2) or hypoxic (1 % O2) conditions and subsequently analysed. Neutrophil survival and energy availability were significantly decreased by tocilizumab in a dose-dependent manner in high-dose TLR2/4-stimulated cells, but to a greater extent under normoxia as compared to hypoxia. We also found high-dose LPS-stimulated oxidative burst and phagocytosis of neutrophils to be higher under hypoxic versus normoxic conditions, but this difference was reduced by tocilizumab. Finally, we observed that tocilizumab affected neutrophil mediator secretion as a function of oxygen availability. Tocilizumab is known for both beneficial effects and a higher incidence of neutropenia when treating RA patients. Our results suggest that both effects can at least in part be explained by a reduction in neutrophil survival, a dose-dependent inhibition of hypoxia-induced NADPH oxidase-mediated oxidative burst and phagocytosis of infiltrating hypoxic neutrophils and an alteration of mediator secretion.

  14. Chemotactic Activity on Human Neutrophils to Streptococcus mutans

    Directory of Open Access Journals (Sweden)

    Tetiana Haniastuti

    2013-07-01

    Full Text Available Objective: The aim of this study was to evaluate chemotactic activity o neutrophil to S. mutans. Chemotaxis assay was performed in blind well chambers. Materials and Methods: Hanks balanced salt solution (HBSS containing 106 S. mutans,  108 S. mutans, 10-8 M fMLP, or HBSS alone were placed in the lower wells of the chamber and covered with polycorbonate membrane filter. Neutrophils suspension (2x105 cells was then placed in the upper compartment. After incubation for 60 mins at 37ºC in a humidified atmosphere with 5% CO2, the filters were removed and stained with Giemsa. Result: ANOVA revealed statistically significant differences among groups (p<0.05, indicating that S. mutans induced neutrophils chemotaxis. The number of neutrophils migration in response to 108 S. mutans and 106 S. mutans were signifiantly greater compared to fMLP (p<0.05. Conclusion: S. mutans may activate human neutrophils, resulting in the chemotaxis of the neutrophils.DOI: 10.14693/jdi.v16i2.99

  15. Age is the work of art? Impact of neutrophil and organism age on neutrophil extracellular trap formation.

    Science.gov (United States)

    Ortmann, Weronika; Kolaczkowska, Elzbieta

    2018-03-01

    Neutrophil extracellular traps or NETs are released by highly activated neutrophils in response to infectious agents, sterile inflammation, autoimmune stimuli and cancer. In the cells, the nuclear envelop disintegrates and decondensation of chromatin occurs that depends on peptidylarginine deiminase 4 (PAD4) and neutrophil elastase (NE). Subsequently, proteins from neutrophil granules (e.g., NE, lactoferrin and myeloperoxidase) and the nucleus (histones) bind to decondensed DNA and the whole structure is ejected from the cell. The DNA decorated with potent antimicrobials and proteases can act to contain dissemination of infection and in sterile inflammation NETs were shown to degrade cytokines and chemokines via serine proteases. On the other hand, overproduction of NETs, or their inadequate removal and prolonged presence in vasculature or tissues, can lead to bystander damage or even initiation of diseases. Considering the pros and cons of NET formation, it is of relevance if the stage of neutrophil maturation (immature, mature and senescent cells) affects the capacity to produce NETs as the cells of different age-related phenotypes dominate in given (pathological) conditions. Moreover, the immune system of neonates and elderly individuals is weaker than in adulthood. Is the same pattern followed when it comes to NETs? The overall importance of individual and neutrophil age on the capacity to release NETs is reviewed in detail and the significance of these facts is discussed.

  16. Ly6G-mediated depletion of neutrophils is dependent on macrophages.

    Science.gov (United States)

    Bruhn, Kevin W; Dekitani, Ken; Nielsen, Travis B; Pantapalangkoor, Paul; Spellberg, Brad

    2016-01-01

    Antibody-mediated depletion of neutrophils is commonly used to study neutropenia. However, the mechanisms by which antibodies deplete neutrophils have not been well defined. We noticed that mice deficient in complement and macrophages had blunted neutrophil depletion in response to anti-Ly6G monoclonal antibody (MAb) treatment. In vitro, exposure of murine neutrophils to anti-Ly6G MAb in the presence of plasma did not result in significant depletion of cells, either in the presence or absence of complement. In vivo, anti-Ly6G-mediated neutrophil depletion was abrogated following macrophage depletion, but not complement depletion, indicating a requirement for macrophages to induce neutropenia by this method. These results inform the use and limitations of anti-Ly6G antibody as an experimental tool for depleting neutrophils in various immunological settings.

  17. Neutrophils, dendritic cells and Toxoplasma.

    Science.gov (United States)

    Denkers, Eric Y; Butcher, Barbara A; Del Rio, Laura; Bennouna, Soumaya

    2004-03-09

    Toxoplasma gondii rapidly elicits strong Type 1 cytokine-based immunity. The necessity for this response is well illustrated by the example of IFN-gamma and IL-12 gene knockout mice that rapidly succumb to the effects of acute infection. The parasite itself is skilled at sparking complex interactions in the innate immune system that lead to protective immunity. Neutrophils are one of the first cell types to arrive at the site of infection, and the cells release several proinflammatory cytokines and chemokines in response to Toxoplasma. Dendritic cells are an important source of IL-12 during infection with T. gondii and other microbial pathogens, and they are also specialized for high-level antigen presentation to T lymphocytes. Tachyzoites express at least two types of molecules that trigger innate immune cell cytokine production. One of these involves Toll-like receptor/MyD88 pathways common to many microbial pathogens. The second pathway is less conventional and involves molecular mimicry between a parasite cyclophilin and host CC chemokine receptor 5-binding ligands. Neutrophils, dendritic cells and Toxoplasma work together to elicit the immune response required for host survival. Cytokine and chemokine cross-talk between parasite-triggered neutrophils and dendritic cells results in recruitment, maturation and activation of the latter. Neutrophil-empowered dendritic cells possess properties expected of highly potent antigen presenting cells that drive T helper 1 generation.

  18. Immune modulation by neutrophil subsets

    NARCIS (Netherlands)

    Kamp, V.M.

    2013-01-01

    We show that human neutrophils can suppress T-cell proliferation in acute systemic inflammation and thus have anti-inflammatory functions, next to their well-known pro-inflammatory functions. The suppression is mediated by ROS production and integrin MAC-1, which are also important for the

  19. Effects of acrolein on leukotriene biosynthesis in human neutrophils.

    Science.gov (United States)

    Berry, Karin A Zemski; Henson, Peter M; Murphy, Robert C

    2008-12-01

    Acrolein is a toxic, highly reactive alpha,beta-unsaturated aldehyde that is present in high concentrations in cigarette smoke. In the current study, the effect of acrolein on eicosanoid synthesis in stimulated human neutrophils was examined. Eicosanoid synthesis in neutrophils was initiated by priming with granulocyte-macrophage colony-stimulating factor (GM-CSF) and subsequent stimulation with formyl-methionyl-leucyl-phenylalanine (fMLP) and 5-lipoxygenase (5-LO) products in addition to small amounts of cyclooxygenase (COX) products were detected using LC/MS/MS. A dose-dependent decrease in the formation of 5-LO products was observed in GM-CSF/fMLP-stimulated neutrophils when acrolein (0-50 microM) was present with almost complete inhibition at > or = 25 microM acrolein. The production of COX products was not affected by acrolein in these cells. The effect of acrolein was examined on key parts of the eicosanoid pathway, such as arachidonic acid release, intracellular calcium ion concentration, and adenosine production. In addition, the direct effect of acrolein on 5-LO enzymatic activity was probed using a recombinant enzyme. Some of these factors were affected by acrolein but did not completely explain the almost complete inhibition of 5-LO product formation in GM-CSF/fMLP-treated cells with acrolein. In addition, the effect of acrolein on different stimuli that initiate the 5-LO pathway [platelet-activating factor (PAF)/fMLP, GM-CSF/PAF, opsonized zymosan, and A23187] was examined. Acrolein had no significant effect on the leukotriene production in neutrophils stimulated with PAF/fMLP, GM-CSF/ PAF, or OPZ. Additionally, 50% inhibition of the 5-LO pathway was observed in A23187-stimulated neutrophils. Our results suggest that acrolein has a profound effect on the 5-LO pathway in neutrophils, which may have implications in disease states, such as chronic obstructive pulmonary disease and other pulmonary disease, where both activated neutrophils and acrolein are

  20. Chemokine Receptor Ccr1 Drives Neutrophil-Mediated Kidney Immunopathology and Mortality in Invasive Candidiasis

    Science.gov (United States)

    Lionakis, Michail S.; Swamydas, Muthulekha; Wan, Wuzhou; Richard Lee, Chyi-Chia; Cohen, Jeffrey I.; Scheinberg, Phillip; Gao, Ji-Liang; Murphy, Philip M.

    2012-01-01

    Invasive candidiasis is the 4th leading cause of nosocomial bloodstream infection in the US with mortality that exceeds 40% despite administration of antifungal therapy; neutropenia is a major risk factor for poor outcome after invasive candidiasis. In a fatal mouse model of invasive candidiasis that mimics human bloodstream-derived invasive candidiasis, the most highly infected organ is the kidney and neutrophils are the major cellular mediators of host defense; however, factors regulating neutrophil recruitment have not been previously defined. Here we show that mice lacking chemokine receptor Ccr1, which is widely expressed on leukocytes, had selectively impaired accumulation of neutrophils in the kidney limited to the late phase of the time course of the model; surprisingly, this was associated with improved renal function and survival without affecting tissue fungal burden. Consistent with this, neutrophils from wild-type mice in blood and kidney switched from Ccr1lo to Ccr1high at late time-points post-infection, when Ccr1 ligands were produced at high levels in the kidney and were chemotactic for kidney neutrophils ex vivo. Further, when a 1∶1 mixture of Ccr1+/+ and Ccr1−/− donor neutrophils was adoptively transferred intravenously into Candida-infected Ccr1+/+ recipient mice, neutrophil trafficking into the kidney was significantly skewed toward Ccr1+/+ cells. Thus, neutrophil Ccr1 amplifies late renal immunopathology and increases mortality in invasive candidiasis by mediating excessive recruitment of neutrophils from the blood to the target organ. PMID:22916017

  1. Chemokine receptor Ccr1 drives neutrophil-mediated kidney immunopathology and mortality in invasive candidiasis.

    Directory of Open Access Journals (Sweden)

    Michail S Lionakis

    Full Text Available Invasive candidiasis is the 4(th leading cause of nosocomial bloodstream infection in the US with mortality that exceeds 40% despite administration of antifungal therapy; neutropenia is a major risk factor for poor outcome after invasive candidiasis. In a fatal mouse model of invasive candidiasis that mimics human bloodstream-derived invasive candidiasis, the most highly infected organ is the kidney and neutrophils are the major cellular mediators of host defense; however, factors regulating neutrophil recruitment have not been previously defined. Here we show that mice lacking chemokine receptor Ccr1, which is widely expressed on leukocytes, had selectively impaired accumulation of neutrophils in the kidney limited to the late phase of the time course of the model; surprisingly, this was associated with improved renal function and survival without affecting tissue fungal burden. Consistent with this, neutrophils from wild-type mice in blood and kidney switched from Ccr1(lo to Ccr1(high at late time-points post-infection, when Ccr1 ligands were produced at high levels in the kidney and were chemotactic for kidney neutrophils ex vivo. Further, when a 1∶1 mixture of Ccr1(+/+ and Ccr1(-/- donor neutrophils was adoptively transferred intravenously into Candida-infected Ccr1(+/+ recipient mice, neutrophil trafficking into the kidney was significantly skewed toward Ccr1(+/+ cells. Thus, neutrophil Ccr1 amplifies late renal immunopathology and increases mortality in invasive candidiasis by mediating excessive recruitment of neutrophils from the blood to the target organ.

  2. Accelerated apoptosis of neutrophils in familial Mediterranean fever

    DEFF Research Database (Denmark)

    Manukyan, Gayane; Aminov, Rustam; Hakobyan, Gagik

    2015-01-01

    The causative mutations for familial Mediterranean fever (FMF) are located in the MEFV gene, which encodes pyrin. Pyrin modulates the susceptibility to apoptosis via its PYD domain, but how the mutated versions of pyrin affect apoptotic processes are poorly understood. Spontaneous and induced rates...... of systemic neutrophil apoptosis as well as the levels of proteins involved in apoptosis were investigated ex vivo in patients with FMF using flow cytometry and RT-qPCR. The freshly collected neutrophils from the patients in FMF remission displayed a significantly larger number of cells spontaneously entering...... apoptosis compared to control (6.27 ± 2.14 vs. 1.69 ± 0.18%). This elevated ratio was retained after 24 h incubation of neutrophils in the growth medium (32.4 ± 7.41 vs. 7.65 ± 1.32%). Correspondingly, the mRNA level for caspase-3 was also significantly increased under these conditions. In response...

  3. Proinflammatory mediators stimulate neutrophil-directed angiogenesis.

    LENUS (Irish Health Repository)

    McCourt, M

    2012-02-03

    BACKGROUND: Vascular endothelial growth factor (VEGF; vascular permeability factor) is one of the most potent proangiogenic cytokines, and it plays a central role in mediating the process of angiogenesis or new blood vessel formation. Neutrophils (PMNs) recently have been shown to produce VEGF. HYPOTHESIS: The acute inflammatory response is a potent stimulus for PMN-directed angiogenesis. METHODS: Neutrophils were isolated from healthy volunteers and stimulated with lipopolysaccharide (LPS), tumor necrosis factor alpha (TNF-alpha), interleukin 6 (IL-6), and anti-human Fas monoclonal antibody. Culture supernatants were assayed for VEGF using enzyme-linked immunosorbent assays. Culture supernatants from LPS- and TNF-alpha-stimulated PMNs were then added to human umbilical vein endothelial cells and human microvessel endothelial cells and assessed for endothelial cell proliferation using 5-bromodeoxyuridine labeling. Tubule formation was also assessed on MATRIGEL basement membrane matrix. Neutrophils were lysed to measure total VEGF release, and VEGF expression was detected using Western blot analysis. RESULTS: Lipopolysaccharide and TNF-alpha stimulation resulted in significantly increased release of PMN VEGF (532+\\/-49 and 484+\\/-80 pg\\/mL, respectively; for all, presented as mean +\\/- SEM) compared with control experiments (32+\\/-4 pg\\/mL). Interleukin 6 and Fas had no effect. Culture supernatants from LPS- and TNF-alpha-stimulated PMNs also resulted in significant increases (P<.005) in macrovascular and microvascular endothelial cell proliferation and tubule formation. Adding anti-human VEGF-neutralizing polyclonal antibody to stimulated PMN supernatant inhibited these effects. Total VEGF release following cell lysis and Western blot analysis suggests that the VEGF is released from an intracellular store. CONCLUSION: Activated human PMNs are directly angiogenic by releasing VEGF, and this has important implications for inflammation, capillary leak syndrome

  4. Formation of neutrophil extracellular traps under low oxygen level

    Directory of Open Access Journals (Sweden)

    Katja Branitzki-Heinemann

    2016-11-01

    Full Text Available Since their discovery, neutrophil extracellular traps (NETs have been characterized as a fundamental host innate immune defense mechanism. Conversely, excessive NET release may have a variety of detrimental consequences for the host. A fine balance between NET formation and elimination is necessary to sustain a protective effect during an infectious challenge. Our own recently published data revealed that stabilization of hypoxia inducible factor 1α (HIF-1α by the iron chelating HIF-1α-agonist desferoxamine or AKB-4924 enhanced the release of phagocyte extracellular traps. Since HIF-1α is a global regulator of the cellular response to low oxygen, we hypothesized that NET formation may be similarly increased under low oxygen conditions. Hypoxia occurs in tissues during infection or inflammation, mostly due to overconsumption of oxygen by pathogens and recruited immune cells. Therefore, experiments were performed to characterize the formation of NETs under hypoxic oxygen conditions compared to normoxia. Human blood-derived neutrophils were isolated and incubated under normoxic (21% oxygen level and compared to hypoxic (1% conditions. Dissolved oxygen levels were monitored in the primary cell culture using a Fibox4-PSt3 measurement system. The formation of NETs was quantified by fluorescence microscopy in response to the known NET-inducer phorbol 12-myristate 13-acetate (PMA or S. aureus wildtype and a nuclease-deficient mutant. In contrast to our hypothesis, spontaneous NET formation of neutrophils incubated under hypoxia was distinctly reduced compared to control neutrophils incubated under normoxia. Furthermore, neutrophils incubated under hypoxia showed significantly reduced formation of NETs in response to PMA. Gene expression analysis revealed that mRNA level of hif-1α as well as hif-1α target genes was not altered. However, in good correlation to the decreased NET formation under hypoxia, the cholesterol content of the neutrophils was

  5. Iodinated contrast media induce neutrophil apoptosis through a mitochondrial and caspase mediated pathway.

    LENUS (Irish Health Repository)

    Fanning, N F

    2012-02-03

    Iodinated contrast media (ICM) can induce apoptosis (programmed cell death) in renal, myocardial and endothelial cells. Following intravascular injection, circulating immune cells are exposed to high concentrations of ICM. As neutrophils constitutively undergo apoptosis we hypothesized that ICM may adversely affect neutrophil survival. Our aim was to investigate the effect of ICM on neutrophil apoptosis. Neutrophils were isolated from healthy subjects and cultured in vitro with ionic (diatrizoate and ioxaglate) and non-ionic (iohexol and iotrolan) ICM. The effect of ICM on neutrophil apoptosis in both unstimulated and lipopolysaccharide-stimulated neutrophils was determined by annexin V flow cytometry. The influence of physicochemical properties of the different ICM on apoptosis of neutrophils was also studied. We further investigated the effects of ICM on key intracellular signal pathways, including p38 mitogen-activated protein kinase (MAPK) by Western blotting, and mitochondrial depolarization and caspase activity by flow cytometry. Isoiodine concentrations (20 mg ml(-1)) of ionic (diatrizoate 69.6+\\/-2.9%; ioxaglate 58.9+\\/-2.0%) and non-ionic (iohexol 57.3+\\/-2.9%; iotrolan 57.1+\\/-2.6%) ICM significantly induced neutrophil apoptosis over control levels (47.7+\\/-1.4%). The apoptotic effect of ICM was influenced by their chemical structure, with ionic ICM having a more significant (p<0.01) apoptotic effect than non-ionic ICM (p<0.05). Furthermore, ICM reversed the anti-apoptotic effect of lipopolysaccharide (1000 ng ml(-1)) treated neutrophils to control levels (23.0+\\/-3.5% to 61.2+\\/-5.3%; n=4; p<0.05). These agents induce apoptosis through a p38 MAPK independent pathway that results in mitochondrial depolarization, and is dependent on caspase activation. As neutrophils play a central role in host response to infection and injury, ICM, through induction of neutrophil apoptosis, could have a significant deleterious effect on host immune defence and

  6. Imaging neutrophil migration dynamics using micro-optical coherence tomography (Conference Presentation)

    Science.gov (United States)

    Chu, Kengyeh K.; Yonker, Lael; Som, Avira; Pazos, Michael; Kusek, Mark E.; Hurley, Bryan P.; Tearney, Guillermo J.

    2016-03-01

    Neutrophils are immune cells that undergo chemotaxis, detecting and migrating towards a chemical signal gradient. Neutrophils actively migrate across epithelial boundaries, interacting with the epithelium to selectively permit passage without compromising the epithelial barrier. In many inflammatory disorders, excessive neutrophil migration can cause damage to the epithelium itself. The signaling pathways and mechanisms that facilitate trans-epithelial migration are not fully characterized. Our laboratory has developed micro-optical coherence tomography (μOCT), which has 2 μm lateral resolution and 1 μm axial resolution. As a high-resolution native contrast modality, μOCT can directly visualize individual neutrophils as they interact with a cell layer cultured on a transwell filter. A chemoattractant can be applied to the apical side of inverted monolayer, and human neutrophils placed in the basolateral compartment, while μOCT captures 3D images of the chemotaxis. μOCT images can also generate quantitative metrics of migration volume to study the dependence of chemotaxis on monolayer cell type, chemoattractant type, and disease state of the neutrophils. For example, a disease known as leukocyte adhesion deficiency (LAD) can be simulated by treating neutrophils with antibodies that interfere with the CD18 receptor, a facilitator of trans-epithelial migration. We conducted a migration study of anti-CD18 treated and control neutrophils using T84 intestinal epithelium as a barrier. After one hour, μOCT time-lapse imaging indicated a strong difference in the fraction of neutrophils that remain attached to the epithelium after migration (0.67 +/- 0.12 attached anti-CD18 neutrophils, 0.23 +/- 0.08 attached control neutrophils, n = 6, p < 0.05), as well as a modest but non-significant decrease in total migration volume for treated neutrophils. We can now integrate μOCT-derived migration metrics with simultaneously acquired measurements of transepithelial electrical

  7. Cytoplasmic lipid bodies of human neutrophilic leukocytes

    International Nuclear Information System (INIS)

    Weller, P.F.; Ackerman, S.J.; Nicholson-Weller, A.; Dvorak, A.M.

    1989-01-01

    The morphology and function of cytoplasmic lipid bodies in human neutrophils were evaluated. By transmission electron microscopy, neutrophil lipid bodies were cytoplasmic inclusions, usually several microns in diameter, that occasionally coalesced to attain a diameter up to 7 microM. Neutrophil lipid bodies were not enveloped by membrane but were often surrounded by a more electron-dense shell at their periphery. Normal peripheral blood neutrophils contained an average of approximately one lipid body per cell. Lipid bodies appeared in greater numbers in neutrophils from inflammatory lesions. Perturbation of neutrophils during conventional methods of cell isolation and purification modestly increased lipid body numbers in neutrophils, whereas incubation of neutrophils with 1 microM oleic acid rapidly induced lipid body formation over 30 to 60 minutes. After granulocytes were incubated for 2 hours with 3H-fatty acids, including arachidonic, oleic, and palmitic acids, electron microscopic autoradiography demonstrated that lipid bodies represented the predominant intracellular sites of localization of each of the three 3H-fatty acids. There was lesser labeling noted in the perinuclear cisterna, but not in cell membranes. Virtually all of each of the three 3H-fatty acids incorporated by the neutrophils were esterified into chromatographically resolved classes of neutral lipids or phospholipids. These findings indicate that cytoplasmic lipid bodies are more prominent in neutrophils in vivo engaged in inflammatory responses and that these organelles in human neutrophils function as sites of deposition of esterified, incorporated fatty acids

  8. Burn injury reduces neutrophil directional migration speed in microfluidic devices.

    Directory of Open Access Journals (Sweden)

    Kathryn L Butler

    2010-07-01

    Full Text Available Thermal injury triggers a fulminant inflammatory cascade that heralds shock, end-organ failure, and ultimately sepsis and death. Emerging evidence points to a critical role for the innate immune system, and several studies had documented concurrent impairment in neutrophil chemotaxis with these post-burn inflammatory changes. While a few studies suggest that a link between neutrophil motility and patient mortality might exist, so far, cumbersome assays have prohibited exploration of the prognostic and diagnostic significance of chemotaxis after burn injury. To address this need, we developed a microfluidic device that is simple to operate and allows for precise and robust measurements of chemotaxis speed and persistence characteristics at single-cell resolution. Using this assay, we established a reference set of migration speed values for neutrophils from healthy subjects. Comparisons with samples from burn patients revealed impaired directional migration speed starting as early as 24 hours after burn injury, reaching a minimum at 72-120 hours, correlated to the size of the burn injury and potentially serving as an early indicator for concurrent infections. Further characterization of neutrophil chemotaxis using this new assay may have important diagnostic implications not only for burn patients but also for patients afflicted by other diseases that compromise neutrophil functions.

  9. Increased lung neutrophil apoptosis and inflammation resolution in nonresponding pneumonia.

    Science.gov (United States)

    Moret, I; Lorenzo, M J; Sarria, B; Cases, E; Morcillo, E; Perpiñá, M; Molina, J M; Menéndez, R

    2011-11-01

    Neutrophil activation state and its relationship with an inflammatory environment in community-acquired pneumonia (CAP) remain insufficiently elucidated. We aimed to evaluate the neutrophil apoptosis and cytokine pattern in CAP patients after 72 h of treatment, and their impact on infection resolution. Apoptosis of blood and bronchoalveolar lavage (BAL) neutrophils was measured in nonresponding CAP (NCAP), in responding CAP (blood only) and in patients without infection (control). Pro-inflammatory (interleukin (IL)-6, IL-8) and anti-inflammatory (IL-10) cytokines were measured. Main outcomes were clinical stability and days of hospitalisation. Basal neutrophil apoptosis was higher in the BAL and blood of NCAP, whereas spontaneous apoptosis (after 24 h culture) was lower. Cytokines in NCAP were higher than in responding CAP and control: IL-6 was increased in BAL and blood, IL-8 in BAL and IL-10 in blood. An increased basal apoptosis (≥20%) in BAL of NCAP was associated with lower systemic IL-10 (p<0.01), earlier clinical stability (p=0.05) and shorter hospital stay (p=0.02). A significant correlation was found for systemic IL-6 and IL-10 with days to reach stability and length of stay. After 72 h of treatment, an increased basal alveolar neutrophil apoptosis might contribute to downregulation of inflammation and to faster clinical stability.

  10. Spontaneous neutrophil activation in HTLV-1 infected patients

    Directory of Open Access Journals (Sweden)

    Jaqueline B. Guerreiro

    Full Text Available Human T cell lymphotropic Virus type-1 (HTLV-1 induces lymphocyte activation and proliferation, but little is known about the innate immune response due to HTLV-1 infection. We evaluated the percentage of neutrophils that metabolize Nitroblue tetrazolium (NBT to formazan in HTLV-1 infected subjects and the association between neutrophil activation and IFN-gamma and TNF-alpha levels. Blood was collected from 35 HTLV-1 carriers, from 8 patients with HAM/TSP (HTLV-1- associated myelopathy; 22 healthy individuals were evaluated for spontaneous and lipopolysaccharide (LPS-stimulated neutrophil activity (reduction of NBT to formazan. The production of IFN-gamma and TNF-alpha by unstimulated mononuclear cells was determined by ELISA. Spontaneous NBT levels, as well as spontaneous IFN-gamma and TNF-alpha production, were significantly higher (p<0.001 in HTLV-1 infected subjects than in healthy individuals. A trend towards a positive correlation was noted, with increasing percentage of NBT positive neutrophils and levels of IFN-gamma. The high IFN-gamma producing HTLV-1 patient group had significantly greater NBT than healthy controls, 43±24% and 17±4.8% respectively (p< 0.001, while no significant difference was observed between healthy controls and the low IFN-gamma-producing HTLV-1 patient group (30±20%. Spontaneous neutrophil activation is another marker of immune perturbation resulting from HTLV-1 infection. In vivo activation of neutrophils observed in HTLV-1 infected subjects is likely to be the same process that causes spontaneous IFN-gamma production, or it may partially result from direct IFN-gamma stimulation.

  11. Enhanced neutrophil chemotactic activity after bronchial challenge in subjects with grain dust-induced asthma.

    Science.gov (United States)

    Park, H S; Jung, K S

    1998-03-01

    There have been few reports suggesting involvement of neutrophils in induction of bronchoconstriction after inhalation of grain dust. To understand the role of neutrophils in pathogenesis of grain dust-induced asthma. We observed serum neutrophil chemotactic activity during grain dust-bronchoprovocation tests in six asthmatic subjects with positive bronchial challenges (group I). They were compared with those of six symptomatic subjects from the same workplace with negative bronchial challenges (group II). After grain dust inhalation, serum neutrophil chemotactic activity significantly increased at 30 minutes (P = .028), and then decreased to baseline level at 240 minutes (P = .028) in five subjects of group I having isolated early asthmatic responses. Enhanced neutrophil chemotactic activity was persistent for up to 240 minutes in one asthmatic subject having both early and late asthmatic responses. There was, however, no significant change in serum neutrophil chemotactic activity during bronchial challenges in subjects of group II. Pre-incubation of sera with anti-interleukin-8 (IL-8) antibody did not affect the neutrophil chemotactic activity results of group I subjects. These results suggest that enhanced neutrophil chemotactic activity distinct from IL-8 may contribute to significant bronchoconstriction induced by grain dust.

  12. Neutrophils are not less sensitive than other blood leukocytes to the genomic effects of glucocorticoids.

    Directory of Open Access Journals (Sweden)

    Gaelle Hirsch

    Full Text Available Neutrophils are generally considered less responsive to glucocorticoids compared to other inflammatory cells. The reported increase in human neutrophil survival mediated by these drugs partly supports this assertion. However, it was recently shown that dexamethasone exerts potent anti-inflammatory effects in equine peripheral blood neutrophils. Few comparative studies of glucocorticoid effects in neutrophils and other leukocytes have been reported and a relative insensitivity of neutrophils to these drugs could not be ruled out.We assessed glucocorticoid-responsiveness in equine and human peripheral blood neutrophils and neutrophil-depleted leukocytes.Blood neutrophils and neutrophil-depleted leukocytes were isolated from 6 healthy horses and 4 human healthy subjects. Cells were incubated for 5 h with or without LPS (100 ng/mL alone or combined with hydrocortisone, prednisolone or dexamethasone (10(-8 M and 10(-6 M. IL-1β, TNF-α, IL-8, glutamine synthetase and GR-α mRNA expression was quantified by qPCR. Equine neutrophils were also incubated for 20 h with or without the three glucocorticoids and cell survival was assessed by flow cytometry and light microscopy on cytospin preparations.We found that glucocorticoids down-regulated LPS-induced pro-inflammatory mRNA expression in both cell populations and species. These drugs also significantly increased glutamine synthetase gene expression in both equine cell populations. The magnitude of glucocorticoid response between cell populations was generally similar in both species. We also showed that dexamethasone had a comparable inhibitory effect on pro-inflammatory gene expression in both human and equine neutrophils. As reported in other species, glucocorticoids significantly increase the survival in equine neutrophils.Glucocorticoids exert genomic effects of similar magnitude on neutrophils and on other blood leukocytes. We speculate that the poor response to glucocorticoids observed in some

  13. Neutrophils Are Not Less Sensitive Than Other Blood Leukocytes to the Genomic Effects of Glucocorticoids

    Science.gov (United States)

    Hirsch, Gaelle; Lavoie-Lamoureux, Anouk; Beauchamp, Guy; Lavoie, Jean-Pierre

    2012-01-01

    Background Neutrophils are generally considered less responsive to glucocorticoids compared to other inflammatory cells. The reported increase in human neutrophil survival mediated by these drugs partly supports this assertion. However, it was recently shown that dexamethasone exerts potent anti-inflammatory effects in equine peripheral blood neutrophils. Few comparative studies of glucocorticoid effects in neutrophils and other leukocytes have been reported and a relative insensitivity of neutrophils to these drugs could not be ruled out. Objective We assessed glucocorticoid-responsiveness in equine and human peripheral blood neutrophils and neutrophil-depleted leukocytes. Methods Blood neutrophils and neutrophil-depleted leukocytes were isolated from 6 healthy horses and 4 human healthy subjects. Cells were incubated for 5 h with or without LPS (100 ng/mL) alone or combined with hydrocortisone, prednisolone or dexamethasone (10−8 M and 10−6 M). IL-1β, TNF-α, IL-8, glutamine synthetase and GR-α mRNA expression was quantified by qPCR. Equine neutrophils were also incubated for 20 h with or without the three glucocorticoids and cell survival was assessed by flow cytometry and light microscopy on cytospin preparations. Results We found that glucocorticoids down-regulated LPS-induced pro-inflammatory mRNA expression in both cell populations and species. These drugs also significantly increased glutamine synthetase gene expression in both equine cell populations. The magnitude of glucocorticoid response between cell populations was generally similar in both species. We also showed that dexamethasone had a comparable inhibitory effect on pro-inflammatory gene expression in both human and equine neutrophils. As reported in other species, glucocorticoids significantly increase the survival in equine neutrophils. Conclusions Glucocorticoids exert genomic effects of similar magnitude on neutrophils and on other blood leukocytes. We speculate that the poor response to

  14. Biomaterial associated impairment of local neutrophil function.

    Science.gov (United States)

    Kaplan, S S; Basford, R E; Kormos, R L; Hardesty, R L; Simmons, R L; Mora, E M; Cardona, M; Griffith, B L

    1990-01-01

    The effect of biomaterials on neutrophil function was studied in vitro to determine if these materials activated neutrophils and to determine the subsequent response of these neutrophils to further stimulation. Two biomaterials--polyurethane, a commonly used substance, and Velcro pile (used in the Jarvik 7 heart)--were evaluated. Two control substances, polyethylene and serum-coated polystyrene, were used for comparison. Neutrophil superoxide release was measured following incubation with these materials for 10, 30, and 120 min in the absence of additional stimulation and after stimulation with formylmethionylleucylphenylalanine (fMLP) or phorbol myristate acetate (PMA). The authors observed that the incubation of neutrophils on both polyurethane and Velcro resulted in substantially increased superoxide release that was greater after the 10 min than after the 30 or 120 min association. These activated neutrophils exhibited a poor additional response to fMLP but responded well to PMA. The effect of implantation of the Novacor left ventricular assist device on peripheral blood neutrophil function was also evaluated. The peripheral blood neutrophils exhibited normal superoxide release and chemotaxis. These studies suggest that biomaterials may have a profound local effect on neutrophils, which may predispose the patient to periprosthetic infection, but that the reactivity of circulating neutrophils is unimpaired.

  15. Tamoxifen induces apoptotic neutrophil efferocytosis in horses.

    Science.gov (United States)

    Olave, C; Morales, N; Uberti, B; Henriquez, C; Sarmiento, J; Ortloff, A; Folch, H; Moran, G

    2018-03-01

    Macrophages and neutrophils are important cellular components in the process of acute inflammation and its subsequent resolution, and evidence increasingly suggests that they play important functions during the resolution of chronic, adaptive inflammatory processes. Exacerbated neutrophil activity can be harmful to surrounding tissues; this is important in a range of diseases, including allergic asthma and chronic obstructive pulmonary disease in humans, and equine asthma (also known as recurrent airway obstruction (RAO). Tamoxifen (TX) is a non-steroidal estrogen receptor modulator with effects on cell growth and survival. Previous studies showed that TX treatment in horses with induced acute pulmonary inflammation promoted early apoptosis of blood and BALF neutrophils, reduction of BALF neutrophils, and improvement in animals' clinical status. The aim of this study was to describe if TX induces in vitro efferocytosis of neutrophils by alveolar macrophages. Efferocytosis assay, myeloperoxidase (MPO) detection and translocation phosphatidylserine (PS) were performed on neutrophils isolated from peripheral blood samples from five healthy horses. In in vitro samples from heathy horses, TX treatment increases the phenomenon of efferocytosis of peripheral neutrophils by alveolar macrophages. Similar increases in supernatant MPO concentration and PS translocation were observed in TX-treated neutrophils, compared to control cells. In conclusion, these results confirm that tamoxifen has a direct effect on equine peripheral blood neutrophils, through stimulation of the engulfment of apoptotic neutrophils by alveolar macrophages.

  16. Neutrophils Compromise Retinal Pigment Epithelial Barrier Integrity

    Directory of Open Access Journals (Sweden)

    Jiehao Zhou

    2010-01-01

    Full Text Available We hypothesized that neutrophils and their secreted factors mediate breakdown of the integrity of the outer blood-retina-barrier by degrading the apical tight junctions of the retinal pigment epithelium (RPE. The effect of activated neutrophils or neutrophil cell lysate on apparent permeability of bovine RPE-Choroid explants was evaluated by measuring [H] mannitol flux in a modified Ussing chamber. The expression of matrix metalloproteinase- (MMP- 9 in murine peritoneal neutrophils, and the effects of neutrophils on RPE tight-junction protein expression were assessed by confocal microscopy and western blot. Our results revealed that basolateral incubation of explants with neutrophils decreased occludin and ZO-1 expression at 1 and 3 hours and increased the permeability of bovine RPE-Choroid explants by >3-fold (P<.05. Similarly, basolateral incubation of explants with neutrophil lysate decreased ZO-1 expression at 1 and 3 hours (P<.05 and increased permeability of explants by 75%. Further, we found that neutrophils prominently express MMP-9 and that incubation of explants with neutrophils in the presence of anti-MMP-9 antibody inhibited the increase in permeability. These data suggest that neutrophil-derived MMP-9 may play an important role in disrupting the integrity of the outer blood-retina barrier.

  17. Mixed species biofilms of Fusobacterium necrophorum and Porphyromonas levii impair the oxidative response of bovine neutrophils in vitro.

    Science.gov (United States)

    Lockhart, Joey S; Buret, Andre G; Ceri, Howard; Storey, Douglas G; Anderson, Stefanie J; Morck, Douglas W

    2017-10-01

    Biofilms composed of anaerobic bacteria can result in persistent infections and chronic inflammation. Host immune cells have difficulties clearing biofilm-related infections and this can result in tissue damage. Neutrophils are a vital component of the innate immune system and help clear biofilms. The comparative neutrophilic response to biofilms versus planktonic bacteria remains incompletely understood, particularly in the context of mixed infections. The objective of this study was to generate mixed species anaerobic bacterial biofilms composed of two opportunistic pathogens, Fusobacterium necrophorum and Porphyromonas levii, and evaluate neutrophil responses to extracellular fractions from both biofilms and planktonic cell co-cultures of the same bacteria. Purified bovine neutrophils exposed to culture supernatants from mixed species planktonic bacteria showed elevated oxidative activity compared to neutrophils exposed to biofilms composed of the same bacteria. Bacterial lipopolysaccharide plays a significant role in the stimulation of neutrophils; biofilms produced substantially more lipopolysaccharide than planktonic bacteria under these experimental conditions. Removal of lipopolysaccharide significantly reduced neutrophil oxidative response to culture supernatants of planktonic bacteria. Oxidative responses to LPS-removed biofilm supernatants and LPS-removed planktonic cell supernatants were similar. The limited neutrophil response to biofilm bacteria observed in this study supports the reduced ability of the innate immune system to eradicate biofilm-associated infections. Lipopolysaccharide is likely important in neutrophil response; however, the presence of other extracellular, immune modifying molecules in the bacterial media also appears to be important in altering neutrophil function. Copyright © 2017 Elsevier Ltd. All rights reserved.

  18. (neutrophil) Activity, Chronic Gastritis, Gastric Atrophy And Intestinal ...

    African Journals Online (AJOL)

    Incidental (early gastric) cancer was found in 3%, dysplasia in 2% and reactive gastropathy in 7% of the cases. A statistically significant relationship was found between Helicobacter pylori colonization intensity and the degrees of neutrophil activity, chronic inflammation and intestinal metaplasia. Conclusion: We concluded ...

  19. Neutrophil Responses to Sterile Implant Materials.

    Directory of Open Access Journals (Sweden)

    Siddharth Jhunjhunwala

    Full Text Available In vivo implantation of sterile materials and devices results in a foreign body immune response leading to fibrosis of implanted material. Neutrophils, one of the first immune cells to be recruited to implantation sites, have been suggested to contribute to the establishment of the inflammatory microenvironment that initiates the fibrotic response. However, the precise numbers and roles of neutrophils in response to implanted devices remains unclear. Using a mouse model of peritoneal microcapsule implantation, we show 30-500 fold increased neutrophil presence in the peritoneal exudates in response to implants. We demonstrate that these neutrophils secrete increased amounts of a variety of inflammatory cytokines and chemokines. Further, we observe that they participate in the foreign body response through the formation of neutrophil extracellular traps (NETs on implant surfaces. Our results provide new insight into neutrophil function during a foreign body response to peritoneal implants which has implications for the development of biologically compatible medical devices.

  20. Neutrophil Reverse Migration Becomes Transparent with Zebrafish

    Directory of Open Access Journals (Sweden)

    Taylor W. Starnes

    2012-01-01

    Full Text Available The precise control of neutrophil-mediated inflammation is critical for both host defense and the prevention of immunopathology. In vivo imaging studies in zebrafish, and more recently in mice, have made the novel observation that neutrophils leave a site of inflammation through a process called neutrophil reverse migration. The application of advanced imaging techniques to the genetically tractable, optically transparent zebrafish larvae was critical for these advances. Still, the mechanisms underlying neutrophil reverse migration and its effects on the resolution or priming of immune responses remain unclear. Here, we review the current knowledge of neutrophil reverse migration, its potential roles in host immunity, and the live imaging tools that make zebrafish a valuable model for increasing our knowledge of neutrophil behavior in vivo.

  1. Neutrophils in Tuberculosis: Heterogeneity Shapes the Way?

    Science.gov (United States)

    2017-01-01

    Infection with M. tuberculosis remains one of the most common infections in the world. The outcome of the infection depends on host ability to mount effective protection and balance inflammatory responses. Neutrophils are innate immune cells implicated in both processes. Accordingly, during M. tuberculosis infection, they play a dual role. Particularly, they contribute to the generation of effector T cells, participate in the formation of granuloma, and are directly involved in tissue necrosis, destruction, and infection dissemination. Neutrophils have a high bactericidal potential. However, data on their ability to eliminate M. tuberculosis are controversial, and the results of neutrophil depletion experiments are not uniform. Thus, the overall roles of neutrophils during M. tuberculosis infection and factors that determine these roles are not fully understood. This review analyzes data on neutrophil defensive and pathological functions during tuberculosis and considers hypotheses explaining the dualism of neutrophils during M. tuberculosis infection and tuberculosis disease. PMID:28626346

  2. Heterogeneity of neutrophil antibodies in patients with primary Sjögren's syndrome.

    Science.gov (United States)

    Lamour, A; Le Corre, R; Pennec, Y L; Cartron, J; Youinou, P

    1995-11-01

    Our aims were to determine the prevalence of neutrophil antibodies in patients with primary Sjögren's syndrome (pSS), identify their target antigen(s), and evaluate their functional significance. Neutrophil antibodies were detected using an indirect immunofluorescence (IIIF) test and an enzyme-linked immunosorbent assay (ELISA), using recombinant human Fc-gamma receptor (Fc gamma RIIIb) as a capture agent. Luminol-dependent chemiluminescence was then measured by an established technique. Antibodies to neutrophils were detected in 30 of 66 patients (45%) and categorized on the basis of positivity for the two assays: IIF+/ELISA+ (group A: five patients), IIF+/ELISA- (group B: five patients), and IFF-/ELISA+ (group C: 20 patients). All positive sera contained antibodies directed to the neutrophil specific Fc gamma RIIIb, and none of them bound to NAnull neutrophils. The titer of neutrophil-reactive antibodies (groups A and B) showed no correlation with the neutrophil count, but these autoantibodies did reduce the cell ability to generate a respiratory burst. Thus, neutrophil antibodies are common in patients with pSS. Their main target appears to be Fc gamma RIII, and this may partly account for the dysfunction in Fc gamma R-mediated clearance by the reticuloendothelial system reported in these patients.

  3. A Neutrophil Phenotype Model for Extracorporeal Treatment of Sepsis.

    Directory of Open Access Journals (Sweden)

    Alexander D Malkin

    2015-10-01

    Full Text Available Neutrophils play a central role in eliminating bacterial pathogens, but may also contribute to end-organ damage in sepsis. Interleukin-8 (IL-8, a key modulator of neutrophil function, signals through neutrophil specific surface receptors CXCR-1 and CXCR-2. In this study a mechanistic computational model was used to evaluate and deploy an extracorporeal sepsis treatment which modulates CXCR-1/2 levels. First, a simplified mechanistic computational model of IL-8 mediated activation of CXCR-1/2 receptors was developed, containing 16 ODEs and 43 parameters. Receptor level dynamics and systemic parameters were coupled with multiple neutrophil phenotypes to generate dynamic populations of activated neutrophils which reduce pathogen load, and/or primed neutrophils which cause adverse tissue damage when misdirected. The mathematical model was calibrated using experimental data from baboons administered a two-hour infusion of E coli and followed for a maximum of 28 days. Ensembles of parameters were generated using a Bayesian parallel tempering approach to produce model fits that could recreate experimental outcomes. Stepwise logistic regression identified seven model parameters as key determinants of mortality. Sensitivity analysis showed that parameters controlling the level of killer cell neutrophils affected the overall systemic damage of individuals. To evaluate rescue strategies and provide probabilistic predictions of their impact on mortality, time of onset, duration, and capture efficacy of an extracorporeal device that modulated neutrophil phenotype were explored. Our findings suggest that interventions aiming to modulate phenotypic composition are time sensitive. When introduced between 3-6 hours of infection for a 72 hour duration, the survivor population increased from 31% to 40-80%. Treatment efficacy quickly diminishes if not introduced within 15 hours of infection. Significant harm is possible with treatment durations ranging from 5

  4. Impaired neutrophil function in intestinal lymphangiectasia.

    OpenAIRE

    Bolton, R P; Cotter, K L; Losowsky, M S

    1986-01-01

    Impaired neutrophil chemotaxis and phagocytosis were shown in three patients with intestinal lymphangiectasia. Abnormalities in cell associated and serum derived activity occurred, and possible mechanisms are suggested.

  5. Complement Activation Induces Neutrophil Adhesion and Neutrophil-Platelet Aggregate Formation on Vascular Endothelial Cells

    Directory of Open Access Journals (Sweden)

    Magdalena Riedl

    2017-01-01

    Discussion: Therefore, our findings of (i neutrophils adhering to complement-activated endothelial cells, (ii the formation of neutrophil-platelet aggregates on endothelial cells, and (iii the ability of aHUS serum to induce similar effects identify a possible role for neutrophils in aHUS manifestation.

  6. Semi-Automatic Rating Method for Neutrophil Alkaline Phosphatase Activity.

    Science.gov (United States)

    Sugano, Kanae; Hashi, Kotomi; Goto, Misaki; Nishi, Kiyotaka; Maeda, Rie; Kono, Keigo; Yamamoto, Mai; Okada, Kazunori; Kaga, Sanae; Miwa, Keiko; Mikami, Taisei; Masauzi, Nobuo

    2017-01-01

    The neutrophil alkaline phosphatase (NAP) score is a valuable test for the diagnosis of myeloproliferative neoplasms, but it has still manually rated. Therefore, we developed a semi-automatic rating method using Photoshop ® and Image-J, called NAP-PS-IJ. Neutrophil alkaline phosphatase staining was conducted with Tomonaga's method to films of peripheral blood taken from three healthy volunteers. At least 30 neutrophils with NAP scores from 0 to 5+ were observed and taken their images. From which the outer part of neutrophil was removed away with Image-J. These were binarized with two different procedures (P1 and P2) using Photoshop ® . NAP-positive area (NAP-PA) and granule (NAP-PGC) were measured and counted with Image-J. The NAP-PA in images binarized with P1 significantly (P < 0.05) differed between images with NAP scores from 0 to 3+ (group 1) and those from 4+ to 5+ (group 2). The original images in group 1 were binarized with P2. NAP-PGC of them significantly (P < 0.05) differed among all four NAP score groups. The mean NAP-PGC with NAP-PS-IJ indicated a good correlation (r = 0.92, P < 0.001) to results by human examiners. The sensitivity and specificity of NAP-PS-IJ were 60% and 92%, which might be considered as a prototypic method for the full-automatic rating NAP score. © 2016 Wiley Periodicals, Inc.

  7. Alpha-1-antitrypsin is produced by human neutrophil granulocytes and their precursors and liberated during granule exocytosis

    DEFF Research Database (Denmark)

    Clemmensen, Stine N; Jacobsen, Lars C; Rørvig, Sara

    2011-01-01

    Alpha-1-antitrypsin (A1AT) is an important inhibitor of neutrophil proteases including elastase, cathepsin G, and proteinase 3. Transcription profiling data suggest that A1AT is expressed by human neutrophil granulocytes during all developmental stages. A1AT has hitherto only been found associate...... significantly to the antiprotease levels in tissues during inflammation. Impaired binding of neutrophil A1AT to serine proteases in patients with (PI)ZZ mutations may enhance their susceptibility to the development of emphysema....

  8. Neutrophil chemotactic activity in bronchoalveolar lavage fluid of patients with AIDS-associated Pneumocystis carinii pneumonia

    DEFF Research Database (Denmark)

    Benfield, T L; Kharazmi, A; Larsen, C G

    1997-01-01

    been shown to confer a poor prognosis in PCP. We therefore investigated the potential of BAL fluid from 17 patients with PCP to induce neutrophil chemotaxis. BAL fluid from patients induced considerable neutrophil chemotactic activity compared to normal controls. Elevated levels of IL-8 were detected...... in patient samples as compared to controls. A specific anti-IL-8 antibody significantly reduced chemotactic activity of patient samples by more than 50%. In conclusion, IL-8 appears to be a significant participant of neutrophil chemotaxis in AIDS-associated PCP, and may participate in the recruitment...

  9. Obesity is associated with more activated neutrophils in African American male youth.

    Science.gov (United States)

    Xu, X; Su, S; Wang, X; Barnes, V; De Miguel, C; Ownby, D; Pollock, J; Snieder, H; Chen, W; Wang, X

    2015-01-01

    There is emerging evidence suggesting the role of peripheral blood leukocytes in the pathogenesis of obesity and related diseases. However, few studies have taken a genome-wide approach to investigating gene expression profiles in peripheral leukocytes between obese and lean individuals with the consideration of obesity-related shifts in leukocyte types. We conducted this study in 95 African Americans (AAs) of both genders (age 14-20 years, 46 lean and 49 obese). Complete blood count with differential test (CBC) was performed in whole blood. Genome-wide gene expression analysis was obtained using the Illumina HumanHT-12 V4 Beadchip with RNA extracted from peripheral leukocytes. Out of the 95 participants, 64 had neutrophils stored. The validation study was based on real-time PCR with RNA extracted from purified neutrophils. CBC test suggested that, in males, obesity was associated with increased neutrophil percentage (P=0.03). Genome-wide gene expression analysis showed that, in males, the majority of the most differentially expressed genes were related to neutrophil activation. Validation of the gene expression levels of ELANE (neutrophil elastase) and MPO (myeloperoxidase) in purified neutrophils demonstrated that the expression of these two genes--important biomarkers of neutrophils activation--were significantly elevated in obese males (P=0.01 and P=0.02, respectively). The identification of increased neutrophil percentage and activation in obese AA males suggests that neutrophils have an essential role in the pathogenesis of obesity-related disease. Further functional and mechanistic studies on neutrophils may contribute to the development of novel intervention strategies reducing the burden associated with obesity-related health problems.

  10. Different innate neutrophil responses in controlled and uncontrolled asthma

    NARCIS (Netherlands)

    Tang, Francesca; Foxley, Gloria; Gibson, Peter; Burgess, Janette; Baines, Katherine; Oliver, Brian

    2015-01-01

    Introduction: Respiratory viruses are a major cause of asthma exacerbations. Neutrophilic inflammation occurs during infections and is associated with difficult to treat asthma. The role of neutrophils in viral infections and whether neutrophil dysfunction contributes to exacerbation pathogenesis

  11. General versus regional anaesthesia for cataract surgery: effects on neutrophil apoptosis and the postoperative pro-inflammatory state.

    LENUS (Irish Health Repository)

    Goto, Y

    2012-02-03

    At clinically relevant concentrations, volatile anaesthetic agents influence neutrophil function. Our hypothesis was that sevoflurane would inhibit neutrophil apoptosis and consequently influence the postoperative pro-inflammatory state. In order to identify selectively the effect of the anaesthetic agent sevoflurane, we studied patients undergoing minimally stimulating (cataract) surgery randomly allocated to receive either sevoflurane (n = 11) or local anaesthesia (n = 12). Venous blood samples were taken immediately prior to anaesthesia and at 1, 8 and 24 h thereafter. The rate of neutrophil apoptosis, plasma concentration of cytokines and differential white cell count were measured. The rates of neutrophil apoptosis and plasma concentrations of IL-1beta, TNF-alpha and IL-8 at each time point were similar in the two groups. IL-6 concentrations increased significantly and to a similar extent compared to preanaesthetic levels at 8 and 24 h. This study demonstrates that sevoflurane does not influence the rate of neutrophil apoptosis, cytokine concentrations and neutrophil count following cataract surgery.

  12. Oral neutrophil responses to acute prolonged exercise may not be representative of blood neutrophil responses.

    Science.gov (United States)

    Davison, Glen; Jones, Arwel Wyn

    2015-03-01

    Neutrophil numbers and function (oxidative burst) were assessed in peripheral blood and oral samples before and after prolonged exercise. Blood neutrophil count increased (∼3.5-fold, P < 0.001) and function decreased (30% ± 19% decrease, P = 0.005) postexercise. Oral neutrophil count (P = 0.392) and function (P = 0.334) were unchanged. Agreement between oral and blood neutrophil function responses to exercise was poor. These findings highlight the importance of studying neutrophils within various compartments/sample types.

  13. Neutrophil migration under normal and sepsis conditions.

    Science.gov (United States)

    Lerman, Yelena V; Kim, Minsoo

    2015-01-01

    Neutrophil migration is critical for pathogen clearance and host survival during severe sepsis. Interaction of neutrophil adhesion receptors with ligands on endothelial cells results in firm adhesion of the circulating neutrophils, followed by neutrophil activation and directed migration to sites of infection through the basement membrane and interstitial extracellular matrix. Proteolytic enzymes and reactive oxygen species are produced and released by neutrophils in response to a variety of inflammatory stimuli. Although these mediators are important for host defense, they also promote tissue damage. Excessive neutrophil migration during the early stages of sepsis may lead to an exaggerated inflammatory response with associated tissue damage and subsequent organ dysfunction. On the other hand, dysregulation of migration and insufficient migratory response that occurs during the latter stages of severe sepsis contributes to neutrophils' inability to contain and control infection and impaired wound healing. This review discusses the major steps and associated molecules involved in the balance of neutrophil trafficking, the precise regulation of which during sepsis spells life or death for the host.

  14. Neutrophil heterogeneity: implications for homeostasis and pathogenesis

    NARCIS (Netherlands)

    Silvestre-Roig, Carlos; Hidalgo, Andres; Soehnlein, Oliver

    2016-01-01

    Neutrophils are polymorphonuclear leukocytes of the phagocytic system that act as first line of host defense against invading pathogens but are also important mediators of inflammation-induced injury. In contrast to other members of the innate immune system, neutrophils are classically considered a

  15. Candida albicans escapes from mouse neutrophils

    DEFF Research Database (Denmark)

    Ermert, David; Niemiec, Maria J; Röhm, Marc

    2013-01-01

    is the most widely used model organism. Neutrophils are essential immune cells to prevent opportunistic mycoses. To explore potential differences between the rodent infection model and the human host, we compared the interactions of C. albicans with neutrophil granulocytes from mice and humans. We revealed...

  16. Neutrophils: potential therapeutic targets in tularemia?

    Directory of Open Access Journals (Sweden)

    Lee-Ann H Allen

    2013-12-01

    Full Text Available The central role of neutrophils in innate immunity and host defense has long been recognized, and the ability of these cells to efficiently engulf and kill invading bacteria has been extensively studied, as has the role of neutrophil apoptosis in resolution of the inflammatory response. In the past few years additional immunoregulatory properties of neutrophils were discovered, and it is now clear that these cells play a much greater role in control of the immune response than was previously appreciated. In this regard, it is noteworthy that Francisella tularensis is one of relatively few pathogens that can successfully parasitize neutrophils as well as macrophages, DC and epithelial cells. Herein we will review the mechanisms used by F. tularensis to evade elimination by neutrophils. We will also reprise effects of this pathogen on neutrophil migration and lifespan as compared with other infectious and inflammatory disease states. In addition, we will discuss the evidence which suggests that neutrophils contribute to disease progression rather than effective defense during tularemia, and consider whether manipulation of neutrophil migration or turnover may be suitable adjunctive therapeutic strategies.

  17. Local anesthetic-induced inhibition of human neutrophil priming: the influence of structure, lipophilicity, and charge

    NARCIS (Netherlands)

    Picardi, Susanne; Cartellieri, Sibylle; Groves, Danja; Hahnenekamp, Klaus; Gerner, Peter; Durieux, Marcel E.; Stevens, Markus F.; Lirk, Philipp; Hollmann, Markus W.

    2013-01-01

    Local anesthetics (LAs) are widely known for inhibition of voltage-gated sodium channels underlying their antiarrhythmic and antinociceptive effects. However, LAs have significant immunomodulatory properties and were shown to affect human neutrophil functions independent of sodium-channel blockade.

  18. Activated Protein C Attenuates Severe Inflammation by Targeting VLA-3high Neutrophil Subpopulation in Mice.

    Science.gov (United States)

    Sarangi, Pranita P; Lee, Hyun-Wook; Lerman, Yelena V; Trzeciak, Alissa; Harrower, Eric J; Rezaie, Alireza R; Kim, Minsoo

    2017-10-15

    The host injury involved in multiorgan system failure during severe inflammation is mediated, in part, by massive infiltration and sequestration of hyperactive neutrophils in the visceral organ. A recombinant form of human activated protein C (rhAPC) has shown cytoprotective and anti-inflammatory functions in some clinical and animal studies, but the direct mechanism is not fully understood. Recently, we reported that, during endotoxemia and severe polymicrobial peritonitis, integrin VLA-3 (CD49c/CD29) is specifically upregulated on hyperinflammatory neutrophils and that targeting the VLA-3 high neutrophil subpopulation improved survival in mice. In this article, we report that rhAPC binds to human neutrophils via integrin VLA-3 (CD49c/CD29) with a higher affinity compared with other Arg-Gly-Asp binding integrins. Similarly, there is preferential binding of activated protein C (PC) to Gr1 high CD11b high VLA-3 high cells isolated from the bone marrow of septic mice. Furthermore, specific binding of rhAPC to human neutrophils via VLA-3 was inhibited by an antagonistic peptide (LXY2). In addition, genetically modified mutant activated PC, with a high affinity for VLA-3, shows significantly improved binding to neutrophils compared with wild-type activated PC and significantly reduced neutrophil infiltration into the lungs of septic mice. These data indicate that variants of activated PC have a stronger affinity for integrin VLA-3, which reveals novel therapeutic possibilities. Copyright © 2017 by The American Association of Immunologists, Inc.

  19. Contribution of neutrophils to acute lung injury.

    Science.gov (United States)

    Grommes, Jochen; Soehnlein, Oliver

    2011-01-01

    Treatment of acute lung injury (ALI) and its most severe form, acute respiratory distress syndrome (ARDS), remain unsolved problems of intensive care medicine. ALI/ARDS are characterized by lung edema due to increased permeability of the alveolar-capillary barrier and subsequent impairment of arterial oxygenation. Lung edema, endothelial and epithelial injury are accompanied by an influx of neutrophils into the interstitium and broncheoalveolar space. Hence, activation and recruitment of neutrophils are regarded to play a key role in progression of ALI/ARDS. Neutrophils are the first cells to be recruited to the site of inflammation and have a potent antimicrobial armour that includes oxidants, proteinases and cationic peptides. Under pathological circumstances, however, unregulated release of these microbicidal compounds into the extracellular space paradoxically can damage host tissues. This review focuses on the mechanisms of neutrophil recruitment into the lung and on the contribution of neutrophils to tissue damage in ALI.

  20. The complex interplay between neutrophils and cancer.

    Science.gov (United States)

    Rakic, Andrea; Beaudry, Paul; Mahoney, Douglas J

    2018-03-01

    Neutrophils are the most abundant type of white blood cell, and are an essential component of the innate immune system. They characteristically arrive rapidly at sites of infection and injury, and release a variety of cytokines and toxic molecules to eliminate pathogens and elicit an acute inflammatory response. Research into the function of neutrophils in cancer suggest they have divergent roles. Indeed, while most studies have found neutrophils to be associated with cancer progression, others have also documented anticancer effects. In this review, we describe the investigations into neutrophil populations that have been implicated in promoting tumor growth and metastasis as well those demonstrating antitumor functions. The collective research suggests a complex role for neutrophils in cancer biology, which raises the prospect of their targeting for the treatment of cancer.

  1. Dynamic interactions of neutrophils and biofilms

    Directory of Open Access Journals (Sweden)

    Josefine Hirschfeld

    2014-12-01

    Full Text Available Background: The majority of microbial infections in humans are biofilm-associated and difficult to treat, as biofilms are highly resistant to antimicrobial agents and protect themselves from external threats in various ways. Biofilms are tenaciously attached to surfaces and impede the ability of host defense molecules and cells to penetrate them. On the other hand, some biofilms are beneficial for the host and contain protective microorganisms. Microbes in biofilms express pathogen-associated molecular patterns and epitopes that can be recognized by innate immune cells and opsonins, leading to activation of neutrophils and other leukocytes. Neutrophils are part of the first line of defense and have multiple antimicrobial strategies allowing them to attack pathogenic biofilms. Objective/design: In this paper, interaction modes of neutrophils with biofilms are reviewed. Antimicrobial strategies of neutrophils and the counteractions of the biofilm communities, with special attention to oral biofilms, are presented. Moreover, possible adverse effects of neutrophil activity and their biofilm-promoting side effects are discussed. Results/conclusion: Biofilms are partially, but not entirely, protected against neutrophil assault, which include the processes of phagocytosis, degranulation, and formation of neutrophil extracellular traps. However, virulence factors of microorganisms, microbial composition, and properties of the extracellular matrix determine whether a biofilm and subsequent microbial spread can be controlled by neutrophils and other host defense factors. Besides, neutrophils may inadvertently contribute to the physical and ecological stability of biofilms by promoting selection of more resistant strains. Moreover, neutrophil enzymes can degrade collagen and other proteins and, as a result, cause harm to the host tissues. These parameters could be crucial factors in the onset of periodontal inflammation and the subsequent tissue breakdown.

  2. Human neutrophils in auto-immunity.

    Science.gov (United States)

    Thieblemont, Nathalie; Wright, Helen L; Edwards, Steven W; Witko-Sarsat, Véronique

    2016-04-01

    Human neutrophils have great capacity to cause tissue damage in inflammatory diseases via their inappropriate activation to release reactive oxygen species (ROS), proteases and other tissue-damaging molecules. Furthermore, activated neutrophils can release a wide variety of cytokines and chemokines that can regulate almost every element of the immune system. In addition to these important immuno-regulatory processes, activated neutrophils can also release, expose or generate neoepitopes that have the potential to break immune tolerance and result in the generation of autoantibodies, that characterise a number of human auto-immune diseases. For example, in vasculitis, anti-neutrophil cytoplasmic antibodies (ANCA) that are directed against proteinase 3 or myeloperoxidase are neutrophil-derived autoantigens and activated neutrophils are the main effector cells of vascular damage. In other auto-immune diseases, these neutrophil-derived neoepitopes may arise from a number of processes that include release of granule enzymes and ROS, changes in the properties of components of their plasma membrane as a result of activation or apoptosis, and via the release of Neutrophil Extracellular Traps (NETs). NETs are extracellular structures that contain chromatin that is decorated with granule enzymes (including citrullinated proteins) that can act as neo-epitopes to generate auto-immunity. This review therefore describes the processes that can result in neutrophil-mediated auto-immunity, and the role of neutrophils in the molecular pathologies of auto-immune diseases such as vasculitis, rheumatoid arthritis (RA) and systemic lupus erythematosus (SLE). We discuss the potential role of NETs in these processes and some of the debate in the literature regarding the role of this phenomenon in microbial killing, cell death and auto-immunity. Copyright © 2016 Elsevier Ltd. All rights reserved.

  3. Treatment with Rutin - A Therapeutic Strategy for Neutrophil-Mediated Inflammatory and Autoimmune Diseases - Anti-inflammatory Effects of Rutin on Neutrophils -

    Directory of Open Access Journals (Sweden)

    Bahareh Abd Nikfarjam

    2017-03-01

    Full Text Available Objectives: Neutrophils represent the front line of human defense against infections. Immediately after stimulation, neutrophilic enzymes are activated and produce toxic mediators such as pro-inflammatory cytokines, nitric oxide (NO and myeloperoxidase (MPO. These mediators can be toxic not only to infectious agents but also to host tissues. Because flavonoids exhibit antioxidant and anti-inflammatory effects, they are subjects of interest for pharmacological modulation of inflammation. In the present study, the effects of rutin on stimulus-induced NO and tumor necrosis factor (TNF-α productions and MPO activity in human neutrophils were investigated. Methods: Human peripheral blood neutrophils were isolated using Ficoll-Hypaque density gradient centrifugation coupled with dextran T500 sedimentation. The cell preparations containing > 98% granulocytes were determined by morphological examination through Giemsa staining. Neutrophils were cultured in complete Roswell Park Memorial Institute (RPMI medium, pre-incubated with or without rutin (25 μM for 45 minutes, and stimulated with phorbol 12-myristate 13-acetate (PMA. Then, the TNF-α, NO and MPO productions were analyzed using enzyme-linked immunosorbent assay (ELISA, Griess Reagent, and MPO assay kits, respectively. Also, the viability of human neutrophils was assessed using tetrazolium salt 3-(4,5-dimethylthiazol-2-yl-2,5-diphenyl tetrazolium bromide (MTT, and neutrophils were treated with various concentrations of rutin (1 - 100 μM, after which MTT was appended and incubated at 37ºC for 4 hour. Results: Rutin at concentrations up to 100 μM did not affect neutrophil viability during the 4-hour incubation period. Rutin significantly decreased the NO and TNF-α productions in human peripheral blood neutrophils compared to PMA-control cells (P < 0.001. Also, MPO activity was significantly reduced by rutin (P < 0.001. Conclusion: In this in vitro study, rutin had an anti-inflammatory effect

  4. Biochemical changes in neutrophils of cervical cancer patients treated with 60Co

    International Nuclear Information System (INIS)

    Krishnamurthy, Vijayalakshmi; Gunalan, Gayathri; Haridas, Sumathy; Thangamani, Vanitha

    2008-01-01

    Cervical carcinoma is the second most common malignancy of the female genital tract in India. The highest incidence occurs at Chennai. This study was conducted on 30 women with biopsy-proved squamous cell carcinoma of the cervix of stage IIb. The neutrophil count increased significantly in cancer patients compared to control subjects. Total protein, glycogen and total lipid increased in neutrophils of cervical cancer patients. The level of cholestrol, triglycerides and fatty acids increased significantly in neutrophils of such patients compared to control subjects. The activity of alkaline phosphatase increased significantly in cervical cancer patients. Upon treatment with cobalt-60, these changes were brought to near-normal levels. This study highlights the impairment in the neutrophil function in cervical cancer patients, which may lead to reduced immune status. (author)

  5. Role of the Mycoplasma pneumoniae/Interleukin-8/Neutrophil Axis in the Pathogenesis of Pneumonia.

    Directory of Open Access Journals (Sweden)

    Zhengrong Chen

    Full Text Available Neutrophil infiltration is the characteristic pathological feature of M. pneumoniae pneumonia (MPP. This study aimed to explore the associations among neutrophil activity, clinical presentation, and role of the M. pneumoniae/interleukin-8 (IL-8/neutrophil axis in the pathogenesis of MPP. A total of 42 patients with MPP were prospectively enrolled in the study. Neutrophil activity, including matrix metalloproteinase-9 (MMP-9, myeloperoxidase (MPO, and neutrophil elastase (NE, were measured. Clinical information was collected for all patients and control group. In vitro, IL-8 production was measured at different time points after M. pneumoniae infection of bronchial epithelial cells, and neutrophil activity was analyzed after IL-8 stimulation. The percentage of neutrophil in the bronchoalveolar lavage fluid was higher in the group of patients with high levels of M. pneumoniae DNA than in those with low levels of M. pneumoniae DNA (P < 0.05. IL-8, MMP-9, and NE in patients with MPP significantly increased compared with controls and decreased after treatment (P < 0.05. MPO and MMP-9 were associated with duration of fever (r = 0.332, P < 0.05 and length of stay (r = 0.342, P < 0.05, respectively. In vitro, M. pneumoniae induced IL-8 production by bronchial epithelial cells in a time dependent manner. MPO, MMP-9 and NE production by neutrophils significantly increased compared with medium controls after IL-8 stimulation. In summary, the M. pneumoniae/IL-8/neutrophil axis likely plays a vital role in the pathogenesis of MPP.

  6. Neutrophil-to-lymphocyte ratio as an independent predictor for survival in patients with localized clear cell renal cell carcinoma after radiofrequency ablation: a propensity score matching analysis.

    Science.gov (United States)

    Chang, Xiaofeng; Zhang, Fan; Liu, Tieshi; Wang, Wei; Guo, Hongqian

    2017-06-01

    To investigate the role of neutrophil-to-lymphocyte ratio as a prognostic indicator in patients with localized clear cell renal cell carcinoma treated with radiofrequency ablation. We retrospectively analyzed data from patients with renal cell carcinoma who underwent radiofrequency ablation from 2006 to 2013. The Kaplan-Meier method was used to generate the survival curves according to different categories of neutrophil-to-lymphocyte ratio. Relationships between preoperative neutrophil-to-lymphocyte ratio or the change of neutrophil-to-lymphocyte ratio and survival were evaluated with multivariable Cox proportional hazards regression analysis. A propensity score matching analysis was carried out to avoid confounding bias. A total of 185 patients were included in present study. When stratified by preoperative neutrophil-to-lymphocyte ratio cutoff value of 2.79, 5-year recurrence-free survival, 5-year disease-free survival, and 5-year overall survival rates of neutrophil-to-lymphocyte ratio analysis, 5-year recurrence-free survival, 5-year disease-free survival, and 5-year overall survival rates of neutrophil-to-lymphocyte ratio ratio with the change of neutrophil-to-lymphocyte ratio, patients with both preoperative neutrophil-to-lymphocyte ratio ≥2.79 and the change of neutrophil-to-lymphocyte ratio ≥0.40 had the worst disease-free survival. Results of multivariable analysis showed that preoperative neutrophil-to-lymphocyte ratio and the change of neutrophil-to-lymphocyte ratio correlated with cancer relapse remarkably. High preoperative neutrophil-to-lymphocyte ratio and elevated postoperative neutrophil-to-lymphocyte ratio are associated with significant increase in risk of local recurrence as well as distant metastasis. The combination of neutrophil-to-lymphocyte ratio with the other prognostic indicators can be applied in the evaluation of relapse risk in patients with clear cell renal cell carcinoma after radiofrequency ablation.

  7. Serum and Glucocorticoid Regulated Kinase 1 (SGK1) Regulates Neutrophil Clearance During Inflammation Resolution

    Science.gov (United States)

    Burgon, Joseph; Robertson, Anne L.; Sadiku, Pranvera; Wang, Xingang; Hooper-Greenhill, Edward; Prince, Lynne R.; Walker, Paul; Hoggett, Emily E.; Ward, Jonathan R.; Farrow, Stuart N.; Zuercher, William J.; Jeffrey, Philip; Savage, Caroline O.; Ingham, Philip W.; Hurlstone, Adam F.; Whyte, Moira K. B.; Renshaw, Stephen A.

    2013-01-01

    The inflammatory response is integral to maintaining health, by functioning to resist microbial infection and repair tissue damage. Large numbers of neutrophils are recruited to inflammatory sites to neutralise invading bacteria through phagocytosis and the release of proteases and reactive oxygen species into the extracellular environment. Removal of the original inflammatory stimulus must be accompanied by resolution of the inflammatory response, including neutrophil clearance, to prevent inadvertent tissue damage. Neutrophil apoptosis and its temporary inhibition by survival signals provides a target for anti-inflammatory therapeutics, making it essential to better understand this process. GM-CSF, a neutrophil survival factor, causes a significant increase in mRNA levels for the known anti-apoptotic protein Serum and Glucocorticoid Regulated Kinase 1 (SGK1). We have characterised the expression patterns and regulation of SGK family members in human neutrophils, and shown that inhibition of SGK activity completely abrogates the anti-apoptotic effect of GM-CSF. Using a transgenic zebrafish model, we have disrupted sgk1 gene function and shown this specifically delays inflammation resolution, without altering neutrophil recruitment to inflammatory sites in vivo. These data suggest SGK1 plays a key role in regulating neutrophil survival signalling, and thus may prove a valuable therapeutic target for the treatment of inflammatory disease. PMID:24431232

  8. Serum and glucocorticoid-regulated kinase 1 regulates neutrophil clearance during inflammation resolution.

    Science.gov (United States)

    Burgon, Joseph; Robertson, Anne L; Sadiku, Pranvera; Wang, Xingang; Hooper-Greenhill, Edward; Prince, Lynne R; Walker, Paul; Hoggett, Emily E; Ward, Jonathan R; Farrow, Stuart N; Zuercher, William J; Jeffrey, Philip; Savage, Caroline O; Ingham, Philip W; Hurlstone, Adam F; Whyte, Moira K B; Renshaw, Stephen A

    2014-02-15

    The inflammatory response is integral to maintaining health by functioning to resist microbial infection and repair tissue damage. Large numbers of neutrophils are recruited to inflammatory sites to neutralize invading bacteria through phagocytosis and the release of proteases and reactive oxygen species into the extracellular environment. Removal of the original inflammatory stimulus must be accompanied by resolution of the inflammatory response, including neutrophil clearance, to prevent inadvertent tissue damage. Neutrophil apoptosis and its temporary inhibition by survival signals provides a target for anti-inflammatory therapeutics, making it essential to better understand this process. GM-CSF, a neutrophil survival factor, causes a significant increase in mRNA levels for the known anti-apoptotic protein serum and glucocorticoid-regulated kinase 1 (SGK1). We have characterized the expression patterns and regulation of SGK family members in human neutrophils and shown that inhibition of SGK activity completely abrogates the antiapoptotic effect of GM-CSF. Using a transgenic zebrafish model, we have disrupted sgk1 gene function and shown this specifically delays inflammation resolution, without altering neutrophil recruitment to inflammatory sites in vivo. These data suggest SGK1 plays a key role in regulating neutrophil survival signaling and thus may prove a valuable therapeutic target for the treatment of inflammatory disease.

  9. Effects of chronic occupational exposure to anaesthetic gases on the rate of neutrophil apoptosis among anaesthetists.

    LENUS (Irish Health Repository)

    Tyther, R

    2012-02-03

    BACKGROUND AND OBJECTIVE: Volatile anaesthetic agents are known to influence neutrophil function. The aim was to determine the effect of chronic occupational exposure to volatile anaesthetic agents on the rate of neutrophil apoptosis among anaesthetists. To test this hypothesis, we compared the rate of neutrophil apoptosis in anaesthetists who had been chronically exposed to volatile anaesthetic agents with that in unexposed volunteers. METHODS: Venous blood (20 mL) was withdrawn from 24 ASA I-II volunteers, from which neutrophils were isolated, and maintained in culture. At 1, 12 and 24 h in culture, the percentage of neutrophil apoptosis was assessed by dual staining with annexin V-FITC and propidium iodide. RESULTS: At 1 h (but not at 12 and 24 h) in culture, the rate of neutrophil apoptosis was significantly less in the anaesthetists--13.8 (12.9%) versus 34.4 (12.1%) (P = 0.001). CONCLUSIONS: Chronic occupational exposure to volatile anaesthetic agents may inhibit neutrophil apoptosis. This may have implications for anaesthetists and similarly exposed healthcare workers in terms of the adequacy of their inflammatory response.

  10. CARD9-Dependent Neutrophil Recruitment Protects against Fungal Invasion of the Central Nervous System.

    Directory of Open Access Journals (Sweden)

    Rebecca A Drummond

    2015-12-01

    Full Text Available Candida is the most common human fungal pathogen and causes systemic infections that require neutrophils for effective host defense. Humans deficient in the C-type lectin pathway adaptor protein CARD9 develop spontaneous fungal disease that targets the central nervous system (CNS. However, how CARD9 promotes protective antifungal immunity in the CNS remains unclear. Here, we show that a patient with CARD9 deficiency had impaired neutrophil accumulation and induction of neutrophil-recruiting CXC chemokines in the cerebrospinal fluid despite uncontrolled CNS Candida infection. We phenocopied the human susceptibility in Card9-/- mice, which develop uncontrolled brain candidiasis with diminished neutrophil accumulation. The induction of neutrophil-recruiting CXC chemokines is significantly impaired in infected Card9-/- brains, from both myeloid and resident glial cellular sources, whereas cell-intrinsic neutrophil chemotaxis is Card9-independent. Taken together, our data highlight the critical role of CARD9-dependent neutrophil trafficking into the CNS and provide novel insight into the CNS fungal susceptibility of CARD9-deficient humans.

  11. Interval and continuous exercise regimens suppress neutrophil-derived microparticle formation and neutrophil-promoted thrombin generation under hypoxic stress.

    Science.gov (United States)

    Chen, Yi-Ching; Ho, Ching-Wen; Tsai, Hsing-Hua; Wang, Jong-Shyan

    2015-04-01

    Acute hypoxic exposure increases vascular thrombotic risk. The release of procoagulant-rich microparticles from neutrophils accelerates the pathogenesis of inflammatory thrombosis. The present study explicates the manner in which interval and continuous exercise regimens affect neutrophil-derived microparticle (NDMP) formation and neutrophil/NDMP-mediated thrombin generation (TG) under hypoxic condition. A total of 60 sedentary males were randomized to perform either aerobic interval training [AIT; 3-min intervals at 40% and 80% V̇O2max (maximal O2 consumption)] or moderate continuous training (MCT; sustained 60% V̇O2max) for 30 min/day, 5 days/week for 5 weeks, or to a control (CTL) group who did not receive any form of training. At rest and immediately after hypoxic exercise test (HE, 100 W under 12% O2 for 30 min), the NDMP characteristics and dynamic TG were measured by flow cytometry and thrombinography respectively. Before the intervention, HE (i) elevated coagulant factor VIII/fibrinogen concentrations and shortened activated partial thromboplastin time (aPTT), (ii) increased total and tissue factor (TF)-rich/phosphatidylserine (PS)-exposed NDMP counts and (iii) enhanced the peak height and rate of TG promoted by neutrophils/NDMPs. Following the 5-week intervention, AIT exhibited higher enhancement of V̇O2max than did MCT. Notably, both MCT and AIT attenuated the extents of HE-induced coagulant factor VIII/fibrinogen elevations and aPTT shortening. Furthermore, the two exercise regimens significantly decreased TF-rich/PS-exposed NDMP formation and depressed neutrophil/NDMP-mediated dynamic TG at rest and following HE. Hence, we conclude that AIT is superior to MCT for enhancing aerobic capacity. Moreover, either AIT or MCT effectively ameliorates neutrophil/NDMP-promoted TG by down-regulating expression of procoagulant factors during HE, which may reduce thrombotic risk evoked by hypoxia. Moreover, either AIT or MCT effectively ameliorates neutrophil

  12. Activation of bovine neutrophils by Brucella spp.

    Science.gov (United States)

    Keleher, Lauren L; Skyberg, Jerod A

    2016-09-01

    Brucellosis is a globally important zoonotic infectious disease caused by gram negative bacteria of the genus Brucella. While many species of Brucella exist, Brucella melitensis, Brucella abortus, and Brucella suis are the most common pathogens of humans and livestock. The virulence of Brucella is largely influenced by its ability to evade host factors, including phagocytic killing mechanisms, which are critical for the host response to infection. The aim of this study was to characterize the bovine neutrophil response to virulent Brucella spp. Here, we found that virulent strains of smooth B. abortus, B. melitensis, B. suis, and virulent, rough, strains of Brucella canis possess similar abilities to resist killing by resting, or IFN-γ-activated, bovine neutrophils. Bovine neutrophils responded to infection with a time-dependent oxidative burst that varied little between Brucella spp. Inhibition of TAK1, or SYK kinase blunted the oxidative burst of neutrophils in response to Brucella infection. Interestingly, Brucella spp. did not induce robust death of bovine neutrophils. These results indicate that bovine neutrophils respond similarly to virulent Brucella spp. In addition, virulent Brucella spp., including naturally rough strains of B. canis, have a conserved ability to resist killing by bovine neutrophils. Copyright © 2016 Elsevier B.V. All rights reserved.

  13. Depression of efficiency of neutrophils for Candida albicans phagocytosis in personnel working in radiation field

    International Nuclear Information System (INIS)

    Hassan, A.A.

    2000-01-01

    The neutrophil functions, chemotaxis (direct and random migration), phagocytosis using Candida albicans (percent, index), phagocytosis by NBT (percent, score) and adherence were studied on 55 persons working in radiation field (group I) and 40 persons as control (group II). The effect of radiation on blood picture of persons working in this field with special references to leucocytic counts and neutrophil functions was studied. White and red cells counts were 6.275 +- 1.723 and 5.475 +- 1.039 (group I) and 6.440 +- 1.556, 4.704 +- 0.734 for group II, respectively with no significant difference, while in neutrophil function there was a statistically significant difference in all functions between two groups (P < 0.01). This indicates the importance of neutrophil functions in following up persons working in radiation field

  14. Predictive value of eosinophils and neutrophils on clinical effects of ICS in COPD

    DEFF Research Database (Denmark)

    Hartjes, Floor J; Vonk, Judith M; Faiz, Alen

    2018-01-01

    BACKGROUND AND OBJECTIVE: Inflammation is present to a variable degree and composition in patients with COPD. This study investigates associations between both eosinophils and neutrophils in blood, sputum, airway wall biopsies and bronchoalveolar lavage (BAL) and their potential use as biomarkers...... and BAL were evaluated at baseline. In addition, at baseline, 6 and 30 months, forced expiratory flow in 1 s (FEV1 ), residual volume/total lung capacity (hyperinflation) and Clinical COPD Questionnaire were evaluated. RESULTS: Cross-sectional analyses at baseline showed that higher blood eosinophils were...... significantly associated with higher eosinophil counts in sputum, biopsies and BAL. However, blood neutrophils did not significantly correlate with neutrophil counts in the other compartments. Baseline eosinophils and neutrophils, in whichever compartment measured, did not predict longitudinal FEV1 changes...

  15. The effect of cigarette smoking on neutrophil kinetics in human lungs [see comments

    International Nuclear Information System (INIS)

    MacNee, W.; Wiggs, B.; Belzberg, A.S.; Hogg, J.C.

    1989-01-01

    Neutrophils may play a part in the pathogenesis of the centrilobular emphysema associated with cigarette smoking. The capillary bed of the lungs concentrates neutrophils approximately 100-fold with respect to erythrocytes, producing a large pool of marginated cells. We examined the effect of cigarette smoking on the kinetics of this pool of cells, using 99mTc-labeled erythrocytes to measure regional blood velocity and 111In-labeled neutrophils to measure the removal of neutrophils during the first passage through the pulmonary circulation, their subsequent washout from the lungs, and the effect of local blood velocity on the number of neutrophils retained in each lung region. We observed no difference in these measurements between subjects who had never smoked (n = 6) and smokers who did not smoke during the study (n = 12). However, subjects who did smoke during the study (n = 12) had a significantly slower rate of washout of radiolabeled neutrophils from the lung (0.08 +/- 0.04 of the total per minute, as compared with 0.13 +/- 0.06 in smokers who did not smoke during the experiment and 0.14 +/- 0.08 in non-smokers) (P = 0.02). We also observed an increase in the regional retention of labeled neutrophils with respect to blood velocity in 5 of the 12 subjects who smoked during the study, but in none of the other subjects. We conclude that the presence of cigarette smoke in the lungs of some subjects increases the local concentration of neutrophils, and suggest that the lesions that characterize emphysema may be a result of the destruction of lung tissue by neutrophils that remain within pulmonary microvessels

  16. N-Formyl-Perosamine Surface Homopolysaccharides Hinder the Recognition of Brucella abortus by Mouse Neutrophils.

    Science.gov (United States)

    Mora-Cartín, Ricardo; Chacón-Díaz, Carlos; Gutiérrez-Jiménez, Cristina; Gurdián-Murillo, Stephany; Lomonte, Bruno; Chaves-Olarte, Esteban; Barquero-Calvo, Elías; Moreno, Edgardo

    2016-06-01

    Brucella abortus is an intracellular pathogen of monocytes, macrophages, dendritic cells, and placental trophoblasts. This bacterium causes a chronic disease in bovines and in humans. In these hosts, the bacterium also invades neutrophils; however, it fails to replicate and just resists the killing action of these leukocytes without inducing significant activation or neutrophilia. Moreover, B. abortus causes the premature cell death of human neutrophils. In the murine model, the bacterium is found within macrophages and dendritic cells at early times of infection but seldom in neutrophils. Based on this observation, we explored the interaction of mouse neutrophils with B. abortus In contrast to human, dog, and bovine neutrophils, naive mouse neutrophils fail to recognize smooth B. abortus bacteria at early stages of infection. Murine normal serum components do not opsonize smooth Brucella strains, and neutrophil phagocytosis is achieved only after the appearance of antibodies. Alternatively, mouse normal serum is capable of opsonizing rough Brucella mutants. Despite this, neutrophils still fail to kill Brucella, and the bacterium induces cell death of murine leukocytes. In addition, mouse serum does not opsonize Yersinia enterocolitica O:9, a bacterium displaying the same surface polysaccharide antigen as smooth B. abortus Therefore, the lack of murine serum opsonization and absence of murine neutrophil recognition are specific, and the molecules responsible for the Brucella camouflage are N-formyl-perosamine surface homopolysaccharides. Although the mouse is a valuable model for understanding the immunobiology of brucellosis, direct extrapolation from one animal system to another has to be undertaken with caution. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

  17. The effect of lidocaine on neutrophil respiratory burst during induction of general anaesthesia and tracheal intubation.

    LENUS (Irish Health Repository)

    Swanton, B J

    2012-02-03

    BACKGROUND AND OBJECTIVE: Respiratory burst is an essential component of the neutrophil\\'s biocidal function. In vitro, sodium thiopental, isoflurane and lidocaine each inhibit neutrophil respiratory burst. The objectives of this study were (a) to determine the effect of a standard clinical induction\\/tracheal intubation sequence on neutrophil respiratory burst and (b) to determine the effect of intravenous lidocaine administration during induction of anaesthesia on neutrophil respiratory burst. METHODS: Twenty ASA I and II patients, aged 18-60 years, undergoing elective surgery were studied. After induction of anaesthesia [fentanyl (2 microg kg-1), thiopental (4-6 mg kg-1), isoflurane (end-tidal concentration 0.5-1.5%) in nitrous oxide (66%) and oxygen], patients randomly received either lidocaine 1.5 mg kg-1 (group L) or 0.9% saline (group S) prior to tracheal intubation. Neutrophil respiratory burst was measured immediately prior to induction of anaesthesia, immediately before and 1 and 5 min after lidocaine\\/saline. RESULTS: Neutrophil respiratory burst decreased significantly after induction of anaesthesia in both groups [87.4 +\\/- 8.2% (group L) and 88.5 +\\/- 13.4% (group S) of preinduction level (P < 0.01 both groups)]. After intravenous lidocaine (but not saline) administration, neutrophil respiratory burst returned towards preinduction levels, both before (97.1 +\\/- 23.6%) and after (94.4 +\\/- 16.6%) tracheal intubation. CONCLUSION: Induction of anaesthesia and tracheal intubation using thiopentone and isoflurane, inhibit neutrophil respiratory burst. This effect may be diminished by the administration of lidocaine.

  18. Grain dust induces IL-8 production from bronchial epithelial cells: effect on neutrophil recruitment.

    Science.gov (United States)

    Park, H S; Suh, J H; Kim, S S; Kwon, O J

    2000-06-01

    There have been several investigations suggesting an involvement of activated neutrophils in the development of grain dust (GD)-induced occupational asthma. Interleukin-8 in the sputa from GD-induced asthmatic patients increased significantly after the exposure to GD. To confirm IL-8 production from bronchial epithelial cells when exposed to GD, and to evaluate the role of IL-8 on neutrophil recruitment. We cultured Beas-2B, a bronchial epithelial cell line. To observe GD-induced responses, four different concentrations ranging from 1 to 200 microg/mL of GD were incubated for 24 hours and compared with those without incubation of GD. To evaluate the effect of pro-inflammatory cytokines on IL-8 production and neutrophil chemotaxis, epithelial cells were incubated with peripheral blood mononuclear cell (PBMC) culture supernatant derived from subjects with GD-induced asthma exposed to 10 microg/mL of GD, and then compared with those without addition of PBMC supernatant. The level of released IL-8 in the supernatant was measured by enzyme-linked immunosorbent assay. Neutrophil chemotactic activity of the culture supernatant was determined by modified Boyden chamber method. Interleukin-8 production and neutrophil chemotactic activity from bronchial epithelial cells significantly increased with additions of GD in a dose-dependent manner (P < .05, respectively), and were significantly augmented with additions of PBMC supernatant (P < .05, respectively) at each concentration. Close correlation was noted between neutrophil chemotactic activity and IL-8 level (r = 0.87, P < .05). Compared with the untreated sample, pre-treatment of anti-IL-8 antibody induced a significant suppression (up to 67.2%) of neutrophil chemotactic activity in a dose-dependent manner. These results suggest that IL-8 produced from bronchial epithelial cells may be a major cytokine, which induces neutrophil migration into the airways when exposed to GD.

  19. Modulation of oxygen-dependent and oxygen-independent metabolism of neutrophilic granulocytes by quantum points.

    Science.gov (United States)

    Pleskova, S N; Mikheeva, E R

    2011-08-01

    Inhibition of neutrophilic granulocyte metabolism under the effect of semiconductor quantum points was demonstrated. The status of the oxidative system was evaluated by the NBT test, nonoxidative status by the lysosomal cationic test. It was found that quantum points in a dose of 0.1 mg/ml irrespective of their core and composition of coating significantly inhibited oxygen-dependent and oxygen-independent metabolism of neutrophilic granulocytes.

  20. Elevated Neutrophil Lymphocyte Ratio in Recurrent Optic Neuritis

    Directory of Open Access Journals (Sweden)

    Hande Guclu

    2015-01-01

    Full Text Available Purpose. To demonstrate the relation between optic neuritis (ON and systemic inflammation markers as neutrophil lymphocyte ratio (N/L ratio, platelet count, mean platelet volume (MPV, and red cell distribution width (RDW and furthermore to evaluate the utilization of these markers to predict the frequency of the ON episodes. Methods. Forty-two patients with acute ON and forty healthy subjects were enrolled into the study. The medical records were reviewed for age, sex, hemoglobin (Hb, Haematocrit (Htc, RDW, platelet count, MPV, white blood cell count (WBC, neutrophil and lymphocyte count, and neutrophil lymphocyte ratio (N/L ratio. Results. The mean N/L ratio, platelet counts, and RDW were significantly higher in ON group (p=0.000, p=0.048, and p=0.002. There was a significant relation between N/L ratio and number of episodes (r=0.492, p=0.001. There was a statistically significant difference for MPV between one episode group and recurrent ON group (p=0.035. Conclusions. Simple and inexpensive laboratory methods could help us show systemic inflammation and monitor ON patients. Higher N/L ratio can be a useful marker for predicting recurrent attacks.

  1. Neutrophil Lymphocyte Ratio Predicts Postoperative Pain after ...

    African Journals Online (AJOL)

    2018-02-07

    Feb 7, 2018 ... between preoperatively measured neutrophil-lymphocyte ratio (NLR) – as an inflammation ... analgesic (tenoxicam – as the first drug of choice, paracetamol, tramadol, or pethidine) usage ... fracture fixation). Age, sex, type of ...

  2. Investigation of urinary neutrophil gelatinase associated lipocalin ...

    African Journals Online (AJOL)

    Investigation of urinary neutrophil gelatinase associated lipocalin (NGAL) for early diagnosis of acute kidney ... African Journal of Urology ... Demographic and clinical data including surgical procedure were recorded in Excel and analyzed by ...

  3. Targeting Neutrophilic Inflammation using Polymersome-Mediated Cellular Delivery

    OpenAIRE

    Robertson, J.D.; Ward, J.R.; Avila-Olias, M.; Battaglia, G.; Renshaw, S.A.

    2017-01-01

    Neutrophils are key effector cells in inflammation and play an important role in neutralizing invading pathogens. During inflammation resolution, neutrophils undergo apoptosis before they are removed by macrophages, but if apoptosis is delayed, neutrophils can cause extensive tissue damage and chronic disease. Promotion of neutrophil apoptosis is a potential therapeutic approach for treating persistent inflammation, yet neutrophils have proven difficult cells to manipulate experimentally. In ...

  4. Neutrophil extracellular trap formation in supragingival biofilms.

    Science.gov (United States)

    Hirschfeld, Josefine; Dommisch, Henrik; Skora, Philipp; Horvath, Gabor; Latz, Eicke; Hoerauf, Achim; Waller, Tobias; Kawai, Toshihisa; Jepsen, Søren; Deschner, James; Bekeredjian-Ding, Isabelle

    2015-01-01

    Oral biofilms are the causative agents of the highly prevalent oral diseases periodontitis and caries. Additionally, the host immune response is thought to play a critical role in disease onset. Neutrophils are known to be a key host response factor to bacterial challenge on host surfaces. Release of neutrophil extracellular traps (NETs) as a novel antimicrobial defense strategy has gained increasing attention in the past years. Here, we investigated the influx of neutrophils into the dental plaque and the ability of oral bacteria to trigger intra-biofilm release of NETs and intracellular proteins. Supragingival biofilms and whole saliva were sampled from systemically healthy subjects participating in an experimental gingivitis study. Biofilms were analysed by immunofluorescence followed by confocal and fluorescence microscopy. Moreover, concentrations of cytokines and immune-associated proteins in biofilm suspensions and saliva were assessed by ELISA. Neutrophils obtained from blood were stimulated with twelve bacterial species isolated from cultured biofilms or with lipopolysaccharide to monitor NET formation. Neutrophils, NETs, neutrophil-associated proteins (myeloperoxidase, elastase-2, cathepsin G, cathelicidin LL-37), interleukin-8, interleukin-1β and tumor necrosis factor were detected within plaque samples and saliva. All tested bacterial species as well as the polymicrobial samples isolated from the plaque of each donor induced release of NETs and interleukin-8. The degree of NET formation varied among different subjects and did not correlate with plaque scores or clinical signs of local inflammation. Our findings indicate that neutrophils are attracted towards dental biofilms, in which they become incorporated and where they are stimulated by microbes to release NETs and immunostimulatory proteins. Thus, neutrophils and NETs may be involved in host biofilm control, although their specific role needs to be further elucidated. Moreover, inter

  5. Neutrophil extracellular traps: double-edged swords of innate immunity.

    Science.gov (United States)

    Kaplan, Mariana J; Radic, Marko

    2012-09-15

    Spectacular images of neutrophils ejecting nuclear chromatin and bactericidal proteins, in response to microbes, were first reported in 2004. As externalized chromatin could entangle bacteria, these structures were named neutrophil extracellular traps (NETs). Subsequent studies identified microorganisms and sterile conditions that stimulate NETs, as well as additional cell types that release extracellular chromatin. The release of NETs is the most dramatic stage in a cell death process called NETosis. Experimental evidence suggests that NETs participate in pathogenesis of autoimmune and inflammatory disorders, with proposed involvement in glomerulonephritis, chronic lung disease, sepsis, and vascular disorders. Exaggerated NETosis or diminished NET clearance likely increases risk of autoreactivity to NET components. The biological significance of NETs is just beginning to be explored. A more complete integration of NETosis within immunology and pathophysiology will require better understanding of NET properties associated with specific disease states and microbial infections. This may lead to the identification of important therapeutic targets.

  6. Association between neutrophil/lymphocyte ratio and coronary collateral circulation

    Directory of Open Access Journals (Sweden)

    Mustafa Oylumlu

    2012-03-01

    Full Text Available Objectives: To investigate relation between neutrophil/lymphocyte ratio and coronary collateral flow.Material and methods: Eighty-two patients admittedDicle University Medical Faculty Hospital Cardiology Departmentwith diagnosis of coronary artery disease anddetected significant stenosis or occlusion at least one ofthe coronary arteries, were included to study. Age, sex,presence of diabetes mellitus and hypertension, acute/stable coronary disease, body mass index, neutrophil/lymphocyte ratio, white blood count, Rentrop scores andnumber of diseased vessel were recorded.Results: Well-developed coronary collateral circulationwas found in 33 of the patients. Forty-nine patients hadpoor coronary collateral circulation. Mean age, sex, bodymass index, presence of diabetes mellitus and hypertensionwere similar in two groups. Mean neutrophil/lymphocyteratio was lower in well-developed coronary collateralcirculation group than poor coronary collateral circulationgroup, but there was no significant differences (2.78 vs2.89, p=0.12.Conclusions: There was no association between neutron/hil lymphocyte ratio and coronary collateral circulationaccording to our data. J Clin Exp Invest 2012; 3(1:29-32

  7. Neutrophil depletion reduces edema formation and tissue loss following traumatic brain injury in mice

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    Kenne Ellinor

    2012-01-01

    Full Text Available Abstract Background Brain edema as a result of secondary injury following traumatic brain injury (TBI is a major clinical concern. Neutrophils are known to cause increased vascular permeability leading to edema formation in peripheral tissue, but their role in the pathology following TBI remains unclear. Methods In this study we used controlled cortical impact (CCI as a model for TBI and investigated the role of neutrophils in the response to injury. The outcome of mice that were depleted of neutrophils using an anti-Gr-1 antibody was compared to that in mice with intact neutrophil count. The effect of neutrophil depletion on blood-brain barrier function was assessed by Evan's blue dye extravasation, and analysis of brain water content was used as a measurement of brain edema formation (24 and 48 hours after CCI. Lesion volume was measured 7 and 14 days after CCI. Immunohistochemistry was used to assess cell death, using a marker for cleaved caspase-3 at 24 hours after injury, and microglial/macrophage activation 7 days after CCI. Data were analyzed using Mann-Whitney test for non-parametric data. Results Neutrophil depletion did not significantly affect Evan's blue extravasation at any time-point after CCI. However, neutrophil-depleted mice exhibited a decreased water content both at 24 and 48 hours after CCI indicating reduced edema formation. Furthermore, brain tissue loss was attenuated in neutropenic mice at 7 and 14 days after injury. Additionally, these mice had a significantly reduced number of activated microglia/macrophages 7 days after CCI, and of cleaved caspase-3 positive cells 24 h after injury. Conclusion Our results suggest that neutrophils are involved in the edema formation, but not the extravasation of large proteins, as well as contributing to cell death and tissue loss following TBI in mice.

  8. Interleukin-17 Promotes Neutrophil-Mediated Immunity by Activating Microvascular Pericytes and Not Endothelium

    Science.gov (United States)

    Liu, Rebecca; Lauridsen, Holly M.; Amezquita, Robert A.; Pierce, Richard W.; Jane-wit, Dan; Fang, Caodi; Pellowe, Amanda S.; Kirkiles-Smith, Nancy C.; Gonzalez, Anjelica L.; Pober, Jordan S.

    2016-01-01

    A classical hallmark of acute inflammation is neutrophil infiltration of tissues, a multi-step process that involves sequential cell-cell interactions of circulating leukocytes with interleukin (IL)-1- or tumor necrosis factor-α (TNF)-activated microvascular endothelial cells (ECs) and pericytes (PCs) that form the wall of the postcapillary venules. The initial infiltrating cells accumulate perivascularly in close proximity to PCs. IL-17, a pro-inflammatory cytokine that acts on target cells via a heterodimeric receptor formed by IL-17RA and IL-17RC subunits, also promotes neutrophilic inflammation but its effects on vascular cells are less clear. We report that both cultured human ECs and PCs strongly express IL-17RC and, while neither cell type expresses much IL-17RA, PCs express significantly more than ECs. IL-17, alone or synergistically with TNF, significantly alters inflammatory gene expression in cultured human PCs but not ECs. RNA-seq analysis identifies many IL-17-induced transcripts in PCs encoding proteins known to stimulate neutrophil-mediated immunity. Conditioned media (CM) from IL-17-activated PCs, but not ECs, induce pertussis toxin-sensitive neutrophil polarization, likely mediated by PC-secreted chemokines, and also stimulate neutrophil production of pro-inflammatory molecules, including TNF, IL-1α, IL-1β, and IL-8. Furthermore, IL-17-activated PCs but not ECs can prolong neutrophil survival by producing G-CSF and GM-CSF, delaying the mitochondria outer membrane permeabilization and caspase 9 activation. Importantly, neutrophils exhibit enhanced phagocytic capacity after activation by CM from IL-17-treated PCs. We conclude that PCs, not ECs, are the major target of IL-17 within the microvessel wall and that IL-17-activated PCs can modulate neutrophil functions within the perivascular tissue space. PMID:27534549

  9. In vivo study of indomethacin in bronchiectasis: effect on neutrophil function and lung secretion.

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    Llewellyn-Jones, C G; Johnson, M M; Mitchell, J L; Pye, A; Okafor, V C; Hill, S L; Stockley, R A

    1995-09-01

    Bronchiectasis is associated with sputum containing high levels of the proteolytic enzyme elastase, which is thought to be involved in the pathogenesis of the disease. Agents which inhibit neutrophil function and interfere with neutrophil elastase release may have a beneficial effect on the development and progression of such diseases. We have studied the effects of the nonsteroidal anti-inflammatory agent indomethacin on neutrophil function in nine patients with clinically stable bronchiectasis. All patients remained clinically stable during the study. We observed a significant reduction in peripheral neutrophil chemotaxis to 10 nmol.L-1 N-formyl-methionyl-leucyl-phenylalanine (FMLP) from a mean of 19.86 (SEM 1.35) to 8.46 (0.68) cells.field-1 after 4 weeks of therapy. There was also a significant reduction in fibronectin degradation both by resting and FMLP-stimulated neutrophils, from a mean of 1.90 (0.19) micrograms x 3 x 10(5) cells at the start of therapy to 0.87 (0.08) micrograms after 4 weeks, and from 3.17 (0.35) micrograms to 1.48 (0.05) micrograms, respectively. There was no effect on spontaneous or stimulated superoxide anion generation by neutrophils. Despite the marked changes in peripheral neutrophil function, no adverse effect was observed on viable bacterial load in the bronchial secretions. In addition, there was no difference in sputum albumin, elastase or myeloperoxidase levels, and only minor changes in the chemotactic activity of the sputum. These results suggest that nonsteroidal anti-inflammatory agents have a major effect on peripheral neutrophil function but do not appear to have an adverse effect on bacterial colonization of the airways.

  10. Effect of neutrophil depletion on gelatinase expression, edema formation and hemorrhagic transformation after focal ischemic stroke

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    Machado Livia S

    2005-08-01

    Full Text Available Abstract Background While gelatinase (MMP-2 and -9 activity is increased after focal ischemia/reperfusion injury in the brain, the relative contribution of neutrophils to the MMP activity and to the development of hemorrhagic transformation remains unknown. Results Anti-PMN treatment caused successful depletion of neutrophils in treated animals. There was no difference in either infarct volume or hemorrhage between control and PMN depleted animals. While there were significant increases in gelatinase (MMP-2 and MMP-9 expression and activity and edema formation associated with ischemia, neutrophil depletion failed to cause any change. Conclusion The main finding of this study is that, in the absence of circulating neutrophils, MMP-2 and MMP-9 expression and activity are still up-regulated following focal cerebral ischemia. Additionally, neutrophil depletion had no influence on indicators of ischemic brain damage including edema, hemorrhage, and infarct size. These findings indicate that, at least acutely, neutrophils are not a significant contributor of gelatinase activity associated with acute neurovascular damage after stroke.

  11. Association of microparticles and neutrophil activation with decompression sickness.

    Science.gov (United States)

    Thom, Stephen R; Bennett, Michael; Banham, Neil D; Chin, Walter; Blake, Denise F; Rosen, Anders; Pollock, Neal W; Madden, Dennis; Barak, Otto; Marroni, Alessandro; Balestra, Costantino; Germonpre, Peter; Pieri, Massimo; Cialoni, Danilo; Le, Phi-Nga Jeannie; Logue, Christopher; Lambert, David; Hardy, Kevin R; Sward, Douglas; Yang, Ming; Bhopale, Veena B; Dujic, Zeljko

    2015-09-01

    Decompression sickness (DCS) is a systemic disorder, assumed due to gas bubbles, but additional factors are likely to play a role. Circulating microparticles (MPs)--vesicular structures with diameters of 0.1-1.0 μm--have been implicated, but data in human divers have been lacking. We hypothesized that the number of blood-borne, Annexin V-positive MPs and neutrophil activation, assessed as surface MPO staining, would differ between self-contained underwater breathing-apparatus divers suffering from DCS vs. asymptomatic divers. Blood was analyzed from 280 divers who had been exposed to maximum depths from 7 to 105 meters; 185 were control/asymptomatic divers, and 90 were diagnosed with DCS. Elevations of MPs and neutrophil activation occurred in all divers but normalized within 24 h in those who were asymptomatic. MPs, bearing the following proteins: CD66b, CD41, CD31, CD142, CD235, and von Willebrand factor, were between 2.4- and 11.7-fold higher in blood from divers with DCS vs. asymptomatic divers, matched for time of sample acquisition, maximum diving depth, and breathing gas. Multiple logistic regression analysis documented significant associations (P < 0.001) between DCS and MPs and for neutrophil MPO staining. Effect estimates were not altered by gender, body mass index, use of nonsteroidal anti-inflammatory agents, or emergency oxygen treatment and were modestly influenced by divers' age, choice of breathing gas during diving, maximum diving depth, and whether repetitive diving had been performed. There were no significant associations between DCS and number of MPs without surface proteins listed above. We conclude that MP production and neutrophil activation exhibit strong associations with DCS. Copyright © 2015 the American Physiological Society.

  12. NR4A orphan nuclear receptor family members, NR4A2 and NR4A3, regulate neutrophil number and survival.

    Science.gov (United States)

    Prince, Lynne R; Prosseda, Svenja D; Higgins, Kathryn; Carlring, Jennifer; Prestwich, Elizabeth C; Ogryzko, Nikolay V; Rahman, Atiqur; Basran, Alexander; Falciani, Francesco; Taylor, Philip; Renshaw, Stephen A; Whyte, Moira K B; Sabroe, Ian

    2017-08-24

    The lifespan of neutrophils is plastic and highly responsive to factors that regulate cellular survival. Defects in neutrophil number and survival are common to both hematologic disorders and chronic inflammatory diseases. At sites of inflammation, neutrophils respond to multiple signals that activate protein kinase A (PKA) signaling, which positively regulates neutrophil survival. The aim of this study was to define transcriptional responses to PKA activation and to delineate the roles of these factors in neutrophil function and survival. In human neutrophil gene array studies, we show that PKA activation upregulates a significant number of apoptosis-related genes, the most highly regulated of these being NR4A2 and NR4A3 Direct PKA activation by the site-selective PKA agonist pair N6/8-AHA (8-AHA-cAMP and N6-MB-cAMP) and treatment with endogenous activators of PKA, including adenosine and prostaglandin E2, results in a profound delay of neutrophil apoptosis and concomitant upregulation of NR4A2/3 in a PKA-dependent manner. NR4A3 expression is also increased at sites of neutrophilic inflammation in a human model of intradermal inflammation. PKA activation also promotes survival of murine neutrophil progenitor cells, and small interfering RNA to NR4A2 decreases neutrophil production in this model. Antisense knockdown of NR4A2 and NR4A3 homologs in zebrafish larvae significantly reduces the absolute neutrophil number without affecting cellular migration. In summary, we show that NR4A2 and NR4A3 are components of a downstream transcriptional response to PKA activation in the neutrophil, and that they positively regulate neutrophil survival and homeostasis. © 2017 by The American Society of Hematology.

  13. Coccidioides Endospores and Spherules Draw Strong Chemotactic, Adhesive, and Phagocytic Responses by Individual Human Neutrophils.

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    Cheng-Yuk Lee

    Full Text Available Coccidioides spp. are dimorphic pathogenic fungi whose parasitic forms cause coccidioidomycosis (Valley fever in mammalian hosts. We use an innovative interdisciplinary approach to analyze one-on-one encounters between human neutrophils and two forms of Coccidioides posadasii. To examine the mechanisms by which the innate immune system coordinates different stages of the host response to fungal pathogens, we dissect the immune-cell response into chemotaxis, adhesion, and phagocytosis. Our single-cell technique reveals a surprisingly strong response by initially quiescent neutrophils to close encounters with C. posadasii, both from a distance (by complement-mediated chemotaxis as well as upon contact (by serum-dependent adhesion and phagocytosis. This response closely resembles neutrophil interactions with Candida albicans and zymosan particles, and is significantly stronger than the neutrophil responses to Cryptococcus neoformans, Aspergillus fumigatus, and Rhizopus oryzae under identical conditions. The vigorous in vitro neutrophil response suggests that C. posadasii evades in vivo recognition by neutrophils through suppression of long-range mobilization and recruitment of the immune cells. This observation elucidates an important paradigm of the recognition of microbes, i.e., that intact immunotaxis comprises an intricate spatiotemporal hierarchy of distinct chemotactic processes. Moreover, in contrast to earlier reports, human neutrophils exhibit vigorous chemotaxis toward, and frustrated phagocytosis of, the large spherules of C. posadasii under physiological-like conditions. Finally, neutrophils from healthy donors and patients with chronic coccidioidomycosis display subtle differences in their responses to antibody-coated beads, even though the patient cells appear to interact normally with C. posadasii endospores.

  14. Neutrophils in Cancer: Two Sides of the Same Coin.

    Science.gov (United States)

    Uribe-Querol, Eileen; Rosales, Carlos

    2015-01-01

    Neutrophils are the most abundant leukocytes in blood and are considered to be the first line of defense during inflammation and infections. In addition, neutrophils are also found infiltrating many types of tumors. Tumor-associated neutrophils (TANs) have relevant roles in malignant disease. Indeed neutrophils may be potent antitumor effector cells. However, increasing clinical evidence shows TANs correlate with poor prognosis. The tumor microenvironment controls neutrophil recruitment and in turn TANs help tumor progression. Hence, TANs can be beneficial or detrimental to the host. It is the purpose of this review to highlight these two sides of the neutrophil coin in cancer and to describe recent studies that provide some light on the mechanisms for neutrophil recruitment to the tumor, for neutrophils supporting tumor progression, and for neutrophil activation to enhance their antitumor functions.

  15. Neutrophils in Cancer: Two Sides of the Same Coin

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    Eileen Uribe-Querol

    2015-01-01

    Full Text Available Neutrophils are the most abundant leukocytes in blood and are considered to be the first line of defense during inflammation and infections. In addition, neutrophils are also found infiltrating many types of tumors. Tumor-associated neutrophils (TANs have relevant roles in malignant disease. Indeed neutrophils may be potent antitumor effector cells. However, increasing clinical evidence shows TANs correlate with poor prognosis. The tumor microenvironment controls neutrophil recruitment and in turn TANs help tumor progression. Hence, TANs can be beneficial or detrimental to the host. It is the purpose of this review to highlight these two sides of the neutrophil coin in cancer and to describe recent studies that provide some light on the mechanisms for neutrophil recruitment to the tumor, for neutrophils supporting tumor progression, and for neutrophil activation to enhance their antitumor functions.

  16. Role of β1 Integrin in Tissue Homing of Neutrophils During Sepsis

    Science.gov (United States)

    Sarangi, Pranita P.; Hyun, Young-Min; Lerman, Yelena V.; Pietropaoli, Anthony P.; Kim, Minsoo

    2012-01-01

    Aberrant activation of neutrophils during sepsis results in the widespread release of pro-inflammatory mediators, leading to multi-organ system failure and death. However, aberrant activation of neutrophils during sepsis results in the widespread release of harmful inflammatory mediators causing host tissue injuries that can lead to multi organ system failure and death. One of the pivotal components of neutrophil migration during inflammation is the expression of surface integrins. In this study, we show that administration of a cyclic analog of RGD peptide (Arg-Gly-Asp) significantly reduced the number of tissue-invading neutrophils and the degree of sepsis-induced lethality in mice as compared to control peptide. Secondly, β1 integrin (CD29) was highly up-regulated on the neutrophils isolated from both septic patients and animals. Finally, conditional genetic ablation of β1 integrin from granulocytes also improved survival and bacterial clearance in septic animals Thus, our results indicate that expression of β1 integrin is important for modulating neutrophil trafficking during sepsis, and that therapeutics designed against β1 integrins may be beneficial. PMID:22683734

  17. Neutrophil targeted nano-drug delivery system for chronic obstructive lung diseases.

    Science.gov (United States)

    Vij, Neeraj; Min, Taehong; Bodas, Manish; Gorde, Aakruti; Roy, Indrajit

    2016-11-01

    The success of drug delivery to target airway cell(s) remains a significant challenge due to the limited ability of nanoparticle (NP) systems to circumvent protective airway-defense mechanisms. The size, density, surface and physical-chemical properties of nanoparticles are the key features that determine their ability to navigate across the airway-barrier. We evaluated here the efficacy of a PEGylated immuno-conjugated PLGA-nanoparticle (PINP) to overcome this challenge and selectively deliver drug to specific inflammatory cells (neutrophils). We first characterized the size, shape, surface-properties and neutrophil targeting using dynamic laser scattering, transmission electron microscopy and flow cytometry. Next, we assessed the efficacy of neutrophil-targeted PINPs in transporting through the airway followed by specific binding and release of drug to neutrophils. Finally, our results demonstrate the efficacy of PINP mediated non-steroidal anti-inflammatory drug-(ibuprofen) delivery to neutrophils in murine models of obstructive lung diseases, based on its ability to control neutrophilic-inflammation and resulting lung disease. Copyright © 2016 Elsevier Inc. All rights reserved.

  18. Neutrophilic respiratory tract inflammation and peripheral blood neutrophilia after grain sorghum dust extract challenge.

    Science.gov (United States)

    Von Essen, S G; O'Neill, D P; McGranaghan, S; Olenchock, S A; Rennard, S I

    1995-11-01

    To determine if inhalation of grain sorghum dust in the laboratory would cause neutrophilic upper and lower respiratory tract inflammation in human volunteers, as well as systemic signs of illness. Prospective. University of Nebraska Medical Center. Thirty normal volunteers. Inhalation challenge with 20 mL of a nebulized solution of filter-sterilized grain sorghum dust extract (GSDE). One group received prednisone, 20 mg for 2 days, prior to the challenge. Bronchoscopy with bronchoalveolar lavage (BAL) was performed 24 h after challenge, with samples collected as bronchial and alveolar fractions. Findings included visible signs of airways inflammation, quantified as the bronchitis index. The percentage of bronchial neutrophils was significantly increased in those challenged with GSDE vs the control solution, Hanks' balanced salt solution (40.3 +/- 4.5% vs 14.3 +/- 5.1%, p grain dust extract. To explain the increase in peripheral blood neutrophil counts, the capacity of the peripheral blood neutrophils to migrate in chemotaxis experiments was examined. The results demonstrate an increase in peripheral blood neutrophils and an increase in chemotactic responsiveness. Inhalation challenge with a grain dust extract causes respiratory tract inflammation and a peripheral blood neutrophilia. One reason for this may be an increase in activated peripheral blood neutrophils.

  19. 5-Lipoxygenase-Dependent Recruitment of Neutrophils and Macrophages by Eotaxin-Stimulated Murine Eosinophils

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    Ricardo Alves Luz

    2014-01-01

    Full Text Available The roles of eosinophils in antimicrobial defense remain incompletely understood. In ovalbumin-sensitized mice, eosinophils are selectively recruited to the peritoneal cavity by antigen, eotaxin, or leukotriene(LTB4, a 5-lipoxygenase (5-LO metabolite. 5-LO blockade prevents responses to both antigen and eotaxin. We examined responses to eotaxin in the absence of sensitization and their dependence on 5-LO. BALB/c or PAS mice and their mutants (5-LO-deficient ALOX; eosinophil-deficient GATA-1 were injected i.p. with eotaxin, eosinophils, or both, and leukocyte accumulation was quantified up to 24 h. Significant recruitment of eosinophils by eotaxin in BALB/c, up to 24 h, was accompanied by much larger numbers of recruited neutrophils and monocytes/macrophages. These effects were abolished by eotaxin neutralization and 5-LO-activating protein inhibitor MK886. In ALOX (but not PAS mice, eotaxin recruitment was abolished for eosinophils and halved for neutrophils. In GATA-1 mutants, eotaxin recruited neither neutrophils nor macrophages. Transfer of eosinophils cultured from bone-marrow of BALB/c donors, or from ALOX donors, into GATA-1 mutant recipients, i.p., restored eotaxin recruitment of neutrophils and showed that the critical step dependent on 5-LO is the initial recruitment of eosinophils by eotaxin, not the secondary neutrophil accumulation. Eosinophil-dependent recruitment of neutrophils in naive BALB/c mice was associated with increased binding of bacteria.

  20. Structural analysis of the receptors for granulocyte colony-stimulating factor on neutrophils

    International Nuclear Information System (INIS)

    Hanazono, Y.; Hosoi, T.; Kuwaki, T.; Matsuki, S.; Miyazono, K.; Miyagawa, K.; Takaku, F.

    1990-01-01

    We investigated granulocyte colony-stimulating factor (G-CSF) receptors on neutrophils from three patients with chronic myelogenous leukemia (CML) in the chronic phase, in comparison with four normal volunteers. Because we experienced some difficulties in radioiodinating intact recombinant human G-CSF, we developed a new derivative of human G-CSF termed YPY-G-CSF. It was easy to iodinate this protein using the lactoperoxidase method because of two additional tyrosine residues, and its radioactivity was higher than that previously reported. The biological activity of YPY-G-CSF as G-CSF was fully retained. Scatchard analysis demonstrated that CML neutrophils had a single class of binding sites (1400 +/- 685/cell) with a dissociation constant (Kd) of 245 +/- 66 pM. The number of sites and Kd value of CML neutrophils were not significantly different from those of normal neutrophils (p greater than 0.9). Cross-linking studies revealed two specifically labeled bands of [125I]YPY-G-CSF-receptor complexes with apparent molecular masses of 160 and 110 kd on both normal and CML neutrophils. This is the first report describing two receptor proteins on neutrophils. According to the analyses of the proteolytic process of these cross-linked complexes and proteolytic mapping, we assume that alternative splicing or processing from a single gene may generate two distinct receptor proteins that bind specifically to G-CSF but have different fates in intracellular metabolism

  1. Synergic production of neutrophil chemotactic activity by colonic epithelial cells and eosinophils.

    Science.gov (United States)

    Dent, Gordon; Loweth, Sam C; Hasan, Anwar Matar; Leslie, Fiona M

    2014-10-01

    The presence of eosinophils in the lumen and mucosa of the intestine is characteristic of both ulcerative colitis (UC) and Crohn's disease (CD). There is evidence of eosinophil activation in the intestine during acute inflammatory episodes of these diseases; these episodes are also characterized by an influx of neutrophils, which have the potential to cause extensive tissue damage. We undertook a study to determine whether eosinophils in contact with colonic epithelial cells produce factors that may attract neutrophils in response to immunological stimulation. Neutrophil chemotactic activity (NCA) and concentrations of three neutrophil-attracting CXC chemokines - CXCL1 (Groα), CXCL5 (Ena78) and CXCL8 (IL8) - were measured in supernatants of T84 colonic epithelial cells and blood eosinophils or eosinophil-like myeloid leukaemia cells (AML14.3D10), alone or in combination. Cells were stimulated with serum-opsonized zymosan (OZ) particles. NCA (Peosinophil co-cultures were significantly higher than in the supernatants of either cell type alone. Release of CXCL1 (Peosinophils but not higher than from OZ-stimulated epithelial cells. Eosinophils and colonic epithelial cells exhibit synergy in production of neutrophil chemoattractants in response to immunological stimulation. This may represent a mechanism for exaggerated recruitment of neutrophils to the intestine in response to acute infection in conditions that are characterized by the presence of eosinophils in the bowel. Copyright © 2014 Elsevier GmbH. All rights reserved.

  2. Survival of Peripheral Blood Neutrophil Following Treatment with Soluble Factors from Rat Mesenchymal Stem Cells

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    S Hamounnavard

    2014-11-01

    Full Text Available Introduction: Mesenchymal stem cells have immunomodulatory properties and own extensive potentials to proliferate and differentiate into different cell lineages. Thus, this study was conducted to investigate the effect of supernatant of rat MSCs on the neutrophils viability. Methods: MSCs was isolated from femoral and tibial bone marrow of rat (6-8 weeks and was cultured in DMEM. After maturation of MSCs, its supernatant was incubated with neutrophils isolated from peripheral blood of rat at 37 ° C for 1 h. Neutrophil survival was measured at 6 and 24 h incubation with supernatant of MSCs by flow cytometric analysis using An/PI. Data were analyzed by one-way ANOVA followed by Tukey test (P˂0.05. Results: 6-hour incubation of neutrophils with supernatant of MSCs significantly increased the healthy cells percentage and significantly decreased the amount of necrosis (P˂0.05, but no significant decrease was observed in regard with apoptosis compared to the controls (P˃0.05. The 24-hour incubation of neutrophils with cell supernatant significantly increased the percentage of healthy cells and apoptosis was significantly reduced compared to the control group (P˂0.05. Moreover, a reduction in cell necrosis was not significant in the treated groups compared to the control (P˃0.05. Conclusions: In addition to the clinical importance of MSCs, their biological aspects are of great potential for cell therapy, such as self-renewal, proliferation and immune modulatory effects.

  3. Human neutrophils facilitate tumor cell transendothelial migration.

    LENUS (Irish Health Repository)

    Wu, Q D

    2012-02-03

    Tumor cell extravasation plays a key role in tumor metastasis. However, the precise mechanisms by which tumor cells migrate through normal vascular endothelium remain unclear. In this study, using an in vitro transendothelial migration model, we show that human polymorphonuclear neutrophils (PMN) assist the human breast tumor cell line MDA-MB-231 to cross the endothelial barrier. We found that tumor-conditioned medium (TCM) downregulated PMN cytocidal function, delayed PMN apoptosis, and concomitantly upregulated PMN adhesion molecule expression. These PMN treated with TCM attached to tumor cells and facilitated tumor cell migration through different endothelial monolayers. In contrast, MDA-MB-231 cells alone did not transmigrate. FACScan analysis revealed that these tumor cells expressed high levels of intercellular adhesion molecule-1 (ICAM-1) but did not express CD11a, CD11b, or CD18. Blockage of CD11b and CD18 on PMN and of ICAM-1 on MDA-MB-231 cells significantly attenuated TCM-treated, PMN-mediated tumor cell migration. These tumor cells still possessed the ability to proliferate after PMN-assisted transmigration. These results indicate that TCM-treated PMN may serve as a carrier to assist tumor cell transendothelial migration and suggest that tumor cells can exploit PMN and alter their function to facilitate their extravasation.

  4. Neutrophil Leukocyte: Combustive Microbicidal Action and Chemiluminescence

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    Robert C. Allen

    2015-01-01

    Full Text Available Neutrophil leukocytes protect against a varied and complex array of microbes by providing microbicidal action that is simple, potent, and focused. Neutrophils provide such action via redox reactions that change the frontier orbitals of oxygen (O2 facilitating combustion. The spin conservation rules define the symmetry barrier that prevents direct reaction of diradical O2 with nonradical molecules, explaining why combustion is not spontaneous. In burning, the spin barrier is overcome when energy causes homolytic bond cleavage producing radicals capable of reacting with diradical O2 to yield oxygenated radical products that further participate in reactive propagation. Neutrophil mediated combustion is by a different pathway. Changing the spin quantum state of O2 removes the symmetry restriction to reaction. Electronically excited singlet molecular oxygen (O2*1 is a potent electrophilic reactant with a finite lifetime that restricts its radius of reactivity and focuses combustive action on the target microbe. The resulting exergonic dioxygenation reactions produce electronically excited carbonyls that relax by light emission, that is, chemiluminescence. This overview of neutrophil combustive microbicidal action takes the perspectives of spin conservation and bosonic-fermionic frontier orbital considerations. The necessary principles of particle physics and quantum mechanics are developed and integrated into a fundamental explanation of neutrophil microbicidal metabolism.

  5. House Dust Mite Allergen Regulates Constitutive Apoptosis of Normal and Asthmatic Neutrophils via Toll-Like Receptor 4.

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    Do Hyung Kim

    Full Text Available House dust mites (HDMs induce allergic diseases such as asthma. Neutrophil apoptosis is an important process of innate immunity, and its dysregulation is associated with asthma. In this study, we examined the effects of HDM on constitutive apoptosis of normal and asthmatic neutrophils. Extract of Dermatophagoides pteronissinus (DP inhibited neutrophil apoptosis, but Dermatophagoides farinae extract had no effect. Anti-apoptotic signaling mediated by DP involves in TLR4, Lyn, PI3K, Akt, ERK, and NF-κB in normal neutrophils. DP delayed cleavage of procaspase 9 and procaspase 3 and the decrease in Mcl-1 expression. Supernatant collected from DP-treated normal neutrophils inhibited the constitutive apoptosis of normal neutrophils, and S100A8 and S100A9 were identified as anti-apoptotic proteins in the supernatant. S100A8 and S100A9 transduced the anti-apoptotic signal via TLR4, Lyn, PI3K, Akt, ERK, and NF-κB. DP also suppressed asthmatic neutrophil apoptosis and induced secretion of S100A8 and S100A9, which delayed the constitutive apoptosis. The anti-apoptotic effects of DP, S100A8 and S100A9 in asthmatic neutrophils are associated with TLR4, Lyn, PI3K, Akt, ERK, and NF-κB. The concentrations of S100A8 and S100A9 were significantly elevated in asthmatic bronchoalveolar lavage fluid (BALF when compared to normal BALF (p<0.01, but not in serum. S100A8 concentration in BALF was positively correlated with the number of BALF neutrophils and negatively correlated with FEV1(%. These findings improve our understanding of the role of HDM in regulation of neutrophil apoptosis in normal individuals and asthmatics and will enable elucidation of asthma pathogenesis.

  6. Mycobacterium tuberculosis Cell Wall Fragments Released upon Bacterial Contact with the Human Lung Mucosa Alter the Neutrophil Response to Infection.

    Science.gov (United States)

    Scordo, Julia M; Arcos, Jesús; Kelley, Holden V; Diangelo, Lauren; Sasindran, Smitha J; Youngmin, Ellie; Wewers, Mark D; Wang, Shu-Hua; Balada-Llasat, Joan-Miquel; Torrelles, Jordi B

    2017-01-01

    In 2016, the World Health Organization reported that one person dies of tuberculosis (TB) every 21 s. A host environment that Mycobacterium tuberculosis ( M.tb ) finds during its route of infection is the lung mucosa bathing the alveolar space located in the deepest regions of the lungs. We published that human lung mucosa, or alveolar lining fluid (ALF), contains an array of hydrolytic enzymes that can significantly alter the M.tb surface during infection by cleaving off parts of its cell wall. This interaction results in two different outcomes: modifications on the M.tb cell wall surface and release of M.tb cell wall fragments into the environment. Typically, one of the first host immune cells at the site of M.tb infection is the neutrophil. Neutrophils can mount an extracellular and intracellular innate immune response to M.tb during infection. We hypothesized that exposure of neutrophils to ALF-induced M.tb released cell wall fragments would prime neutrophils to control M.tb infection better. Our results show that ALF fragments activate neutrophils leading to an increased production of inflammatory cytokines and oxidative radicals. However, neutrophil exposure to these fragments reduces production of chemoattractants (i.e., interleukin-8), and degranulation, with the subsequent reduction of myeloperoxidase release, and does not induce cytotoxicity. Unexpectedly, these ALF fragment-derived modulations in neutrophil activity do not further, either positively or negatively, contribute to the intracellular control of M.tb growth during infection. However, secreted products from neutrophils primed with ALF fragments are capable of regulating the activity of resting macrophages. These results indicate that ALF-induced M.tb fragments could further contribute to the control of M.tb growth and local killing by resident neutrophils by switching on the total oxidative response and limiting migration of neutrophils to the infection site.

  7. Th-17 regulatory cytokines IL-21, IL-23, and IL-6 enhance neutrophil production of IL-17 cytokines during asthma.

    Science.gov (United States)

    Halwani, Rabih; Sultana, Asma; Vazquez-Tello, Alejandro; Jamhawi, Amer; Al-Masri, Abeer A; Al-Muhsen, Saleh

    2017-11-01

    In a subset of severe asthma patients, chronic airway inflammation is associated with infiltration of neutrophils, Th-17 cells and elevated expression of Th-17-derived cytokines (e.g., interleukin [IL]-17, IL-21, IL-22). Peripheral neutrophils from allergic asthmatics are known to express higher IL-17 cytokine levels than those from healthy subjects, but the regulatory mechanisms involved are not well understood. We hypothesize that Th-17 regulatory cytokines could modulate IL-17 expression in neutrophils. Peripheral blood neutrophils isolated from asthmatics were stimulated with IL-21, IL-23, and IL-6 cytokines and their ability to produce IL-17A and IL-17F was determined relative to healthy controls. Signal transducer and activator of transcription 3 (STAT3) phosphorylation levels were measured in stimulated neutrophil using flow cytometry. The requirement for STAT3 phosphorylation was determined by blocking its activation using a specific chemical inhibitor. Stimulating asthmatic neutrophils with IL-21, 23, and 6 enhanced the production of IL-17A and IL-17F at significantly higher levels comparatively to healthy controls. Stimulating neutrophils with IL-21, IL-23, and IL-6 cytokines enhanced STAT3 phosphorylation, in all cases. Interestingly, inhibiting STAT3 phosphorylation using a specific chemical inhibitor dramatically blocked the ability of neutrophils to produce IL-17, demonstrating that STAT3 activation is the major factor mediating IL-17 gene expression. These findings suggest that neutrophil infiltration in lungs of severe asthmatics may represent an important source of pro-inflammatory IL-17A and -F cytokines, a production enhanced by Th-17 regulatory cytokines, and thus providing a feedback mechanism that sustains inflammation. Our results suggest that STAT3 pathway could be a potential target for regulating neutrophilic inflammation during severe asthma.

  8. Neutrophils in Homeostasis, Immunity, and Cancer.

    Science.gov (United States)

    Nicolás-Ávila, José Ángel; Adrover, José M; Hidalgo, Andrés

    2017-01-17

    Neutrophils were among the first leukocytes described and visualized by early immunologists. Prominent effector functions during infection and sterile inflammation classically placed them low in the immune tree as rapid, mindless aggressors with poor regulatory functions. This view is currently under reassessment as we uncover new aspects of their life cycle and identify transcriptional and phenotypic diversity that endows them with regulatory properties that extend beyond their lifetime in the circulation. These properties are revealing unanticipated roles for neutrophils in supporting homeostasis, as well as complex disease states such as cancer. We focus this review on these emerging functions in order to define the true roles of neutrophils in homeostasis, immunity, and disease. Copyright © 2017 Elsevier Inc. All rights reserved.

  9. Quantitative proteomics reveals differential biological processes in healthy neonatal cord neutrophils and adult neutrophils

    KAUST Repository

    Zhu, Jiang; Zhang, Huoming; Guo, Tiannan; Li, Wenying; Li, Huiyu; Zhu, Yi; Huang, Shiang

    2014-01-01

    Neonatal neutrophils are characterized by the immaturity of bactericidal mechanisms that contributes largely to neonatal mortality. However, underlying molecular mechanism associated with the immaturity remains incompletely understood. In this study, we performed comparative proteomic analysis on neonatal neutrophils derived from human cord blood and adult peripheral neutrophils. A total of 1332 proteins were identified and quantified, and 127 proteins were characterized as differentially expressed between adult and cord neutrophils. The differentially expressed proteins are mapped in KEGG pathways into five clusters and indicated impaired functions of neonatal neutrophils in proteasome, lysosome, phagosome, and leukocyte transendothelial migration. In particular, many proteins associated with NETosis, a critical mechanism for antimicrobial process and auto-clearance, were also found to be downregulated in cord neutrophils. This study represents a first comparative proteome profiling of neonatal and adult neutrophils, and provides a global view of differentially expressed proteome for enhancing our understanding of their various functional difference. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  10. GROUP B STREPTOCOCCUS CIRCUMVENTS NEUTROPHILS AND NEUTROPHIL EXTRACELLULAR TRAPS DURING AMNIOTIC CAVITY INVASION AND PRETERM LABOR

    Science.gov (United States)

    Boldenow, Erica; Gendrin, Claire; Ngo, Lisa; Bierle, Craig; Vornhagen, Jay; Coleman, Michelle; Merillat, Sean; Armistead, Blair; Whidbey, Christopher; Alishetti, Varchita; Santana-Ufret, Veronica; Ogle, Jason; Gough, Michael; Srinouanprachanh, Sengkeo; MacDonald, James W; Bammler, Theo K; Bansal, Aasthaa; Liggitt, H. Denny; Rajagopal, Lakshmi; Waldorf, Kristina M Adams

    2016-01-01

    Preterm birth is a leading cause of neonatal morbidity and mortality. Although microbial invasion of the amniotic cavity (MIAC) is associated with the majority of early preterm births, the temporal events that occur during MIAC and preterm labor are not known. Group B Streptococci (GBS) are β-hemolytic, gram-positive bacteria, which commonly colonize the vagina but have been recovered from the amniotic fluid in preterm birth cases. To understand temporal events that occur during MIAC, we utilized a unique chronically catheterized nonhuman primate model that closely emulates human pregnancy. This model allows monitoring of uterine contractions, timing of MIAC and immune responses during pregnancy-associated infections. Here, we show that adverse outcomes such as preterm labor, MIAC, and fetal sepsis were observed more frequently during infection with hemolytic GBS when compared to nonhemolytic GBS. Although MIAC was associated with systematic progression in chorioamnionitis beginning with chorionic vasculitis and progressing to neutrophilic infiltration, the ability of the GBS hemolytic pigment toxin to induce neutrophil cell death and subvert killing by neutrophil extracellular traps (NETs) in placental membranes in vivo facilitated MIAC and fetal injury. Furthermore, compared to maternal neutrophils, fetal neutrophils exhibit decreased neutrophil elastase activity and impaired phagocytic functions to GBS. Collectively, our studies demonstrate how a unique bacterial hemolytic lipid toxin enables GBS to circumvent neutrophils and NETs in placental membranes to induce fetal injury and preterm labor. PMID:27819066

  11. Quantitative proteomics reveals differential biological processes in healthy neonatal cord neutrophils and adult neutrophils

    KAUST Repository

    Zhu, Jiang

    2014-06-11

    Neonatal neutrophils are characterized by the immaturity of bactericidal mechanisms that contributes largely to neonatal mortality. However, underlying molecular mechanism associated with the immaturity remains incompletely understood. In this study, we performed comparative proteomic analysis on neonatal neutrophils derived from human cord blood and adult peripheral neutrophils. A total of 1332 proteins were identified and quantified, and 127 proteins were characterized as differentially expressed between adult and cord neutrophils. The differentially expressed proteins are mapped in KEGG pathways into five clusters and indicated impaired functions of neonatal neutrophils in proteasome, lysosome, phagosome, and leukocyte transendothelial migration. In particular, many proteins associated with NETosis, a critical mechanism for antimicrobial process and auto-clearance, were also found to be downregulated in cord neutrophils. This study represents a first comparative proteome profiling of neonatal and adult neutrophils, and provides a global view of differentially expressed proteome for enhancing our understanding of their various functional difference. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  12. Pathogenic Bacterium Acinetobacter baumannii Inhibits the Formation of Neutrophil Extracellular Traps by Suppressing Neutrophil Adhesion

    Science.gov (United States)

    Kamoshida, Go; Kikuchi-Ueda, Takane; Nishida, Satoshi; Tansho-Nagakawa, Shigeru; Ubagai, Tsuneyuki; Ono, Yasuo

    2018-01-01

    Hospital-acquired infections caused by Acinetobacter baumannii have become problematic because of high rates of drug resistance. A. baumannii is usually harmless, but it may cause infectious diseases in an immunocompromised host. Although neutrophils are the key players of the initial immune response against bacterial infection, their interactions with A. baumannii remain largely unknown. A new biological defense mechanism, termed neutrophil extracellular traps (NETs), has been attracting attention. NETs play a critical role in bacterial killing by bacterial trapping and inactivation. Many pathogenic bacteria have been reported to induce NET formation, while an inhibitory effect on NET formation is rarely reported. In the present study, to assess the inhibition of NET formation by A. baumannii, bacteria and human neutrophils were cocultured in the presence of phorbol 12-myristate 13-acetate (PMA), and NET formation was evaluated. NETs were rarely observed during the coculture despite neutrophil PMA stimulation. Furthermore, A. baumannii prolonged the lifespan of neutrophils by inhibiting NET formation. The inhibition of NET formation by other bacteria was also investigated. The inhibitory effect was only apparent with live A. baumannii cells. Finally, to elucidate the mechanism of this inhibition, neutrophil adhesion was examined. A. baumannii suppressed the adhesion ability of neutrophils, thereby inhibiting PMA-induced NET formation. This suppression of cell adhesion was partly due to suppression of the surface expression of CD11a in neutrophils. The current study constitutes the first report on the inhibition of NET formation by a pathogenic bacterium, A. baumannii, and prolonging the neutrophil lifespan. This novel pathogenicity to inhibit NET formation, thereby escaping host immune responses might contribute to a development of new treatment strategies for A. baumannii infections. PMID:29467765

  13. Pathogenic Bacterium Acinetobacter baumannii Inhibits the Formation of Neutrophil Extracellular Traps by Suppressing Neutrophil Adhesion

    Directory of Open Access Journals (Sweden)

    Go Kamoshida

    2018-02-01

    Full Text Available Hospital-acquired infections caused by Acinetobacter baumannii have become problematic because of high rates of drug resistance. A. baumannii is usually harmless, but it may cause infectious diseases in an immunocompromised host. Although neutrophils are the key players of the initial immune response against bacterial infection, their interactions with A. baumannii remain largely unknown. A new biological defense mechanism, termed neutrophil extracellular traps (NETs, has been attracting attention. NETs play a critical role in bacterial killing by bacterial trapping and inactivation. Many pathogenic bacteria have been reported to induce NET formation, while an inhibitory effect on NET formation is rarely reported. In the present study, to assess the inhibition of NET formation by A. baumannii, bacteria and human neutrophils were cocultured in the presence of phorbol 12-myristate 13-acetate (PMA, and NET formation was evaluated. NETs were rarely observed during the coculture despite neutrophil PMA stimulation. Furthermore, A. baumannii prolonged the lifespan of neutrophils by inhibiting NET formation. The inhibition of NET formation by other bacteria was also investigated. The inhibitory effect was only apparent with live A. baumannii cells. Finally, to elucidate the mechanism of this inhibition, neutrophil adhesion was examined. A. baumannii suppressed the adhesion ability of neutrophils, thereby inhibiting PMA-induced NET formation. This suppression of cell adhesion was partly due to suppression of the surface expression of CD11a in neutrophils. The current study constitutes the first report on the inhibition of NET formation by a pathogenic bacterium, A. baumannii, and prolonging the neutrophil lifespan. This novel pathogenicity to inhibit NET formation, thereby escaping host immune responses might contribute to a development of new treatment strategies for A. baumannii infections.

  14. Phenotypic Diversity and Plasticity in Circulating Neutrophil Subpopulations in Cancer

    Directory of Open Access Journals (Sweden)

    Jitka Y. Sagiv

    2015-02-01

    Full Text Available Controversy surrounds neutrophil function in cancer because neutrophils were shown to provide both pro- and antitumor functions. We identified a heterogeneous subset of low-density neutrophils (LDNs that appear transiently in self-resolving inflammation but accumulate continuously with cancer progression. LDNs display impaired neutrophil function and immunosuppressive properties, characteristics that are in stark contrast to those of mature, high-density neutrophils (HDNs. LDNs consist of both immature myeloid-derived suppressor cells (MDSCs and mature cells that are derived from HDNs in a TGF-β-dependent mechanism. Our findings identify three distinct populations of circulating neutrophils and challenge the concept that mature neutrophils have limited plasticity. Furthermore, our findings provide a mechanistic explanation to mitigate the controversy surrounding neutrophil function in cancer.

  15. Effects of Ingestion of Different Amounts of Carbohydrate after Endurance Exercise on Circulating Cytokines and Markers of Neutrophil Activation

    Directory of Open Access Journals (Sweden)

    Kumpei Tanisawa

    2018-04-01

    Full Text Available We aimed to examine the effects of ingestion of different amounts of carbohydrate (CHO after endurance exercise on neutrophil count, circulating cytokine levels, and the markers of neutrophil activation and muscle damage. Nine participants completed three separate experimental trials consisting of 1 h of cycling exercise at 70% V · O2 max, followed by ingestion of 1.2 g CHO·kg body mass−1·h−1 (HCHO trial, 0.2 g CHO·kg body mass−1·h−1 (LCHO trial, or placebo (PLA trial during the 2 h recovery phase in random order. Circulating glucose, insulin, and cytokine levels, blood cell counts, and the markers of neutrophil activation and muscle damage were measured. The concentrations of plasma glucose and serum insulin at 1 h after exercise were higher in the HCHO trial than in the LCHO and PLA trials. Although there were significant main effects of time on several variables, including neutrophil count, cytokine levels, and the markers of neutrophil activation and muscle damage, significant time × trial interactions were not observed for any variables. These results suggest that CHO ingestion after endurance exercise does not enhance exercise-induced increase in circulating neutrophil and cytokine levels and markers of neutrophil activation and muscle damage, regardless of the amount of CHO ingested.

  16. Enhanced {sup 18}F-FDG uptake in activated neutrophils is unaffected by respiratory burst inhibition with RGD

    Energy Technology Data Exchange (ETDEWEB)

    Paik, J. Y.; Lee, K. H.; Go, B. H.; Jeong, K. H.; Kim, H. K.; Choi, J. S.; Choi, Y.; Kim, P. T [Samsung Medical Center, Seoul (Korea, Republic of)

    2004-07-01

    Respiratory burst generation is an important response of activated neutrophils and is associated with enhanced glucose metabolism. Since such activation in dependent on adhesion through integrins, we investigated how integrin occupation with RGD influences respiratory burst response and {sup 18}F-FDG uptake in neutrophils. Human neutrophils separated from healthy volunteers were incubated in RPMI media. For RGD peptide inhibitory experiments, neutrophils were preincubated with 200 {mu} g/ml of cRGD peptides ad 37.deg. for 2 hr prior. Respiratory burst generation and uptake of {sup 18}F-FDG was then measured with or without PMA stimulation. Cellular total hexokinase levels were assayed with a colorimetric method. Treatment with RGD in the basal state resulted in a significant but relatively small increase in neutrophil superoxide release to 1.5{+-}0.25 fold o control levels (p<0.005). Whereas PMA stimulation caused a marked increase in superoxide generation, pretreatment with RGD caused a significant attenuation of this response to 35.6{+-}0.2% (p<0.005). PMA stimulation resulted in a significant increase in {sup 18}F-FDG uptake. However, unlike the attenution of superoxide generation, neutrophils pretreated with RGD before PMA stimulation showed an identical magnitude of enhanced {sup 18}F-FDG uptake (201.8{+-}20.5 of controls, p=0.0001). In addition, hexokinase levels were increased to comparable levels of approximately 1.5 fold for PMA stimulated neutrophils irrespective of RGD pretreatment. In conclusion, soluble RGD blocks stimulation of respiratory burst activation in neutrophils but does not inhibit stimulation of cellular glucose metabolism. This dissociation may contribute to maximally enhanced neutrophil FDG uptake in inflammatory lesions regardless of the occupancy of their integrin receptors.

  17. Yersinia pestis subverts the dermal neutrophil response in a mouse model of bubonic plague.

    Science.gov (United States)

    Shannon, Jeffrey G; Hasenkrug, Aaron M; Dorward, David W; Nair, Vinod; Carmody, Aaron B; Hinnebusch, B Joseph

    2013-08-27

    The majority of human Yersinia pestis infections result from introduction of bacteria into the skin by the bite of an infected flea. Once in the dermis, Y. pestis can evade the host's innate immune response and subsequently disseminate to the draining lymph node (dLN). There, the pathogen replicates to large numbers, causing the pathognomonic bubo of bubonic plague. In this study, several cytometric and microscopic techniques were used to characterize the early host response to intradermal (i.d.) Y. pestis infection. Mice were infected i.d. with fully virulent or attenuated strains of dsRed-expressing Y. pestis, and tissues were analyzed by flow cytometry. By 4 h postinfection, there were large numbers of neutrophils in the infected dermis and the majority of cell-associated bacteria were associated with neutrophils. We observed a significant effect of the virulence plasmid (pCD1) on bacterial survival and neutrophil activation in the dermis. Intravital microscopy of i.d. Y. pestis infection revealed dynamic interactions between recruited neutrophils and bacteria. In contrast, very few bacteria interacted with dendritic cells (DCs), indicating that this cell type may not play a major role early in Y. pestis infection. Experiments using neutrophil depletion and a CCR7 knockout mouse suggest that dissemination of Y. pestis from the dermis to the dLN is not dependent on neutrophils or DCs. Taken together, the results of this study show a very rapid, robust neutrophil response to Y. pestis in the dermis and that the virulence plasmid pCD1 is important for the evasion of this response. Yersinia pestis remains a public health concern today because of sporadic plague outbreaks that occur throughout the world and the potential for its illegitimate use as a bioterrorism weapon. Since bubonic plague pathogenesis is initiated by the introduction of Y. pestis into the skin, we sought to characterize the response of the host's innate immune cells to bacteria early after

  18. Effect of Weightlessness on Neutrophils and Lymphocytes of Rats

    Directory of Open Access Journals (Sweden)

    Khusi Muhammad Saqib, Zia-ur-Rahman1 and Saeed Ahmad Nagra2

    2012-01-01

    Full Text Available In the present study, two hundreds and forty healthy albino young (n=120 and old (n=120 rats were used during winter and summer season. Rats were divided into four groups in each season i.e. young and old, consisting of male (n=30 and female (n=30 in each age category. In each age  sex matched rats, three subgroups were made and have been given the name as cage control (CC group, horizontal restrained group (HR and head down suspended (HDS group. For winter season, the room temperature of experimental period ranged from 20 to 23°C and for summer season, the experimental room temperature ranged from 30 to 33°C. A 12 hours light/12 hours dark cycle with ad libitum food offered each day to an individual rats as well as fresh water (at normal temperature were provided every day from 9-10 h (morning Rats were decapitated on day 7th (n=5 and day 28th (n=5 of experimental period from all groups to collected the blood in a hepranized tubes for the estimation of lymphocytes and neutrophils. Appropriate statistical analysis was performed to estimate the difference between age, days, treatments and their possible interactions during each season. During winter and summer seasons, male and female rats did show a significant decrease in lymphocytes, however a significant increase in the neutrophils percent was also observed in the HR and HDS groups. During summer, a significant increase in neutrophils and a decrease in lymphocytes were observed in male and female rats of HR and HDS groups.

  19. The role of phagocytosis, oxidative burst and neutrophil extracellular traps in the interaction between neutrophils and the periodontal pathogen Porphyromonas gingivalis.

    Science.gov (United States)

    Jayaprakash, K; Demirel, I; Khalaf, H; Bengtsson, T

    2015-10-01

    Neutrophils are regarded as the sentinel cells of innate immunity and are found in abundance within the gingival crevice. Discovery of neutrophil extracellular traps (NETs) within the gingival pockets prompted us to probe the nature of the interactions of neutrophils with the prominent periopathogen Porphyromonas gingivalis. Some of the noted virulence factors of this Gram-negative anaerobe are gingipains: arginine gingipains (RgpA/B) and lysine gingipain (Kgp). The aim of this study was to evaluate the role of gingipains in phagocytosis, formation of reactive oxygen species, NETs and CXCL8 modulation by using wild-type strains and isogenic gingipain mutants. Confocal imaging showed that gingipain mutants K1A (Kgp) and E8 (RgpA/B) induced extracellular traps in neutrophils, whereas ATCC33277 and W50 were phagocytosed. The viability of both ATCC33277 and W50 dwindled as the result of phagocytosis and could be salvaged by cytochalasin D, and the bacteria released high levels of lipopolysaccharide in the culture supernatant. Porphyromonas gingivalis induced reactive oxygen species and CXCL8 with the most prominent effect being that of the wild-type strain ATCC33277, whereas the other wild-type strain W50 was less effective. Quantitative real-time polymerase chain reaction revealed a significant CXCL8 expression by E8. All the tested P. gingivalis strains increased cytosolic free calcium. In conclusion, phagocytosis is the primary neutrophil response to P. gingivalis, although NETs could play an accessory role in infection control. Although gingipains do not seem to directly regulate phagocytosis, NETs or oxidative burst in neutrophils, their proteolytic properties could modulate the subsequent outcomes such as nutrition acquisition and survival by the bacteria. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  20. Neutrophil beta-2 microglobulin: an inflammatory mediator

    DEFF Research Database (Denmark)

    Bjerrum, O W; Nissen, Mogens Holst; Borregaard, N

    1990-01-01

    Beta-2 microglobulin (beta 2m) constitutes the light invariant chain of HLA class I antigen, and is a constituent of mobilizable compartments of neutrophils. Two forms of beta 2m exist: native beta 2m and proteolytically modified beta 2m (Des-Lys58-beta 2m), which shows alpha mobility in crossed ...

  1. Neutrophils in atherosclerosis. A brief overview

    NARCIS (Netherlands)

    Hartwig, H.; Silvestre Roig, C.; Daemen, M.; Lutgens, E.; Soehnlein, O.

    2015-01-01

    Atherosclerosis is a chronic inflammation of the arterial wall and the continuous infiltration of leukocytes into the plaque enhances the progression of the lesion. Because of the scarce detection of neutrophils in atherosclerotic plaques compared to other immune cells, their contribution was

  2. Myeloperoxidase attracts neutrophils by physical forces

    Czech Academy of Sciences Publication Activity Database

    Klinke, A.; Nussbaum, C.; Kubala, Lukáš; Friedrichs, K.; Rudolph, T.K.; Rudolph, V.; Paust, H.-J.; Schröder, Ch.; Benten, D.; Lau, D.; Szocs, K.; Furtmüller, P.G.; Heeringa, P.; Sydow, K.; Duchstein, H.-J.; Ehmke, H.; Schumacher, U.; Meinertz, T.; Sperandio, M.; Baldus, S.

    2011-01-01

    Roč. 117, č. 4 (2011), s. 1350-1358 ISSN 0006-4971 Institutional research plan: CEZ:AV0Z50040507; CEZ:AV0Z50040702 Keywords : myeloperoxidase * polymorphonuclear neutrophils * glycocalyx Subject RIV: BO - Biophysics Impact factor: 9.898, year: 2011

  3. Rat Neutrophil Phagocytosis Following Feed Restriction

    Czech Academy of Sciences Publication Activity Database

    Slapničková, Martina; Berger, J.

    2002-01-01

    Roč. 11, č. 3 (2002), s. 172-177 ISSN 0938-7714 Institutional research plan: CEZ:AV0Z5052915 Keywords : circulating neutrophil * diet restriction * phagocytosis Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 0.167, year: 2001

  4. Circumventing Y. pestis Virulence by Early Recruitment of Neutrophils to the Lungs during Pneumonic Plague.

    Directory of Open Access Journals (Sweden)

    Yaron Vagima

    2015-05-01

    Full Text Available Pneumonic plague is a fatal disease caused by Yersinia pestis that is associated with a delayed immune response in the lungs. Because neutrophils are the first immune cells recruited to sites of infection, we investigated the mechanisms responsible for their delayed homing to the lung. During the first 24 hr after pulmonary infection with a fully virulent Y. pestis strain, no significant changes were observed in the lungs in the levels of neutrophils infiltrate, expression of adhesion molecules, or the expression of the major neutrophil chemoattractants keratinocyte cell-derived chemokine (KC, macrophage inflammatory protein 2 (MIP-2 and granulocyte colony stimulating factor (G-CSF. In contrast, early induction of chemokines, rapid neutrophil infiltration and a reduced bacterial burden were observed in the lungs of mice infected with an avirulent Y. pestis strain. In vitro infection of lung-derived cell-lines with a YopJ mutant revealed the involvement of YopJ in the inhibition of chemoattractants expression. However, the recruitment of neutrophils to the lungs of mice infected with the mutant was still delayed and associated with rapid bacterial propagation and mortality. Interestingly, whereas KC, MIP-2 and G-CSF mRNA levels in the lungs were up-regulated early after infection with the mutant, their protein levels remained constant, suggesting that Y. pestis may employ additional mechanisms to suppress early chemoattractants induction in the lung. It therefore seems that prevention of the early influx of neutrophils to the lungs is of major importance for Y. pestis virulence. Indeed, pulmonary instillation of KC and MIP-2 to G-CSF-treated mice infected with Y. pestis led to rapid homing of neutrophils to the lung followed by a reduction in bacterial counts at 24 hr post-infection and improved survival rates. These observations shed new light on the virulence mechanisms of Y. pestis during pneumonic plague, and have implications for the

  5. Streptococcus sanguinis induces neutrophil cell death by production of hydrogen peroxide.

    Science.gov (United States)

    Sumioka, Ryuichi; Nakata, Masanobu; Okahashi, Nobuo; Li, Yixuan; Wada, Satoshi; Yamaguchi, Masaya; Sumitomo, Tomoko; Hayashi, Mikako; Kawabata, Shigetada

    2017-01-01

    Streptococcus is the dominant bacterial genus in the human oral cavity and a leading cause of infective endocarditis. Streptococcus sanguinis belongs to the mitis group of streptococci and produces hydrogen peroxide (H2O2) by the action of SpxB, a pyruvate oxidase. In this study, we investigated the involvement of SpxB in survival of S. sanguinis in human blood and whether bacterial H2O2 exhibits cytotoxicity against human neutrophils. Results of a bactericidal test with human whole blood revealed that the spxB mutation in S. sanguinis is detrimental to its survival in blood. When S. sanguinis strains were exposed to isolated neutrophils, the bacterial survival rate was significantly decreased by spxB deletion. Furthermore, human neutrophils exposed to the S. sanguinis wild-type strain, in contrast to those exposed to an spxB mutant strain, underwent cell death with chromatin de-condensation and release of web-like extracellular DNA, reflecting induction of neutrophil extracellular traps (NETs). Since reactive oxygen species-mediated NET induction requires citrullination of arginine residues in histone proteins and subsequent chromatin de-condensation, we examined citrullination levels of histone in infected neutrophils. It is important to note that the citrullinated histone H3 was readily detected in neutrophils infected with the wild-type strain, as compared to infection with the spxB mutant strain. Moreover, decomposition of streptococcal H2O2 with catalase reduced NET induction. These results suggest that H2O2 produced by S. sanguinis provokes cell death of neutrophils and NET formation, thus potentially affecting bacterial survival in the bloodstream.

  6. Streptococcus sanguinis induces neutrophil cell death by production of hydrogen peroxide.

    Directory of Open Access Journals (Sweden)

    Ryuichi Sumioka

    Full Text Available Streptococcus is the dominant bacterial genus in the human oral cavity and a leading cause of infective endocarditis. Streptococcus sanguinis belongs to the mitis group of streptococci and produces hydrogen peroxide (H2O2 by the action of SpxB, a pyruvate oxidase. In this study, we investigated the involvement of SpxB in survival of S. sanguinis in human blood and whether bacterial H2O2 exhibits cytotoxicity against human neutrophils. Results of a bactericidal test with human whole blood revealed that the spxB mutation in S. sanguinis is detrimental to its survival in blood. When S. sanguinis strains were exposed to isolated neutrophils, the bacterial survival rate was significantly decreased by spxB deletion. Furthermore, human neutrophils exposed to the S. sanguinis wild-type strain, in contrast to those exposed to an spxB mutant strain, underwent cell death with chromatin de-condensation and release of web-like extracellular DNA, reflecting induction of neutrophil extracellular traps (NETs. Since reactive oxygen species-mediated NET induction requires citrullination of arginine residues in histone proteins and subsequent chromatin de-condensation, we examined citrullination levels of histone in infected neutrophils. It is important to note that the citrullinated histone H3 was readily detected in neutrophils infected with the wild-type strain, as compared to infection with the spxB mutant strain. Moreover, decomposition of streptococcal H2O2 with catalase reduced NET induction. These results suggest that H2O2 produced by S. sanguinis provokes cell death of neutrophils and NET formation, thus potentially affecting bacterial survival in the bloodstream.

  7. Nicotine and caffeine alter the effects of the LPS- primed mesenchymal stem cells on the co-cultured neutrophils.

    Science.gov (United States)

    Abbasi, Ardeshir; Kukia, Nasim Rahmani; Froushani, Seyyed Meysam Abtahi; Hashemi, Seyed Mahmoud

    2018-04-15

    Mesenchymal stem cells (MSCs) express some of the nicotinic receptor subunits and adenosine receptors. The communication between tissue MSCs with neutrophils has been shown in previous studies. The aim of the present study is to determine the role of nicotine or caffeine on MSCs and its effects on neutrophils. After the isolation, MSCs were pulsed with LPS (10 ng/ml) for 1 h. Then, MSCs were incubated with different concentrations of caffeine (0.1, 0.5 and 1 mM) and or with different concentrations of nicotine (0.1, 0.5, and 1 μM) for 48 h. Afterwards, the medium was aspirated and the cells were used for co-culture experiment with neutrophil. The obtained data showed that LPS primed MSCs could decrease neutrophil vitality, whereas the treatment of MSCs with nicotine and/or especially a treatment with caffeine reverse this effect. Obtained data showed that when the LPS-primed MSCs were treated with nicotine or caffeine, the vitality of co-cultured neutrophils was significantly increased. The rate of the respiratory burst of neutrophils after co-culture by LPS-primed MSCs was decreased compared to the respiratory burst of neutrophil alone. Nicotine and/or caffeine treatment could reverse this reduction. Generally, these findings provide a new insight into understanding the anti-inflammatory and immunomodulatory effects of nicotine and caffeine. Copyright © 2018 Elsevier Inc. All rights reserved.

  8. Flow Cytometric Evaluation of Human Neutrophil Apoptosis During Nitric Oxide Generation In Vitro: The Role of Exogenous Antioxidants

    Directory of Open Access Journals (Sweden)

    Zofia Sulowska

    2005-01-01

    in vitro. The effect of exogenous supply of NO donors such as SNP, SIN-1, and GEA-3162 on the course of human neutrophil apoptosis and the role of extracellular antioxidants in this process was investigated. Isolated from peripheral blood, neutrophils were cultured in the presence or absence of NO donor compounds and antioxidants for 8, 12, and 20 hours. Apoptosis of neutrophils was determined in vitro by flow cytometric analysis of cellular DNA content and Annexin V protein binding to the cell surface. Exposure of human neutrophils to GEA-3162 and SIN-1 significantly accelerates and enhances their apoptosis in vitro in a time-dependent fashion. In the presence of SNP, intensification of apoptosis has not been revealed until 12 hours after the culture. The inhibition of GEA-3162- and SIN-1-mediated neutrophil apoptosis by superoxide dismutase (SOD but not by catalase (CAT was observed. Our results show that SOD and CAT can protect neutrophils against NO-donors-induced apoptosis and suggest that the interaction of NO and oxygen metabolites signals may determine the destructive or protective role of NO donor compounds during apoptotic neutrophil death.

  9. Distinct cellular sources of hepoxilin A3 and leukotriene B4 are used to coordinate bacterial-induced neutrophil transepithelial migration.

    Science.gov (United States)

    Pazos, Michael A; Pirzai, Waheed; Yonker, Lael M; Morisseau, Christophe; Gronert, Karsten; Hurley, Bryan P

    2015-02-01

    Neutrophilic infiltration is a leading contributor to pathology in a number of pulmonary disease states, including cystic fibrosis. Hepoxilin A3 (HXA3) is a chemotactic eicosanoid shown to mediate the transepithelial passage of neutrophils in response to infection in several model systems and at multiple mucosal surfaces. Another well-known eicosanoid mediating general neutrophil chemotaxis is leukotriene B4 (LTB4). We sought to distinguish the roles of each eicosanoid in the context of infection of lung epithelial monolayers by Pseudomonas aeruginosa. Using human and mouse in vitro transwell model systems, we used a combination of biosynthetic inhibitors, receptor antagonists, as well as mutant sources of neutrophils to assess the contribution of each chemoattractant in driving neutrophil transepithelial migration. We found that following chemotaxis to epithelial-derived HXA3 signals, neutrophil-derived LTB4 is required to amplify the magnitude of neutrophil migration. LTB4 signaling is not required for migration to HXA3 signals, but LTB4 generation by migrated neutrophils plays a significant role in augmenting the initial HXA3-mediated migration. We conclude that HXA3 and LTB4 serve independent roles to collectively coordinate an effective neutrophilic transepithelial migratory response. Copyright © 2015 by The American Association of Immunologists, Inc.

  10. Changes in Neutrophil Functions in Astronauts

    Science.gov (United States)

    Kaur, Indreshpal; Simons, Elizabeth R.; Castro, Victoria; Pierson, Duane L.

    2002-01-01

    Neutrophil functions (phagocytosis, oxidative burst, degranulation) and expression of surface markers involved in these functions were studied in 25 astronauts before and after 4 space shuttle missions. Space flight duration ranged from 5 to 11 days. Blood specimens were obtained 10 days before launch (preflight or L-10), immediately after landing (landing or R+0), and again at 3 days after landing (postflight or R+3). Blood samples were also collected from 9 healthy low-stressed subjects at 3 time points simulating a 10-day shuttle mission. The number of neutrophils increased at landing by 85 percent when compared to the preflight numbers. Neutrophil functions were studied in whole blood using flow cytometric methods. Phagocytosis of E.coli-FITC and oxidative burst capacity of the neutrophils following the 9 to 11 day missions were lower at all three sampling points than the mean values for control subjects. Phagocytosis and oxidative burst capacity of the astronauts was decreased even 10-days before space flight. Mission duration appears to be a factor in phagocytic and oxidative functions. In contrast, following the short-duration (5-days) mission, these functions were unchanged from control values. No consistent changes in degranulation were observed following either short or medium length space missions. The expression of CD16, CD32, CD11a, CD11b, CD11c, L-selectin and CD36 was measured and found to be variable. Specifically, CD16 and CD32 did not correlate with the changes in oxidative burst and phagocytosis. We can conclude from this study that the stresses associated with space flight can alter the important functions of neutrophils.

  11. In vivo effects of dexamethasone and indomethacin on neutrophil-induced alterations of nasal epithelial mucosubstances

    International Nuclear Information System (INIS)

    Hotchkiss, J.A.; Portereiko, J.V.; Harkema, J.R.

    1988-01-01

    Previous studies have shown that neutrophils migrating through rat nasal mucosal epithelium, in response to intranasal instillation of endotoxin, induce a transient decrease in stored epithelial mucosubstances. Prostaglandins and leukotrienes can either increase or decrease mucous secretion of airway epithelia in vitro. In this study, rats were treated with indomethacin a specific inhibitor of prostaglandin synthesis, or with dexamethasone, a general inhibitor of arachidonic acid metabolism, and challenged with intranasally instilled endotoxin. Dexamethasone alone or in combination with indomethacin, but not indomethacin alone, significantly altered the neutrophil response to intranasally instilled endotoxin and may have inhibited the neutrophil-induced decrease in stored mucosubstances. These data suggest that leukotrienes and possibly prostaglandins play a significant role in the coordinated response of the nasal mucosal epitholium to an acute inflammatory stimulus. (author)

  12. In vivo effects of dexamethasone and indomethacin on neutrophil-induced alterations of nasal epithelial mucosubstances

    Energy Technology Data Exchange (ETDEWEB)

    Hotchkiss, J A; Portereiko, J V; Harkema, J R

    1988-12-01

    Previous studies have shown that neutrophils migrating through rat nasal mucosal epithelium, in response to intranasal instillation of endotoxin, induce a transient decrease in stored epithelial mucosubstances. Prostaglandins and leukotrienes can either increase or decrease mucous secretion of airway epithelia in vitro. In this study, rats were treated with indomethacin a specific inhibitor of prostaglandin synthesis, or with dexamethasone, a general inhibitor of arachidonic acid metabolism, and challenged with intranasally instilled endotoxin. Dexamethasone alone or in combination with indomethacin, but not indomethacin alone, significantly altered the neutrophil response to intranasally instilled endotoxin and may have inhibited the neutrophil-induced decrease in stored mucosubstances. These data suggest that leukotrienes and possibly prostaglandins play a significant role in the coordinated response of the nasal mucosal epitholium to an acute inflammatory stimulus. (author)

  13. Staphylococcus epidermidis strategies to avoid killing by human neutrophils.

    Directory of Open Access Journals (Sweden)

    Gordon Y C Cheung

    2010-10-01

    Full Text Available Staphylococcus epidermidis is a leading nosocomial pathogen. In contrast to its more aggressive relative S. aureus, it causes chronic rather than acute infections. In highly virulent S. aureus, phenol-soluble modulins (PSMs contribute significantly to immune evasion and aggressive virulence by their strong ability to lyse human neutrophils. Members of the PSM family are also produced by S. epidermidis, but their role in immune evasion is not known. Notably, strong cytolytic capacity of S. epidermidis PSMs would be at odds with the notion that S. epidermidis is a less aggressive pathogen than S. aureus, prompting us to examine the biological activities of S. epidermidis PSMs. Surprisingly, we found that S. epidermidis has the capacity to produce PSMδ, a potent leukocyte toxin, representing the first potent cytolysin to be identified in that pathogen. However, production of strongly cytolytic PSMs was low in S. epidermidis, explaining its low cytolytic potency. Interestingly, the different approaches of S. epidermidis and S. aureus to causing human disease are thus reflected by the adaptation of biological activities within one family of virulence determinants, the PSMs. Nevertheless, S. epidermidis has the capacity to evade neutrophil killing, a phenomenon we found is partly mediated by resistance mechanisms to antimicrobial peptides (AMPs, including the protease SepA, which degrades AMPs, and the AMP sensor/resistance regulator, Aps (GraRS. These findings establish a significant function of SepA and Aps in S. epidermidis immune evasion and explain in part why S. epidermidis may evade elimination by innate host defense despite the lack of cytolytic toxin expression. Our study shows that the strategy of S. epidermidis to evade elimination by human neutrophils is characterized by a passive defense approach and provides molecular evidence to support the notion that S. epidermidis is a less aggressive pathogen than S. aureus.

  14. Effect of moderate exercise on peritoneal neutrophils from juvenile rats.

    Science.gov (United States)

    Braz, Glauber Ruda; Ferreira, Diorginis Soares; Pedroza, Anderson Apolonio; da Silva, Aline Isabel; Sousa, Shirley Maria; Pithon-Curi, Tania Cristina; Lagranha, Claudia

    2015-09-01

    Previous studies showed that moderate exercise in adult rats enhances neutrophil function, although no studies were performed in juvenile rats. We evaluated the effects of moderate exercise on the neutrophil function in juvenile rats. Viability and neutrophils function were evaluated. Moderate exercise did not impair the viability and mitochondrial transmembrane potential of neutrophils, whereas there was greater reactive oxygen species production (164%; p < 0.001) and phagocytic capacity (29%; p < 0.05). Our results suggest that moderate exercise in juvenile rats improves neutrophil function, similar to adults.

  15. Effect of smoking on neutrophil apoptosis in chronic periodontitis: An immunohistochemical study

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    Sachin S Shivanaikar

    2013-01-01

    Full Text Available Context: Periodontal disease is caused by chronic infection inducing an inflammatory reaction leading to breakdown of tooth-supporting tissues. There are various risk factors for the disease, and smoking is one of them. Apoptosis plays a critical role in the regulation of inflammation and host immune response which helps in tissue homeostasis, and a disturbance in this is often associated with disease. The imbalance between the apoptosis and proliferation in the periodontal tissue results in periodontal disease. Neutrophils play an important role in the defense mechanism and are the most abundant immune cells in gingival inflammatory infiltrate in patients suffering from periodontal disease. Neutrophil disorders are associated with rapid destruction of periodontal tissues. Aim: To study the influence of smoking on apoptosis of neutrophils by quantifying them in the gingival connective tissue of smoking and nonsmoking subjects suffering from chronic periodontitis. Materials and Methods: Thirty gingival biopsies were harvested from 15 smoking and 15 nonsmoking subjects who suffered from chronic periodontitis. The apoptosis of neutrophils was assessed and quantified using p53 monoclonal mouse antihuman antibody. Statistical Analysis Used: Chi-square/Fisher′sexact test was used to find the significance of study parameters on a categorical scale between the two groups. Results: Neutrophil apoptosis was significantly more in the group of nonsmokers. There was no statistical difference between plaque and bleeding index, but there was a significant increase in clinical attachment loss among smokers. Conclusions: The study reveals that smoking plays a significant role in the inhibition of neutrophil apoptosis, thereby contributing to the destruction of periodontal tissues in periodontitis.

  16. Modulation and Apoptosis of Neutrophil Granulocytes by Extracorporeal Photopheresis in the Treatment of Chronic Graft-Versus-Host Disease.

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    Cindy Franklin

    Full Text Available Chronic graft-versus-host disease (cGVHD is a common side effect of allogeneic stem cell transplantation and a major cause of morbidity and mortality in affected patients. Especially skin, eyes and oral mucosa are affected. This can lead to pain and functional impairment. Extracorporeal photopheresis (ECP is an effective immunomodulatory therapy with minimal side effects but its mode of action is still largely unknown. The objective of the present study was to examine the effects of ECP on neutrophil granulocytes in patients with cGVHD. Analysis of leukocytes from cGVHD patients obtained from the ECP device during treatment showed that neutrophil granulocytes account for the majority of cells treated during ECP. Neutrophils from healthy donors treated in vitro with 8-methoxypsoralen and UVA light as well as neutrophils from buffy coats of patients with cGVHD treated by ECP showed increased apoptosis and decreased half-life. In remaining non-apoptotic cells chemoirradiation resulted in loss of activation markers and reduced effector functions. This was accompanied by an increase in extracellular arginase-1 activity. Additional comparison of neutrophils isolated from blood of cGVHD patients before and 24h after ECP revealed a decreased half-life and reduction of effector functions of post-ECP neutrophils ex vivo. These observations strongly suggest that ECP induces both apoptosis and physiological changes in neutrophils and that these changes also take place in vivo. This study is the first to show that ECP modulates apoptosis and inflammatory activity in neutrophil granulocytes, indicating that neutrophils may significantly contribute to the overall immunomodulatory effects attributed to this treatment.

  17. Targeting Neutrophilic Inflammation Using Polymersome-Mediated Cellular Delivery.

    Science.gov (United States)

    Robertson, James D; Ward, Jon R; Avila-Olias, Milagros; Battaglia, Giuseppe; Renshaw, Stephen A

    2017-05-01

    Neutrophils are key effector cells in inflammation and play an important role in neutralizing invading pathogens. During inflammation resolution, neutrophils undergo apoptosis before they are removed by macrophages, but if apoptosis is delayed, neutrophils can cause extensive tissue damage and chronic disease. Promotion of neutrophil apoptosis is a potential therapeutic approach for treating persistent inflammation, yet neutrophils have proven difficult cells to manipulate experimentally. In this study, we deliver therapeutic compounds to neutrophils using biocompatible, nanometer-sized synthetic vesicles, or polymersomes, which are internalized by binding to scavenger receptors and subsequently escape the early endosome through a pH-triggered disassembly mechanism. This allows polymersomes to deliver molecules into the cell cytosol of neutrophils without causing cellular activation. After optimizing polymersome size, we show that polymersomes can deliver the cyclin-dependent kinase inhibitor (R)-roscovitine into human neutrophils to promote apoptosis in vitro. Finally, using a transgenic zebrafish model, we show that encapsulated (R)-roscovitine can speed up inflammation resolution in vivo more efficiently than the free drug. These results show that polymersomes are effective intracellular carriers for drug delivery into neutrophils. This has important consequences for the study of neutrophil biology and the development of neutrophil-targeted therapeutics. Copyright © 2017 The Authors.

  18. Spontaneous neutrophil migration patterns during sepsis after major burns.

    Science.gov (United States)

    Jones, Caroline N; Moore, Molly; Dimisko, Laurie; Alexander, Andrew; Ibrahim, Amir; Hassell, Bryan A; Warren, H Shaw; Tompkins, Ronald G; Fagan, Shawn P; Irimia, Daniel

    2014-01-01

    Finely tuned to respond quickly to infections, neutrophils have amazing abilities to migrate fast and efficiently towards sites of infection and inflammation. Although neutrophils ability to migrate is perturbed in patients after major burns, no correlations have yet been demonstrated between altered migration and higher rate of infections and sepsis in these patients when compared to healthy individuals. To probe if such correlations exist, we designed microfluidic devices to quantify the neutrophil migration phenotype with high precision. Inside these devices, moving neutrophils are confined in channels smaller than the neutrophils and forced to make directional decisions at bifurcations and around posts. We employed these devices to quantify neutrophil migration across 18 independent parameters in 74 blood samples from 13 patients with major burns and 3 healthy subjects. Blinded, retrospective analysis of clinical data and neutrophil migration parameters revealed that neutrophils isolated from blood samples collected during sepsis migrate spontaneously inside the microfluidic channels. The spontaneous neutrophil migration is a unique phenotype, typical for patients with major burns during sepsis and often observed one or two days before the diagnosis of sepsis is confirmed. The spontaneous neutrophil migration phenotype is rare in patients with major burns in the absence of sepsis, and is not encountered in healthy individuals. Our findings warrant further studies of neutrophils and their utility for early diagnosing and monitoring sepsis in patients after major burns.

  19. Involvement of purinergic signaling on nitric oxide production by neutrophils stimulated with Trichomonas vaginalis.

    Science.gov (United States)

    Frasson, Amanda Piccoli; De Carli, Geraldo Attilio; Bonan, Carla Denise; Tasca, Tiana

    2012-03-01

    Trichomonas vaginalis is a parasite from the human urogenital tract that causes trichomonosis, the most prevalent non-viral sexually transmitted disease. The neutrophil infiltration has been considered to be primarily responsible for cytological changes observed at infection site, and the chemoattractants can play an important role in this leukocytic recruitment. Nitric oxide (NO) is one of the most widespread mediator compounds, and it is implicated in modulation of immunological mechanisms. Extracellular nucleotides and nucleosides are signaling molecules involved in several processes, including immune responses and control of leukocyte trafficking. Ectonucleoside triphosphate diphosphohydrolase members, ecto-5'-nucleotidase, and adenosine deaminase (ectoADA) have been characterized in T. vaginalis. Herein, we investigated the effects of purinergic system on NO production by neutrophils stimulated with T. vaginalis. The trophozoites were able to induce a high NO synthesis by neutrophils through iNOS pathway. The extracellular nucleotides ATP, ADP, and ATPγS (a non-hydrolyzable ATP analog) showed no significant change in NO secretion. In contrast, adenosine and its degradation product, inosine, promoted a low production of the compound. The immunosuppressive effect of adenosine upon NO release by neutrophils occurred due to adenosine A(2A) receptor activation. The ecto-5'-nucleotidase activity displayed by T. vaginalis was shown to be important in adenosine generation, indicating the efficiency of purinergic cascade. Our data suggest the influence of purinergic signaling, specifically adenosinergic system, on NO production by neutrophils in T. vaginalis infection, contributing to the immunological aspects of disease.

  20. Regulation of Discrete Functional Responses by Syk and Src Family Tyrosine Kinases in Human Neutrophils

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    Thornin Ear

    2017-01-01

    Full Text Available Neutrophils play a critical role in innate immunity and also influence adaptive immune responses. This occurs in good part through their production of inflammatory and immunomodulatory cytokines, in conjunction with their prolonged survival at inflamed foci. While a picture of the signaling machinery underlying these neutrophil responses is now emerging, much remains to be uncovered. In this study, we report that neutrophils constitutively express various Src family isoforms (STKs, as well as Syk, and that inhibition of these protein tyrosine kinases selectively hinders inflammatory cytokine generation by acting posttranscriptionally. Accordingly, STK or Syk inhibition decreases the phosphorylation of signaling intermediates (e.g., eIF-4E, S6K, and MNK1 involved in translational control. By contrast, delayed apoptosis appears to be independent of either STKs or Syk. Our data therefore significantly extend our understanding of which neutrophil responses are governed by STKs and Syk and pinpoint some signaling intermediates that are likely involved. In view of the foremost role of neutrophils in several chronic inflammatory conditions, our findings identify potential molecular targets that could be exploited for future therapeutic intervention.

  1. Solar ultraviolet irradiation induces decorin degradation in human skin likely via neutrophil elastase.

    Science.gov (United States)

    Li, Yong; Xia, Wei; Liu, Ying; Remmer, Henriette A; Voorhees, John; Fisher, Gary J

    2013-01-01

    Exposure of human skin to solar ultraviolet (UV) irradiation induces matrix metalloproteinase-1 (MMP-1) activity, which degrades type I collagen fibrils. Type I collagen is the most abundant protein in skin and constitutes the majority of skin connective tissue (dermis). Degradation of collagen fibrils impairs the structure and function of skin that characterize skin aging. Decorin is the predominant proteoglycan in human dermis. In model systems, decorin binds to and protects type I collagen fibrils from proteolytic degradation by enzymes such as MMP-1. Little is known regarding alterations of decorin in response to UV irradiation. We found that solar-simulated UV irradiation of human skin in vivo stimulated substantial decorin degradation, with kinetics similar to infiltration of polymorphonuclear (PMN) cells. Proteases that were released from isolated PMN cells degraded decorin in vitro. A highly selective inhibitor of neutrophil elastase blocked decorin breakdown by proteases released from PMN cells. Furthermore, purified neutrophil elastase cleaved decorin in vitro and generated fragments with similar molecular weights as those resulting from protease activity released from PMN cells, and as observed in UV-irradiated human skin. Cleavage of decorin by neutrophil elastase significantly augmented fragmentation of type I collagen fibrils by MMP-1. Taken together, these data indicate that PMN cell proteases, especially neutrophil elastase, degrade decorin, and this degradation renders collagen fibrils more susceptible to MMP-1 cleavage. These data identify decorin degradation and neutrophil elastase as potential therapeutic targets for mitigating sun exposure-induced collagen fibril degradation in human skin.

  2. Cervico-vaginal immunoglobulin G levels increase post-ovulation independently of neutrophils.

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    Katrein Schaefer

    Full Text Available The prevalence of sexually transmitted infections (STIs is often higher in females than in males. Although the reproductive cycle profoundly modulates local immunity in the female reproductive tract (FRT system, significant gaps in our knowledge of the immunobiology of the FRT still exist. An intriguing and frequently observed characteristic of the FRT is the predominant presence of immunoglobulin (Ig G in cervico-vaginal secretions. We show here that in the mouse, IgG accumulation was enhanced approximately 5-fold post-ovulation, and was accompanied by an influx of neutrophils into the FRT. To determine whether these two events were causally related, we performed short-term neutrophil depletion experiments at individual stages throughout the estrous cycle. Our results demonstrate that neutrophils were not necessary for cycle-dependent tissue remodeling and cycle progression and that cycle-dependent IgG accumulation occurred independent of neutrophils. We thus conclude that neutrophil influx and IgG accumulation are independent events that occur in the FRT during the reproductive cycle.

  3. Development of an experimental model of neutrophilic pulmonary response induction in mice

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    Leonardo Araújo Pinto

    2003-08-01

    Full Text Available BACKGROUND: Several lung diseases are characterized by a predominantly neutrophilic inflammation. A better understanding of the mechanisms of action of some drugs on the airway inflammation of such diseases may bring advances to the treatment. OBJECTIVE: To develop a method to induce pulmonary neutrophilic response in mice, without active infection. METHODS: Eight adult Swiss mice were used. The study group (n = 4 received an intranasal challenge with 1 x 10(12 CFU/ml of Pseudomonas aeruginosa (Psa, frozen to death. The control group (n = 4 received an intranasal challenge with saline solution. Two days after the intranasal challenge, a bron­choalveolar lavage (BAL was performed with total cell and differential cellularity counts. RESULTS: The total cell count was significantly higher in the group with Psa, as compared to the control group (median of 1.17 x 10(6 and 0.08 x 10(6, respectively, p = 0.029. In addition to this, an absolute predominance of neutrophils was found in the differential cellularity of the mice that had received the Psa challenge. CONCLUSIONS: The model of inducing a neutrophilic pulmonary disease using frost-dead bacteria was successfully developed. This neutrophilic inflammatory response induction model in Swiss mice lungs may be an important tool for testing the anti-inflammatory effect of some antimicrobial drugs on the inflammation of the lower airways.

  4. The cystic fibrosis neutrophil: a specialized yet potentially defective cell.

    LENUS (Irish Health Repository)

    Hayes, Elaine

    2012-02-01

    Cystic fibrosis (CF) is one of the commonest genetically inherited diseases in the world. It is characterized by recurrent respiratory tract infections eventually leading to respiratory failure. One of the hallmarks of this disease is a persistent and predominantly neutrophil driven inflammation. Neutrophils provide the first line of defence by killing and digesting phagocytosed bacteria and fungi, yet despite advances in our understanding of the molecular and cellular basis of CF, there remains a paradox of why recruited CF neutrophils fail to eradicate bacterial infections in the lung. This review describes mechanisms involved in neutrophil migration, microbial killing and apoptosis leading to inflammatory resolution. We discuss dysregulated neutrophil activity and consider genetic versus inflammatory neutrophil reprogramming in CF and ultimately pharmacological modulation of the CF neutrophil for therapeutic intervention.

  5. The cystic fibrosis neutrophil: a specialized yet potentially defective cell.

    LENUS (Irish Health Repository)

    Hayes, Elaine

    2011-04-01

    Cystic fibrosis (CF) is one of the commonest genetically inherited diseases in the world. It is characterized by recurrent respiratory tract infections eventually leading to respiratory failure. One of the hallmarks of this disease is a persistent and predominantly neutrophil driven inflammation. Neutrophils provide the first line of defence by killing and digesting phagocytosed bacteria and fungi, yet despite advances in our understanding of the molecular and cellular basis of CF, there remains a paradox of why recruited CF neutrophils fail to eradicate bacterial infections in the lung. This review describes mechanisms involved in neutrophil migration, microbial killing and apoptosis leading to inflammatory resolution. We discuss dysregulated neutrophil activity and consider genetic versus inflammatory neutrophil reprogramming in CF and ultimately pharmacological modulation of the CF neutrophil for therapeutic intervention.

  6. Hydrogen sulfide reduces neutrophil recruitment in hind-limb ischemia-reperfusion injury in an L-selectin and ADAM-17 dependent manner

    Science.gov (United States)

    Ball, Carissa J.; Reiffel, Alyssa J.; Chintalapani, Sathvika; Kim, Minsoo; Spector, Jason A.; King, Michael R.

    2012-01-01

    Background Reperfusion following ischemia leads to neutrophil recruitment injured tissue. Selectins and β2 integrins regulate neutrophil interaction with the endothelium during neutrophil rolling and firm adhesion. Excessive neutrophil infiltration into tissue is thought to contribute to IRI damage. NaHS mitigates the damage caused by ischemia-reperfusion injury (IRI). This study's objective was to determine the effect of hydrogen sulfide (NaHS) on neutrophil adhesion receptor expression. Methods Human neutrophils were either left untreated or incubated in 20 μM NaHS, and/or 50 μg/mL pharmacological ADAM-17 inhibitor TAPI-0; activated by IL-8, fMLP, or TNF-α; and labeled against PSGL-1, LFA-1, Mac-1 α, L-selectin and β2 integrin epitopes CBRM1/5 or KIM127 for flow cytometry. Cohorts of 3 C57BL/6 mice received an intravenous dose of saline vehicle, or 20 μM NaHS with or without 50 μg/mL TAPI-0 before unilateral tourniquet induced hind-limb ischemia for 3 hours followed by 3 hours of reperfusion. Bilateral gastrocnemius muscles were processed for histology before neutrophil infiltration quantification. Results NaHS treatment significantly increased L-selectin shedding from human neutrophils following activation by fMLP and IL-8 in an ADAM-17 dependent manner. Mice treated with NaHS to raise bloodstream concentration by 20 μM prior to ischemia or reperfusion showed a significant reduction in neutrophil recruitment into skeletal muscle tissue following tourniquet-induced hindlimb IRI. Conclusions NaHS administration results in the downregulation of L-selectin expression in activated human neutrophils. This leads to a reduction in neutrophil extravasation and tissue infiltration and may partially account for the protective effects of NaHS seen in the setting of IRI. PMID:23446563

  7. Multiple lupus-associated ITGAM variants alter Mac-1 functions on neutrophils.

    Science.gov (United States)

    Zhou, Yebin; Wu, Jianming; Kucik, Dennis F; White, Nathan B; Redden, David T; Szalai, Alexander J; Bullard, Daniel C; Edberg, Jeffrey C

    2013-11-01

    Multiple studies have demonstrated that single-nucleotide polymorphisms (SNPs) in the ITGAM locus (including the nonsynonymous SNPs rs1143679, rs1143678, and rs1143683) are associated with systemic lupus erythematosus (SLE). ITGAM encodes the protein CD11b, a subunit of the β2 integrin Mac-1. The purpose of this study was to determine the effects of ITGAM genetic variation on the biologic functions of neutrophil Mac-1. Neutrophils from ITGAM-genotyped and -sequenced healthy donors were isolated for functional studies. The phagocytic capacity of neutrophil ITGAM variants was probed with complement-coated erythrocytes, serum-treated zymosan, heat-treated zymosan, and IgG-coated erythrocytes. The adhesion capacity of ITGAM variants, in adhering to either purified intercellular adhesion molecule 1 or tumor necrosis factor α-stimulated endothelial cells, was assessed in a flow chamber. Expression levels of total CD11b and activation of CD11b were assessed by flow cytometry. Mac-1-mediated neutrophil phagocytosis, determined in cultures with 2 different complement-coated particles, was significantly reduced in individuals with nonsynonymous variant alleles of ITGAM. This reduction in phagocytosis was related to variation at either rs1143679 (in the β-propeller region) or rs1143678/rs1143683 (highly linked SNPs in the cytoplasmic/calf-1 regions). Phagocytosis mediated by Fcγ receptors was also significantly reduced in donors with variant ITGAM alleles. Similarly, firm adhesion of neutrophils was significantly reduced in individuals with variant ITGAM alleles. These functional alterations were not attributable to differences in total receptor expression or activation. The nonsynonymous ITGAM variants rs1143679 and rs1143678/rs113683 contribute to altered Mac-1 function on neutrophils. These results underscore the need to consider multiple nonsynonymous SNPs when assessing the functional consequences of ITGAM variation on immune cell processes and the risk of SLE

  8. Ozone-induced airway hyperresponsiveness in patients with asthma: role of neutrophil-derived serine proteinases.

    Science.gov (United States)

    Hiltermann, T J; Peters, E A; Alberts, B; Kwikkers, K; Borggreven, P A; Hiemstra, P S; Dijkman, J H; van Bree, L A; Stolk, J

    1998-04-01

    Proteinase inhibitors may be of potential therapeutic value in the treatment of respiratory diseases such as chronic obstructive pulmonary disease (COPD) or asthma. Our aim was to study the role of neutrophils, and neutrophil-derived serine proteinases in an acute model in patients with asthma. Exposure to ozone induces an acute neutrophilic inflammatory reaction accompanied by an increase in airway hyperresponsiveness. It is thought that these two effects of ozone are linked, and that neutrophil-derived serine proteinases (i.e. elastase) may play a role in the ozone-induced airway hyperresponsiveness. Therefore, we examined the effect of recombinant antileukoprotease (rALP), one of the major serine proteinase inhibitors in the lung, on ozone-induced changes in airway hyperresponsiveness in this model. We observed that 16 h after exposure to ozone, airway hyperresponsiveness to methacholine was increased both following placebo and rALP treatment. There was no significant difference between placebo and rALP treatment (change in area under the dose-response curve to methacholine: 117.3+/-59.0 vs 193.6+/-59.6 % fall x DD; p=.12). Moreover, the immediate decrease in FEV1 after ozone exposure was not significantly different between the two groups (placebo: -29.6+/-6.7%; rALP: -20.9+/-3.8%; p=.11). In addition, no significant differences were observed in plasma levels of fibrinogen degradation products generated by neutrophil serine proteinases before and after exposure to ozone. We conclude that neutrophil-derived serine proteinases are not important mediators for ozone-induced hyperresponsiveness.

  9. Neutrophil-to-lymphocyte ratio, calprotectin and YKL-40 in patients with chronic obstructive pulmonary disease

    DEFF Research Database (Denmark)

    Sørensen, Allan Klitgaard; Holmgaard, Dennis Back; Mygind, Lone Hagens

    2015-01-01

    BACKGROUND: Chronic obstructive pulmonary disease (COPD) is characterized by chronic inflammation and progressive decline in pulmonary function. Neutrophil-to-lymphocyte ratio (NLR), YKL-40 and calprotectin are biomarkers of inflammation and predict mortality in patients with different inflammatory...... diseases. We aimed to investigate the correlation between levels of these three biomarkers and neutrophil granulocyte and lymphocyte count in patients with moderate to very severe COPD stratified by use of systemic glucocorticoids. Furthermore, we studied the ability of these biomarkers to predict all......- and multivariate Cox regression analyses with hazard ratios (HR) and 95% confidence intervals (CI). RESULTS: Plasma calprotectin was positively correlated with neutrophil granulocyte count and NLR. No significant association was found between plasma YKL-40 and the cellular biomarkers, irrespective...

  10. Vitamin C: A Novel Regulator of Neutrophil Extracellular Trap Formation

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    Ramesh Natarajan

    2013-08-01

    Full Text Available Introduction: Neutrophil extracellular trap (NET formation was recently identified as a novel mechanism to kill pathogens. However, excessive NET formation in sepsis can injure host tissues. We have recently shown that parenteral vitamin C (VitC is protective in sepsis. Whether VitC alters NETosis is unknown. Methods: We used Gulo−/− mice as they lack the ability to synthesize VitC. Sepsis was induced by intraperitoneal infusion of a fecal stem solution (abdominal peritonitis, FIP. Some VitC deficient Gulo−/− mice received an infusion of ascorbic acid (AscA, 200 mg/kg 30 min after induction of FIP. NETosis was assessed histologically and by quantification for circulating free DNA (cf-DNA in serum. Autophagy, histone citrullination, endoplasmic reticulum (ER stress, NFκB activation and apoptosis were investigated in peritoneal PMNs. Results: Sepsis produced significant NETs in the lungs of VitC deficient Gulo−/− mice and increased circulating cf-DNA. This was attenuated in the VitC sufficient Gulo−/− mice and in VitC deficient Gulo−/− mice infused with AscA. Polymorphonuclear neutrophils (PMNs from VitC deficient Gulo−/− mice demonstrated increased activation of ER stress, autophagy, histone citrullination, and NFκB activation, while apoptosis was inhibited. VitC also significantly attenuated PMA induced NETosis in PMNs from healthy human volunteers.

  11. The neutrophil-to-lymphocyte ratio as a marker of systemic endothelial dysfunction in asymptomatic subjects.

    Science.gov (United States)

    Martínez-Urbistondo, Diego; Beltrán, Almudena; Beloqui, Oscar; Huerta, Ana

    2016-01-01

    The neutrophil-to-lymphocyte ratio has demonstrated to be a prognostic inflammatory marker in cardiovascular disease. The objective of this study is to evaluate the association between neutrophil-to-lymphocyte ratio and pathologic urinary albumin/creatinine ratio as an early marker of cardiovascular risk and systemic endothelial dysfunction, associated with microvascular disease, in asymptomatic subjects. A unicenter cross-sectional study was conducted, including 1816 asymptomatic subjects. Patients with previous cardiovascular disease, those who were treated with ACE inhibitors and/or angiotensin II receptor blockers and patients with albumin/creatinine ratio over 300mg/g were excluded. The outcome of the study was the presence of a pathologic urinary albumin/creatinine ratio. The neutrophil-to-lymphocyte ratio was significantly associated with altered urinary albumin/creatinine ratio in the univariate analysis and after adjustment for other known endothelial and cardiovascular risk factors (age, hypertension, dyslipidaemia, diabetes or altered glomerular filtration rate). Based on the sensitivity and specificity of different neutrophil-to-lymphocyte ratio thresholds, 3 risk groups were created for altered urinary albumin/creatinine ratio: low risk in those with neutrophil-to-lymphocyte ratio 3. These groups were found to have a statistically significant and independent prognostic power for altered urinary albumin/creatinine ratio in asymptomatic patients. The neutrophil-to-lymphocyte ratio appears to be a cost-efficient, non-invasive and independent potential marker of systemic endothelial dysfunction in asymptomatic subjects. Copyright © 2015 Sociedad Española de Nefrología. Published by Elsevier España, S.L.U. All rights reserved.

  12. PARTICIPATION OF TLR4 IN ENGULFMENT OF ESCHERICHIA COLI BY HUMAN BLOOD NEUTROPHILS IN PRESENCE OF LIPOPOLYSACCHARIDES

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    S. V. Zubova

    2012-01-01

    Full Text Available Abstract. TLR4 is a key player in signaling system of host cells. Possible role of TLR4 is actively discussed, e.g. its significance for phagocytosis. A capacity of neutrophils to engulf FITC-labeled E. coli bacteria upon activation with LPS of different origin was studied in presence of anti-TLR4 Mab’s (HTA125 clone. It was shown that, in whole blood, TLR4 does not play any essential role in engulfment of bacteria by the neutrophils. Phagocytic activity of neutrophils in blood increases increased after their priming with E. coli endotoxins. LPS from Rb. сapsulatus did not affect phagocytosis. In presence of endotoxins, the degree of TLR4 involvement in neutrophil phagocytosis depends on LPS structure.

  13. Variations in neutrophil count in preterm infants with respiratory distress syndrome who subsequently developed chronic lung disease.

    Science.gov (United States)

    Kohelet, D; Arbel, E; Ballin, A; Goldberg, M

    2000-01-01

    Neutrophil counts were studied in 62 preterm infants receiving mechanical ventilation for neonatal respiratory distress syndrome (NRDS). Exploratory analysis indicated that the severity of NRDS, as demonstrated by fractional inspired oxygen (FiO2), mean airway pressure (MAP), arterial-alveolar PO2 ratio (a/APO2) and oxygenation index (OI), was correlated with percentage change of neutrophil counts during the first 5 days of life. Further analysis demonstrated that infants with NRDS who subsequently developed chronic lung disease (CLD) (n = 21) had statistically significant differences in variation of neutrophil counts when compared with the remainder (n = 41) without CLD (-35.0% +/- 4.3 vs. -16.9% +/- 5.8, p variations in neutrophil counts during the first 5 days of life may be found in infants with NRDS who subsequently develop CLD and that these changes may have predictive value regarding the development of CLD.

  14. Relative uptake of technetium 99m stannous colloid by neutrophils and monocytes is altered by gram-negative infection

    International Nuclear Information System (INIS)

    Ramsay, Stuart C.; Maggs, Jacqueline A.; Ketheesan, Natkunam; Norton, Robert; LaBrooy, Justin

    2005-01-01

    Gram-negative infection alters phagocytic cell function; hence, it could affect phagocytic uptake of inorganic colloids by these cells. Neutrophil and monocyte uptake of technetium 99m stannous colloid ( 99m Tc SnC) in whole blood was measured in 10 patients with gram-negative infection (Burkholderia pseudomallei) and 7 controls. Mean uptake per individual neutrophil was reduced in infection. Uptake per monocyte was not significantly different. Blood from six normal individuals was incubated with lysed B. pseudomallei and colloid, which showed reduced neutrophil uptake, but increased monocyte uptake. These results indicate that uptake of 99m Tc SnC stannous colloid can be used to measure alteration in phagocytic cell function. They suggest that infection with B. pseudomallei is associated with reduced phagocytosis by individual neutrophils, possibly through toxic effects of bacterial products. This could have immunopathogenic consequences for this gram-negative infection and may explain why it responds to granulocyte colony-stimulating factor

  15. Contribution of Neutrophils to Acute Lung Injury

    OpenAIRE

    Grommes, Jochen; Soehnlein, Oliver

    2010-01-01

    Treatment of acute lung injury (ALI) and its most severe form, acute respiratory distress syndrome (ARDS), remain unsolved problems of intensive care medicine. ALI/ARDS are characterized by lung edema due to increased permeability of the alveolar-capillary barrier and subsequent impairment of arterial oxygenation. Lung edema, endothelial and epithelial injury are accompanied by an influx of neutrophils into the interstitium and broncheoalveolar space. Hence, activation and recruitment of neut...

  16. Leukotriene B4-Neutrophil Elastase Axis Drives Neutrophil Reverse Transendothelial Cell Migration In Vivo

    Science.gov (United States)

    Colom, Bartomeu; Bodkin, Jennifer V.; Beyrau, Martina; Woodfin, Abigail; Ody, Christiane; Rourke, Claire; Chavakis, Triantafyllos; Brohi, Karim; Imhof, Beat A.; Nourshargh, Sussan

    2015-01-01

    Summary Breaching endothelial cells (ECs) is a decisive step in the migration of leukocytes from the vascular lumen to the extravascular tissue, but fundamental aspects of this response remain largely unknown. We have previously shown that neutrophils can exhibit abluminal-to-luminal migration through EC junctions within mouse cremasteric venules and that this response is elicited following reduced expression and/or functionality of the EC junctional adhesion molecule-C (JAM-C). Here we demonstrate that the lipid chemoattractant leukotriene B4 (LTB4) was efficacious at causing loss of venular JAM-C and promoting neutrophil reverse transendothelial cell migration (rTEM) in vivo. Local proteolytic cleavage of EC JAM-C by neutrophil elastase (NE) drove this cascade of events as supported by presentation of NE to JAM-C via the neutrophil adhesion molecule Mac-1. The results identify local LTB4-NE axis as a promoter of neutrophil rTEM and provide evidence that this pathway can propagate a local sterile inflammatory response to become systemic. PMID:26047922

  17. Leukotriene B4-Neutrophil Elastase Axis Drives Neutrophil Reverse Transendothelial Cell Migration In Vivo.

    Science.gov (United States)

    Colom, Bartomeu; Bodkin, Jennifer V; Beyrau, Martina; Woodfin, Abigail; Ody, Christiane; Rourke, Claire; Chavakis, Triantafyllos; Brohi, Karim; Imhof, Beat A; Nourshargh, Sussan

    2015-06-16

    Breaching endothelial cells (ECs) is a decisive step in the migration of leukocytes from the vascular lumen to the extravascular tissue, but fundamental aspects of this response remain largely unknown. We have previously shown that neutrophils can exhibit abluminal-to-luminal migration through EC junctions within mouse cremasteric venules and that this response is elicited following reduced expression and/or functionality of the EC junctional adhesion molecule-C (JAM-C). Here we demonstrate that the lipid chemoattractant leukotriene B4 (LTB4) was efficacious at causing loss of venular JAM-C and promoting neutrophil reverse transendothelial cell migration (rTEM) in vivo. Local proteolytic cleavage of EC JAM-C by neutrophil elastase (NE) drove this cascade of events as supported by presentation of NE to JAM-C via the neutrophil adhesion molecule Mac-1. The results identify local LTB4-NE axis as a promoter of neutrophil rTEM and provide evidence that this pathway can propagate a local sterile inflammatory response to become systemic. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.

  18. Prevention of vascular inflammation by nanoparticle targeting of adherent neutrophils

    Science.gov (United States)

    Wang, Zhenjia; Li, Jing; Cho, Jaehyung; Malik, Asrar B.

    2014-03-01

    Inflammatory diseases such as acute lung injury and ischaemic tissue injury are caused by the adhesion of a type of white blood cell--polymorphonuclear neutrophils--to the lining of the circulatory system or vascular endothelium and unchecked neutrophil transmigration. Nanoparticle-mediated targeting of activated neutrophils on vascular endothelial cells at the site of injury may be a useful means of directly inactivating neutrophil transmigration and hence mitigating vascular inflammation. Here, we report a method employing drug-loaded albumin nanoparticles, which efficiently deliver drugs into neutrophils adherent to the surface of the inflamed endothelium. Using intravital microscopy of tumour necrosis factor-α-challenged mouse cremaster post-capillary venules, we demonstrate that fluorescently tagged albumin nanoparticles are largely internalized by neutrophils adherent to the activated endothelium via cell surface Fcɣ receptors. Administration of albumin nanoparticles loaded with the spleen tyrosine kinase inhibitor, piceatannol, which blocks `outside-in' β2 integrin signalling in leukocytes, detached the adherent neutrophils and elicited their release into the circulation. Thus, internalization of drug-loaded albumin nanoparticles into neutrophils inactivates the pro-inflammatory function of activated neutrophils, thereby offering a promising approach for treating inflammatory diseases resulting from inappropriate neutrophil sequestration and activation.

  19. Sexy again: the renaissance of neutrophils in psoriasis.

    Science.gov (United States)

    Schön, Michael P; Broekaert, Sigrid M C; Erpenbeck, Luise

    2017-04-01

    Notwithstanding their prominent presence in psoriatic skin, the functional role of neutrophilic granulocytes still remains somewhat enigmatic. Sparked by exciting scientific discoveries regarding neutrophil functions within the last years, the interest in these short-lived cells of the innate immune system has been boosted recently. While it had been known for some time that neutrophils produce and respond to a number of inflammatory mediators, recent research has linked neutrophils with the pathogenic functions of IL-17, possibly in conjunction with the formation of NETs (neutrophil extracellular traps). Antipsoriatic therapies exert their effects, at least in part, through interference with neutrophils. Neutrophils also appear to connect psoriasis with comorbid diseases. However, directly tampering with neutrophil functions is not trivial as evinced by the failure of therapeutic approaches targeting redundantly regulated cellular communication networks. It has also become apparent that neutrophils link important pathogenic functions of the innate and the adaptive immune system and that they are intricately involved in regulatory networks underlying the pathophysiology of psoriasis. In order to advocate intensified research into the role of this interesting cell population, we here highlight some features of neutrophils and put them into perspective with our current view of the pathophysiology of psoriasis. © 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  20. Effect of Isolation Techniques on Viability of Bovine Blood Neutrophils

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    P. Sláma

    2006-01-01

    Full Text Available The effect of selected isolation methods on the viability of neutrophil granulocytes (neutrophils from the blood of healthy Holstein x Bohemian Red Pied crossbred heifers was evaluated. Two methods of neutrophil isolation were used: a neutrophil isolation on the basis of hypotonic erythrocyte lysis (in two variants: after the erythrocyte lysis proper, the cells were centrifuged at either 200 g or 1000 g, and b neutrophil isolation with FACS Lysing Solution as the lysing agent. The viability of the isolated neutrophils was evaluated on the basis of apoptosis and necrosis. The results obtained with flow cytometry (FCM suggest that, from the isolation techniques used, the method based on FACS Lysing Solution impaired the neutrophil viability least. After the application of this method, 5.36 ± 2.15% of neutrophils were apoptotic and 0.51 ± 0.12% were necrotic. In contrast, when the hypotonic erythrocyte lysis was used, the proportion of apoptotic neutrophils amounted to 42.14 ± 7.12% and 49.00 ± 14.70%, respectively, and 41.12 ± 5.55% and 36.91 ± 24.38% respectively of necrotic neutrophils (P < 0.01. This was also confirmed by the light microscopy. After the isolation with FASC Lysing Solution, 1.92 ± 1.74% of neutrophils were apoptotic and 1.05 ± 0.76% were necrotic, as distinct from after the hypotonic erythrocyte lysis where 9.43 ± 3.69% of neutrophils were apoptotic and 12.67 ± 4.74% of necrotic after centrifugation at 200 g, while 12.60 ± 4.35 were apoptotic and 14.96 ± 12.64% were necrotic after centrifugation at 1000 g. It follows from the above-mentioned data that hypotonic lysis is not a suitable method for the isolation of neutrophils, as the method itself markedly affects cell viability.

  1. Expression of IL-17A concentration and effector functions of peripheral blood neutrophils in food allergy hypersensitivity patients.

    Science.gov (United States)

    Żbikowska-Gotz, Magdalena; Pałgan, Krzysztof; Gawrońska-Ukleja, Ewa; Kuźmiński, Andrzej; Przybyszewski, Michał; Socha, Ewa; Bartuzi, Zbigniew

    2016-03-01

    Lymphocytes Th17 and other types of immune system cells produce IL17. By induction of cytokines and chemokines, the IL17 cytokine is involved in mechanisms of allergic reaction with participation of neutrophil granulocytes. It affects activation, recruitment, and migration of neutrophils to the tissues, regulating inflammatory reaction intensity. Excited neutrophils secrete inter alia elastase and reactive oxygen species (ROS)--significant mediators of inflammation process responsible for tissues damage.The aim of the study was to evaluate the concentrations of serum interleukin 17A, serum neutrophil elastase, and ROS production by neutrophils in patients with food allergy.The study included 30 patients with food allergy diagnosed based on interview, clinical symptoms, positive SPT, placebo controlled double-blind oral provocation trial, and the presence of asIgE in blood serum against selected food allergens using fluoro-immuno-enzymatic method FEIA UNICap 100. The control group consisted of 10 healthy volunteers. The concentrations of IL17A were determined in all patients using ELISA method with eBioscience kits, and elastase using BenderMed Systems kits. Chemiluminescence of non-stimulated neutrophils was evaluated using luminol-dependent kinetic method for 40 min on Luminoskan (Labsystems luminometer).The results of serum IL-17A concentrations and the values of chemiluminescence obtained by non-activated neutrophils, as well as elastase concentrations, were higher in patients with food allergic hypersensitivity compared to healthy volunteers.This study demonstrates a significance of IL-17A and activated neutrophil granulocytes in the course of diseases with food allergic hypersensitivity. © The Author(s) 2015.

  2. Impact of ABCB1 variants on neutrophil depression: a prospective study

    DEFF Research Database (Denmark)

    Bergmann, Troels Korshøj; Andersen, Charlotte Brasch; Gréen, Henrik

    2010-01-01

    toxicity was registered. Patients carrying one or two variant alleles of ABCB1 C3435T had progressively more pronounced neutrophil decrease at nadir (P-value 0.03). The same association was found for ABCB1 C1236T and G2677T/A with P-values of 0.06 and 0.02. No statistically significant correlations were...

  3. Nontypeable Haemophilus influenzae initiates formation of neutrophil extracellular traps.

    Science.gov (United States)

    Juneau, Richard A; Pang, Bing; Weimer, Kristin E D; Armbruster, Chelsie E; Swords, W Edward

    2011-01-01

    Nontypeable Haemophilus influenzae (NTHI) is a leading cause of otitis media infections, which are often chronic and/or recurrent in nature. NTHI and other bacterial species persist in vivo within biofilms during otitis media and other persistent infections. These biofilms have a significant host component that includes neutrophil extracellular traps (NETs). These NETs do not mediate clearance of NTHI, which survives within NET structures by means of specific subpopulations of lipooligosaccharides on the bacterial surface that are determinants of biofilm formation in vitro. In this study, the ability of NTHI and NTHI components to initiate NET formation was examined using an in vitro model system. Both viable and nonviable NTHI strains were shown to promote NET formation, as did preparations of bacterial DNA, outer membrane proteins, and lipooligosaccharide (endotoxin). However, only endotoxin from a parental strain of NTHI exhibited equivalent potency in NET formation to that of NTHI. Additional studies showed that NTHI entrapped within NET structures is resistant to both extracellular killing within NETs and phagocytic killing by incoming neutrophils, due to oligosaccharide moieties within the lipooligosaccharides. Thus, we concluded that NTHI elicits NET formation by means of multiple pathogen-associated molecular patterns (most notably endotoxin) and is highly resistant to killing within NET structures. These data support the conclusion that, for NTHI, formation of NET structures may be a persistence determinant by providing a niche within the middle-ear chamber.

  4. Effector Mechanisms of Neutrophils within the Innate Immune System in Response to Mycobacterium tuberculosis Infection

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    Eric Warren

    2017-02-01

    Full Text Available Neutrophils have a significant yet controversial role in the innate immune response to Mycobacterium tuberculosis (M. tb infection, which is not yet fully understood. In addition to neutrophils’ well-known effector mechanisms, they may also help control infection of M. tb through the formation of neutrophil extracellular traps (NETs, which are thought to further promote the killing of M. tb by resident alveolar macrophages. Cytokines such as IFN-γ have now been shown to serve an immunomodulatory role in neutrophil functioning in conjunction to its pro-inflammatory function. Additionally, the unique transcriptional changes of neutrophils may be used to differentiate between infection with M. tb and other bacterial and chronic rheumatological diseases such as Systemic Lupus Erythematosus. Adversely, during the innate immune response to M. tb, inappropriate phagocytosis of spent neutrophils can result in nonspecific damage to host cells due to necrotic lysis. Furthermore, some individuals have been shown to be more genetically susceptible to tuberculosis (TB due to a “Trojan Horse” phenomenon whereby neutrophils block the ability of resident macrophages to kill M. tb. Despite these aforementioned negative consequences, through the scope of this review we will provide evidence to support the idea that neutrophils, while sometimes damaging, can also be an important component in warding off M. tb infection. This is exemplified in immunocompromised individuals, such as those with human immunodeficiency virus (HIV infection or Type 2 diabetes mellitus. These individuals are at an increased risk of developing tuberculosis (TB due to a diminished innate immune response associated with decreased levels of glutathione. Consequently, there has been a worldwide effort to limit and contain M. tb infection through the use of antibiotics and vaccinations. However, due to several significant limitations, the current bacille Calmette-Guerin vaccine (BCG

  5. Correlation of Leukocyte Count and Percentage of Segmented Neutrophils with Pathohistological Findings of Appendix in Children

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    Marko Baskovic

    2018-01-01

    Full Text Available BackgroundAppendicitis is the most common indication for an emergency operation in children's age. Although none of the laboratory values has not high sensitivity and specificity for the diagnosis of appendicitis, leukocyte count and the percentage of segmented neutrophils are most commonly used. The aim of this study was to determine whether there is a statistically significant correlation between leukocyte count and the percentage of segmented neutrophils compared to the pathohistological finding of appendix in children. Materials and MethodsWe retrospectively analyzed the data in the period from 1 January 2016 to 31 December 2016. The analysis was made on 211 patients. Spearman's correlation coefficient (rs was calculated. We determined the specificity and sensitivity of leukocyte count and the percentage of segmented neutrophils used in the calculation of Alvorado and Pediatric Appendicitis score.ResultsThe results of the research have shown that the correlation between leukocyte count and the pathohistological findings is weak (rs = 0.29, p = 3.61*10-8, while there is no correlation between the percentage of segmented neutrophils and pathohistological findings (rs = 0.18, p = 7.08 *10-5. The sensitivity of leukocyte count is 93% and the specificity is 30%, while the sensitivity to the percentage of segmented neutrophils is 71% and the specificity is 50%. ROC analysis for leukocytes shows area under the curve of 0.648, while for segmented neutrophils of 0.574.ConclusionGiven the correlation results obtained, the clinical experience of physicians will still have one of the leading roles in diagnosing acute appendicitis in children.

  6. Use of CFSE staining of borreliae in studies on the interaction between borreliae and human neutrophils

    Directory of Open Access Journals (Sweden)

    Hytönen Jukka

    2006-10-01

    Full Text Available Abstract Background Species of the tick-transmitted spirochete group Borrelia burgdorferi sensu lato (B. burgdorferi cause Lyme borreliosis. Acute borrelial infection of the skin has unusual characteristics with only a mild local inflammatory response suggesting that the interaction between borreliae and the cells of the first-line defence might differ from that of other bacteria. It has been reported that human neutrophils phagocytose motile borreliae through an unconventional mechanism (tube phagocytosis which is not observed with non-motile borreliae. Therefore, it would be of great interest to visualise the bacteria by a method not affecting motility and viability of borreliae to be able to study their interaction with the cells of the innate immunity. Carboxyfluorescein diacetate, succinimidyl ester (CFSE labelling has been previously used for studying the adhesion of labelled bacteria to host cells and the uptake of labelled substrates by various cells using flow cytometry. Results In this study, CFSE was shown to efficiently stain different genospecies of B. burgdorferi without affecting bacterial viability or motility. Use of CFSE staining allowed subsequent quantification of borreliae associated with human neutrophils with flow cytometry and confocal microscopy. As a result, no difference in association between different borrelial genospecies (Borrelia burgdorferi sensu stricto, Borrelia afzelii, Borrelia garinii, or between borreliae and the pyogenic bacterium Streptococcus pyogenes, with neutrophils could be detected. Borrelial virulence, on the other hand, affected association with neutrophils, with significantly higher association of a non-virulent mutant B. burgdorferi sensu stricto strain compared to the parental virulent wild type strain. Conclusion These results suggest that the flow cytometric assay using CFSE labelled borreliae is a valuable tool in the analysis of the interaction between borreliae and human neutrophils. The

  7. Effects of Porphyromonas gingivalis LipopolysaccharideTolerized Monocytes on Inflammatory Responses in Neutrophils.

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    Xiang-Qing Zhu

    Full Text Available Periodontitis is a chronic inflammatory disease induced by bacteria. Exposure of the host to periodontal pathogens and their virulence factors induces a state of hyporesponsiveness to subsequent stimulations, which is termed endotoxin tolerance. The role and mechanism of lipopolysaccharide (LPS-tolerized monocytes in inflammatory responses in neutrophils are currently unclear. Here, conditioned supernatants were collected from THP-1 cells treated with or without repeated 1 μg/ml Porphyromonas gingivalis (P.gingivalis LPS. The chemotactic response of freshly isolated neutrophils recruited by supernatants was determined by a transwell migration assay, which demonstrated a reduced migration of neutrophils stimulated with supernatants from tolerized THP-1 cells in comparison to non-tolerized THP-1 cells. In addition, there was a marked increase in reactive oxygen species (ROS generation and a significant decrease in Caspase 3 activities in neutrophils treated with supernatants from THP-1 cells that were treated repeatedly with P.gingivalis LPS in comparison to single treatment. A cytokine antibody array was then used to assess cytokine expression patterns in THP-1 cells. In tolerized THP-1 cells, 43 cytokine (43/170 expression levels were decreased, including chemokine ligand 23 (CCL23 and IFN-γ, while 11 cytokine (11/170 expression levels were increased, such as death receptor 6 (DR6. Furthermore, there was decreased production of IFN-γ and epithelial neutrophil activating peptide-78 (ENA-78 in THP-1 cells after stimulation with repeated P. gingivalis LPS in comparison to single challenge, which was confirmed by ELISA. Therefore, P.gingivalis LPS- tolerized THP-1 cells were able to depress neutrophil chemotaxis and apoptosis, and contribute to respiratory burst, which might be related to the changes in cytokine expression patterns in THP-1 cells.

  8. Mycobacteria attenuate nociceptive responses by formyl peptide receptor triggered opioid peptide release from neutrophils.

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    Heike L Rittner

    2009-04-01

    Full Text Available In inflammation, pain is regulated by a balance of pro- and analgesic mediators. Analgesic mediators include opioid peptides which are secreted by neutrophils at the site of inflammation, leading to activation of opioid receptors on peripheral sensory neurons. In humans, local opioids and opioid peptides significantly downregulate postoperative as well as arthritic pain. In rats, inflammatory pain is induced by intraplantar injection of heat inactivated Mycobacterium butyricum, a component of complete Freund's adjuvant. We hypothesized that mycobacterially derived formyl peptide receptor (FPR and/or toll like receptor (TLR agonists could activate neutrophils, leading to opioid peptide release and inhibition of inflammatory pain. In complete Freund's adjuvant-induced inflammation, thermal and mechanical nociceptive thresholds of the paw were quantified (Hargreaves and Randall-Selitto methods, respectively. Withdrawal time to heat was decreased following systemic neutrophil depletion as well as local injection of opioid receptor antagonists or anti-opioid peptide (i.e. Met-enkephalin, beta-endorphin antibodies indicating an increase in pain. In vitro, opioid peptide release from human and rat neutrophils was measured by radioimmunoassay. Met-enkephalin release was triggered by Mycobacterium butyricum and formyl peptides but not by TLR-2 or TLR-4 agonists. Mycobacterium butyricum induced a rise in intracellular calcium as determined by FURA loading and calcium imaging. Opioid peptide release was blocked by intracellular calcium chelation as well as phosphoinositol-3-kinase inhibition. The FPR antagonists Boc-FLFLF and cyclosporine H reduced opioid peptide release in vitro and increased inflammatory pain in vivo while TLR 2/4 did not appear to be involved. In summary, mycobacteria activate FPR on neutrophils, resulting in tonic secretion of opioid peptides from neutrophils and in a decrease in inflammatory pain. Future therapeutic strategies may aim

  9. Neutrophil depletion causes a fatal defect in murine pulmonary Staphylococcus aureus clearance.

    Science.gov (United States)

    Robertson, Charles M; Perrone, Erin E; McConnell, Kevin W; Dunne, W Michael; Boody, Barrett; Brahmbhatt, Tejal; Diacovo, M Julia; Van Rooijen, Nico; Hogue, Lisa A; Cannon, Carolyn L; Buchman, Timothy G; Hotchkiss, Richard S; Coopersmith, Craig M

    2008-12-01

    Staphylococcus aureus is the most common cause of healthcare-associated pneumonia. Despite the significant morbidity and mortality associated with the disease, animal models of S. aureus pneumonia are rare. We examined the pathogenicity of four different strains of S. aureus (both methicillin-sensitive and -resistant as well as Panton-Valentine leukocidin-positive and -negative) in four strains of immunocompetent inbred and outbred mice (FVB/N, C57Bl/6, BALB/c, ND4; n = 148). The immunological basis for the development of murine S. aureus pneumonia was then determined by selectively depleting neutrophils, lymphocytes, or pulmonary macrophages prior to the onset of infection. An additional cohort of animals was rendered immunosuppressed by induction of abdominal sepsis via cecal ligation and puncture 2, 4, or 7 d prior to the onset of pneumonia. Nearly all immunocompetent mice survived, regardless of which strain of S. aureus was used or which strain of mouse was infected. Among animals with immune depletion or prior immunosuppression, survival was decreased only following neutrophil depletion (26% versus 90% alive at 7 d, P < 0.0001). Compared to immunocompetent animals, neutrophil-depleted mice with S. aureus pneumonia had delayed pulmonary bacterial clearance at 16 and 40 h but had no difference in levels of bacteremia. Neutrophil-depleted mice also had elevated levels of pulmonary monocyte chemotactic protein-1 (822 pg/mL versus 150 pg/mL, P < 0.05). In contrast, pulmonary histological appearance was similar in both groups as was dry/wet lung weight. These results suggest that neutrophils play a critical role in the host response to S. aureus pneumonia, and the survival differences observed in neutrophil-depleted mice are associated with alterations in bacterial clearance and pulmonary cytokine response.

  10. Down-regulated resistin level in consequence of decreased neutrophil counts in untreated Grave's disease.

    Science.gov (United States)

    Peng, Ying; Qi, Yicheng; Huang, Fengjiao; Chen, Xinxin; Zhou, Yulin; Ye, Lei; Wang, Weiqing; Ning, Guang; Wang, Shu

    2016-11-29

    Resistin, belongs to cysteine-rich secretory protein, is mainly produced by circulating leukocytes, such as neutrophils monocytes and macrophages in humans. To date, few but controversial studies have reported about resistin concentrations in hyperthyroid patients, especially in Graves' disease (GD). We undertaked a controlled, prospective study to explore the serum resistin concentration in GD patients before and after -MMI treatment. In addition, we also investigated the main influencing factor on serum resistin level and discuessed the potential role of serum resistin plays in GD patients. 39 untreated GD (uGD) patients, including 8 males and 31 females, were enrolled in our investigation. All of these patients were prescribed with MMI treatment, in addition to 25 healthy controls. Anthropometric parameters and hormone assessment were measured. Enzyme-linked immunosorbent assay was used to detect serum resistin concentration in different stages of GD patients. Furthermore, neutrophil cell line NB4 with or without T3 treatment to detect the effect of thyroid hormones on resistin expression. The serum resistin level and neutrophil counts in untreated GD patients were significantly declined. And all of these parameters were recovered to normal after MMI treatment in ethyroid GD (eGD) and TRAb-negative conversion (nGD) patients. Resistin concentration exhibited a negative correlation with FT3 and FT4, but a positive correlation with absolute number of neutrophiles in uGD patients, whereas did not correlate with thyroid autoimmune antibodies and BMI. Neutrophile cell line, NB4, produced decreased expression of resistin when stimulated with T3. Our study showed a decrease of serum resistin level in GD patients and we suggested that the serum resistin might primarily secreted from circulating neutrophils and down-regulated by excessive thyroid hormones in GD patients.

  11. Advanced Role of Neutrophils in Common Respiratory Diseases

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    Jinping Liu

    2017-01-01

    Full Text Available Respiratory diseases, always being a threat towards the health of people all over the world, are most tightly associated with immune system. Neutrophils serve as an important component of immune defense barrier linking innate and adaptive immunity. They participate in the clearance of exogenous pathogens and endogenous cell debris and play an essential role in the pathogenesis of many respiratory diseases. However, the pathological mechanism of neutrophils remains complex and obscure. The traditional roles of neutrophils in severe asthma, chronic obstructive pulmonary diseases (COPD, pneumonia, lung cancer, pulmonary fibrosis, bronchitis, and bronchiolitis had already been reviewed. With the development of scientific research, the involvement of neutrophils in respiratory diseases is being brought to light with emerging data on neutrophil subsets, trafficking, and cell death mechanism (e.g., NETosis, apoptosis in diseases. We reviewed all these recent studies here to provide you with the latest advances about the role of neutrophils in respiratory diseases.

  12. Localization and Functionality of the Inflammasome in Neutrophils

    DEFF Research Database (Denmark)

    Bakele, Martina; Joos, Melanie; Burdi, Sofia

    2014-01-01

    Neutrophils represent the major fraction of circulating immune cells and are rapidly recruited to sites of infection and inflammation. The inflammasome is a multiprotein complex that regulates the generation of IL-1 family proteins. The precise subcellular localization and functionality...... of the inflammasome in human neutrophils are poorly defined. Here we demonstrate that highly purified human neutrophils express key components of the NOD-like receptor family, pyrin domain containing 3 (NLRP3), and absent in melanoma 2 (AIM2) inflammasomes, particularly apoptosis-associated speck-like protein...... and released as protein, highly purified neutrophils neither expressed nor released IL-1α at baseline or upon stimulation. Upon inflammasome activation, highly purified neutrophils released substantially lower levels of IL-1β protein compared with partially purified neutrophils. Serine proteases and caspases...

  13. Differential Effect of Viable Versus Necrotic Neutrophils on Mycobacterium tuberculosis Growth and Cytokine Induction in Whole Blood

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    David M. Lowe

    2018-04-01

    Full Text Available Neutrophils exert both positive and negative influences on the host response to tuberculosis, but the mechanisms by which these differential effects are mediated are unknown. We studied the impact of live and dead neutrophils on the control of Mycobacterium tuberculosis using a whole blood bioluminescence-based assay, and assayed supernatant cytokine concentrations using Luminex™ technology and ELISA. CD15+ granulocyte depletion from blood prior to infection with M. tuberculosis-lux impaired control of mycobacteria by 96 h, with a greater effect than depletion of CD4+, CD8+, or CD14+ cells (p < 0.001. Augmentation of blood with viable granulocytes significantly improved control of mycobacteria by 96 h (p = 0.001, but augmentation with necrotic granulocytes had the opposite effect (p = 0.01. Both augmentations decreased supernatant concentrations of tumor necrosis factor and interleukin (IL-12 p40/p70, but necrotic granulocyte augmentation also increased concentrations of IL-10, G-CSF, GM-CSF, and CCL2. Necrotic neutrophil augmentation reduced phagocytosis of FITC-labeled M. bovis BCG by all phagocytes, whereas viable neutrophil augmentation specifically reduced early uptake by CD14+ cells. The immunosuppressive effect of dead neutrophils required necrotic debris rather than supernatant. We conclude that viable neutrophils enhance control of M. tuberculosis in blood, but necrotic neutrophils have the opposite effect—the latter associated with induction of IL-10, growth factors, and chemoattractants. Our findings suggest a mechanism by which necrotic neutrophils may exert detrimental effects on the host response in active tuberculosis.

  14. Release of Active Peptidyl Arginine Deiminases by Neutrophils Can Explain Production of Extracellular Citrullinated Autoantigens in Rheumatoid Arthritis Synovial Fluid

    Science.gov (United States)

    Spengler, Julia; Lugonja, Božo; Jimmy Ytterberg, A.; Zubarev, Roman A.; Creese, Andrew J.; Pearson, Mark J.; Grant, Melissa M.; Milward, Michael; Lundberg, Karin; Buckley, Christopher D.; Filer, Andrew; Raza, Karim; Cooper, Paul R.; Chapple, Iain L.

    2015-01-01

    Objective In the majority of patients with rheumatoid arthritis (RA), antibodies specifically recognize citrullinated autoantigens that are generated by peptidylarginine deiminases (PADs). Neutrophils express high levels of PAD and accumulate in the synovial fluid (SF) of RA patients during disease flares. This study was undertaken to test the hypothesis that neutrophil cell death, induced by either NETosis (extrusion of genomic DNA–protein complexes known as neutrophil extracellular traps [NETs]) or necrosis, can contribute to production of autoantigens in the inflamed joint. Methods Extracellular DNA was quantified in the SF of patients with RA, patients with osteoarthritis (OA), and patients with psoriatic arthritis (PsA). Release of PAD from neutrophils was investigated by Western blotting, mass spectrometry, immunofluorescence staining, and PAD activity assays. PAD2 and PAD4 protein expression, as well as PAD enzymatic activity, were assessed in the SF of patients with RA and those with OA. Results Extracellular DNA was detected at significantly higher levels in RA SF than in OA SF (P < 0.001) or PsA SF (P < 0.05), and its expression levels correlated with neutrophil concentrations and PAD activity in RA SF. Necrotic neutrophils released less soluble extracellular DNA compared to NETotic cells in vitro (P < 0.05). Higher PAD activity was detected in RA SF than in OA SF (P < 0.05). The citrullinated proteins PAD2 and PAD4 were found attached to NETs and also freely diffused in the supernatant. PAD enzymatic activity was detected in supernatants of neutrophils undergoing either NETosis or necrosis. Conclusion Release of active PAD isoforms into the SF by neutrophil cell death is a plausible explanation for the generation of extracellular autoantigens in RA. PMID:26245941

  15. Diverse novel functions of neutrophils in immunity, inflammation, and beyond

    OpenAIRE

    Mocsai, A.

    2013-01-01

    Neutrophils have long been considered simple suicide killers at the bottom of the hierarchy of the immune response. That view began to change 10–20 yr ago, when the sophisticated mechanisms behind how neutrophils locate and eliminate pathogens and regulate immunity and inflammation were discovered. The last few years witnessed a new wave of discoveries about additional novel and unexpected functions of these cells. Neutrophils have been proposed to participate in protection against intracellu...

  16. Neutrophilic dermatoses in a patient with collagenous colitis

    OpenAIRE

    Didac Barco; Maria A. Barnadas; Esther Roé; Francisco J. Sancho; Elena Ricart; Agustín Alomar

    2010-01-01

    We report the case of a 75-year old woman with collagenous colitis who presented with erythematous and edematous plaques on the periorbital and eyelid regions, accompanied by oral ulcers. Histopathology showed a dermal neutrophilic infiltrate plus mild septal and lobular panniculitis with lymphocytes, neutrophils and eosinophils. Five years earlier she had presented a flare of papules and vesicles on the trunk, together with oral ulcers; a skin biopsy revealed a neutrophilic dermal infiltrate...

  17. Phagocytosis, bacterial killing, and cytokine activation of circulating blood neutrophils in horses with severe equine asthma and control horses.

    Science.gov (United States)

    Vanderstock, Johanne M; Lecours, Marie-Pier; Lavoie-Lamoureux, Annouck; Gottschalk, Marcelo; Segura, Mariela; Lavoie, Jean-Pierre; Jean, Daniel

    2018-04-01

    OBJECTIVE To evaluate in vitro phagocytosis and bactericidal activity of circulating blood neutrophils in horses with severe equine asthma and control horses and to determine whether circulating blood neutrophils in horses with severe equine asthma have an increase in expression of the proinflammatory cytokine tumor necrosis factor (TNF)-α and the chemokine interleukin (IL)-8 and a decrease in expression of the anti-inflammatory cytokine IL-10 in response to bacteria. ANIMALS 6 horses with severe equine asthma and 6 control horses. PROCEDURES Circulating blood neutrophils were isolated from horses with severe equine asthma and control horses. Phagocytosis was evaluated by use of flow cytometry. Bactericidal activity of circulating blood neutrophils was assessed by use of Streptococcus equi and Streptococcus zooepidemicus as targets, whereas the cytokine mRNA response was assessed by use of a quantitative PCR assay. RESULTS Circulating blood neutrophils from horses with severe equine asthma had significantly lower bactericidal activity toward S zooepidemicus but not toward S equi, compared with results for control horses. Phagocytosis and mRNA expression of TNF-α, IL-8, and IL-10 were not different between groups. CONCLUSIONS AND CLINCAL RELEVANCE Impairment of bactericidal activity of circulating blood neutrophils in horses with severe equine asthma could contribute to an increased susceptibility to infections.

  18. Neutrophil elastase inhibitor, ONO-5046, modulates acid-induced lung and systemic injury in rabbits.

    Science.gov (United States)

    Kaneko, K; Kudoh, I; Hattori, S; Yamada, H; Ohara, M; Wiener-Kronish, J; Okumura, F

    1997-09-01

    Acid instillation leads to direct lung and to secondary systemic organ injury, probably via activated macrophages and neutrophils. This study investigated the effects of neutrophil elastase on organ injury after unilateral lung acid instillation by administrating a specific neutrophil elastase inhibitor, ONO-5046, before acid instillation. Three groups of anesthetized rabbits (n = 12 in each group) underwent tracheostomies, and instillations were made into their right lower lobe airspaces with either phosphate buffered saline (pH, 7.4; volume, 1.2 ml/kg; n = 12) or HCl (pH, 1.25; volume, 1.2 ml/kg; n = 24). In half of the acid-instilled rabbits, ONO-5046, 10 mg/kg, was given intravenously 15 min before the HCl instillation, and then 10 mg x kg(-1) x h(-1) of the drug was continuously infused throughout the experiment. The other groups of animals received the vehicle intravenously. Anesthesia and mechanical ventilation was continued for 8 h, whereas arterial blood gases were sampled intermittently. Eight hours after saline or acid instillation, the animals were killed, and their lungs, heart, kidneys, liver, and small intestines were harvested. Wet-to-dry weight ratios (W/ D) and myeloperoxidase (MPO) assays of these organs were done, and elastase assays on the bronchoalveolar lavage fluids (BALF) obtained from each lung also were performed. Pretreatment with ONO-5046 attenuated the physiologic changes seen in the vehicle-treated animals. Significant decreases in W/D of the noninstilled lungs and of the small intestine and normalization of the oxygenation of the experimental animals occurred. The ONO-5046 pretreatment did not affect the neutrophil sequestration in the lungs or in the other organs as determined by neutrophil counts in BALF and by the MPO assays. A neutrophil elastase inhibitor, ONO-5046, administered immediately before acid instillation attenuated the physiologic changes seen in the vehicle-treated animals. The drug blocked neutrophil elastase but

  19. Neutrophils in traumatic brain injury (TBI): friend or foe?

    Science.gov (United States)

    Liu, Yang-Wuyue; Li, Song; Dai, Shuang-Shuang

    2018-05-17

    Our knowledge of the pathophysiology about traumatic brain injury (TBI) is still limited. Neutrophils, as the most abundant leukocytes in circulation and the first-line transmigrated immune cells at the sites of injury, are highly involved in the initiation, development, and recovery of TBI. Nonetheless, our understanding about neutrophils in TBI is obsolete, and mounting evidences from recent studies have challenged the conventional views. This review summarizes what is known about the relationships between neutrophils and pathophysiology of TBI. In addition, discussions are made on the complex roles as well as the controversial views of neutrophils in TBI.

  20. Hypoxia upregulates neutrophil degranulation and potential for tissue injury

    Science.gov (United States)

    Hoenderdos, Kim; Lodge, Katharine M; Hirst, Robert A; Chen, Cheng; Palazzo, Stefano G C; Emerenciana, Annette; Summers, Charlotte; Angyal, Adri; Porter, Linsey; Juss, Jatinder K; O'Callaghan, Christopher; Chilvers, Edwin R

    2016-01-01

    Background The inflamed bronchial mucosal surface is a profoundly hypoxic environment. Neutrophilic airway inflammation and neutrophil-derived proteases have been linked to disease progression in conditions such as COPD and cystic fibrosis, but the effects of hypoxia on potentially harmful neutrophil functional responses such as degranulation are unknown. Methods and results Following exposure to hypoxia (0.8% oxygen, 3 kPa for 4 h), neutrophils stimulated with inflammatory agonists (granulocyte-macrophage colony stimulating factor or platelet-activating factor and formylated peptide) displayed a markedly augmented (twofold to sixfold) release of azurophilic (neutrophil elastase, myeloperoxidase), specific (lactoferrin) and gelatinase (matrix metalloproteinase-9) granule contents. Neutrophil supernatants derived under hypoxic but not normoxic conditions induced extensive airway epithelial cell detachment and death, which was prevented by coincubation with the antiprotease α-1 antitrypsin; both normoxic and hypoxic supernatants impaired ciliary function. Surprisingly, the hypoxic upregulation of neutrophil degranulation was not dependent on hypoxia-inducible factor (HIF), nor was it fully reversed by inhibition of phospholipase C signalling. Hypoxia augmented the resting and cytokine-stimulated phosphorylation of AKT, and inhibition of phosphoinositide 3-kinase (PI3K)γ (but not other PI3K isoforms) prevented the hypoxic upregulation of neutrophil elastase release. Conclusion Hypoxia augments neutrophil degranulation and confers enhanced potential for damage to respiratory airway epithelial cells in a HIF-independent but PI3Kγ-dependent fashion. PMID:27581620

  1. The Effects of Exercise on Judoists’ Circulating Blood Neutrophils

    Directory of Open Access Journals (Sweden)

    A Zar

    2012-05-01

    Full Text Available

    Background and Objectives: Type, intensity and duration of exercises exert pivotal effects on athletes’ immune system and probably athletes’ susceptibility to upper respiratory tract infections. In this study we examined the effects of one session of moderate-intensity exercise on male judoists’ circulating blood neutrophil counts (BNC and respiratory burst, and self-reported upper respiratory clinical infections 24 hours after the exercise and during the sport seasons. Methods: Ten male judoists after obtaining informed consent were included in the study. The athletes took part in a session of moderate-intensity exercise (60 minutes running on a treadmill at 60% of maximum heart rate. Blood samples were drawn at rest immediately after the exercise. Blood neutrophil count and percentage of Phorbol Myristate Acetate (PMA stimulated neutrophils in whole blood were assessed [as a marker of oxidative burst (OB quality]. Athletes were asked about any signs of upper respiratory infections 24 hours after the exercise and during sport seasons. Paired-t test was used for statistical analysis and statistical significance was set at p<0.05. Results: BNC were in normal range at rest, and meaningfully increased immediately after the exercise (p<0.05. At rest, the OB activity was in normal range, and increased immediately after the exercise (not significant. During 24 hours after the exercise, athletes showed no signs of upper respiratory system infections. Also they mentioned no history of increased susceptibility of upper respiratory infections during sport seasons. Conclusion: Continuous judo exercises have no negative effects on BNC and OB activity. This finding is in accordance with the absence of self-reported upper respiratory infections in judoists during sport seasons. Significant increase in BNC after a session of exercise was a

  2. Degree of neutrophil, atrophy, and metaplasia intestinal were associate with malondialdehyde level in gastritis patients

    Science.gov (United States)

    Siregar, G. A.; Sari, D. K.; Sungkar, T.

    2018-03-01

    The main pathogenesis of gastritis is inflammation that closely related to free radicals. Malondialdehyde (MDA) is a free radical biomarker and is found to increase in gastritis patients. However, these studies are generally performed on experimental animals as well as MDA examination in gastric mucosa. This study aim was to determine the association of degrees of gastritis (degree of lymphocyte infiltration, neutrophil activity, atrophy, and intestinal metaplasia) with plasma MDA level. A cross-sectional study of 80 consecutive gastritis patients who came to an endoscopic unit of Adam Malik General Hospital in Medan, Indonesia, from May–September 2017. Assessed for severity of chronic inflammatory, neutrophil activity, atrophy, and intestinal metaplasia refers to Updated Sydney System. Plasma MDA levels were examined using an HPLC MDA kit. Univariate analysis, bivariate (chi-square and Fisher exact test), and multivariate (binary logistic regression test) were programmed with SPSS version 22. There was no significant association between degree of lymphocyte infiltration with MDA level. There were significant associations between degree of neutrophil activity, atrophy, and intestinal metaplasia with MDA level (p=0.039, 0.003, 0.021; respectively). The moderate+severe degree of neutrophil activity, atrophy, and intestinal metaplasia were associated with high level of MDA.

  3. CYP4F18-Deficient Neutrophils Exhibit Increased Chemotaxis to Complement Component C5a

    Directory of Open Access Journals (Sweden)

    Rachel Vaivoda

    2015-01-01

    Full Text Available CYP4Fs were first identified as enzymes that catalyze hydroxylation of leukotriene B4 (LTB4. CYP4F18 has an unusual expression in neutrophils and was predicted to play a role in regulating LTB4-dependent inflammation. We compared chemotaxis of wild-type and Cyp4f18 knockout neutrophils using an in vitro assay. There was no significant difference in the chemotactic response to LTB4, but the response to complement component C5a increased 1.9–2.25-fold in knockout cells compared to wild-type (P < 0.01. This increase was still observed when neutrophils were treated with inhibitors of eicosanoid synthesis. There were no changes in expression of other CYP4 enzymes in knockout neutrophils that might compensate for loss of CYP4F18 or lead to differences in activity. A mouse model of dextran sodium sulfate colitis was used to investigate the consequences of increased C5a-dependent chemotaxis in vivo, but there was no significant difference in weight loss, disease activity, or colonic tissue myeloperoxidase between wild-type and Cyp4f18 knockout mice. This study demonstrates the limitations of inferring CYP4F function based on an ability to use LTB4 as a substrate, points to expanding roles for CYP4F enzymes in immune regulation, and underscores the in vivo challenges of CYP knockout studies.

  4. Heightened systemic levels of neutrophil and eosinophil granular proteins in pulmonary tuberculosis and reversal following treatment.

    Science.gov (United States)

    Moideen, Kadar; Kumar, Nathella Pavan; Nair, Dina; Banurekha, Vaithilingam V; Bethunaickan, Ramalingam; Babu, Subash

    2018-04-09

    Granulocytes are activated during tuberculosis (TB) infection and act as immune effector cells and granulocyte responses are implicated in TB pathogenesis. Plasma levels of neutrophil and eosinophil granular proteins provide an indirect measure of degranulation. In this study, we wanted to examine the levels of neutrophil and eosinophil granular proteins in individuals with pulmonary tuberculosis (PTB) and to compare them with the levels in latent TB (LTB) individuals. Hence, we measured the plasma levels of myeloperoxidase (MPO), neutrophil elastase, and proteinase-3; major basic protein (MBP), eosinophil derived neurotoxin (EDN), eosinophil cationic protein (ECP) and eosinophil peroxidase (EPX) in these individuals. Finally, we also measured the levels of all of these parameters in PTB individuals following anti-tuberculosis (ATT) treatment. Our data reveal that PTB individuals are characterized by significantly higher plasma levels of MPO, elastase, human proteinase 3 as well as MBP and EDN in comparison to LTB individuals. Our data also reveal that ATT resulted in reversal of all of these changes, indicating an association with TB disease. Finally, our data show that the systemic levels of MPO and proteinase-3 can significantly discriminate PTB from LTB individuals. Thus, our data suggest that neutrophil and eosinophil granular proteins could play a potential role in the innate immune response and therefore, the pathogenesis of pulmonary TB. Copyright © 2018 American Society for Microbiology.

  5. Differential Use of Human Neutrophil Fcγ Receptors for Inducing Neutrophil Extracellular Trap Formation.

    Science.gov (United States)

    Alemán, Omar Rafael; Mora, Nancy; Cortes-Vieyra, Ricarda; Uribe-Querol, Eileen; Rosales, Carlos

    2016-01-01

    Neutrophils (PMN) are the most abundant leukocytes in the blood. PMN migrate from the circulation to sites of infection, where they are responsible for antimicrobial functions. PMN use phagocytosis, degranulation, and formation of neutrophil extracellular traps (NETs) to kill microbes. NETs are fibers composed of chromatin and neutrophil-granule proteins. Several pathogens, including bacteria, fungi, and parasites, and also some pharmacological stimuli such as phorbol 12-myristate 13-acetate (PMA) are efficient inducers of NETs. Antigen-antibody complexes are also capable of inducing NET formation. However the particular Fcγ receptor involved in triggering this function is a matter of controversy. In order to provide some insight into what Fcγ receptor is responsible for NET formation, each of the two human Fcγ receptors was stimulated individually by specific monoclonal antibodies and NET formation was evaluated. FcγRIIa cross-linking did not promote NET formation. Cross-linking other receptors such as integrins also did not promote NET formation. In contrast FcγRIIIb cross-linking induced NET formation similarly to PMA stimulation. NET formation was dependent on NADPH-oxidase, PKC, and ERK activation. These data show that cross-linking FcγRIIIb is responsible for NET formation by the human neutrophil.

  6. Differential Use of Human Neutrophil Fcγ Receptors for Inducing Neutrophil Extracellular Trap Formation

    Directory of Open Access Journals (Sweden)

    Omar Rafael Alemán

    2016-01-01

    Full Text Available Neutrophils (PMN are the most abundant leukocytes in the blood. PMN migrate from the circulation to sites of infection, where they are responsible for antimicrobial functions. PMN use phagocytosis, degranulation, and formation of neutrophil extracellular traps (NETs to kill microbes. NETs are fibers composed of chromatin and neutrophil-granule proteins. Several pathogens, including bacteria, fungi, and parasites, and also some pharmacological stimuli such as phorbol 12-myristate 13-acetate (PMA are efficient inducers of NETs. Antigen-antibody complexes are also capable of inducing NET formation. However the particular Fcγ receptor involved in triggering this function is a matter of controversy. In order to provide some insight into what Fcγ receptor is responsible for NET formation, each of the two human Fcγ receptors was stimulated individually by specific monoclonal antibodies and NET formation was evaluated. FcγRIIa cross-linking did not promote NET formation. Cross-linking other receptors such as integrins also did not promote NET formation. In contrast FcγRIIIb cross-linking induced NET formation similarly to PMA stimulation. NET formation was dependent on NADPH-oxidase, PKC, and ERK activation. These data show that cross-linking FcγRIIIb is responsible for NET formation by the human neutrophil.

  7. Rac1 deletion in mouse neutrophils has selective effects on neutrophil functions

    NARCIS (Netherlands)

    Glogauer, Michael; Marchal, Christophe C.; Zhu, Fei; Worku, Aelaf; Clausen, Björn E.; Foerster, Irmgard; Marks, Peter; Downey, Gregory P.; Dinauer, Mary; Kwiatkowski, David J.

    2003-01-01

    Defects in myeloid cell function in Rac2 knockout mice underline the importance of this isoform in activation of NADPH oxidase and cell motility. However, the specific role of Rac1 in neutrophil function has been difficult to assess since deletion of Rac1 results in embryonic lethality in mice. To

  8. Gβ1 is required for neutrophil migration in zebrafish.

    Science.gov (United States)

    Ke, Wenfan; Ye, Ding; Mersch, Kacey; Xu, Hui; Chen, Songhai; Lin, Fang

    2017-08-01

    Signaling mediated by G protein-coupled receptors (GPCRs) is essential for the migration of cells toward chemoattractants. The recruitment of neutrophils to injured tissues in zebrafish larvae is a useful model for studying neutrophil migration and trafficking in vivo. Indeed, the study of this process led to the discovery that PI3Kγ is required for the polarity and motility of neutrophils, features that are necessary for the directed migration of these cells to wounds. However, the mechanism by which PI3Kγ is activated remains to be determined. Here we show that signaling by specifically the heterotrimeric G protein subunit Gβ1 is critical for neutrophil migration in response to wounding. In embryos treated with small-molecule inhibitors of Gβγ signaling, neutrophils failed to migrate to wound sites. Although both the Gβ1 and Gβ4 isoforms are expressed in migrating neutrophils, only deficiency for the former (morpholino-based knockdown) interfered with the directed migration of neutrophils towards wounds. The Gβ1 deficiency also impaired the ability of cells to change cell shape and reduced their general motility, defects that are similar to those in neutrophils deficient for PI3Kγ. Transplantation assays showed that the requirement for Gβ1 in neutrophil migration is cell autonomous. Finally, live imaging revealed that Gβ1 is required for polarized activation of PI3K, and for the actin dynamics that enable neutrophil migration. Collectively, our data indicate that Gβ1 signaling controls proper neutrophil migration by activating PI3K and modulating actin dynamics. Moreover, they illustrate a role for a specific Gβ isoform in chemotaxis in vivo. Copyright © 2017 Elsevier Inc. All rights reserved.

  9. Significance of the neutrophil-lymphocyte ratio in elderly type 2 diabetic patients with coronary heart disease%老年2型糖尿病并发冠心病患者血中性粒细胞与淋巴细胞比值的变化及意义

    Institute of Scientific and Technical Information of China (English)

    何茜; 谭洪文; 于瑞萍; 罗建华; 徐淑静

    2016-01-01

    目的 探讨老年2型糖尿病并发冠心病患者血中性粒细胞与淋巴细胞比值(NLR)的变化及意义. 方法 采用回顾性病例分析方法,收集2014年4月至2015年7月在贵州省人民医院心内科行冠状动脉造影的老年患者228例,其中糖尿病组77例、冠心病组72例和糖尿病+冠心病组79例,另选70例老年健康体检者为对照组,检测各组白细胞计数、超敏C反应蛋白(hs-CRP)等指标,并计算NLR.Logistic回归分析糖尿病并发冠心病的危险因素,多元逐步回归分析NLR的影响因素. 结果 糖尿病组、冠心病组及糖尿病+冠心病组与对照组比较,白细胞计数[(7.48±1.81)×109/L、(7.72±1.89)×109/L、(7.98±2.12)×109/L与(6.22±1.61)×109/L]、中性粒细胞计数[(4.49±1.38)×109/L、(4.88±1.56)×109/L、(5.35±1.40)×109/L与(3.52±0.84)×109/L]、NLR(2.84±0.77、3.07±0.79、3.34±0.83与1.58±0.42)、hs-CRP[(2.92±0.65) mg/L、(3.20±0.86) mg/L、(4.98±1.10) mg/L与(1.15±0.23)mg/L],均明显升高(P<0.05或P<0.01);淋巴细胞计数降低[(1.57±0.41)×109/L、(1.58±0.40)×109/L、(1.61±0.48)×109/L与(2.22±0.51)×109/L,P<0.05];糖尿病+冠心病组NLR、hs-CRP高于糖尿病组和冠心病组(P<0.05).Pearson相关分析显示,NLR与Gensini积分、hs-CRP正相关(r值分别为0.7455、0.7792,P<0.01).Logistic回归分析显示,NLR、hs-CRP、糖化血红蛋白(HbA1c)是糖尿病并发冠心病的危险因素(OR值分别为4.331、3.997、2.928,P<0.05).多元逐步回归分析显示,NLR与空腹血糖、HbA1c、收缩压正相关(标准化回归系数分别为0.3133、0.4720、0.3069,P<0.05). 结论 NLR可能是老年糖尿病患者并发冠心病的一个较好预测指标.%Objective To evaluate the value of the neutrophil-lymphocyte ratio (NLR) in elderly type 2 diabetic patients (T2DM) with coronary heart disease (CHD).Methods We performed a retrospective observational study on 228 patients undergoing coronary angiography in Guizhou

  10. Neutrophil-Derived Proteases Escalate Inflammation through Activation of IL-36 Family Cytokines.

    Science.gov (United States)

    Henry, Conor M; Sullivan, Graeme P; Clancy, Danielle M; Afonina, Inna S; Kulms, Dagmar; Martin, Seamus J

    2016-02-02

    Recent evidence has strongly implicated the IL-1 family cytokines IL-36α, IL-36β, and IL-36γ as key initiators of skin inflammation. Similar to the other members of the IL-1 family, IL-36 cytokines are expressed as inactive precursors and require proteolytic processing for activation; however, the responsible proteases are unknown. Here, we show that IL-36α, IL-36β, and IL-36γ are activated differentially by the neutrophil granule-derived proteases cathepsin G, elastase, and proteinase-3, increasing their biological activity ~500-fold. Active IL-36 promoted a strong pro-inflammatory signature in primary keratinocytes and was sufficient to perturb skin differentiation in a reconstituted 3D human skin model, producing features resembling psoriasis. Furthermore, skin eluates from psoriasis patients displayed significantly elevated cathepsin G-like activity that was sufficient to activate IL-36β. These data identify neutrophil granule proteases as potent IL-36-activating enzymes, adding to our understanding of how neutrophils escalate inflammatory reactions. Inhibition of neutrophil-derived proteases may therefore have therapeutic benefits in psoriasis. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

  11. Neutrophil-Derived Proteases Escalate Inflammation through Activation of IL-36 Family Cytokines

    Directory of Open Access Journals (Sweden)

    Conor M. Henry

    2016-02-01

    Full Text Available Recent evidence has strongly implicated the IL-1 family cytokines IL-36α, IL-36β, and IL-36γ as key initiators of skin inflammation. Similar to the other members of the IL-1 family, IL-36 cytokines are expressed as inactive precursors and require proteolytic processing for activation; however, the responsible proteases are unknown. Here, we show that IL-36α, IL-36β, and IL-36γ are activated differentially by the neutrophil granule-derived proteases cathepsin G, elastase, and proteinase-3, increasing their biological activity ∼500-fold. Active IL-36 promoted a strong pro-inflammatory signature in primary keratinocytes and was sufficient to perturb skin differentiation in a reconstituted 3D human skin model, producing features resembling psoriasis. Furthermore, skin eluates from psoriasis patients displayed significantly elevated cathepsin G-like activity that was sufficient to activate IL-36β. These data identify neutrophil granule proteases as potent IL-36-activating enzymes, adding to our understanding of how neutrophils escalate inflammatory reactions. Inhibition of neutrophil-derived proteases may therefore have therapeutic benefits in psoriasis.

  12. The effect of the anaesthetic agent isoflurane on the rate of neutrophil apoptosis in vitro.

    LENUS (Irish Health Repository)

    Tyther, R

    2012-02-03

    BACKGROUND: Volatile anaesthetic agents influence neutrophil function, and potentially, the inflammatory response to surgery. AIM: The objective of this study was to determine the effect of isoflurane (1-4%) on human polymorphonuclear neutrophil apoptosis in vitro. METHODS: Venous blood from 12 healthy volunteers was exposed to 0, 1, and 4% isoflurane delivered via a 14G Wallace flexihub internal jugular cannula, at a fresh gas flow of 0.51\\/min for 5 minutes. Isolated neutrophils were assessed for apoptosis at 1, 12, and 24 hours in culture using dual staining with annexin V-FITC and propidium iodide (Annexin-V FITC assay). Data were analysed using paired, one-tailed Student\\'s t-tests. p<0.05 was considered significant. RESULTS: At 1 hour apoptosis was inhibited in the 1% (5.1 [6.8]%; p=0.017) and 4% (4.8 [4.5]%; p=0.008) isoflurane groups compared to control (11.3 [6.9]%). At 12 and 24 hours, a dose-dependent inhibition of apoptosis was demonstrated, i.e. 4% > 1% > 0%. CONCLUSION: Human neutrophil apoptosis is inhibited in a concentration-dependent manner in vitro by isoflurane in clinical concentrations.

  13. Active nuclear transcriptome analysis reveals inflammasome-dependent mechanism for early neutrophil response to Mycobacterium marinum.

    Science.gov (United States)

    Kenyon, Amy; Gavriouchkina, Daria; Zorman, Jernej; Napolitani, Giorgio; Cerundolo, Vincenzo; Sauka-Spengler, Tatjana

    2017-07-26

    The mechanisms governing neutrophil response to Mycobacterium tuberculosis remain poorly understood. In this study we utilise biotagging, a novel genome-wide profiling approach based on cell type-specific in vivo biotinylation in zebrafish to analyse the initial response of neutrophils to Mycobacterium marinum, a close genetic relative of M. tuberculosis used to model tuberculosis. Differential expression analysis following nuclear RNA-seq of neutrophil active transcriptomes reveals a significant upregulation in both damage-sensing and effector components of the inflammasome, including caspase b, NLRC3 ortholog (wu: fb15h11) and il1β. Crispr/Cas9-mediated knockout of caspase b, which acts by proteolytic processing of il1β, results in increased bacterial burden and less infiltration of macrophages to sites of mycobacterial infection, thus impairing granuloma development. We also show that a number of immediate early response genes (IEGs) are responsible for orchestrating the initial neutrophil response to mycobacterial infection. Further perturbation of the IEGs exposes egr3 as a key transcriptional regulator controlling il1β transcription.

  14. Increase in neutrophil Fc gamma receptor I expression following interferon gamma treatment in rheumatoid arthritis.

    Science.gov (United States)

    Goulding, N J; Knight, S M; Godolphin, J L; Guyre, P M

    1992-04-01

    The therapeutic potential of interferon gamma (IFN gamma) in a number of disease states is still being explored, but progress is hampered by the lack of a suitable measure of in vivo biological activity. To assess the in vivo biological effects of recombinant human IFN gamma (rhIFN gamma), 14 patients were studied in a randomised, prospective, double blind, placebo controlled trial of this cytokine for the treatment of rheumatoid arthritis. The levels of Fc gamma receptors on peripheral blood neutrophils were measured at baseline and after 21 days of once daily, subcutaneous injections of rhIFN gamma or placebo. An induction of neutrophil Fc gamma receptor type I (Fc gamma RI) was seen in the group of patients receiving recombinant human rhIFN gamma but not in those receiving placebo. No change in the expression of Fc gamma RII or Fc gamma RIII was detected. The amount of induction of Fc gamma RI detected on the neutrophils of patients receiving rhIFN gamma did not correlate with clinical measures of response at either 21 days or at the end of the study (24 weeks). No significant clinical responses were observed in the rhIFN gamma group at these times. These data confirm that the reported in vitro effect of IFN gamma on human neutrophil Fc receptor expression can be reproduced in vivo.

  15. Neutrophil Extracellular DNA Traps Induce Autoantigen Production by Airway Epithelial Cells

    Directory of Open Access Journals (Sweden)

    Youngwoo Choi

    2017-01-01

    Full Text Available The hypothesis of autoimmune involvement in asthma has received much recent interest. Autoantibodies, such as anti-cytokeratin (CK 18, anti-CK19, and anti-α-enolase antibodies, react with self-antigens and are found at high levels in the sera of patients with severe asthma (SA. However, the mechanisms underlying autoantibody production in SA have not been fully determined. The present study was conducted to demonstrate that neutrophil extracellular DNA traps (NETs, cytotoxic molecules released from neutrophils, are a key player in the stimulation of airway epithelial cells (AECs to produce autoantigens. This study showed that NETs significantly increased the intracellular expression of tissue transglutaminase (tTG but did not affect that of CK18 in AECs. NETs induced the extracellular release of both tTG and CK18 in a concentration-dependent manner. Moreover, NETs directly degraded intracellular α-enolase into small fragments. However, antibodies against neutrophil elastase (NE or myeloperoxidase (MPO attenuated the effects of NETs on AECs. Furthermore, each NET isolated from healthy controls (HC, nonsevere asthma (NSA, and SA had different characteristics. Taken together, these findings suggest that AECs exposed to NETs may exhibit higher autoantigen production, especially in SA. Therefore, targeting of NETs may represent a new therapy for neutrophilic asthma with a high level of autoantigens.

  16. Diverse novel functions of neutrophils in immunity, inflammation, and beyond.

    Science.gov (United States)

    Mócsai, Attila

    2013-07-01

    Neutrophils have long been considered simple suicide killers at the bottom of the hierarchy of the immune response. That view began to change 10-20 yr ago, when the sophisticated mechanisms behind how neutrophils locate and eliminate pathogens and regulate immunity and inflammation were discovered. The last few years witnessed a new wave of discoveries about additional novel and unexpected functions of these cells. Neutrophils have been proposed to participate in protection against intracellular pathogens such as viruses and mycobacteria. They have been shown to intimately shape the adaptive immune response at various levels, including marginal zone B cells, plasmacytoid dendritic cells and T cell populations, and even to control NK cell homeostasis. Neutrophils have been shown to mediate an alternative pathway of systemic anaphylaxis and to participate in allergic skin reactions. Finally, neutrophils were found to be involved in physiological and pathological processes beyond the immune system, such as diabetes, atherosclerosis, and thrombus formation. Many of those functions appear to be related to their unique ability to release neutrophil extracellular traps even in the absence of pathogens. This review summarizes those novel findings on versatile functions of neutrophils and how they change our view of neutrophil biology in health and disease.

  17. Regulation of calcium homeostasis in activated human neutrophils ...

    African Journals Online (AJOL)

    Objectives. The objectives of the current study were to: (i) present an integrated model for the restoration of calcium homeostasis in activated human neutrophils based on current knowledge and recent research; and (ii) identify potential targets for the modulation of calcium fluxes in activated neutrophils based on this model ...

  18. Associations Between Neutrophil Gelatinase Associated Lipocalin, Neutrophil-to-Lymphocyte Ratio, Atrial Fibrillation and Renal Dysfunction in Chronic Heart Failure

    Science.gov (United States)

    Argan, Onur; Ural, Dilek; Kozdag, Guliz; Sahin, Tayfun; Bozyel, Serdar; Aktas, Mujdat; Karauzum, Kurtulus; Yılmaz, Irem; Dervis, Emir; Agir, Aysen

    2016-01-01

    Background Atrial fibrillation (AF) and renal dysfunction are two common comorbidities in patients with chronic heart failure with reduced ejection fraction (HFrEF). This study evaluated the effect of permanent AF on renal function in HFrEF and investigated the associations of atrial fibrillation, neutrophil gelatinase-associated lipocalin (NGAL), and neutrophil-to-lymphocyte ratio (NLR) with adverse clinical outcome. Material/Methods Serum NGAL levels measured by ELISA and NLR were compared between patients with sinus rhythm (HFrEF-SR, n=68), with permanent AF (HFrEF-AF, n=62), and a healthy control group (n=50). Results Mean eGFR levels were significantly lower, and NLR and NGAL levels were significantly higher in the HFrEF patients than in the control patients but the difference between HFrEF-SR and HFrEF-AF was not statistically significant (NGAL: 95 ng/mL in HFrEF-SR, 113 ng/mL in HFrEF-AF and 84 ng/mL in the control group; pfailure, C-reactive protein, NLR, triiodothyronine, and hemoglobin. In ROC analysis, a NLR >3 had a 68% sensitivity and 75% specificity to predict progression of kidney disease (AUC=0.72, 95% CI 0.58–0.85, p=0.001). Conclusions Presence of AF in patients with HFrEF was not an independent contributor of adverse clinical outcome (i.e., all-cause death, re-hospitalization) or progression of renal dysfunction. Renal dysfunction in HFrEF was associated with both NLR and NGAL levels, but systemic inflammation reflected by NLR seemed to be a more important determinant of progression of kidney dysfunction. PMID:27918494

  19. Neutrophil Extracellular Traps are Involved in the Innate Immune Response to Infection with Leptospira

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    Scharrig, Emilia; Carestia, Agostina; Ferrer, María F.; Cédola, Maia; Pretre, Gabriela; Drut, Ricardo; Picardeau, Mathieu; Schattner, Mirta; Gómez, Ricardo M.

    2015-01-01

    NETosis is a process by which neutrophils extrude their DNA together with bactericidal proteins that trap and/or kill pathogens. In the present study, we evaluated the ability of Leptospira spp. to induce NETosis using human ex vivo and murine in vivo models. Microscopy and fluorometric studies showed that incubation of human neutrophils with Leptospira interrogans serovar Copenhageni strain Fiocruz L1-130 (LIC) resulted in the release of DNA extracellular traps (NETs). The bacteria number, pathogenicity and viability were relevant factors for induction of NETs, but bacteria motility was not. Entrapment of LIC in the NETs resulted in LIC death; however, pathogenic but not saprophytic Leptospira sp. exerted nuclease activity and degraded DNA. Mice infected with LIC showed circulating NETs after 2 days post-infection (dpi). Depletion of neutrophils with mAb1A8 significantly reduced the amount of intravascular NETs in LIC-infected mice, increasing bacteremia at 3 dpi. Although there was a low bacterial burden, scarce neutrophils and an absence of inflammation in the early stages of infection in the kidney and liver, at the beginning of the leptospiruric phase, the bacterial burden was significantly higher in kidneys of neutrophil-depleted-mice compared to non-depleted and infected mice. Surprisingly, interstitial nephritis was of similar intensity in both groups of infected mice. Taken together, these data suggest that LIC triggers NETs, and that the intravascular formation of these DNA traps appears to be critical not only to prevent early leptospiral dissemination but also to preclude further bacterial burden. PMID:26161745

  20. Oxidative burst of circulating neutrophils following traumatic brain injury in human.

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    Yiliu Liao

    Full Text Available Besides secondary injury at the lesional site, Traumatic brain injury (TBI can cause a systemic inflammatory response, which may cause damage to initially unaffected organs and potentially further exacerbate the original injury. Here we investigated plasma levels of important inflammatory mediators, oxidative activity of circulating leukocytes, particularly focusing on neutrophils, from TBI subjects and control subjects with general trauma from 6 hours to 2 weeks following injury, comparing with values from uninjured subjects. We observed increased plasma level of inflammatory cytokines/molecules TNF-α, IL-6 and CRP, dramatically increased circulating leukocyte counts and elevated expression of TNF-α and iNOS in circulating leukocytes from TBI patients, which suggests a systemic inflammatory response following TBI. Our data further showed increased free radical production in leukocyte homogenates and elevated expression of key oxidative enzymes iNOS, COX-2 and NADPH oxidase (gp91(phox in circulating leukocytes, indicating an intense induction of oxidative burst following TBI, which is significantly greater than that in control subjects with general trauma. Furthermore, flow cytometry assay proved neutrophils as the largest population in circulation after TBI and showed significantly up-regulated oxidative activity and suppressed phagocytosis rate for circulating neutrophils following brain trauma. It suggests that the highly activated neutrophils might play an important role in the secondary damage, even outside the injured brain. Taken together, the potent systemic inflammatory response induced by TBI, especially the intensively increase oxidative activity of circulating leukocytes, mainly neutrophils, may lead to a systemic damage, dysfunction/damage of bystander tissues/organs and even further exacerbate secondary local damage. Controlling these pathophysiological processes may be a promising therapeutic strategy and will protect unaffected

  1. 18F-fluoro-2-deoxyglucose PET informs neutrophil accumulation and activation in lipopolysaccharide-induced acute lung injury.

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    Rodrigues, Rosana S; Bozza, Fernando A; Hanrahan, Christopher J; Wang, Li-Ming; Wu, Qi; Hoffman, John M; Zimmerman, Guy A; Morton, Kathryn A

    2017-05-01

    Molecular imaging of the earliest events related to the development of acute lung injury (ALI)/acute respiratory distress syndrome (ARDS) could facilitate therapeutic development and patient management. We previously reported that 18 F-fluoro-2-deoxyglucose ( 18 F-FDG) PET identifies ALI/ARDS prior to radiographic abnormalities. The purpose of this study was to establish the time courses of 18 F-FDG uptake, edema and neutrophil recruitment in an endotoxin-induced acute lung injury model and to examine molecular events required for 14 C-2DG uptake in activated neutrophils. Lung uptake of 18 F-FDG was measured by PET in control male Sprague Dawley rats and at 2, 6 and 24h following the intraperitoneal injection of 10mg/kg LPS. Lung edema (attenuation) was measured by microCT. Neutrophil influx into the lungs was measured by myeloperoxidase assay. Control and activated human donor neutrophils were compared for uptake of 14 C-2DG, transcription and content of hexokinase and GLUT isoforms and for hexokinase (HK) activity. Significant uptake of 18 F-FDG occurred by 2h following LPS, and progressively increased to 24h. Lung uptake of 18 F-FDG preceded increased CT attenuation (lung edema). Myeloperoxidase activity in the lungs, supporting neutrophil influx, paralleled 18 F-FDG uptake. Activation of isolated human neutrophils resulted in increased uptake of 14 C-2DG, expression of GLUT 3 and GLUT 4 and expression and increased HK1 activity. Systemic endotoxin-induced ALI results in very early and progressive uptake of 18 F-FDG, parallels neutrophil accumulation and occurs earlier than lung injury edema. Activated neutrophils show increased uptake of 14 C-2DG, expression of specific GLUT3, GLUT4 and HK1 protein and HK activity. ADVANCES IN KNOWLEDGE AND IMPLICATIONS FOR PATIENT CARE: 18 F-FDG pulmonary uptake is an early biomarker of neutrophil recruitment in ALI and is associated with specific molecular events that mediate 14 C-2DG uptake in activated neutrophils. 18 F

  2. Neutrophilic dermatoses in a patient with collagenous colitis

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    Didac Barco

    2010-01-01

    Full Text Available We report the case of a 75-year old woman with collagenous colitis who presented with erythematous and edematous plaques on the periorbital and eyelid regions, accompanied by oral ulcers. Histopathology showed a dermal neutrophilic infiltrate plus mild septal and lobular panniculitis with lymphocytes, neutrophils and eosinophils. Five years earlier she had presented a flare of papules and vesicles on the trunk, together with oral ulcers; a skin biopsy revealed a neutrophilic dermal infiltrate and Sweet’s syndrome was diagnosed. Both the neutrophilic panniculitis and the Sweet’s syndrome were accompanied by fever, malaise and diarrhea. Cutaneous and intestinal symptoms disappeared with corticoid therapy. The two types of neutrophilic dermatoses that appeared in periods of colitis activity suggest that intestinal and cutaneous manifestations may be related.

  3. Swell activated chloride channel function in human neutrophils

    Energy Technology Data Exchange (ETDEWEB)

    Salmon, Michael D. [Leukocyte and Ion Channel Research Laboratory, School of Health and Biosciences, University of East London, Stratford Campus, London E15 4LZ (United Kingdom); Ahluwalia, Jatinder, E-mail: j.ahluwalia@uel.ac.uk [Leukocyte and Ion Channel Research Laboratory, School of Health and Biosciences, University of East London, Stratford Campus, London E15 4LZ (United Kingdom)

    2009-04-17

    Non-excitable cells such as neutrophil granulocytes are the archetypal inflammatory immune cell involved in critical functions of the innate immune system. The electron current generated (I{sub e}) by the neutrophil NADPH oxidase is electrogenic and rapidly depolarises the membrane potential. For continuous function of the NADPH oxidase, I{sub e} has to be balanced to preserve electroneutrality, if not; sufficient depolarisation would prevent electrons from leaving the cell and neutrophil function would be abrogated. Subsequently, the depolarisation generated by the neutrophil NADPH oxidase I{sub e} must be counteracted by ion transport. The finding that depolarisation required counter-ions to compensate electron transport was followed by the observation that chloride channels activated by swell can counteract the NADPH oxidase membrane depolarisation. In this mini review, we discuss the research findings that revealed the essential role of swell activated chloride channels in human neutrophil function.

  4. Neutrophils, a candidate biomarker and target for radiation therapy?

    Science.gov (United States)

    Schernberg, Antoine; Blanchard, Pierre; Chargari, Cyrus; Deutsch, Eric

    2017-11-01

    Neutrophils are the most abundant blood-circulating white blood cells, continuously generated in the bone marrow. Growing evidence suggests they regulate the innate and adaptive immune system during tumor evolution. This review will first summarize the recent findings on neutrophils as a key player in cancer evolution, then as a potential biomarker, and finally as therapeutic targets, with respective focuses on the interplay with radiation therapy. A complex interplay: Neutrophils have been associated with tumor progression through multiple pathways. Ionizing radiation has cytotoxic effects on cancer cells, but the sensitivity to radiation therapy in vivo differ from isolated cancer cells in vitro, partially due to the tumor microenvironment. Different microenvironmental states, whether baseline or induced, can modulate or even attenuate the effects of radiation, with consequences for therapeutic efficacy. Inflammatory biomarkers: Inflammation-based scores have been widely studied as prognostic biomarkers in cancer patients. We have performed a large retrospective cohort of patients undergoing radiation therapy (1233 patients), with robust relationship between baseline blood neutrophil count and 3-year's patient's overall survival in patients with different cancer histologies. (Pearson's correlation test: p = .001, r = -.93). Therapeutic approaches: Neutrophil-targeting agents are being developed for the treatment of inflammatory and autoimmune diseases. Neutrophils either can exert antitumoral (N1 phenotype) or protumoral (N2 phenotype) activity, depending on the Tumor Micro Environment. Tumor associated N2 neutrophils are characterized by high expression of CXCR4, VEGF, and gelatinase B/MMP9. TGF-β within the tumor microenvironment induces a population of TAN with a protumor N2 phenotype. TGF-β blockade slows tumor growth through activation of CD8 + T cells, macrophages, and tumor associated neutrophils with an antitumor N1 phenotype. This supports

  5. Visceral leishmaniasis patients display altered composition and maturity of neutrophils as well as impaired neutrophil effector functions

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    Endalew Yizengaw

    2016-11-01

    Full Text Available Immunologically, active visceral leishmaniasis (VL is characterised by profound immunosuppression, severe systemic inflammatory responses and an impaired capacity to control parasite replication. Neutrophils are highly versatile cells, which play a crucial role in the induction as well as the resolution of inflammation, the control of pathogen replication and the regulation of immune responses. Neutrophil functions have been investigated in human cutaneous leishmaniasis, however, their role in human visceral leishmaniasis is poorly understood.In the present study we evaluated the activation status and effector functions of neutrophils in patients with active VL and after successful anti-leishmanial treatment. Our results show that neutrophils are highly activated and have degranulated; high levels of arginase, myeloperoxidase and elastase, all contained in neutrophils’ granules, were found in the plasma of VL patients. In addition, we show that a large proportion of these cells are immature. We also analysed effector functions of neutrophils that are essential for pathogen clearance and show that neutrophils have an impaired capacity to release neutrophil extracellular traps, produce reactive oxygen species and phagocytose bacterial particles, but not Leishmania parasites.Our results suggest that impaired effector functions, increased activation and immaturity of neutrophils play a key role in the pathogenesis of VL.

  6. Hypertonic Saline Suppresses NADPH Oxidase-Dependent Neutrophil Extracellular Trap Formation and Promotes Apoptosis

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    Ajantha Nadesalingam

    2018-03-01

    Full Text Available Tonicity of saline (NaCl is important in regulating cellular functions and homeostasis. Hypertonic saline is administered to treat many inflammatory diseases, including cystic fibrosis. Excess neutrophil extracellular trap (NET formation, or NETosis, is associated with many pathological conditions including chronic inflammation. Despite the known therapeutic benefits of hypertonic saline, its underlying mechanisms are not clearly understood. Therefore, we aimed to elucidate the effects of hypertonic saline in modulating NETosis. For this purpose, we purified human neutrophils and induced NETosis using agonists such as diacylglycerol mimetic phorbol myristate acetate (PMA, Gram-negative bacterial cell wall component lipopolysaccharide (LPS, calcium ionophores (A23187 and ionomycin from Streptomyces conglobatus, and bacteria (Pseudomonas aeruginosa and Staphylococcus aureus. We then analyzed neutrophils and NETs using Sytox green assay, immunostaining of NET components and apoptosis markers, confocal microscopy, and pH sensing reagents. This study found that hypertonic NaCl suppresses nicotinamide adenine dinucleotide phosphate oxidase (NADPH2 or NOX2-dependent NETosis induced by agonists PMA, Escherichia coli LPS (0111:B4 and O128:B12, and P. aeruginosa. Hypertonic saline also suppresses LPS- and PMA- induced reactive oxygen species production. It was determined that supplementing H2O2 reverses the suppressive effect of hypertonic saline on NOX2-dependent NETosis. Many of the aforementioned suppressive effects were observed in the presence of equimolar concentrations of choline chloride and osmolytes (d-mannitol and d-sorbitol. This suggests that the mechanism by which hypertonic saline suppresses NOX2-dependent NETosis is via neutrophil dehydration. Hypertonic NaCl does not significantly alter the intracellular pH of neutrophils. We found that hypertonic NaCl induces apoptosis while suppressing NOX2-dependent NETosis. In contrast, hypertonic

  7. The tripeptide feG regulates the production of intracellular reactive oxygen species by neutrophils

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    Davison Joseph S

    2006-06-01

    Full Text Available Abstract Background The D-isomeric form of the tripeptide FEG (feG is a potent anti-inflammatory agent that suppresses type I hypersensitivity (IgE-mediated allergic reactions in several animal species. One of feG's primary actions is to inhibit leukocyte activation resulting in loss of their adhesive and migratory properties. Since activation of neutrophils is often associated with an increase in respiratory burst with the generation of reactive oxygen species (ROS, we examined the effect of feG on the respiratory burst in neutrophils of antigen-sensitized rats. A role for protein kinase C (PKC in the actions of feG was evaluated by using selective isoform inhibitors for PKC. Results At 18h after antigen (ovalbumin challenge of sensitized Sprague-Dawley rats a pronounced neutrophilia occurred; a response that was reduced in animals treated with feG (100 μg/kg. With antigen-challenged animals the protein kinase C (PKC activator, PMA, significantly increased intracellular ROS of circulating neutrophils, as determined by flow cytometry using the fluorescent probe dihydrorhodamine-123. This increase was prevented by treatment with feG at the time of antigen challenge. The inhibitor of PKCδ, rottlerin, which effectively prevented intracellular ROS production by circulating neutrophils of animals receiving a naïve antigen, failed to inhibit PMA-stimulated ROS production if the animals were challenged with antigen. feG treatment, however, re-established the inhibitory effects of the PKCδ inhibitor on intracellular ROS production. The extracellular release of superoxide anion, evaluated by measuring the oxidative reduction of cytochrome C, was neither modified by antigen challenge nor feG treatment. However, hispidin, an inhibitor of PKCβ, inhibited the release of superoxide anion from circulating leukocytes in all groups of animals. feG prevented the increased expression of the β1-integrin CD49d on the circulating neutrophils elicited by antigen

  8. Neutrophil proteomic analysis reveals the participation of antioxidant enzymes, motility and ribosomal proteins in the prevention of ischemic effects by preconditioning

    DEFF Research Database (Denmark)

    Arshid, S; Tahir, M; Fontes, B

    2017-01-01

    therapeutic application; however the exact underlying mechanism is not clear. Neutrophils play an important role in the mechanism of injuries caused by ischemia and reperfusion while IPC led to a decrease in neutrophil stimulation and activation. The effect of preconditioning on the neutrophil proteome...... in such conditions, there is no report of a proteomic study providing a broader view of this scenario. Here we describe a group of proteins significantly regulated by ischemia and reperfusion being such regulation prevented by preconditioning. Such finding may provide relevant information for a deeper understanding...

  9. Gene deletion of P-Selectin and ICAM-1 does not inhibit neutrophil infiltration into peritoneal cavity following cecal ligation-puncture

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    Hess Karen

    2004-07-01

    Full Text Available Abstract Background Neutrophil infiltration is one of the critical cellular components of an inflammatory response during peritonitis. The adhesion molecules, P-selectin and intercellular adhesion molecule (ICAM-1, mediate neutrophil-endothelial cell interactions and the subsequent neutrophil transendothelial migration during the inflammatory response. Despite very strong preclinical data, recent clinical trials failed to show a protective effect of anti-adhesion therapy, suggesting that the length of injury might be a critical factor in neutrophil infiltration. Therefore, the objective of this study was to determine the role of P-selectin and ICAM-1 in neutrophil infiltration into the peritoneal cavity during early and late phases of peritonitis. Methods Peritonitis was induced in both male wild-type and P-selectin/ICAM-1 double deficient (P/I null mice by cecal ligation-puncture (CLP. Peripheral blood and peritoneal lavage were collected at 6 and 24 hours after CLP. The total leukocyte and neutrophil contents were determined, and neutrophils were identified with the aid of in situ immunohistochemical staining. Comparisons between groups were made by applying ANOVA and student t-test analysis. Results CLP induced a severe inflammatory response associated with a significant leukopenia in both wild-type and P/I null mice. Additionally, CLP caused a significant neutrophil infiltration into the peritoneal cavity that was detected in both groups of mice. However, neutrophil infiltration in the P/I null mice at 6 hours of CLP was significantly lower than the corresponding wild-type mice, which reached a similar magnitude at 24 hours of CLP. In contrast, in peritonitis induced by intraperitoneal inoculation of 2% glycogen, no significant difference in neutrophil infiltration was observed between the P/I null and wild-type mice at 6 hours of peritonitis. Conclusions The data suggest that alternative adhesion pathway(s independent of P-selectin and ICAM

  10. Potent inhibition of human neutrophil activations by bractelactone, a novel chalcone from Fissistigma bracteolatum

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    Wu, Yang-Chang [Graduate Institute of Natural Products, Kaohsiung Medical University, Kaohsiung 807, Taiwan (China); Graduate Institute of Integrated Medicine, College of Chinese Medicine, China Medical University, Taichung 404, Taiwan (China); Sureshbabu, Munisamy; Fang, Yao-Ching; Wu, Yi-Hsiu [Graduate Institute of Natural Products, College of Medicine, Chang Gung University, Taoyuan 333, Taiwan (China); Lan, Yu-Hsuan [School of Pharmacy, China Medical University, Taichung 404, Taiwan (China); Chang, Fang-Rong [Graduate Institute of Natural Products, Kaohsiung Medical University, Kaohsiung 807, Taiwan (China); Chang, Ya-Wen [Graduate Institute of Natural Products, College of Medicine, Chang Gung University, Taoyuan 333, Taiwan (China); Hwang, Tsong-Long, E-mail: htl@mail.cgu.edu.tw [Graduate Institute of Natural Products, College of Medicine, Chang Gung University, Taoyuan 333, Taiwan (China); Chinese Herbal Medicine Research Team, Healthy Aging Research Center, Chang Gung University, Kweishan, Taoyuan 333, Taiwan (China)

    2013-02-01

    Fissistigma bracteolatum is widely used in traditional medicine to treat inflammatory diseases. However, its active components and mechanisms of action remain unclear. In this study, (3Z)-6,7-dihydroxy-4-methoxy-3-(phenylmethylidene)-5-(3-phenylpropanoyl) -1-benzofuran-2(3H) (bractelactone), a novel chalcone from F. bracteolatum, showed potent inhibitory effects against superoxide anion (O{sub 2}{sup ·−}) production, elastase release, and CD11b expression in formyl-L-methionyl-L-leucyl-L-phenylalanine (FMLP)-induced human neutrophils. However, bractelactone showed only weak inhibition of phorbol myristate acetate-caused O{sub 2}{sup ·−} production. The peak cytosolic calcium concentration ([Ca{sup 2+}]{sub i}) was unaltered by bractelactone in FMLP-induced neutrophils, but the decay time of [Ca{sup 2+}]{sub i} was significantly shortened. In a calcium-free solution, changes in [Ca{sup 2+}]{sub i} caused by the addition of extracellular Ca{sup 2+} were inhibited by bractelactone in FMLP-activated cells. In addition, bractelactone did not alter the phosphorylation of p38 MAPK, ERK, JNK, or AKT or the concentration of cAMP. These results suggest that bractelactone selectively inhibits store-operated calcium entry (SOCE). In agreement with this concept, bractelactone suppressed sustained [Ca{sup 2+}]{sub i} changes in thapsigargin-activated neutrophils. Furthermore, bractelactone did not alter FMLP-induced formation of inositol 1,4,5-triphosphate. Taken together, our results demonstrate that the anti-inflammatory effects of bractelactone, an active ingredient of F. bracteolatum, in human neutrophils are through the selective inhibition of SOCE. Highlights: ► Bractelactone isolated from Fissistigma bracteolatum. ► Bractelactone inhibited FMLP-induced human neutrophil activations. ► Bractelactone had no effect on IP3 formation. ► Bractelactone did not alter MAPKs, AKT, and cAMP pathways. ► Bractelactone inhibited store-operated calcium entry.

  11. Novel insight into neutrophil immune responses by dry mass determination of Candida albicans morphotypes.

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    Ava Hosseinzadeh

    Full Text Available The common fungal pathogen Candida albicans has the ability to grow as a yeast or as a hypha and can alternate between these morphotypes. The overall biomass of both morphotypes increases with growth. However, only yeasts, but not hyphae, exist as discrete cellular entities. Multiplicity of infection (MOI is a useful parameter to determine the initial inoculum of yeasts for in vitro infection assays. Since the amount of hyphae is difficult to quantify, comparable starting conditions in such assays cannot be determined accurately for yeasts and hyphae using MOI. To circumvent this problem, we have established a set of correlation coefficients to convert fungal metabolic activity and optical density to dry mass. Using these correlations, we were able to accurately compare ROS production and IL-8 release by polymorphonuclear neutrophils upon infection with equal dry mass amounts of yeast and hyphal morphotypes. Neutrophil responses depended on the initial form of infection, irrespective of C. albicans wild-type yeasts transforming to hyphal growth during the assay. Infection with a high mass of live C. albicans yeasts resulted in lower neutrophil ROS and this decrease stems from efficient ROS detoxification by C. albicans without directly affecting the phagocyte ROS machinery. Moreover, we show that dead C. albicans induces significantly less ROS and IL-8 release than live fungi, but thimerosal-killed C. albicans were still able to detoxify neutrophil ROS. Thus, the dry mass approach presented in this study reveals neutrophil responses to different amounts and morphotypes of C. albicans and serves as a template for studies that aim to identify morphotype-specific responses in a variety of immune cells.

  12. Sepsis Induces a Dysregulated Neutrophil Phenotype That Is Associated with Increased Mortality

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    Jaimin M. Patel

    2018-01-01

    Full Text Available Background. Neutrophil dysfunction in sepsis has been implicated in the pathogenesis of multiorgan failure; however, the role of neutrophil extracellular traps (NETs remains uncertain. We aimed to determine the sequential changes in ex vivo NETosis and its relationship with mortality in patients with sepsis and severe sepsis. Methods. This was a prospective observational cohort study enrolling 21 healthy age-matched controls and 39 sepsis and 60 severe sepsis patients from acute admissions to two UK hospitals. Patients had sequential bloods for the ex vivo assessment of NETosis in response to phorbol-myristate acetate (PMA using a fluorometric technique and chemotaxis using time-lapse video microscopy. Continuous data was tested for normality, with appropriate parametric and nonparametric tests, whilst categorical data was analysed using a chi-squared test. Correlations were performed using Spearman’s rho. Results. Ex vivo NETosis was reduced in patients with severe sepsis, compared to patients with sepsis and controls (p=0.002. PMA NETosis from patients with septic shock was reduced further (p<0.001 compared to controls. The degree of metabolic acidosis correlated with reduced NETosis (p<0.001, and this was replicated when neutrophils from healthy donors were incubated in acidotic media. Reduced NETosis at baseline was associated with an increased 30-day (p=0.002 and 90-day mortality (p=0.014 in sepsis patients. These findings were accompanied by defects in neutrophil migration and delayed apoptosis. Resolution of sepsis was not associated with the return to baseline levels of NETosis or migration. Conclusions. Sepsis induces significant changes in neutrophil function with the degree of dysfunction corresponding to the severity of the septic insult which persists beyond physiological recovery from sepsis. The changes induced lead to the failure to effectively contain and eliminate the invading pathogens and contribute to sepsis

  13. Quantifying oral inflammatory load: oral neutrophil counts in periodontal health and disease.

    Science.gov (United States)

    Landzberg, M; Doering, H; Aboodi, G M; Tenenbaum, H C; Glogauer, M

    2015-06-01

    Neutrophils are the primary white blood cells that are recruited to fight the initial phases of microbial infections. While healthy norms have been determined for circulating blood neutrophil counts in order to identify patients with suspected systemic infections, the levels of oral neutrophils (oPMNs) in oral health and in the presence of periodontal diseases have not been described. It is important to address this deficiency in our knowledge as neutrophils are the primary immune cell present in the crevicular fluid and oral environment and previous work has suggested that they may be good indicators of overall oral inflammation and periodontal disease severity. The objective of this study was to measure oPMN counts obtained in a standardized oral rinse from healthy patients and from those with chronic periodontal disease in order to determine if oPMN levels have clinical relevance as markers of periodontal inflammation. A parallel goal of this investigation was to introduce the concept of 'oral inflammatory load', which constitutes the inflammatory burden experienced by the body as a consequence of oral inflammatory disease. Periodontal examinations of patients with a healthy periodontium and chronic periodontal disease were performed (n = 124). Two standardized consecutive saline rinses of 30 s each were collected before patient examination and instrumentation. Neutrophils were quantified in the rinse samples and correlated with the clinical parameters and periodontal diagnosis. Average oPMN counts were determined for healthy patients and for those with mild, moderate and severe chronic periodontal diseases. A statistically significant correlation was found between oPMN counts and deep periodontal probing, sites with bleeding on probing and overall severity of periodontal disease. oPMN counts obtained through a 30-s oral rinse are a good marker of oral inflammatory load and correlate with measures of periodontal disease severity. © 2014 John Wiley & Sons A

  14. Elevated Systemic Levels of Eosinophil, Neutrophil, and Mast Cell Granular Proteins in Strongyloides Stercoralis Infection that Diminish following Treatment.

    Science.gov (United States)

    Rajamanickam, Anuradha; Munisankar, Saravanan; Bhootra, Yukthi; Dolla, Chandra Kumar; Nutman, Thomas B; Babu, Subash

    2018-01-01

    Infection with the helminth parasite Strongyloides stercoralis ( Ss ) is commonly clinically asymptomatic that is often accompanied by peripheral eosinophilia. Granulocytes are activated during helminth infection and can act as immune effector cells. Plasma levels of eosinophil and neutrophil granular proteins convey an indirect measure of granulocyte degranulation and are prominently augmented in numerous helminth-infected patients. In this study, we sought to examine the levels of eosinophil, neutrophil, and mast cell activation-associated granule proteins in asymptomatic Ss infection and to understand their kinetics following anthelmintic therapy. To this end, we measured the plasma levels of eosinophil cationic protein, eosinophil-derived neurotoxin, eosinophil peroxidase, eosinophil major basic protein, neutrophil elastase, myeloperoxidase, neutrophil proteinase-3, mast cell tryptase, leukotriene C4, and mast cell carboxypeptidase-A3 in individuals with asymptomatic Ss infection or without Ss infection [uninfected (UN)]. We also estimated the levels of all of these analytes in infected individuals following definitive treatment of Ss infection. We demonstrated that those infected individuals have significantly enhanced plasma levels of eosinophil cationic protein, eosinophil-derived neurotoxin, eosinophil peroxidase, eosinophil major basic protein, elastase, myeloperoxidase, mast cell tryptase, leukotriene C4, and carboxypeptidase-A3 compared to UN individuals. Following the treatment of Ss infection, each of these granulocyte-associated proteins drops significantly. Our data suggest that eosinophil, neutrophil, and mast cell activation may play a role in the response to Ss infection.

  15. The Application of Dextran Sedimentation as an Initial Step in Neutrophil Purification Promotes Their Stimulation, due to the Presence of Monocytes

    Science.gov (United States)

    Ferrante, Antonio

    2017-01-01

    The purification of human neutrophils for in vitro studies is challenging as they are easily activated through ex vivo manipulations. The technique of erythrocyte sedimentation combined with density gradient centrifugation remains widely practiced and was the subject of this study. Since in the sedimentation step the leukocytes are incubated with dextran, we have raised the likelihood that cellular activation would occur with mediator release leading to neutrophil activation. By comparing the activity of neutrophils purified from whole blood by the classical 2-step method of dextran sedimentation followed by low-density Ficoll-Hypaque (1.077 g/mL) medium, and the 1-step high-density Ficoll-Hypaque (1.114 g/mL) gradient centrifugation, we found that neutrophils from the 2-step method had a significant increase in cell surface CD11b expression and CD62L shedding and a marked increase in adhesion. Decreased random migration and chemotaxis and raised baseline oxidative burst activity were also observed. The effect was not specific to dextran, as using Ficoll for erythrocyte sedimentation replicated the elevated neutrophil adherence. Through the depletion of monocytes, lymphocytes, and platelets prior to sedimentation, we deduced that monocytes were responsible for the neutrophil activation. Our findings suggest that care needs to be exercised in choosing the method of neutrophil purification for functional studies. PMID:29164154

  16. On the mechanism of oscillations in neutrophils

    DEFF Research Database (Denmark)

    Brasen, Jens Christian; Barington, Torben; Olsen, Lars Folke

    2010-01-01

    We have investigated the regulation of the oscillatory generation of H(2)O(2) and oscillations in shape and size in neutrophils in suspension. The oscillations are independent of cell density and hence do not represent a collective phenomena. Furthermore, the oscillations are independent...... of the external glucose concentration and the oscillations in H(2)O(2) production are 180 degrees out of phase with the oscillations in NAD(P)H. Cytochalasin B blocked the oscillations in shape and size whereas it increased the period of the oscillations in H(2)O(2) production. 1- and 2-butanol also blocked...... the oscillations in shape and size, but only 1-butanol inhibited the oscillations in H(2)O(2) production. We conjecture that the oscillations are likely to be due to feedback regulations in the signal transduction cascade involving phosphoinositide 3-kinases (PI3K). We have tested this using a simple mathematical...

  17. An Archaeosome-Adjuvanted Vaccine and Checkpoint Inhibitor Therapy Combination Significantly Enhances Protection from Murine Melanoma

    Directory of Open Access Journals (Sweden)

    Felicity C. Stark

    2017-10-01

    Full Text Available Archaeosomes constitute archaeal lipid vesicle vaccine adjuvants that evoke a strong CD8+ T cell response to antigenic cargo. Therapeutic treatment of murine B16-ovalbumin (B16-OVA melanoma with archaeosome-OVA eliminates small subcutaneous solid tumors; however, they eventually resurge despite an increased frequency of circulating and tumor infiltrating OVA-CD8+ T cells. Herein, a number of different approaches were evaluated to improve responses, including dose number, interval, and the combination of vaccine with checkpoint inhibitors. Firstly, we found that tumor protection could not be enhanced by repetitive and/or delayed boosting to maximize the CD8+ T cell number and/or phenotype. The in vivo cytotoxicity of vaccine-induced OVA-CD8+ T cells was impaired in tumor-bearing mice. Additionally, tumor-infiltrating OVA-CD8+ T cells had an increased expression of programmed cell death protein-1 (PD-1 compared to other organ compartments, suggesting impaired function. Combination therapy of tumor-bearing mice with the vaccine archaeosome-OVA, and α-CTLA-4 administered concurrently as well as α-PD-1 and an α-PD-L1 antibody administered starting 9 days after tumor challenge given on a Q3Dx4 schedule (days 9, 12, 15 and 18, significantly enhanced survival. Following multi-combination therapy ~70% of mice had rapid tumor recession, with no detectable tumor mass after >80 days in comparison to a median survival of 17–22 days for untreated or experimental groups receiving single therapies. Overall, archaeosomes offer a powerful platform for delivering cancer antigens when used in combination with checkpoint inhibitor immunotherapies.

  18. Neutrophil transmigration mediated by the neutrophil-specific antigen CD177 is influenced by the endothelial S536N dimorphism of platelet endothelial cell adhesion molecule-1.

    Science.gov (United States)

    Bayat, Behnaz; Werth, Silke; Sachs, Ulrich J H; Newman, Debra K; Newman, Peter J; Santoso, Sentot

    2010-04-01

    The human neutrophil-specific adhesion molecule CD177 (also known as the NB1 alloantigen) becomes upregulated on the cell surface in a number of inflammatory settings. We recently showed that CD177 functions as a novel heterophilic counterreceptor for the endothelial junctional protein PECAM-1 (CD31), an interaction that is mediated by membrane-proximal PECAM-1 IgD 6, which is known to harbor an S(536)N single nucleotide polymorphism of two major isoforms V(98)N(536)G(643) and L(98)S(536)R(643) and a yet-to-be-determined region on CD177. In vitro transendothelial migration experiments revealed that CD177(+) neutrophils migrated significantly faster through HUVECs expressing the LSR, compared with the VNG, allelic variant of PECAM-1 and that this correlated with the decreased ability of anti-PECAM-1 Ab of ITIM tyrosine phosphorylation in HUVECs expressing the LSR allelic variant relative to the VNG allelic variant. Moreover, engagement of PECAM-1 with rCD177-Fc (to mimic heterophilic CD177 binding) suppressed Ab-induced tyrosine phosphorylation to a greater extent in cells expressing the LSR isoform compared with the VNG isoform, with a corresponding increased higher level of beta-catenin phosphorylation. These data suggest that heterophilic PECAM-1/CD177 interactions affect the phosphorylation state of PECAM-1 and endothelial cell junctional integrity in such a way as to facilitate neutrophil transmigration in a previously unrecognized allele-specific manner.

  19. Pseudomonas aeruginosa ExoU augments neutrophil transepithelial migration.

    Science.gov (United States)

    Pazos, Michael A; Lanter, Bernard B; Yonker, Lael M; Eaton, Alex D; Pirzai, Waheed; Gronert, Karsten; Bonventre, Joseph V; Hurley, Bryan P

    2017-08-01

    Excessive neutrophil infiltration of the lungs is a common contributor to immune-related pathology in many pulmonary disease states. In response to pathogenic infection, airway epithelial cells produce hepoxilin A3 (HXA3), initiating neutrophil transepithelial migration. Migrated neutrophils amplify this recruitment by producing a secondary gradient of leukotriene B4 (LTB4). We sought to determine whether this two-step eicosanoid chemoattractant mechanism could be exploited by the pathogen Pseudomonas aeruginosa. ExoU, a P. aeruginosa cytotoxin, exhibits phospholipase A2 (PLA2) activity in eukaryotic hosts, an enzyme critical for generation of certain eicosanoids. Using in vitro and in vivo models of neutrophil transepithelial migration, we evaluated the impact of ExoU expression on eicosanoid generation and function. We conclude that ExoU, by virtue of its PLA2 activity, augments and compensates for endogenous host neutrophil cPLA2α function, leading to enhanced transepithelial migration. This suggests that ExoU expression in P. aeruginosa can circumvent immune regulation at key signaling checkpoints in the neutrophil, resulting in exacerbated neutrophil recruitment.

  20. Pseudomonas aeruginosa ExoU augments neutrophil transepithelial migration.

    Directory of Open Access Journals (Sweden)

    Michael A Pazos

    2017-08-01

    Full Text Available Excessive neutrophil infiltration of the lungs is a common contributor to immune-related pathology in many pulmonary disease states. In response to pathogenic infection, airway epithelial cells produce hepoxilin A3 (HXA3, initiating neutrophil transepithelial migration. Migrated neutrophils amplify this recruitment by producing a secondary gradient of leukotriene B4 (LTB4. We sought to determine whether this two-step eicosanoid chemoattractant mechanism could be exploited by the pathogen Pseudomonas aeruginosa. ExoU, a P. aeruginosa cytotoxin, exhibits phospholipase A2 (PLA2 activity in eukaryotic hosts, an enzyme critical for generation of certain eicosanoids. Using in vitro and in vivo models of neutrophil transepithelial migration, we evaluated the impact of ExoU expression on eicosanoid generation and function. We conclude that ExoU, by virtue of its PLA2 activity, augments and compensates for endogenous host neutrophil cPLA2α function, leading to enhanced transepithelial migration. This suggests that ExoU expression in P. aeruginosa can circumvent immune regulation at key signaling checkpoints in the neutrophil, resulting in exacerbated neutrophil recruitment.

  1. Biomaterial-induced alterations of neutrophil superoxide production.

    Science.gov (United States)

    Kaplan, S S; Basford, R E; Mora, E; Jeong, M H; Simmons, R L

    1992-08-01

    Because periprosthetic infection remains a vexing problem for patients receiving implanted devices, we evaluated the effect of several materials on neutrophil free radical production. Human peripheral blood neutrophils were incubated with several sterile, lipopolysaccharide (LPS)-free biomaterials used in surgically implantable prosthetic devices: polyurethane, woven dacron, and velcro. Free radical formation as the superoxide (O2-) anion was evaluated by cytochrome c reduction in neutrophils that were exposed to the materials and then removed and in neutrophils allowed to remain in association with the materials. Neutrophils exposed to polyurethane or woven dacron for 30 or 60 min and then removed consistently exhibited an enhanced release of O2- after simulation via receptor engagement with formyl methionyl-leucyl-phenylalanine. Enhanced reactivity to stimulation via protein kinase C with phorbol myristate acetate, however, was not consistently observed. The cells evaluated for O2- release during continuous association with the biomaterials showed enhanced metabolic activity during short periods of association (especially with polyurethane and woven dacron). Although O2- release by neutrophils in association with these materials decreased with longer periods of incubation, it was not obliterated. These studies, therefore, show that several commonly used biomaterials activate neutrophils soon after exposure and that this activated state diminishes with prolonged exposure but nevertheless remains measurable. The diminishing level of activity with prolonged exposure, however, suggests that ultimately a depletion of reactivity may occur and may result in increased susceptibility to periprosthetic infection.

  2. Neutrophils in oral paracoccidioidomycosis and the involvement of Nrf2.

    Directory of Open Access Journals (Sweden)

    Vera Cavalcanti Araújo

    Full Text Available Neutrophils have been implicated in granuloma formation in several infectious diseases, in addition to their main phagocytic and pathogen destruction role. It has been demonstrated that Nrf2 regulates antioxidant protection in neutrophils, attenuating inflammation without compromising the hosts bacterial defense. In this study, we analyzed the presence of neutrophils in Paracoccidioides brasiliensis mycosis (PCM, as well as the immunoexpression of Nrf2. Thirty-nine cases of oral PCM were classified according to quantity of fungi and to the presence of loose or well-organized granulomas and microabscesses. An Nrf2 antibody was used for immunohistochemical analysis. The results showed that neutrophils are present in microabscesses and loose granulomas, but were absent in structured granulomas. A greater quantity of fungi was shown in cases with only loose granulomas when compared to loose and well organized granulomas. Nrf2 was observed in the nuclei of neutrophils of loose granulomas and abscesses, with its expression in loose granulomas maintained despite the additional presence of well organized granulomas in the same specimen. This study suggests that neutrophils participate in P. brasiliensis granuloma formation and that Nrf2 has a possible role in neutrophil survival, via modulation of the inflammatory response.

  3. Tissue-transglutaminase contributes to neutrophil granulocyte differentiation and functions.

    Science.gov (United States)

    Balajthy, Zoltán; Csomós, Krisztián; Vámosi, György; Szántó, Attila; Lanotte, Michel; Fésüs, László

    2006-09-15

    Promyelocytic NB4 leukemia cells undergo differentiation to granulocytes following retinoic acid treatment. Here we report that tissue transglutaminase (TG2), a protein cross-linking enzyme, was induced, then partially translocated into the nucleus, and became strongly associated with the chromatin during the differentiation process. The transglutaminase-catalyzed cross-link content of both the cytosolic and the nuclear protein fractions increased while NB4 cells underwent cellular maturation. Inhibition of cross-linking activity of TG2 by monodansylcadaverin in these cells led to diminished nitroblue tetrazolium (NBT) positivity, production of less superoxide anion, and decreased expression of GP91PHOX, the membrane-associated subunit of NADPH oxidase. Neutrophils isolated from TG2(-/-) mice showed diminished NBT reduction capacity, reduced superoxide anion formation, and down-regulation of the gp91phox subunit of NADPH oxidase, compared with wild-type cells. It was also observed that TG2(-/-) mice exhibited increased neutrophil phagocytic activity, but had attenuated neutrophil chemotaxis and impaired neutrophil extravasation with higher neutrophil counts in their circulation during yeast extract-induced peritonitis. These results clearly suggest that TG2 may modulate the expression of genes related to neutrophil functions and is involved in several intracellular and extracellular functions of extravasating neutrophil.

  4. Activated signature of antiphospholipid syndrome neutrophils reveals potential therapeutic target

    Science.gov (United States)

    Knight, Jason S.; Meng, He; Coit, Patrick; Yalavarthi, Srilakshmi; Sule, Gautam; Gandhi, Alex A.; Grenn, Robert C.; Mazza, Levi F.; Ali, Ramadan A.; Renauer, Paul; Wren, Jonathan D.; Bockenstedt, Paula L.; Wang, Hui; Eitzman, Daniel T.; Sawalha, Amr H.

    2017-01-01

    Antiphospholipid antibodies, present in one-third of lupus patients, increase the risk of thrombosis. We recently reported a key role for neutrophilsneutrophil extracellular traps (NETs), in particular — in the thrombotic events that define antiphospholipid syndrome (APS). To further elucidate the role of neutrophils in APS, we performed a comprehensive transcriptome analysis of neutrophils isolated from patients with primary APS. Moreover, APS-associated venous thrombosis was modeled by treating mice with IgG prepared from APS patients, followed by partial restriction of blood flow through the inferior vena cava. In patients, APS neutrophils demonstrated a proinflammatory signature with overexpression of genes relevant to IFN signaling, cellular defense, and intercellular adhesion. For in vivo studies, we focused on P-selectin glycoprotein ligand-1 (PSGL-1), a key adhesion molecule overexpressed in APS neutrophils. The introduction of APS IgG (as compared with control IgG) markedly potentiated thrombosis in WT mice, but not PSGL-1–KOs. PSGL-1 deficiency was also associated with reduced leukocyte vessel wall adhesion and NET formation. The thrombosis phenotype was restored in PSGL-1–deficient mice by infusion of WT neutrophils, while an anti–PSGL-1 monoclonal antibody inhibited APS IgG–mediated thrombosis in WT mice. PSGL-1 represents a potential therapeutic target in APS. PMID:28931754

  5. Phenotypic changes in neutrophils related to anti-inflammatory therapy.

    Science.gov (United States)

    Barton, A E; Bayley, D L; Mikami, M; Llewellyn-Jones, C G; Stockley, R A

    2000-01-03

    Previous work from the group has shown that non-steroidal anti-inflammatory agents given to volunteers and patients inhibit PMN function possibly by affecting the developing neutrophil during the differentiation process. In this study indomethacin treatment in vivo reduced neutrophil chemotaxis and proteolytic degradation of fibronectin, with a maximal effect after 14 days. Stimulated neutrophil adherence to fibronectin was also reduced but this was not due to quantitative changes in beta(2) integrin expression or function. L-Selectin expression on resting and stimulated neutrophils was increased after 14 days and there was a small decrease in plasma levels of soluble L-selectin. These effects, however, could not be reproduced by treatment of neutrophils with indomethacin in vitro, suggesting they are due to effects on differentiating/maturing PMNs. In an attempt to interpret these changes, studies were performed with dexamethasone, which is known to alter neutrophil function and kinetics. Dexamethasone treatment reduced chemotaxis and increased superoxide generation after 1 day and was associated with increased expression of activated beta(2) integrins and reduced L-selectin expression on resting neutrophils. This suggests the appearance of mainly 'activated' cells as a result of demargination and indicates that the effects of indomethacin are distinctive and not related to changes in compartmentalisation.

  6. Heterogeneity in Neutrophil Microparticles Reveals Distinct Proteome and Functional Properties*

    Science.gov (United States)

    Dalli, Jesmond; Montero-Melendez, Trinidad; Norling, Lucy V; Yin, Xiaoke; Hinds, Charles; Haskard, Dorian; Mayr, Manuel; Perretti, Mauro

    2013-01-01

    Altered plasma neutrophil microparticle levels have recently been implicated in a number of vascular and inflammatory diseases, yet our understanding of their actions is very limited. Herein, we investigate the proteome of neutrophil microparticles in order to shed light on their biological actions. Stimulation of human neutrophils, either in suspension or adherent to an endothelial monolayer, led to the production of microparticles containing >400 distinct proteins with only 223 being shared by the two subsets. For instance, postadherent microparticles were enriched in alpha-2 macroglobulin and ceruloplasmin, whereas microparticles produced by neutrophils in suspension were abundant in heat shock 70 kDa protein 1. Annexin A1 and lactotransferrin were expressed in both microparticle subsets. We next determined relative abundance of these proteins in three types of human microparticle samples: healthy volunteer plasma, plasma of septic patients and skin blister exudates finding that these proteins were differentially expressed on neutrophil microparticles from these samples reflecting in part the expression profiles we found in vitro. Functional assessment of the neutrophil microparticles subsets demonstrated that in response to direct stimulation neutrophil microparticles produced reactive oxygen species and leukotriene B4 as well as locomoted toward a chemotactic gradient. Finally, we investigated the actions of the two neutrophil microparticles subsets described herein on target cell responses. Microarray analysis with human primary endothelial cells incubated with either microparticle subset revealed a discrete modulation of endothelial cell gene expression profile. These findings demonstrate that neutrophil microparticles are heterogenous and can deliver packaged information propagating the activation status of the parent cell, potentially exerting novel and fundamental roles both under homeostatic and disease conditions. PMID:23660474

  7. Influence of exogenous leptin on redox homeostasis in neutrophils and lymphocytes cultured in synovial fluid isolated from patients with rheumatoid arthritis

    Directory of Open Access Journals (Sweden)

    Michał Gajewski

    2016-07-01

    Full Text Available Objectives : Leptin is an adipose cells derived hormone that regulates energy homeostasis within the body. Energy metabolism of immune cells influences their activity within numerous pathological states, but the effect of leptin on these cells in unclear. On the one hand, it was observed that leptin induces neutrophils chemotaxis and modulates phagocytosis. On the other hand, neutrophils exposed to leptin did not display detectable Ca 2+ ions mobilization or β 2 -integrin upregulation. In this study, we investigated the effect of leptin on the redox homeostasis in lymphocytes and neutrophils. Material and methods : Neutrophils and lymphocytes were isolated by density-gradient centrifugation of blood from healthy volunteers. Cells were cultured with or without leptin (100 ng/ml for lymphocytes and 500 ng/ml for neutrophils or with or without synovial fluid (85% for 0–72 h. Culture media were not changed during incubation. Cells were homogenized and homogenate was frozen until laboratory measurements. Redox homeostasis was assessed by the reduced glutathione (GSH vs. oxidized glutathione (GSSG ratio and membrane lipid peroxidation evaluation. Results : Lymphocytes cultured with leptin and synovial fluid showed a significant increase of the GSSG level. The GSSG/GSH ratio increased by 184 ±37%. In neutrophils incubated in a similar environment, the GSSG/GSH ratio increased by just 21 ±7%, and the effect was observed irrespectively of whether they were exposed to leptin or synovial fluid or both together. Neither leptin nor synovial fluid influenced lipid peroxidation in neutrophils, but in lymphocytes leptin intensified lipid peroxidation. Conclusions : Leptin altered the lymphocytes, but not neutrophils redox state. Because firstly neutrophils are anaerobic cells and have just a few mitochondria and secondly lymphocytes have typical aerobic metabolism, the divergence of our data supports the hypothesis that leptin induces oxidative stress by

  8. Predictors of neutrophilic airway inflammation in young smokers with asthma

    DEFF Research Database (Denmark)

    Westergaard, Christian Grabow; Munck, Christian; Helby, Jens

    2014-01-01

    by a higher degree of neutrophilic inflammation than in non-smokers. A state of neutrophilic inflammation may lead to increased steroid resistance and an accelerated loss of lung function owing to tissue destruction. The aim of this study was to elucidate predictors of neutrophilic inflammation in young...... asthmatic smokers not on steroid treatment, including analysis of tobacco history and bacterial colonization. Methods: In a cross-sectional study, 52 steroid-free, current smokers with asthma were examined with induced sputum, fractional exhaled nitric oxide (FeNO), lung function, ACQ6 score, mannitol...... smokers, neutrophilia may be induced when a certain threshold of tobacco consumption is reached....

  9. Effects of Acer okamotoanum sap on the function of polymorphonuclear neutrophilic leukocytes in vitro and in vivo.

    Science.gov (United States)

    An, Beum-Soo; Kang, Ji-Houn; Yang, Hyun; Yang, Mhan-Pyo; Jeung, Eui-Bae

    2013-02-01

    Sap is a plant fluid that primarily consists of water and small amounts of mineral elements, sugars, hormones and other nutrients. Acer mono (A. mono) is an endemic Korean mono maple which was recently suggested to have health benefits due to its abundant calcium and magnesium ion content. In the present study, we examined the effects of sap from Acer okamotoanum (A. okamotoanum) on the phagocytic response of mouse neutrophils in vivo and rat and canine neutrophils in vitro. We tested the regulation of phagocytic activity, oxidative burst activity (OBA) and the levels of filamentous polymeric actin (F-actin) in the absence and presence of dexamethasone (DEX) in vitro and in vivo. Our results showed that DEX primarily reduced OBA in the mouse neutrophils, and that this was reversed in the presence of the sap. By contrast, the phagocytic activity of the mouse cells was not regulated by either DEX or the sap. Rat and canine polymorphonuclear neutrophilic leukocytes (PMNs) responded in vitro to the sap in a similar manner by increasing OBA. However, regulation of phagocytic activity by the sap was different between the species. In canine PMNs, phagocytic activity was enhanced by the sap at a high dose, while it did not significantly modulate this activity in rat PMNs. These findings suggest that the sap of A. okamotoanum stimulates neutrophil activity in the mouse, rat and canine by increasing OBA in vivo and in vitro, and thus may have a potential antimicrobial effect in the PMNs of patients with infections.

  10. The Response of Macrophages and Neutrophils to Hypoxia in the Context of Cancer and Other Inflammatory Diseases

    Directory of Open Access Journals (Sweden)

    Antje Egners

    2016-01-01

    Full Text Available Lack of oxygen (hypoxia is a hallmark of a multitude of acute and chronic diseases and can be either beneficial or detrimental for organ restitution and recovery. In the context of inflammation, hypoxia is particularly important and can significantly influence the course of inflammatory diseases. Macrophages and neutrophils, the chief cellular components of innate immunity, display distinct properties when exposed to hypoxic conditions. Virtually every aspect of macrophage and neutrophil function is affected by hypoxia, amongst others, morphology, migration, chemotaxis, adherence to endothelial cells, bacterial killing, differentiation/polarization, and protumorigenic activity. Prominent arenas of macrophage and neutrophil function, for example, acute/chronic inflammation and the microenvironment of solid tumors, are characterized by low oxygen levels, demonstrating the paramount importance of the hypoxic response for proper function of these cells. Members of the hypoxia-inducible transcription factor (HIF family emerged as pivotal molecular regulators of macrophages and neutrophils. In this review, we will summarize the molecular responses of macrophages and neutrophils to hypoxia in the context of cancer and other chronic inflammatory diseases and discuss the potential avenues for therapeutic intervention that arise from this knowledge.

  11. Oxidative response of neutrophils to platelet-activating factor is altered during acute ruminal acidosis induced by oligofructose in heifers.

    Science.gov (United States)

    Concha, Claudia; Carretta, María Daniella; Alarcón, Pablo; Conejeros, Ivan; Gallardo, Diego; Hidalgo, Alejandra Isabel; Tadich, Nestor; Cáceres, Dante Daniel; Hidalgo, María Angélica; Burgos, Rafael Agustín

    2014-01-01

    Reactive oxygen species (ROS) production is one of the main mechanisms used to kill microbes during innate immune response. D-lactic acid, which is augmented during acute ruminal acidosis, reduces platelet activating factor (PAF)-induced ROS production and L-selectin shedding in bovine neutrophils in vitro. This study was conducted to investigate whether acute ruminal acidosis induced by acute oligofructose overload in heifers interferes with ROS production and L-selectin shedding in blood neutrophils. Blood neutrophils and plasma were obtained by jugular venipuncture, while ruminal samples were collected using rumenocentesis. Lactic acid from plasma and ruminal samples was measured by HPLC. PAF-induced ROS production and L-selectin shedding were measured in vitro in bovine neutrophils by a luminol chemiluminescence assay and flow cytometry, respectively. A significant increase in ruminal and plasma lactic acid was recorded in these animals. Specifically, a decrease in PAF-induced ROS production was observed 8 h after oligofructose overload, and this was sustained until 48 h post oligofructose overload. A reduction in PAF-induced L-selectin shedding was observed at 16 h and 32 h post oligofructose overload. Overall, the results indicated that neutrophil PAF responses were altered in heifers with ruminal acidosis, suggesting a potential dysfunction of the innate immune response.

  12. Frontline Science: HMGB1 induces neutrophil dysfunction in experimental sepsis and in patients who survive septic shock.

    Science.gov (United States)

    Grégoire, Murielle; Tadié, Jean-Marc; Uhel, Fabrice; Gacouin, Arnaud; Piau, Caroline; Bone, Nathaniel; Le Tulzo, Yves; Abraham, Edward; Tarte, Karin; Zmijewski, Jaroslaw W

    2017-06-01

    Sepsis is accompanied by the initial activation of proinflammatory pathways and long-lasting immunosuppression that appears to contribute to late-occurring mortality. Although high-mobility group box 1 (HMGB1) is involved in many aspects of inflammation, its role in sepsis-induced immune suppression remains unclear. In this study, we examined HMGB1's contribution to neutrophil NADPH oxidase activity dysfunction and associated neutrophil-dependent bacterial clearance in mice subjected to sepsis and in patients who survive septic shock. Using a murine model of polymicrobial septic peritonitis, we demonstrated that treatment with anti-HMGB1 Ab significantly diminished sepsis-induced dysfunction of neutrophil NADPH oxidase activity. In a subsequent set of experiments, we found that blocking HMGB1 preserved the ability of neutrophils from patients recovering from septic shock to activate NADPH oxidase. Taken together, our data suggest that HMGB1 accumulation in the late phase of sepsis plays a specific role in the development of postsepsis immunosuppression and specifically affects neutrophil-dependent antibacterial defense mechanisms. Thus, blocking HMGB1 may be a promising therapeutic intervention to diminish the adverse effects of sepsis-induced immunosuppression. © Society for Leukocyte Biology.

  13. Mean platelet volume, neutrophil to lyphocyte ratio and platelet to lymphocyte ratio in psoriasis

    Directory of Open Access Journals (Sweden)

    Mehmet Ünal

    2015-06-01

    Full Text Available Background and Design: It has been demonstrated that ratio of neutrophil and platelet count systemic inflammation and is associated with prognosis of many cardiovascular diseases, malignates and chronic inflammatory diseases.As far as it is known, there are no studies investigating neutrophil/lymphocyeratio(NLR, platelet/lymphocyte ratio(PLR and mean platelet volume(MPV values together within the context of psoriasis, a chronic and systemic inflammatory disease. Materials and Methods: 320 patients followed up in our polyclinic with psoriasis vulgaris and 200 healthy persons were evaluated in the study. Results: Leukocyte, neutrophil, platelet, MPV, NLR and PLR values in patients with psoriasis were significantly higher, and lymphocyte count, on the other hand, was significantly lower than those of the control group. No significant difference was found between MPV, NLR and PLR values of patients with or without a family history, nail and joint involvement. Conclusions: These parameters may be made use of as cheap and easily applicable methods in predicting which psoriasis patients are under the risk of cardiovascular disease. PLR is a better inflammation marker than MPV and NLR in patients with psoriasis. We did not observe a significant relationship between MPV, NLR and PLR values and such disease characteristics as severity of disease, joint involvement, nail involvement and duration of disease in patients with psoriasis. So, we believe that there is little information on the extent to which MPV,NLR and PLR might be useful regarding these characteristics.

  14. Characterization of neutrophil adhesion to different titanium surfaces

    Indian Academy of Sciences (India)

    Scanning electron microscopy and flow cytometry were used to measure .... discs containing neutrophils were inverted and placed in a tube assembly that ... Representative AFM images of smooth (a) and rough (b) Ti surfaces. The arithmetic ...

  15. Systems biology of neutrophil differentiation and immune response

    DEFF Research Database (Denmark)

    Theilgaard-Mönch, Kim; Porse, Bo T; Borregaard, Niels

    2005-01-01

    Systems biology has emerged as a new scientific field, which aims at investigating biological processes at the genomic and proteomic levels. Recent studies have unravelled aspects of neutrophil differentiation and immune responses at the systems level using high-throughput technologies. These stu......Systems biology has emerged as a new scientific field, which aims at investigating biological processes at the genomic and proteomic levels. Recent studies have unravelled aspects of neutrophil differentiation and immune responses at the systems level using high-throughput technologies....... These studies have identified a plethora of novel effector proteins stored in the granules of neutrophils. In addition, these studies provide evidence that neutrophil differentiation and immune response are governed by a highly coordinated transcriptional programme that regulates cellular fate and function...

  16. Cryptococcal capsular glucuronoxylomannan reduces ischaemia-related neutrophil influx

    NARCIS (Netherlands)

    Ellerbroek, PM; Schoemaker, RG; van Veghel, R; Hoepelman, AIM; Coenjaerts, FEJ

    Background The capsular polysaccharide glucuronoxylomannan (GXM) of Cryptococcus neoformans interferes with the chemotaxis and transendothelial migration of neutrophils. Intravenous administration of purified GXM has been shown to reduce the influx of inflammatory cells in an animal model of

  17. Human neutrophil peptide-1 promotes alcohol-induced hepatic fibrosis and hepatocyte apoptosis.

    Directory of Open Access Journals (Sweden)

    Rie Ibusuki

    Full Text Available Neutrophil infiltration of the liver is a typical feature of alcoholic liver injury. Human neutrophil peptide (HNP-1 is an antimicrobial peptide secreted by neutrophils. The aim of this study was to determine if HNP-1 affects ethanol-induced liver injury and to examine the mechanism of liver injury induced by HNP-1.Transgenic (TG mice expressing HNP-1 under the control of a β-actin-based promoter were established. Ethanol was orally administered to HNP-1 TG or wild-type C57BL/6N (WT mice. SK-Hep1 hepatocellular carcinoma cells were used to investigate the effect of HNP-1 on hepatocytes in vitro.After 24 weeks of ethanol intake, hepatic fibrosis and hepatocyte apoptosis were significantly more severe in TG mice than in WT mice. Levels of CD14, TLR4, and IL-6 in liver tissues were higher in TG mice than in WT mice. Apoptosis was accompanied by higher protein levels of caspase-3, caspase-8, and cleaved PARP in liver tissue. In addition, phosphorylated ASK1, ASK1, phosphorylated JNK, JNK1, JNK2, Bax, Bak and Bim were all more abundant in TG mice than in WT mice. In contrast, the level of anti-apoptotic Bcl2 in the liver was significantly lower in TG mice than in WT mice. Analysis of microRNAs in liver tissue showed that miR-34a-5p expression was significantly higher in TG mice than in WT mice. Furthermore, in the presence of ethanol, HNP-1 increased the apoptosis with the decreased level of Bcl2 in a concentration-dependent manner in vitro.HNP-1 secreted by neutrophils may exacerbate alcohol-induced hepatic fibrosis and hepatocyte apoptosis with a decrease in Bcl2 expression and an increase in miR-34a-5p expression.

  18. Intramuscular administration of a synthetic CpG-oligodeoxynucleotide modulates functional responses of neutrophils of neonatal foals.

    Directory of Open Access Journals (Sweden)

    Noah D Cohen

    Full Text Available Neutrophils play an important role in protecting against infection. Foals have age-dependent deficiencies in neutrophil function that may contribute to their predisposition to infection. Thus, we investigated the ability of a CpG-ODN formulated with Emulsigen to modulate functional responses of neutrophils in neonatal foals. Eighteen foals were randomly assigned to receive either a CpG-ODN with Emulsigen (N = 9 or saline intramuscularly at ages 1 and 7 days. At ages 1, 3, 9, 14, and 28, blood was collected and neutrophils were isolated from each foal. Neutrophils were assessed for basal and Rhodococcus equi-stimulated mRNA expression of the cytokines interferon-γ (IFN-γ, interleukin (IL-4, IL-6, and IL-8 using real-time PCR, degranulation by quantifying the amount of β-D glucuronidase activity, and reactive oxygen species (ROS generation using flow cytometry. In vivo administration of the CpG-ODN formulation on days 1 and 7 resulted in significantly (P<0.05 increased IFN-γ mRNA expression by foal neutrophils on days 3, 9, and 14. Degranulation was significantly (P<0.05 lower for foals in the CpG-ODN-treated group than the control group at days 3 and 14, but not at other days. No effect of treatment on ROS generation was detected. These results indicate that CpG-ODN administration to foals might improve innate and adaptive immune responses that could protect foals against infectious diseases and possibly improve responses to vaccination.

  19. Osthol attenuates neutrophilic oxidative stress and hemorrhagic shock-induced lung injury via inhibition of phosphodiesterase 4.

    Science.gov (United States)

    Tsai, Yung-Fong; Yu, Huang-Ping; Chung, Pei-Jen; Leu, Yann-Lii; Kuo, Liang-Mou; Chen, Chun-Yu; Hwang, Tsong-Long

    2015-12-01

    Oxidative stress caused by neutrophils is an important pathogenic factor in trauma/hemorrhagic (T/H)-induced acute lung injury (ALI). Osthol, a natural coumarin found in traditional medicinal plants, has therapeutic potential in various diseases. However, the pharmacological effects of osthol in human neutrophils and its molecular mechanism of action remain elusive. In this study, our data showed that osthol potently inhibited the production of superoxide anion (O2(•-)) and reactive oxidants derived therefrom as well as expression of CD11b in N-formylmethionylleucylphenylalanine (FMLP)-activated human neutrophils. However, osthol inhibited neutrophil degranulation only slightly and it failed to inhibit the activity of subcellular NADPH oxidase. FMLP-induced phosphorylation of extracellular signal-regulated kinase (ERK) and protein kinase B (Akt) was inhibited by osthol. Notably, osthol increased the cAMP concentration and protein kinase A (PKA) activity in activated neutrophils. PKA inhibitors reversed the inhibitory effects of osthol, suggesting that these are mediated through cAMP/PKA-dependent inhibition of ERK and Akt activation. Furthermore, the activity of cAMP-specific phosphodiesterase (PDE) 4, but not PDE3 or PDE7, was significantly reduced by osthol. In addition, osthol reduced myeloperoxidase activity and pulmonary edema in rats subjected to T/H shock. In conclusion, our data suggest that osthol has effective anti-inflammatory activity in human neutrophils through the suppression of PDE4 and protects significantly against T/H shock-induced ALI in rats. Osthol may have potential for future clinical application as a novel adjunct therapy to treat lung inflammation caused by adverse circulatory conditions. Copyright © 2015 Elsevier Inc. All rights reserved.

  20. Effects of Acrolein on Leukotriene Biosynthesis in Human Neutrophils

    OpenAIRE

    Zemski Berry, Karin A.; Henson, Peter M.; Murphy, Robert C.

    2008-01-01

    Acrolein is a toxic, highly reactive α,β-unsaturated aldehyde that is present in high concentrations in cigarette smoke. In the current study, the effect of acrolein on eicosanoid synthesis in stimulated human neutrophils was examined. Eicosanoid synthesis in neutrophils was initiated by priming with granulocyte-macrophage colony-stimulating factor (GM-CSF) and subsequent stimulation with formyl-methionyl-leucyl-phenylalanine (fMLP) and 5-LO products in addition to small amounts of COX produc...

  1. Tumor-Associated Macrophages and Neutrophils in Tumor Microenvironment

    Directory of Open Access Journals (Sweden)

    Jaehong Kim

    2016-01-01

    Full Text Available Distinct tumor microenvironment forms in each progression step of cancer and has diverse capacities to induce both adverse and beneficial consequences for tumorigenesis. It is now known that immune cells can be activated to favor tumor growth and progression, most probably influenced by the tumor microenvironment. Tumor-associated macrophages and tumor-associated neutrophils can exert protumoral functions, enhancing tumor cell invasion and metastasis, angiogenesis, and extracellular matrix remodeling, while inhibiting the antitumoral immune surveillance. Considering that neutrophils in inflammatory environments recruit macrophages and that recruited macrophages affect neutrophil functions, there may be various degrees of interaction between tumor-associated macrophages and tumor-associated neutrophils. Platelets also play an important role in the recruitment and regulation of monocytic and granulocytic cells in the tumor tissues, suggesting that platelet function may be essential for generation of tumor-associated macrophages and tumor-associated neutrophils. In this review, we will explore the biology of tumor-associated macrophages and tumor-associated neutrophils and their possible interactions in the tumor microenvironment. Special attention will be given to the recruitment and activation of these tumor-associated cells and to the roles they play in maintenance of the tumor microenvironment and progression of tumors.

  2. Pathophysiology of neutrophil-mediated extracellular redox reactions.

    Science.gov (United States)

    Jaganjac, Morana; Cipak, Ana; Schaur, Rudolf Joerg; Zarkovic, Neven

    2016-01-01

    Neutrophil granulocyte leukocytes (neutrophils) play fundamental role in the innate immune response. In the presence of adequate stimuli, neutrophils release excessive amount of reactive oxygen species (ROS) that may induce cell and tissue injury. Oxidative burst of neutrophils acts as a double-edged sword. It may contribute to the pathology of atherosclerosis and brain injury but is also necessary in resolving infections. Moreover, neutrophil-derived ROS may also have both a tumor promoting and tumor suppressing role. ROS have a specific activities and diffusion distance, which is related to their short lifetime. Therefore, the manner in which ROS will act depends on the cells targeted and the intra- and extracellular levels of individual ROS, which can further cause production of reactive aldehydes like 4-hydroxynonenal (HNE) that act as a second messengers of ROS. In this review we discuss the influence of neutrophil mediated extracellular redox reactions in ischemia reperfusion injury, transplant rejection and chronic diseases (atherosclerosis, inflammatory bowel diseases and cancer). At the end a brief overview of cellular mechanisms to maintain ROS homeostasis is given.

  3. Granule protein processing and regulated secretion in neutrophils

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    Avinash eSheshechalam

    2014-09-01

    Full Text Available Neutrophils are part of a family of granulocytes that, together with eosinophils and basophils, play an essential role in innate immunity. Neutrophils are the most abundant circulating leukocytes and are vital for rapid immune responses, being recruited to sites of injury or infection within minutes, where they can act as specialized phagocytic cells. However, another prominent function of neutrophils is the release of pro-inflammatory compounds, including cytokines, chemokines and digestive enzymes, which are stored in intracellular compartments and released through regulated exocytosis. Hence, an important feature that contributes to rapid immune responses is capacity of neutrophils to synthesize and store pre-formed pro-inflammatory mediators in specialized intracellular vesicles and thus no new synthesis is required. This review will focus on advancement in three topics relevant to neutrophil secretion. First we will examine what is known about basal level pro-inflammatory mediator synthesis, trafficking and storage in secretory compartments. Second, we will review recent advancements in the mechanisms that control vesicle mobilization and the release of pre-formed mediators. Third, we will examine the upregulation and de novo synthesis of pro-inflammatory mediators by neutrophils engaged at sites of infection.

  4. The role of neutrophilic mediators in acute inflammation of the gut

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    Ritter, C. von.

    1988-01-01

    Activation of granulocytes within the lamina propria by luminally derived bacterial products may represent an important mechanism in the pathogenesis of inflammatory bowel disease. One objective of this thesis was to determine the effects of luminal perfusion with N-formyl-methionyl-leucyl-phenylalanine (FMLP), a bacterial product that attracts and activates granulocytes, on mucosal permeability in different regions of the rat small intestine and colon. Mucosal permeability was measured using the blood-to-lumen clearance of 51 Cr-EDTA during luminal perfusion with FMLP dissolved in Tyrode's solution. Of the bowel segments studied, mucosal permeability was significantly increased only in the distal 10 cm of the ileum. In order to define the role of neutrophilic oxidants in FMLP-induced ileitis, we evaluated the protective effect of several free radical scavengers and antioxidant enzymes. Pretreatment with the either superoxide dismutase or catalase had no effect on the FMLP-induced increase in mucosal permeability. However, treatment with either Mn-desferrioxamine, PZ51, desferrioxamine, or dimethylsulfoxide significantly attenuated FMLP-induced mucosal damage. Non-oxidative toxins released from activated neutrophil may be another mechanism by which FMLP increases mucosal permeability. In order to investigate the role of neutrophilic proteases in FMLP-induced ileitis, the effects of the nonspecific protease inhibitor soybean trypsin inhibitor, and the elastase inhibitors MeOSuc-Ala-Ala-Pro-Val-CH 2 Cl(MAAPV) and Eglin C on the FMLP-induced increases in 51 Cr-EDTA clearance were determined

  5. [Lymphocyte transformation test following stimulation with a protein factor from neutrophilic granulocytes (PMNL) in psoriasis patients].

    Science.gov (United States)

    Ruszczak, Z; Ciborska, L; Kaszuba, A

    1988-12-01

    The lymphocyte transformation test (LTT) was given to 20 healthy subjects and 43 patients with generalized psoriasis vulgaris: it was given right after stimulation with PHA (spontaneous) and after stimulation with allogenic and autogenic protein factor (NPF). NPF was isolated from secondary lysosome granules of peripheral blood neutrophils. The results were analyzed using computer statistic tests. No distinct differences were noticed between the spontaneous transformation test in psoriatic patients compared to the controls. After stimulation with PHA, the percentage of blast cells was significantly lower in patients with psoriasis. When allogenic and autogenic NPF was used for stimulation, the LTT values were significantly higher in the psoriasis group than in the control subjects. This fact points out the increase in sensitivity of lymphocytes to NPF in active psoriasis and the possibility of abnormal neutrophil-lymphocyte interactions in vivo. This phenomenon may be intensified when under the influence of bacterial or viral agents, or medicaments; the degranulation of secondary lysosome granules of neutrophils occurs, causing the release of NPF. These investigations support our opinion that psoriasis is a systemic disease and that NPF plays a considerable role in the psoriatic reaction.

  6. Chlamydia pneumoniae hides inside apoptotic neutrophils to silently infect and propagate in macrophages.

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    Jan Rupp

    Full Text Available BACKGROUND: Intracellular pathogens have developed elaborate strategies for silent infection of preferred host cells. Chlamydia pneumoniae is a common pathogen in acute infections of the respiratory tract (e.g. pneumonia and associated with chronic lung sequelae in adults and children. Within the lung, alveolar macrophages and polymorph nuclear neutrophils (PMN are the first line of defense against bacteria, but also preferred host phagocytes of chlamydiae. METHODOLOGY/PRINCIPAL FINDINGS: We could show that C. pneumoniae easily infect and hide inside neutrophil granulocytes until these cells become apoptotic and are subsequently taken up by macrophages. C. pneumoniae infection of macrophages via apoptotic PMN results in enhanced replicative activity of chlamydiae when compared to direct infection of macrophages, which results in persistence of the pathogen. Inhibition of the apoptotic recognition of C. pneumoniae infected PMN using PS- masking Annexin A5 significantly lowered the transmission of chlamydial infection to macrophages. Transfer of apoptotic C. pneumoniae infected PMN to macrophages resulted in an increased TGF-ss production, whereas direct infection of macrophages with chlamydiae was characterized by an enhanced TNF-alpha response. CONCLUSIONS/SIGNIFICANCE: Taken together, our data suggest that C. pneumoniae uses neutrophil granulocytes to be silently taken up by long-lived macrophages, which allows for efficient propagation and immune protection within the human host.

  7. Short communication: Relationship between urinary neutrophil gelatinase-associated lipocalin and noninfectious pyuria in dogs.

    Science.gov (United States)

    Proverbio, D; Spada, E; Baggiani, L; Bagnagatti De Giorgi, G; Ferro, E; Martino, P A; Perego, R

    2015-01-01

    Neutrophil gelatinase-associated lipocalin (NGAL) is a neutrophil-derived protein whose concentration increases in plasma and urine with ongoing renal damage. Urinary leucocytes can be a potential source of urinary NGAL. The aim of this study is to investigate the effects of urinary neutrophil count and other urinary parameters on urinary NGAL values in urine with negative culture. Urinalysis, urine culture, and determination of urinary NGAL were performed on 33 clinically healthy nonproteinuric dogs with negative urinoculture. The median uNGAL concentration in dogs in this study population was 9.74 ng/mL (IQR 1.93-25.43 ng/mL). In samples with WBCs > 5 hpf (mean 15.9, 6-50 leucocytes/hpf), median uNGAL value was significantly higher than that in samples with WBCs dogs with negative urinoculture. The present study suggests that noninfectious pyuria is significantly correlated with urinary NGAL values and might influence uNGAL values.

  8. In Vitro Evaluation of the Link Between Cell Activation State and Its Rheological Impact on the Microscale Flow of Neutrophil Suspensions

    Science.gov (United States)

    Akenhead, Michael L.; Horrall, Nolan M.; Rowe, Dylan; Sethu, Palaniappan; Shin, Hainsworth Y.

    2015-01-01

    Activated neutrophils have been reported to affect peripheral resistance, for example, by plugging capillaries or adhering to the microvasculature. In vivo and ex vivo data indicate that activated neutrophils circulating in the blood also influence peripheral resistance. We used viscometry and microvascular mimics for in vitro corroboration. The rheological impact of differentiated neutrophil-like HL-60 promyelocytes (dHL60s) or human neutrophil suspensions stimulated with 10 nM fMet-Leu-Phe (fMLP) was quantified using a cone-plate rheometer (450 s−1 shear rate). To evaluate their impact on microscale flow resistance, we used 10-μm Isopore® membranes to model capillaries as well as single 200 × 50 μm microchannels and networks of twenty 20 × 50 μm microfluidic channels to mimic noncapillary microvasculature. Stimulation of dHL60 and neutrophil populations significantly altered their flow behavior as evidenced by their impact on suspension viscosity. Notably, hematocrit abrogated the impact of leukocyte activation on blood cell suspension viscosity. In micropore filters, activated cell suspensions enhanced flow resistance. This effect was further enhanced by the presence of erythrocytes. The resistance of our noncapillary microvascular mimics to flow of activated neutrophil suspensions was significantly increased only with hematocrit. Notably, it was elevated to a higher extent within the micronetwork chambers compared to the single-channel chambers. Collectively, our findings provide supportive evidence that activated neutrophils passing through the microcirculation may alter hemodynamic resistance due to their altered rheology in the noncapillary microvasculature. This effect is another way neutrophil activation due to chronic inflammation may, at least in part, contribute to the elevated hemodynamic resistance associated with cardiovascular diseases (e.g., hypertension and hypercholesterolemia). PMID:26065495

  9. In Vitro Evaluation of the Link Between Cell Activation State and Its Rheological Impact on the Microscale Flow of Neutrophil Suspensions.

    Science.gov (United States)

    Akenhead, Michael L; Horrall, Nolan M; Rowe, Dylan; Sethu, Palaniappan; Shin, Hainsworth Y

    2015-09-01

    Activated neutrophils have been reported to affect peripheral resistance, for example, by plugging capillaries or adhering to the microvasculature. In vivo and ex vivo data indicate that activated neutrophils circulating in the blood also influence peripheral resistance. We used viscometry and microvascular mimics for in vitro corroboration. The rheological impact of differentiated neutrophil-like HL-60 promyelocytes (dHL60s) or human neutrophil suspensions stimulated with 10 nM fMet-Leu-Phe (fMLP) was quantified using a cone-plate rheometer (450 s(-1) shear rate). To evaluate their impact on microscale flow resistance, we used 10-μm Isopore® membranes to model capillaries as well as single 200 × 50 μm microchannels and networks of twenty 20 × 50 μm microfluidic channels to mimic noncapillary microvasculature. Stimulation of dHL60 and neutrophil populations significantly altered their flow behavior as evidenced by their impact on suspension viscosity. Notably, hematocrit abrogated the impact of leukocyte activation on blood cell suspension viscosity. In micropore filters, activated cell suspensions enhanced flow resistance. This effect was further enhanced by the presence of erythrocytes. The resistance of our noncapillary microvascular mimics to flow of activated neutrophil suspensions was significantly increased only with hematocrit. Notably, it was elevated to a higher extent within the micronetwork chambers compared to the single-channel chambers. Collectively, our findings provide supportive evidence that activated neutrophils passing through the microcirculation may alter hemodynamic resistance due to their altered rheology in the noncapillary microvasculature. This effect is another way neutrophil activation due to chronic inflammation may, at least in part, contribute to the elevated hemodynamic resistance associated with cardiovascular diseases (e.g., hypertension and hypercholesterolemia).

  10. Sphingosine 1-phosphate induces neutrophil chemoattractant IL-8: repression by steroids.

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    Md Mostafizur Rahman

    Full Text Available The bioactive sphingolipid sphingosine 1-phosphate (S1P is found in increased amounts in the airways of asthmatics. S1P can regulate airway smooth muscle functions associated with asthmatic inflammation and remodeling, including cytokine secretion. To date however, whether S1P induces secretion of an important chemokine responsible for neutrophilia in airway inflammation--IL-8--was unexplored. The aim of this study was to investigate whether S1P induces IL-8 gene expression and secretion to enhance neutrophil chemotaxis in vitro, as well as examine the molecular mechanisms responsible for repression by the corticosteroid dexamethasone. We show that S1P upregulates IL-8 secretion from ASM cells and enhance neutrophil chemotaxis in vitro. The corticosteroid dexamethasone significantly represses IL-8 mRNA expression and protein secretion in a concentration- and time-dependent manner. Additionally, we reveal that S1P-induced IL-8 secretion is p38 MAPK and ERK-dependent and that these key phosphoproteins act on the downstream effector mitogen- and stress-activated kinase 1 (MSK1 to control secretion of the neutrophil chemoattractant cytokine IL-8. The functional relevance of this in vitro data was demonstrated by neutrophil chemotaxis assays where S1P-induced effects can be significantly attenuated by pretreatment with dexamethasone, pharmacological inhibition of p38 MAPK- or ERK-mediated pathways, or by knocking down MSK-1 with siRNA. Taken together, our study reveals the molecular pathways responsible for IL-8 secretion from ASM cells in response to S1P and indicates ways in which the impact on IL-8-driven neutrophilia may be lessened.

  11. Oxidative stress augments toll-like receptor 8 mediated neutrophilic responses in healthy subjects

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    Matsunaga Kazuto

    2009-06-01

    Full Text Available Abstract Background Excessive oxidative stress has been reported to be generated in inflamed tissues and contribute to the pathogenesis of inflammatory lung diseases, exacerbations of which induced by viral infections are associated with toll-like receptor (TLR activation. Among these receptors, TLR8 has been reported as a key receptor that recognizes single-strand RNA virus. However, it remains unknown whether TLR8 signaling is potentiated by oxidative stress. The aim of this study is to examine whether oxidative stress modulates TLR8 signaling in vitro. Methods Human peripheral blood neutrophils were obtained from healthy non-smokers and stimulated with TLR 7/8 agonist imidazoquinoline resiquimod (R848 in the presence or absence of hydrogen peroxide (H2O2. Neutrophilic responses including cytokine release, superoxide production and chemotaxis were examined, and the signal transduction was also analyzed. Results Activation of TLR8, but not TLR7, augmented IL-8 release. The R848-augmented IL-8 release was significantly potentiated by pretreatment with H2O2 (p L-cysteine reversed this potentiation. The combination of H2O2 and R848 significantly potentiated NF-kB phosphorylation and IkBα degradation. The H2O2-potentiated IL-8 release was suppressed by MG-132, a proteosome inhibitor, and by dexamethasone. The expressions of TLR8, myeloid differentiation primary response gene 88 (MyD88, and tumor necrosis factor receptor-associated factor 6 (TRAF6 were not affected by H2O2. Conclusion TLR8-mediated neutrophilic responses were markedly potentiated by oxidative stress, and the potentiation was mediated by enhanced NF-kB activation. These results suggest that oxidative stress might potentiate the neutrophilic inflammation during viral infection.

  12. Kidney injury molecule-1 and neutrophil gelatinase-associated lipocalin as prognostic markers in idiopathic membranous nephropathy.

    Science.gov (United States)

    Maas, Rutger Jh; van den Brand, Jan Ajg; Waanders, Femke; Meijer, Esther; Goor van, Harry; Peters, Hilde P; Hofstra, Julia M; Wetzels, Jack Fm

    2016-01-01

    Urinary excretion of alpha-1-microglobulin and beta-2-microglobulin reflects tubular damage and predicts outcome in patients with idiopathic membranous nephropathy with reasonable accuracy. Urinary kidney injury molecule-1 and neutrophil gelatinase-associated lipocalin are novel biomarkers of tubular damage. We investigated if these markers could improve prediction of outcome in idiopathic membranous nephropathy. We measured kidney injury molecule-1 and neutrophil gelatinase-associated lipocalin in urine samples from patients with idiopathic membranous nephropathy, who had nephrotic proteinuria and normal renal function. Excretion of alpha-1-microglobulin and beta-2-microglobulin had been measured previously. Progression was defined as a serum creatinine rise >30%, a rise in serum creatinine to an absolute value of ≥135 µmol/L, or a clinical decision to start immunosuppressive therapy. Remission was defined as proteinuria 50% reduction from baseline. Sixty-nine patients were included. Median follow-up was 35 months (interquartile range 18-63 months). Progression occurred in 30 patients (44%), and spontaneous remission in 36 (52%). Kidney injury molecule-1 and neutrophil gelatinase-associated lipocalin excretion rates were significantly correlated with each other, and with alpha-1-microglobulin and beta-2-microglobulin. The areas under the receiver operating characteristic curves for progression were 0.75 (0.62-0.87) for kidney injury molecule-1 and 0.74 (0.62-0.87) for neutrophil gelatinase-associated lipocalin. In multivariate analysis with either alpha-1-microglobulin and beta-2-microglobulin, kidney injury molecule-1 and neutrophil gelatinase-associated lipocalin did not independently predict outcome. Kidney injury molecule-1 and neutrophil gelatinase-associated lipocalin excretion rates correlated with excretion rates of other tubular damage markers and predicted outcome in patients with idiopathic membranous nephropathy. They did not add prognostic value

  13. Staphylococcus aureus penetrate the interkeratinocyte spaces created by skin-infiltrating neutrophils in a mouse model of impetigo.

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    Imanishi, Ichiro; Hattori, Shinpei; Hisatsune, Junzo; Ide, Kaori; Sugai, Motoyuki; Nishifuji, Koji

    2017-02-01

    Impetigo is a bacterial skin disease characterized by intraepidermal neutrophilic pustules. Previous studies have demonstrated that exfoliative toxin producing staphylococci are isolated in the cutaneous lesions of human and canine impetigo. However, the mechanisms of intraepidermal splitting in impetigo remain poorly understood. To determine how staphylococci penetrate the living epidermis and create intraepidermal pustules in vivo using a mouse model of impetigo. Three Staphylococcus aureus strains harbouring the etb gene and three et gene negative strains were epicutaneously inoculated onto tape-stripped mouse skin. The skin samples were subjected to time course histopathological and immunofluorescence analyses to detect intraepidermal neutrophils and infiltrating staphylococci. To determine the role of neutrophils on intraepidermal bacterial invasion, cyclophosphamide (CPA) was injected intraperitoneally into the mice to cause leucopenia before the inoculation of etb gene positive strains. In mice inoculated with etb gene positive S. aureus, intraepidermal pustules resembling impetigo were detected as early as 4 h post-inoculation (hpi). Neutrophils in the epidermis were detected from 4 hpi, whereas intraepidermal staphylococci was detected from 6 hpi. The dimensions of the intraepidermal clefts created in mice inoculated with etb gene positive strains at 6 hpi were significantly larger than those in mice inoculated with et gene negative strains. In CPA treated mice, staphylococci or neutrophils were not detected in the deep epidermis until 6 hpi. Our findings indicate that intraepidermal neutrophils play an important role in S. aureus invasion into the living epidermis in a mouse model of impetigo. © 2016 ESVD and ACVD.

  14. The LPS-induced neutrophil recruitment into rat air pouches is mediated by TNFα: likely macrophage origin

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    C-D. Arreto

    1997-01-01

    Full Text Available The role of resident cells during the lipopolysaccharide (LPS-induced neutrophil recruitment into rat air pouches was investigated. In this model, LPS (Escherichia coli, O55: B5 strain; 2–2000 ng induced a dose– and time-dependent neutrophil recruitment accompanied by the generation of a tumour necrosis factor-α (TNFα-like activity. Dexamethasone (0.05–5 mug and cycloheximide (6 ng, injected 2 h before LPS into the pouches, inhibited the neutrophil recruitment and the generation of the TNFα-like activity, while the H1-receptor antagonist mepyramine (1 and 4 mg/kg, i.p., 0.5 h before LPS and the PAF-receptor antagonist WEB 2170 (0.05 and 1 mg/kg, i.p., 0.5 h before LPS had no effect. Purified alveolar macrophages (AM were used to replenish the pouches of cycloheximide-treated recipient rats. AM provided by PBS-treated animals led to the recovery of the LPS-induced neutrophil recruitment and of the TNFα-like formation contrasting with those from cycloheximide-treated animals (1 mg/kg, i.p.. When delivered in situ, liposome-encapsulated clodronate, a macrophage depletor, significantly impaired both the LPSinduced neutrophil recruitment and the TNFα-like activity. An anti-murine TNFα polyclonal antibody (0.5 h before LPS was also effective. These results emphasize the pivotal role of macrophages for LPS-induced neutrophil recruitment via the formation of TNFα.

  15. Platelet modulation of human neutrophil functions

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    McGarrity, S.T.; Hyers, T.M.; Webster, R.O.

    1986-03-01

    The combined presence of platelets (PLTS) and neutrophils (PMN) at inflammatory sites has led to examination of the hypothesis that interaction of these cells modulates their responses to stimuli. Gel-filtered human PLTS (GFP) were found to inhibit N-formyl-met-leu-phe (FMLP) and phorbol myristate acetate (PMA) stimulated PMN O/sub 2//sup -/ generation in a concentration-dependent fashion. The heat-stable inhibitory activity was present in the supernatants of GFP after incubation with FMLP (10/sup -7/M), thrombin (0.5 U/ml) or ADP (20 ..mu..M), suggesting a role for PLT release products. PLT lysates added to PMN produced up to 80% inhibition of O/sub 2//sup -/ generation for PMA and 40% for FMLP. Like GFP, lysates failed to scavenge O/sub 2/..pi.. produced by xanthine oxidase-hypoxanthine. The inhibitory activity could not be ascribed to serotonin or adenosine. PLT lysates failed to compete with /sup 3/H-FMLP for binding to PMN. Sephadex G-200 fractionation of PLT lysates releaved two peaks of inhibitory activity with apparent Mr > 200,000 and < 14,000 Daltons. Pretreatment of PMN with PLT lysates also results in a concentration-dependent inhibition of degranulation provoked by FMLP (2 x 10/sup -7/M) or PMA (2 ng/ml) and PMN chemotaxis to FMLP (10/sup -8/M). These studies indicate that preformed PLT mediator(s) released in response to physiological stimuli may limit tissue damage by PMN at sites of inflammation.

  16. Potentially pathogenic airway bacteria and neutrophilic inflammation in treatment resistant severe asthma.

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    Green, Benjamin J; Wiriyachaiporn, Surasa; Grainge, Christopher; Rogers, Geraint B; Kehagia, Valia; Lau, Laurie; Carroll, Mary P; Bruce, Kenneth D; Howarth, Peter H

    2014-01-01

    Molecular microbiological analysis of airway samples in asthma has demonstrated an altered microbiome in comparison to healthy controls. Such changes may have relevance to treatment-resistant severe asthma, particularly those with neutrophilic airway inflammation, as bacteria might be anticipated to activate the innate immune response, a process that is poorly steroid responsive. An understanding of the relationship between airway bacterial presence and dominance in severe asthma may help direct alternative treatment approaches. We aimed to use a culture independent analysis strategy to describe the presence, dominance and abundance of bacterial taxa in induced sputum from treatment resistant severe asthmatics and correlate findings with clinical characteristics and airway inflammatory markers. Induced sputum was obtained from 28 stable treatment-resistant severe asthmatics. The samples were divided for supernatant IL-8 measurement, cytospin preparation for differential cell count and Terminal Restriction Fragment Length Polymorphism (T-RFLP) profiling for bacterial community analysis. In 17/28 patients, the dominant species within the airway bacterial community was Moraxella catarrhalis or a member of the Haemophilus or Streptococcus genera. Colonisation with these species was associated with longer asthma disease duration (mean (SD) 31.8 years (16.7) vs 15.6 years (8.0), p = 0.008), worse post-bronchodilator percent predicted FEV1 (68.0% (24.0) vs 85.5% (19.7), p = 0.025) and higher sputum neutrophil differential cell counts (median (IQR) 80% (67-83) vs 43% (29-67), p = 0.001). Total abundance of these organisms significantly and positively correlated with sputum IL-8 concentration and neutrophil count. Airway colonisation with potentially pathogenic micro-organisms in asthma is associated with more severe airways obstruction and neutrophilic airway inflammation. This altered colonisation may have a role in the development of an asthma phenotype that

  17. Histopathological aspects of neutrophilic dermatoses: Investigation of 38 cases and review of the literature

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    Amir-Hoshang Ehsani

    2016-01-01

    Full Text Available Introduction: Neutrophilic dermatosis (ND is a heterogeneous group of diseases with various etiologies and clinical presentations. NDs may clinically present as papule, vesiculopustule, plaque, and nodule of the skin, but they all share the common feature of neutrophilic predominance in the skin. Histological examination of patients with suspected ND is a key step for making the proper diagnosis. Patients and Methods: The aim of this article was to investigate histopathological aspects of different NDs. We obtained our data from medical records of patients at Razi dermatology hospital, between 2012 and 2014. Thirty-eight biopsy records coded under the term of any ND, including Sweet's syndrome (SS, pyoderma gangrenosum (PG, skin lesions of Behcet's disease, neutrophilic drug eruption, amicrobial pustulosis of the folds, pustular vasculitis of the hands, and undetermined ND were recruited in our study. The specimens were evaluated regarding inflammatory reaction pattern, epidermal/adnexal changes, and dermal changes. Results: Most common NDs in our study were PG (42.1% followed by SS (21.1%. The most common pattern of inflammatory reaction was superficial perivascular and interstitial dermal inflammation in 44.7% of the patients. Exocytosis of neutrophils into epidermis, hair follicle, and eccrine gland was seen in 71%, 18.5%, and 28.9% of the specimens, respectively. Ulceration was only seen in ten PG specimens. Dermal fibrosis and vascular proliferation were reported in all PG patients. Conclusion: The prevalence of some histopathological findings in different types of ND was significantly different. These features seem helpful in distinguishing between different NDs.

  18. Recruitment of neutrophils across the blood-brain barrier: the role of posttraumatic hepatic ischemia

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    Mantovani Mario

    2003-01-01

    Full Text Available PURPOSE: To study the effects of total hepatic ischemia, and reperfusion on the accumulation of neutrophils in the brain of rats submitted to normovolemic conditions as well as to controlled hemorrhagic shock state. METHODS: Thirty two adult male Wistar rats, were divided into four groups: the Control group, was submitted to the standard procedures for a period of 60 min of observation; Shock group, was submitted to controlled hemorrhagic shock (mean arterial blood pressure=40mmHg, 20min followed by volemic resuscitation (lactated Ringer's solution + blood, 3:1 and reperfusion for 60min; Pringle group, was submitted to total hepatic ischemia for 15min and reperfusion for 60min. The total group was submitted to controlled hemorrhagic shock for 20min followed by volemic resuscitation (lactated Ringer's solution + blood, 3:1, total hepatic ischemia for 15min and reperfusion for 60min. Measurements of serum lactate and base excess were used to characterize the hemorrhagic shock state with low tissue perfusion. The counting of neutrophils on the brain was performed after the euthanasia of animals. RESULTS: The values for the counting of neutrophils on the brain indicate that did not occur difference among studied groups (p=0.196 (Control 0.12± 0.11, Shock 0.12± 0.13, Pringle 0.02± 0.04, Total 0.14± 0.16. CONCLUSION: Hemorrhagic shock associated to total hepatic ischemia for 15 minutes, followed by 60 minutes of reperfusion, did not causes significant neutrophils accumulation in the brain of rats.

  19. An 'inflammatory' variant of solar purpura: a simulant of leukocytoclastic vasculitis and neutrophilic dermatoses.

    Science.gov (United States)

    Wood, Benjamin A; LeBoit, Philip E

    2013-08-01

    To study the clinical and pathological features of cases of apparent solar purpura, with attention to the recently described phenomenon of inflammatory changes within otherwise typical lesions. We studied 95 cases diagnosed as solar purpura and identified 10 cases (10.5%) in which significant neutrophilic inflammation was present, potentially simulating a leukocytoclastic vasculitis or neutrophilic dermatosis. An additional three cases were identified in subsequent routine practice. The clinical features, including follow-up for subsequent development of vasculitis and histological features were studied. In all cases the histological features were typical of solar purpura, with the exception of inflammatory changes, typically associated with clefting of elastotic stroma. Clinical follow-up information was available for all patients and none developed subsequent evidence of a cutaneous or systemic vasculitis or neutrophilic dermatosis. Inflammatory changes appear to be more frequent in solar purpura than is generally recognised. Awareness of this histological variation and correlation with the clinical findings and evolution is important in avoiding misdiagnosis.

  20. Human neutrophil peptides and complement factor Bb in pathogenesis of acquired thrombotic thrombocytopenic purpura.

    Science.gov (United States)

    Cao, Wenjing; Pham, Huy P; Williams, Lance A; McDaniel, Jenny; Siniard, Rance C; Lorenz, Robin G; Marques, Marisa B; Zheng, X Long

    2016-11-01

    Acquired thrombotic thrombocytopenic purpura is primarily caused by the deficiency of plasma ADAMTS13 activity resulting from autoantibodies against ADAMTS13. However, ADAMTS13 deficiency alone is often not sufficient to cause acute thrombotic thrombocytopenic purpura. Infections or systemic inflammation may precede acute bursts of the disease, but the underlying mechanisms are not fully understood. Herein, 52 patients with acquired autoimmune thrombotic thrombocytopenic purpura and 30 blood donor controls were recruited for the study. The plasma levels of human neutrophil peptides 1-3 and complement activation fragments (i.e. Bb, iC3b, C4d, and sC5b-9) were determined by enzyme-linked immunosorbent assays. Univariate analyses were performed to determine the correlation between each biomarker and clinical outcomes. We found that the plasma levels of human neutrophil peptides 1-3 and Bb in patients with acute thrombotic thrombocytopenic purpura were significantly higher than those in the control (Ppurpura patients and the control. We conclude that innate immunity, i.e. neutrophil and complement activation via the alternative pathway, may play a role in the pathogenesis of acute autoimmune thrombotic thrombocytopenic purpura, and a therapy targeted at these pathways may be considered in a subset of these patients. Copyright© Ferrata Storti Foundation.

  1. The effect of manual acupuncture on blood neutrophil counts in moderate intensity exercise

    Science.gov (United States)

    Ciang, C. Y.; Simadibrata, C.; Tobing, A.; Srilestari, A.

    2017-08-01

    Exercise, even though it has a beneficial effect, can cause muscle damage and trigger inflammatory responses, as evidenced by increased neutrophils in the blood. Acupuncture is a therapeutic modality that is expected to reduce acute inflammatory responses due to exercise. Thirty untrained men were divided randomly into two groups. The manual acupuncture group (n = 15) received stimulation at acupoints ST36 and SP6 bilateral by needle insertion, while the placebo group (n = 15) received insertion of needles on plaster without penetrating the skin. Therapy was done once for 30 minutes immediately after the subjects completed the exercise. Blood neutrophil counts were assessed before exercise and one hour after exercise ended. The results show there is a statistically significant difference in the number of neutrophils before and after exercise between the manual acupuncture group and the placebo group (0.08±0.91 and 0.97±0.70 p = 0.006). Acupuncture therapy effectively mitigates the acute inflammatory response triggered by exercise.

  2. Serum progranulin as an indicator of neutrophilic airway inflammation and asthma severity.

    Science.gov (United States)

    Park, So Young; Hong, Gyong Hwa; Park, Sunjoo; Shin, Bomi; Yoon, Sun-Young; Kwon, Hyouk-Soo; Kim, Tae-Bum; Moon, Hee-Bom; Cho, You Sook

    2016-12-01

    Progranulin, a protein secreted from the airway epithelium, is known to attenuate the downstream cascade of neutrophilic inflammation in particular. We hypothesized that progranulin may have a role in inflammatory regulation in asthma. To investigate the association between serum progranulin levels and various clinical features in patients with asthma. Serum samples and clinical data of 475 patients with asthma and 35 healthy controls at a tertiary referral hospital and its affiliated health promotion center were collected. Serum progranulin levels were compared between patients with asthma and healthy controls and then were compared within the patients with asthma in terms of pulmonary function and measures of inflammatory status. Univariate and multivariate analyses were performed to identify factors associated with severity of asthma. Serum progranulin levels were significantly lower in the asthma group than in healthy group and were positively correlated with prebronchodilator forced expiratory volume in 1 second predicted within patients with asthma. We found a negative correlation between serum progranulin levels and blood neutrophil counts. Multivariate analysis revealed that higher serum progranulin levels were associated with a lower risk of severe asthma (odds ratio, 0.888; 95% confidence interval, 0.846-0.932; P progranulin remains unknown, we suggest that serum progranulin may be an indicator of severe asthma with airflow limitation. Future studies with comprehensive airway sampling strategies are warranted to clarify its role, particularly in neutrophilic asthma. Copyright © 2016 American College of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.

  3. Protein cross-linking by chlorinated polyamines and transglutamylation stabilizes neutrophil extracellular traps.

    Science.gov (United States)

    Csomós, Krisztián; Kristóf, Endre; Jakob, Bernadett; Csomós, István; Kovács, György; Rotem, Omri; Hodrea, Judit; Bagoly, Zsuzsa; Muszbek, Laszlo; Balajthy, Zoltán; Csősz, Éva; Fésüs, László

    2016-08-11

    Neutrophil extracellular trap (NET) ejected from activated dying neutrophils is a highly ordered structure of DNA and selected proteins capable to eliminate pathogenic microorganisms. Biochemical determinants of the non-randomly formed stable NETs have not been revealed so far. Studying the formation of human NETs we have observed that polyamines were incorporated into the NET. Inhibition of myeloperoxidase, which is essential for NET formation and can generate reactive chlorinated polyamines through hypochlorous acid, decreased polyamine incorporation. Addition of exogenous primary amines that similarly to polyamines inhibit reactions catalyzed by the protein cross-linker transglutaminases (TGases) has similar effect. Proteomic analysis of the highly reproducible pattern of NET components revealed cross-linking of NET proteins through chlorinated polyamines and ɛ(γ-glutamyl)lysine as well as bis-γ-glutamyl polyamine bonds catalyzed by the TGases detected in neutrophils. Competitive inhibition of protein cross-linking by monoamines disturbed the cross-linking pattern of NET proteins, which resulted in the loss of the ordered structure of the NET and significantly reduced capacity to trap bacteria. Our findings provide explanation of how NETs are formed in a reproducible and ordered manner to efficiently neutralize microorganisms at the first defense line of the innate immune system.

  4. Relationship between Neutrophil-to-Lymphocyte Ratio and Prognosis in Patients with Deep Neck Space Infection

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    Engin Şengül

    2016-03-01

    Full Text Available Objective: The aim of this study was to demonstrate if there is any relation between the neutrophil-to-lym­phocyte ratio and prognosis in patients with deep neck space infections Methods: One hundred eight patients who were interned and treated due to deep neck infection in department of otolaryngology between January 2010 and January 2015 were analyzed. Demographics, clini­cal data, complications and treatment results of patients were evaluated. Pediatric patients between the age of 0 and 14, the patients who had only peritonsillary abscess, infection secondary to trauma were excluded from study. During follow-up and treatment duration the patients with deep neck infections were divided into two groups ac­cording to whether they had any complication (group 1 or not (group 2. Results: Fourteen (13% patients with deep neck in­fections had life-threatening complications whereas 94 (87% did not have any complication during follow-up and treatment duration. The mean neutrophil-to-lymphocyte ratio in patients with complications was significantly high­er than the patients without any complications (p0.05. Conclusion: This study showed that the neutrophil-to-lymphocyte ratio can be used as a cheap, easily obtained and predictive prognostic factor in patients with deep neck infections.

  5. In vivo characterization of neutrophil extracellular traps in various organs of a murine sepsis model.

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    Koji Tanaka

    Full Text Available Neutrophil extracellular traps (NETs represent extracellular microbial trapping and killing. Recently, it has been implicated in thrombogenesis, autoimmune disease, and cancer progression. The aim of this study was to characterize NETs in various organs of a murine sepsis model in vivo and to investigate their associations with platelets, leukocytes, or vascular endothelium. NETs were classified as two distinct forms; cell-free NETs that were released away from neutrophils and anchored NETs that were anchored to neutrophils. Circulating cell-free NETs were characterized as fragmented or cotton-like structures, while anchored NETs were characterized as linear, reticular, membranous, or spot-like structures. In septic mice, both anchored and cell-free NETs were significantly increased in postcapillary venules of the cecum and hepatic sinusoids with increased leukocyte-endothelial interactions. NETs were also observed in both alveolar space and pulmonary capillaries of the lung. The interactions of NETs with platelet aggregates, leukocyte-platelet aggregates or vascular endothelium of arterioles and venules were observed in the microcirculation of septic mice. Microvessel occlusions which may be caused by platelet aggregates or leukocyte-platelet aggregates and heterogeneously decreased blood flow were also observed in septic mice. NETs appeared to be associated with the formation of platelet aggregates or leukocyte-platelet aggregates. These observational findings may suggest the adverse effect of intravascular NETs on the host during a sepsis.

  6. Cytotoxicity and DNA damage in the neutrophils of patients with sickle cell anaemia treated with hydroxyurea

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    Alano Martins Pedrosa

    2014-04-01

    Full Text Available Hydroxyurea (HU is the most important advance in the treatment of sickle cell anaemia (SCA for preventing complications and improving quality of life for patients. However, some aspects of treatment with HU remain unclear, including their effect on and potential toxicity to other blood cells such as neutrophils. This study used the measurement of Lactate Dehydrogenase (LDH and Methyl ThiazolTetrazolium (MTT and the comet assay to investigate the cytotoxicity and damage index (DI of the DNA in the neutrophils of patients with SCA using HU.In the LDH and MTT assays, a cytoprotective effect was observed in the group of patients treated, as well as an absence of toxicity. When compared to patients without the treatment, the SS group (n=20, 13 women and 07 men, aged 18-69 years, and the group of healthy individuals (AA used as a control group (n=52, 28 women and 24 men, aged 19-60 years, The SSHU group (n=21, 11 women and 10 men, aged 19-63 years showed a significant reduction (p20 months, demonstrating that despite the cytoprotective effects in terms of cell viability, the use of HU can induce DNA damage in neutrophils.

  7. Adenosine triphosphate hydrolysis reduces neutrophil infiltration and necrosis in partial-thickness scald burns in mice.

    Science.gov (United States)

    Bayliss, Jill; Delarosa, Sara; Wu, Jianfeng; Peterson, Jonathan R; Eboda, Oluwatobi N; Su, Grace L; Hemmila, Mark; Krebsbach, Paul H; Cederna, Paul S; Wang, Stewart C; Xi, Chuanwu; Levi, Benjamin

    2014-01-01

    Extracellular adenosine triphosphate (ATP), present in thermally injured tissue, modulates the inflammatory response and causes significant tissue damage. The authors hypothesize that neutrophil infiltration and ensuing tissue necrosis would be mitigated by removing ATP-dependent signaling at the burn site. Mice were subjected to 30% TBSA partial-thickness scald burn by dorsal skin immersion in a water bath at 60 or 20°C (nonburn controls). In the treatment arm, an ATP hydrolyzing enzyme, apyrase, was applied directly to the site immediately after injury. Skin was harvested after 24 hours and 5 days for hematoxylin and eosin stain, elastase, and Ki-67 staining. Tumor necrosis factor (TNF)-α and interferon (IFN)-β expression were measured through quantitative real-time polymerase chain reaction. At 24 hours, the amount of neutrophil infiltration was different between the burn and burn + apyrase groups (P burn group at 24 hours and 5 days. TNF-α and IFN-β expression at 24 hours in the apyrase group was lower than in the burn group (P burn site allays the neutrophil response to thermal injury and reduces tissue necrosis. This decrease in inflammation and tissue necrosis is at least partially because of TNF-α and IFN-β signaling. Apyrase could be used as topical inflammatory regulators to quell the injury caused by inflammation.

  8. Microbicidal activity of neutrophils is inhibited by isolates from recurrent vaginal candidiasis (RVVC) caused by Candida albicans through fungal thioredoxin reductase.

    Science.gov (United States)

    Ratti, Bianca Altrão; Godoy, Janine Silva Ribeiro; de Souza Bonfim Mendonça, Patrícia; Bidóia, Danielle Lazarin; Nakamura, Tânia Ueda; Nakamura, Celso Vataru; Lopes Consolaro, Marcia Edilaine; Estivalet Svidzinski, Terezinha Inez; de Oliveira Silva, Sueli

    2015-01-01

    Vulvovaginal candidiasis (VVC) is characterized by an infection of the vulva and vagina, mainly caused by Candida albicans, a commensal microorganism that inhabits the vaginal, digestive, and respiratory mucosae. Vulvovaginal candidiasis affects approximately 75% of women, and 5% develop the recurrent form (RVVC). The aim of the present study was to evaluate whether neutrophils microbicidal response is triggered when activated with RVVC isolates caused by C. albicans. Our results showed that RVVC isolates induced neutrophil migration but significantly decrease the microbicidal activity of neutrophils, compared with VVC and ASS isolates. The microbicidal activity of neutrophils is highly dependent on the production of reactive oxygen species/reactive nitrogen species (ROS/RNS). However, this isolate induced detoxification of ROS/RNS produced by neutrophils, reflected by the high level of thiol groups and by the oxygen consumption. Therefore, RVVC isolates induced biochemical changes in the inflammatory response triggered by neutrophils, and these effects were mainly related to the detoxification of ROS/RNS through the thioredoxin reductase (TR), a key antioxidant enzyme in fungi. This might be one of the resistance mechanisms triggered by RVVC caused by C. albicans. Copyright © 2014 Elsevier Inc. All rights reserved.

  9. Acral manifestations of Sweet syndrome (neutrophilic dermatosis of the hands).

    Science.gov (United States)

    Wolf, Ronni; Tüzün, Yalçın

    Neutrophilic dermatosis of the hand (NDH) is a rare localized variant of the syndrome, originally described two decades ago by Strutton et al. The lesions of NDH and Sweet syndrome are similar, as indicated in the first report of NDH. Both diagnoses are characterized by an acute onset of fever, leukocytosis, and tender, erythematous infiltrated plaques. There are also bullae and ulceration in NDH, in contrast to Sweet syndrome, in which bullae are quite uncommon, especially at the early stages. Similar to Sweet syndrome, the majority of NDH patients are women (69%). Patients with NDH present with fever, peripheral neutrophilia, leukocytosis, and/or an elevated erythrocyte sedimentation rate or C-reactive protein level, but at a significantly lower rate than those in Sweet syndrome (33%). Similar to Sweet syndrome, NDH has been associated with the following conditions: Malignancies (particularly hematological [21%], most common of which is acute myelogenous leukemia, but many other malignancies as well), inflammatory bowel disease (19%), medication and vaccination-related eruptions, bacterial and viral infections, rheumatologic diseases, and others. The clues to the diagnosis of NDH are the same as for Sweet syndrome. Awareness of this diagnosis is important not only to avoid unnecessary medical and surgical therapy and to expediently initiate the administration of steroids for this highly responsive dermatosis, but also to conduct an appropriate workup to exclude associated diseases, especially malignancies. Copyright © 2017 Elsevier Inc. All rights reserved.

  10. The Fc Receptor Polymorphisms and Expression of Neutrophil Activation Markers in Patients with Sickle Cell Disease from Western India

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    Harshada K. Kangne

    2013-01-01

    Full Text Available Objective. Sickle cell disease has variable clinical manifestations. Activation of neutrophils plays an important role in the initiation and propagation of vaso occlusive crises which can be analysed by determining the expression of neutrophil antigens such as CD16, CD32, and CD62L. The common FcγR polymorphisms (FcγRIIA and FcγRIIIB are considered to influence clinical presentation. This study focuses on distribution of FcγR polymorphisms and their association with neutrophil activity among the patients from western India. Methods. In this paper 127 sickle cell anemia patients and 58 patients with sickle-β-thalassemia (median age 12±8.58 years with variable clinical phenotypes along with 175 normals were investigated. FcγRs polymorphisms were analysed by RFLP and AS-PCR. Activation of neutrophils was measured by flow cytometry. Results. The genotypic frequency of the H/R genotype of FcγRIIA and the NA1/NA1 genotype of FcγRIIIB was significantly decreased in patients compared to normals (P-0.0074, P-0.0471, resp.. We found a significant difference in the expression of CD32 and CD62L among the patients as against normals. A significantly higher expression of CD32 was seen in the milder patients with the H/H genotype (P-0.0231, whereas the expression of CD16 was higher in severe patients with the NA2/NA2 genotype (P-0.0312. Conclusion. The two FcγR polymorphisms had significant association with variable phenotypes of sickle cell disease. The expression of CD62L decreased in our patients indicating activation of neutrophils.

  11. Estrogen supplementation failed to attenuate biochemical indices of neutrophil infiltration or damage in rat skeletal muscles following ischemia.

    Science.gov (United States)

    Tiidus, Peter M; Deller, Mirada; Bombardier, Eric; Gül, Mustafa; Liu, X Linda

    2005-01-01

    This study examined the effects of estrogen supplementation on markers of neutrophil infiltration and damage in skeletal muscle of rats following ischemia. Male and female gonad-intact rats, with or without 14 days of estrogen supplementation were subjected to two hours of hind-limb ischemia and sacrificed at 24, 48 or 72 hours post-ischemia. Control animals were sacrificed without ischemia. Plantaris and red and white gastrocneimus muscles were removed and assayed for myeloperoxidase (MPO), a marker of neutrophil infiltration, and glucose-6-phosphate dehydrogenase (G6PD) and beta-glucuronidase (betaGLU), as markers of muscle damage. Significant elevations of MPO, G6PD and betaGLU activities were observed at various time points post-ischemia. No systematic differences between genders were noted in any of the measures. Estrogen supplementation in both male and female animals failed to significantly attenuate post-ischemia increases in MPO, G6PD and betaGLU activities in any of the muscles studied and in some cases accentuated activities of some of these measures. Unlike previous findings following exercise in skeletal muscle, this study failed to demonstrate estrogen-induced attenuation of indices of neutrophil infiltration or damage in skeletal muscles of rats up to 72 hours following ischemia. This demonstrates that estrogen may not consistently attenuate neutrophil infiltration and that a number of variables including damage modality, tissue or estrogen level may influence this.

  12. Estrogen supplementation failed to attenuate biochemical indices of neutrophil infiltration or damage in rat skeletal muscles following ischemia

    Directory of Open Access Journals (Sweden)

    PETER M TIIDUS

    2005-01-01

    Full Text Available This study examined the effects of estrogen supplementation on markers of neutrophil infiltration and damage in skeletal muscle of rats following ischemia. Male and female gonad-intact rats, with or without 14 days of estrogen supplementation were subjected to two hours of hind-limb ischemia and sacrificed at 24, 48 or 72 hours post-ischemia. Control animals were sacrificed without ischemia. Plantaris and red and white gastrocneimus muscles were removed and assayed for myeloperoxidase (MPO, a marker of neutrophil infiltration, and glucose-6-phosphate dehydrogenase (G6PD and ß-glucuronidase (GLU, as markers of muscle damage. Significant elevations of MPO, G6PD and GLU activities were observed at various time points post-ischemia. No systematic differences between genders were noted in any of the measures. Estrogen supplementation in both male and female animals failed to significantly attenuate post-ischemia increases in MPO, G6PD and GLU activities in any of the muscles studied and in some cases accentuated activities of some of these measures. Unlike previous findings following exercise in skeletal muscle, this study failed to demonstrate estrogen-induced attenuation of indices of neutrophil infiltration or damage in skeletal muscles of rats up to 72 hours following ischemia. This demonstrates that estrogen may not consistently attenuate neutrophil infiltration and that a number of variables including damage modality, tissue or estrogen level may influence this.

  13. Demodex-associated bacterial proteins induce neutrophil activation.

    LENUS (Irish Health Repository)

    2012-02-01

    Background: Patients with rosacea demonstrate a higher density of Demodex mites in their skin than controls. A bacterium isolated from a Demodex mite from a patient with papulopustular rosacea (PPR) was previously shown to provoke an immune response in patients with PPR or ocular rosacea thus suggesting a possible role for bacterial proteins in the etiology of this condition. Objectives: To examine the response of neutrophils to proteins derived from a bacterium isolated from a Demodex mite. Methods: Bacterial cells were lysed and proteins were partially purified by AKTA-FPLC. Isolated neutrophils were exposed to bacterial proteins and monitored for alterations in migration, degranulation and cytokine production. Results: Neutrophils exposed to proteins from Bacillus cells demonstrated increased levels of migration and elevated release of MMP-9, an enzyme known to degrade collagen and cathelicidin, an antimicrobial peptide. In addition neutrophils exposed to the bacterial proteins demonstrated elevated rates of Il-8 and TNF-alpha production. Conclusions: Proteins produced by a bacterium isolated from a Demodex mite have the ability to increase the migration, degranulation and cytokine production abilities of neutrophils. These results suggest that bacteria may play a role in the inflammatory erythema associated with rosacea.

  14. Peptide secreted by human alveolar macrophages releases neutrophil granule contents

    International Nuclear Information System (INIS)

    MacArthur, C.K.; Miller, E.J.; Cohen, A.B.

    1987-01-01

    A monoclonal antibody was developed against an 8000-kDa enzyme-releasing peptide (ERP) released from human alveolar macrophages. ERP was isolated on an immunoaffinity column containing the antibody bound to staphylococcal protein A-Sepharose, and by autoradiography. Release of ERP from the macrophages is not changed by plastic adherence, phagocytosis, calcium ionophore, or phorbol esters. The peptide was not antigenically similar to interferon-γ, tumor necrosis factor, or interleukin lα or 1β. The release of constituents from azurophilic and specific granules was the main identified biologic function of ERP. ERP was a more effective secretagogue in the untreated neutrophils and f-met-leu-phe was more effective in the cytochalasin B-treated neutrophils. Absorption of ERP from macrophage-conditioned medium removed a small amount of the chemotactic activity; however, the immunopurified peptide was not chemotactic or chemokinetic for neutrophils, and at high concentrations, it suppressed base line chemokinesis. Treatment of washed macrophages with trypsin released active ERP of approximately the same m.w. of spontaneously secreted ERP. These studies showed that human alveolar macrophages release a peptide which is a secretagogue for human neutrophils under conditions which may be encountered in the lungs during certain disease states. Proteolytic enzymes which are free in the lungs may release the peptide and lead to the secretion of neutrophil enzymes

  15. Radiation-induced muscositis and neutrophil granulocytes in oral mucosa; Strahleninduzierte Mukositis und neutrophile Granulozyten in der Mundschleimhaut

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    Schmidberger, H.; Rave-Fraenk, M.; Kim, S.; Hille, A.; Pradier, O.; Hess, C.F. [Klinik fuer Strahlentherapie und Radioonkologie, Univ. Goettingen (Germany)

    2003-10-01

    Background: Chemotherapy-induced mucositis can be related to a decrease in oral neutrophils. We tested the relationship between radiation-induced mucositis and oral neutrophil counts. Patients and Methods: Oral neutrophil counts were obtained for ten patients with head and neck cancer who received radiotherapy of the pharynx and oral cavity. Four patients received additional chemotherapy (5-FU, Mitomycin). Counts were obtained before and during treatment; four healthy volunteers were included in the study as well. For evaluation, a quantitative mouth rinse assay, including neutrophil-staining with acridin-orange, was applied. Results: We observed large inter-individual variations with respect to neutrophil counts for patients and control persons (Table 1). During treatment (irradiation or chemoirradiation), large intra-individual variations were seen additionally (Figure 1). We found a correlation between neutrophil counts and clinical reaction grade. Neutrophil counts increased with increasing mucositis (Figure 2). This increase was more pronounced for patients treated with chemoirradiation compared to radiation alone. Treatment breaks at weekends had no clear influence on neutrophil counts. Conclusions: We observed a weak correlation between neutrophil counts and clinical reaction grade. However, the variations in neutrophil counts are too large to utilize this parameter as a surrogate for clinical mucositis grading. The assumption that a decrease in oral neutrophils is associated with radiation-induced mucositis was clearly negated. (orig.) [German] Hintergrund: Die chemotherapieinduzierte Mukositis kann mit einer Verarmung der Mundschleimhaut an neutrophilen Granulozyten vergesellschaftet sein. Wir ueberprueften den Zusammenhang zwischen der radiogenen Mukositis und der Anzahl neutrophiler Granulozyten. Patienten und Methoden: Bei zehn Patienten mit Tumoren der Kopf-Hals-Region, die sich einer Strahlentherapie unterzogen, wurde die Anzahl enoraler neutrophiler

  16. Neutrophil depletion in the early inflammatory phase delayed cutaneous wound healing in older rats: improvements due to the use of un-denatured camel whey protein.

    Science.gov (United States)

    Ebaid, Hossam

    2014-03-04

    While it is known that advanced age alters the recruitment of neutrophils during wound healing, thereby delaying the wound healing process, little is known about prolonged wound healing in advanced ages. Thus, we investigated the correlation of neutrophil recruitment with healing events, and the impact of whey protein (WP) on neutrophil activation. The animals were allocated into wounded young group, wounded older group and wounded older rats with daily treatment of WP at a dose of 100 mg/kg of body weight. Our results pointed to a marked deficiency in the number of neutrophils in the wounds of older rats, which was accompanied with impairment of the healing process. In the group of older rats, phagocytic activity, as tested by fluorescence microscopy, declined throughout the first 24 hours after wounding. Both the neutrophil number and the phagocytic activity recovered in older rats which received WP supplementation. Interestingly, WP was found to significantly up-regulate the MIP-1α and CINC-1 mRNA expression in old rats. On the other hand, the wound size in older rats was significantly higher than that in younger ones. Blood angiogenesis was also significantly delayed in the older group as opposed to the young rats. WP, however, was found to return these indices to normal levels in the older rats. Proliferation and epidermal migration of the keratinocytes and the collagen deposition were also returned to the normal rates. This data confirms the critical role of neutrophil recruitment in the early inflammatory phase of wound healing in older rats. In addition, WP protein was used to improve neutrophil function in older rats, healing events returned to a more normal profile. The virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/2100966986117779.

  17. Doxycycline induced photodamage to human neutrophils and tryptophan

    International Nuclear Information System (INIS)

    Sandberg, S.; Glette, J.; Hopen, G.; Solberg, C.O.

    1984-01-01

    Neutrophil function were studied following irradiation (340-380 nm) of the cells in the presence of 22 μM doxycycline. At increasing light fluence the locomotion, chemiluminescence and glucose oxidation (by the hexose monophosphate shunt) of the neutrophils steadily decreased. The photodamage increased with increasing preincubation temperature and time and was enhanced in D 2 O, reduced in azide and abolished in anaerobiosis. Superoxide dismutase, catalase or mannitol did not influence the photodamage. Photooxidation of tryptophan in the presence of doxycycline was increased 9-10-fold in D 2 O and nearly abolished in the presence of 0.25 mM NaN 3 , indicating that singlet oxygen is the most important reactive oxygen species in the doxycycline-induced photodamage. The results may explain some of the features of tetracycline-induced photosensitivity and why other authors have obtained diverging results when studying the influence of tetracyclines on neutrophil functions. (author)

  18. Neutrophilic dermatosis resembling pyoderma gangrenosum in a dog with polyarthritis.

    Science.gov (United States)

    Bardagí, M; Lloret, A; Fondati, A; Ferrer, L

    2007-04-01

    This report describes a case of neutrophilic dermatosis in a dog, with a number of clinical and pathological similarities to human pyoderma gangrenosum. A seven-year-old, female German shepherd dog with a history of non-erosive idiopathic polyarthritis was presented with severe facial swelling, bilateral erosivoulcerative lesions on the muzzle and multiple, eroded, dermal-subcutaneous nodules on the cranial trunk. Histopathological examination of skin biopsies revealed a necrotising neutrophilic dermatitis. No infectious agents could be detected using specific stains, immunohistochemistry, serology and bacterial aerobic, anaerobic or fungal cultures. A sterile neutrophilic dermatosis resembling human pyoderma gangrenosum was presumptively diagnosed, and the patient showed an excellent response to treatment with prednisone and ciclosporin.

  19. Localized Subcutaneous Acute Febrile Neutrophilic Dermatosis in a Dog

    Directory of Open Access Journals (Sweden)

    Karolin Schoellhorn

    2012-01-01

    Full Text Available A two-year-old spayed female mixed-breed dog was presented with a five-day history of hemorrhagic gastroenteritis and fever. On physical examination, the dog was lethargic and clinically dehydrated. The skin of the entire ventral abdomen extending to both flanks was erythematous, swollen and painful on palpation. Histopathological examination of skin biopsies revealed a severe diffuse neutrophilic dermatitis and panniculitis, resembling the subcutaneous form of Sweet’s syndrome in humans. A large part of the skin lesion developed full-thickness necrosis. After intensive care, three surgical wound debridements and wound adaptations, the wound healed by secondary intention within ten weeks. In the absence of infection of the skin or neoplasia, a diagnosis of neutrophilic dermatosis and panniculitis, resembling the subcutaneous form of acute febrile neutrophilic dermatosis, was made.

  20. Interleukin-8: an expanding universe beyond neutrophil chemotaxis and activation.

    Science.gov (United States)

    Mukaida, N

    2000-12-01

    Since the discovery 13 years ago of interleukin (IL)-8 as a potent neutrophil chemotactic factor, accumulating evidence has established it as a crucial mediator in neutrophil-dependent acute inflammation. Numerous observations have demonstrated that various types of cells can produce a large amount of IL-8, either in response to various stimuli or constitutively, after malignant transformation. Recent studies of IL-8-mediated signaling have revealed that IL-8 activates a wide range of signaling molecules in a coordinate manner. IL-8 has been proven to have diverse actions on various types of leukocytic and nonleukocytic cells besides neutrophils. The author reviews recent progress in IL-8 signal transduction and biological actions on nonneutrophilic leukocytes, including T lymphocytes, monocytes, and hematopoietic progenitor cells. Potential involvement of IL-8 in viral infections and tumor progression is also discussed.

  1. Predictive contribution of neutrophil/lymphocyte ratio in diagnosis of brucellosis.

    Science.gov (United States)

    Olt, Serdar; Ergenç, Hasan; Açıkgöz, Seyyid Bilal

    2015-01-01

    Here we wanted to investigate predictive value of neutrophil/lymphocyte ratio (NLR) and platelet/lymphocyte ratio (PLR) in the diagnosis of brucellosis. Thirty-two brucellosis patients diagnosed with positive serum agglutination test and thirty-two randomized healthy subjects were enrolled in this study retrospectively. Result with ROC analyzes the baseline NLR and hemoglobin values were found to be significantly associated with brucellosis (P = 0.01, P = 0.01, resp.). Herein we demonstrated for the first time that NLR values were significantly associated with brucellosis. This situation can help clinicians during diagnosis of brucellosis.

  2. Anti-Inflammatory benefits of antibiotic-induced neutrophil apoptosis: tulathromycin induces caspase-3-dependent neutrophil programmed cell death and inhibits NF-kappaB signaling and CXCL8 transcription.

    Science.gov (United States)

    Fischer, Carrie D; Beatty, Jennifer K; Zvaigzne, Cheryl G; Morck, Douglas W; Lucas, Merlyn J; Buret, A G

    2011-01-01

    Clearance of apoptotic neutrophils is a central feature of the resolution of inflammation. Findings indicate that immuno-modulation and induction of neutrophil apoptosis by macrolide antibiotics generate anti-inflammatory benefits via mechanisms that remain obscure. Tulathromycin (TUL), a new antimicrobial agent for bovine respiratory disease, offers superior clinical efficacy for reasons not fully understood. The aim of this study was to identify the immuno-modulating effects of tulathromycin and, in this process, to establish tulathromycin as a new model for characterizing the novel anti-inflammatory properties of antibiotics. Bronchoalveolar lavage specimens were collected from Holstein calves 3 and 24 h postinfection, challenged intratracheally with live Mannheimia haemolytica (2 × 10(7) CFU), and treated with vehicle or tulathromycin (2.5 mg/kg body weight). Terminal deoxynucleotidyltransferase-mediated dUTP-biotin nick end labeling (TUNEL) staining and enzyme-linked immunosorbent assay (ELISA) revealed that tulathromycin treatment significantly increased leukocyte apoptosis and reduced levels of proinflammatory leukotriene B(4) in M. haemolytica-challenged calves. In vitro, tulathromycin concentration dependently induced apoptosis in freshly isolated bovine neutrophils from healthy steers in a capase-3-dependent manner but failed to induce apoptosis in bovine fibroblasts, epithelial cells, and endothelial cells, as well as freshly isolated bovine blood monocytes and monocyte-derived macrophages. The proapoptotic effects of TUL were also, in part, drug specific; equimolar concentrations of penicillin G, oxytetracycline, and ceftiofur failed to cause apoptosis in bovine neutrophils. In addition, tulathromycin significantly reduced levels of phosphorylated IκBα, nuclear translocation of NF-κB p65, and mRNA levels of proinflammatory interleukin-8 in lipopolysaccharide (LPS)-stimulated bovine neutrophils. The findings illustrate novel mechanisms through which

  3. High resolution of heterogeneity among human neutrophil granules: physical, biochemical, and ultrastructural properties of isolated fractions.

    Science.gov (United States)

    Rice, W G; Kinkade, J M; Parmley, R T

    1986-08-01

    Previous studies on the fractionation of human neutrophil granules have identified two major populations: myeloperoxidase (MPO)-containing azurophil, or primary, granules and MPO-deficient specific, or secondary, granules. Peripheral blood neutrophils from individual donors were lysed in sucrose-free media by either hypotonic shock or nitrogen cavitation. Using a novel two-gradient Percoll density centrifugation system, the granule-rich postnuclear supernatant was rapidly (ten minutes) and reproducibly resolved into 13 granule fractions (L1 through L8 and H1 through H5). Granule flotation and recentrifugation experiments on both continuous, self-generated and multiple-step gradients using individual and mixed isolated fractions demonstrated that the banding patterns were isopycnic and nonartifactual. Isolated granules were intact based on the findings that biochemical latency of several granule enzymes was greater than 95%, and thin-sectioned electron micrographs demonstrated intact granule profiles. Biochemical analyses of the granule marker proteins MPO, beta-glucuronidase, lysozyme, and lactoferrin indicated that a number of the fractions were related to the major azurophil and specific granule populations. Lactoferrin was found in ten of 13 fractions (L1 through L8, H1 to H2), whereas MPO was found in every fraction. Consistent with these biochemical data, all fractions exhibited varying degrees of heterogeneity based on ultrastructural morphology and cytochemistry, including diaminobenzidine (DAB) reactivity for peroxidase and periodate-thiocarbohydrazide-silver proteinate (PA-TCH-SP) staining for complex glycoconjugates. A variable but significant percentage (23% to 70%) of the granules in fractions L1 through L8 and H1 and H2 showed DAB reactivity, while about 90% of the granules in fractions H3 through H5 were peroxidase positive. These results demonstrated that DAB-reactive granules spanned the entire range of granule size and density. Ultrastructural PA

  4. The Effects of Plantago major on the Activation of the Neutrophil Respiratory Burst

    Directory of Open Access Journals (Sweden)

    Elaine Reina

    2013-10-01

    Full Text Available Plantago major is a common plant that grows worldwide in temperate zones and is found in fields, lawns, and on the roadsides. Its leaves and seeds have been used in almost all parts of the world for centuries as a wound healer, analgesic, antioxidant, and antibiotic, as well as an immune system modulator, antiviral, antifungal, and anti-inflammatory agent. Baicalein and aucubin are the two most biologically active components of P. major, and both have been shown to have antioxidant, anti-inflammatory, and anticancer properties. Neutrophils have a pivotal role in wound healing and inflammation. Their principal mechanism of host defense is the killing of pathogens via the production of reactive oxygen species (ROS. The aim of the present study was to determine the in vitro effects of P. major extract, baicalein, and aucubin on human neutrophil respiratory burst activity. The cytotoxicity of the agents was assessed by lactate dehydrogenase (LDH assays. A standard luminol-dependent chemiluminescence (CL assay was utilized to monitor the respiratory burst of the neutrophils after exposure to P. major extract and its two active ingredients, baicalein and aucubin. Three replicates per group were included in each of the three runs of the experiments and analysis of variance (ANOVA was used for statistical analysis. P. major and baicalein were not toxic to the cells at any of the concentrations examined. Aucubin was toxic to the cells only at the highest concentration tested (P=0.0081. However, genistein was toxic to the cells at all of the concentrations examined except for the lowest concentration of 16.9 μg/ml (P=0.985. P. major (−0.10±0.11, aucubin (0.06±0.16, baicalein (−0.10±0.11, and genistein (−0.18±0.07 all significantly (P<0.0001 inhibited ROS production from the neutrophils. P. major extract inhibited neutrophil ROS production, as did aucubin and baicalein. Therefore, these components should be investigated further with relation to

  5. Protective Effect of Phillyrin on Lethal LPS-Induced Neutrophil Inflammation in Zebrafish

    Directory of Open Access Journals (Sweden)

    Liling Yang

    2017-10-01

    Full Text Available Background/Aims: Forsythia suspensa Vahl. (Oleaceae fruits are widely used in traditional Chinese medicine to treat pneumonia, typhoid, dysentery, ulcers and oedema. Antibacterial and anti-inflammatory activities have been reported for phillyrin (PHN, the main ingredient in Forsythia suspensa Vahl fruits, in vitro. However, the underlying mechanisms in vivo remain poorly defined. In this study, we discovered that PHN exerted potent anti-inflammatory effects in lethal LPS-induced neutrophil inflammation by suppressing the MyD88-dependent signalling pathway in zebrafish. Methods: LPS-yolk microinjection was used to induce a lethal LPS-infected zebrafish model. The effect of PHN on the survival of zebrafish challenged with lethal LPS was evaluated using survival analysis. The effect of PHN on neutrophil inflammation grading in vivo was assessed by tracking neutrophils with a transgenic line. The effects of PHN on neutrophil production and migration were analysed by SB+ cell counts during consecutive hours after modelling. Additionally, key cytokines and members of the MyD88 signalling pathway that are involved in inflammatory response were detected using quantitative RT-PCR. To assess gene expression changes during consecutive hours after modelling, the IL-1β, IL-6, TNF-α, MyD88, TRIF, ERK1/2, JNK, IκBa and NF-κB expression levels were measured. Results: PHN could protect zebrafish against a lethal LPS challenge in a dose-dependent manner, as indicated by decreased neutrophil infltration, reduced tissue necrosis and increased survival rates. Up-regulated IL-1β, IL-6 and TNF-α expression also showed the same tendencies of depression by PHN. Critically, PHN significantly inhibited the LPS-induced activation of MyD88, IκBa, and NF-κB but did not affect the expression of ERK1/2 MAPKs or JNK MAPKs in LPS-stimulated zebrafish. Additionally, PHN regulated the MyD88/IκBα/NF-κB signalling pathway by controlling IκBα, IL-1β, IL-6, and TNF

  6. Innate Defense against Influenza A Virus: Activity of Human Neutrophil Defensins and Interactions of Defensins with Surfactant Protein D

    DEFF Research Database (Denmark)

    Hartshorn, Kevan L.; White, Mitchell R.; Tecle, Tesfaldet

    2006-01-01

    Surfactant protein D (SP-D) plays important roles in innate host defense against influenza A virus (IAV) infection, in part by modifying interactions with neutrophils. Human neutrophil defensins (HNPs) inhibit infectivity of enveloped viruses, including IAV. Our goal in this study was to characte......Surfactant protein D (SP-D) plays important roles in innate host defense against influenza A virus (IAV) infection, in part by modifying interactions with neutrophils. Human neutrophil defensins (HNPs) inhibit infectivity of enveloped viruses, including IAV. Our goal in this study...... was to characterize antiviral interactions between SP-D and HNPs. Recombinant and/or natural forms of SP-D and related collectins and HNPs were tested for antiviral activity against two different strains of IAV. HNPs 1 and 2 did not inhibit viral hemagglutination activity, but they interfered...... with the hemagglutination-inhibiting activity of SP-D. HNPs had significant viral neutralizing activity against divergent IAV strains. However, the HNPs generally had competitive effects when combined with SP-D in assays using an SP-D-sensitive IAV strain. In contrast, cooperative antiviral effects were noted in some...

  7. May the variable magnetic field and pulse red light induce synergy effects in respiratory burst of neutrophils in vitro?

    International Nuclear Information System (INIS)

    Nawrocka - Bogusz, H; Jaroszyk, F

    2011-01-01

    We investigated the effect of the red light (R) (630 nm), magnetic field (MF) and magnetic field combined with the red light (MF+R) upon reactive oxygen species (ROS) production by neutrophils in vitro. The object of the research was hydrogen peroxide (H 2 O 2 ) formation during neutrophils respiratory burst or within steady-state. Blood from healthy volunteers was used for the purpose of the study. Flow cytometry method, using transformation of DCFH-DA (2'7'-dichlorofluorescin diacetate) to the fluorescent DCF (2'7'-dichlorofluorescin), was used for estimation of hydrogen peroxide production. The variable magnetic field of ELF range of the mean induction equals 26.7(μT), the red light at the energy density of 1.17(J/cm 2 ) and their combination were applied for 30 minutes each. The fundamental frequency of pulses was 180÷ 195 Hz. A statistically significant decrease of H 2 O 2 production by neutrophils was observed. The level of the decrease was in the range of 10-30% and was dependent on the kind of applied physical factors and whether neutrophils were stimulated or not. The observation showed that the variable magnetic field combined with red light do not induce the synergy effect.

  8. Ir-LBP, an ixodes ricinus tick salivary LTB4-binding lipocalin, interferes with host neutrophil function.

    Directory of Open Access Journals (Sweden)

    Jérôme Beaufays

    Full Text Available BACKGROUND: During their blood meal, ticks secrete a wide variety of proteins that can interfere with their host's defense mechanisms. Among these proteins, lipocalins play a major role in the modulation of the inflammatory response. METHODOLOGY/PRINCIPAL FINDINGS: We previously identified 14 new lipocalin genes in the tick Ixodes ricinus. One of them codes for a protein that specifically binds leukotriene B4 with a very high affinity (Kd: +/-1 nM, similar to that of the neutrophil transmembrane receptor BLT1. By in silico approaches, we modeled the 3D structure of the protein and the binding of LTB4 into the ligand pocket. This protein, called Ir-LBP, inhibits neutrophil chemotaxis in vitro and delays LTB4-induced apoptosis. Ir-LBP also inhibits the host inflammatory response in vivo by decreasing the number and activation of neutrophils located at the tick bite site. Thus, Ir-LBP participates in the tick's ability to interfere with proper neutrophil function in inflammation. CONCLUSIONS/SIGNIFICANCE: These elements suggest that Ir-LBP is a "scavenger" of LTB4, which, in combination with other factors, such as histamine-binding proteins or proteins inhibiting the classical or alternative complement pathways, permits the tick to properly manage its blood meal. Moreover, with regard to its properties, Ir-LBP could possibly be used as a therapeutic tool for illnesses associated with an increased LTB4 production.

  9. Azurophil granule proteins constitute the major mycobactericidal proteins in human neutrophils and enhance the killing of mycobacteria in macrophages.

    Directory of Open Access Journals (Sweden)

    Prajna Jena

    Full Text Available Pathogenic mycobacteria reside in, and are in turn controlled by, macrophages. However, emerging data suggest that neutrophils also play a critical role in innate immunity to tuberculosis, presumably by their different antibacterial granule proteins. In this study, we purified neutrophil azurophil and specific granules and systematically analyzed the antimycobacterial activity of some purified azurophil and specific granule proteins against M. smegmatis, M. bovis-BCG and M. tuberculosis H37Rv. Using gel overlay and colony forming unit assays we showed that the defensin-depleted azurophil granule proteins (AZP were more active against mycobacteria compared to other granule proteins and cytosolic proteins. The proteins showing antimycobacterial activity were identified by MALDI-TOF mass spectrometry. Electron microscopic studies demonstrate that the AZP disintegrate bacterial cell membrane resulting in killing of mycobacteria. Exogenous addition of AZP to murine macrophage RAW 264.7, THP-1 and peripheral blood monocyte-derived macrophages significantly reduced the intracellular survival of mycobacteria without exhibiting cytotoxic activity on macrophages. Immunofluorescence studies showed that macrophages actively endocytose neutrophil granular proteins. Treatment with AZP resulted in increase in co-localization of BCG containing phagosomes with lysosomes but not in increase of autophagy. These data demonstrate that neutrophil azurophil proteins may play an important role in controlling intracellular survival of mycobacteria in macrophages.

  10. Chronic obstructive pulmonary disease (COPD): neutrophils, macrophages and lymphocytes in patients with anterior tuberculosis compared to tobacco related COPD.

    Science.gov (United States)

    Guiedem, Elise; Ikomey, George Mondinde; Nkenfou, Céline; Walter, Pefura-Yone Eric; Mesembe, Martha; Chegou, Novel Njweipi; Jacobs, Graeme Brendon; Okomo Assoumou, Marie Claire

    2018-03-27

    The inflammatory profile of chronic obstructive pulmonary disease (COPD) related to tobacco is known in certain studies while that of the post tuberculosis form is not yet known. This study aimed to evaluate the levels of neutrophils, macrophages and lymphocytes cells in sputum of COPD patients with history of smoking or anterior tuberculosis. Enumeration of cells in samples was analyzed using standard microscopy. We enrolled 92 participants, 46 (50%) were COPD subjects comprising 22 (47.83%) smokers and 24 (52.17%) with anterior tuberculosis while 46 (50%) healthy persons constituted the control group. The levels of neutrophils, lymphocytes and monocytes were statistically higher in COPD patients compared to the control group with p-values of 0.0001 respectively. Neutrophils levels were higher in COPD patients with history of tobacco than in COPD patients with anterior tuberculosis with a mean rate of 4.72 × 10 6 /ml and 2.48 × 10 6 /ml respectively (p = 0.04). The monocytes and lymphocytes levels were not statistically different between the two sub-groups of COPD patients with p-value of 0.052 and 0.91 respectively. Neutrophils are the only inflammatory cells that were significantly higher in COPD patients with history of smoking as compared to COPD patients with anterior tuberculosis.

  11. The emerging role of neutrophils in thrombosis – The journey of TF through NETs

    Directory of Open Access Journals (Sweden)

    Konstantinos eKambas

    2012-12-01

    Full Text Available The production of TF by neutrophils and their contribution in thrombosis was until recently a matter of scientific debate. Experimental data suggested the de novo TF production by neutrophils under inflammatory stimuli, while others proposed that these cells acquired microparticle-derived TF. Recent experimental evidence revealed the critical role of neutrophils in thrombotic events. Neutrophil derived TF has been implicated in this process in several human and animal models. Additionally, neutrophil extracellular trap (NET release has emerged as a major contributor in neutrophil-driven thrombogenicity in disease models including sepsis, deep venous thrombosis and malignancy. It is suggested that NETs provide the scaffold for fibrin deposition and platelet entrapment and subsequent activation. The recently reported autophagy-dependent extracellular delivery of TF in NETs further supports the involvement of neutrophils in thrombosis. Herein, we seek to review novel data regarding the role of neutrophils in thrombosis, emphasizing the implication of TF and NETs.

  12. CXCR2 mediates NADPH oxidase-independent neutrophil extracellular trap formation in cystic fibrosis airway inflammation

    NARCIS (Netherlands)

    Marcos, Veronica; Zhou, Zhe; Yildirim, Ali Onder; Bohla, Alexander; Hector, Andreas; Vitkov, Ljubomir; Wiedenbauer, Eva-Maria; Krautgartner, Wolf Dietrich; Stoiber, Walter; Belohradsky, Bernd H.; Rieber, Nikolaus; Kormann, Michael; Koller, Barbara; Roscher, Adelbert; Roos, Dirk; Griese, Matthias; Eickelberg, Oliver; Döring, Gerd; Mall, Marcus A.; Hartl, Dominik

    2010-01-01

    Upon activation, neutrophils release DNA fibers decorated with antimicrobial proteins, forming neutrophil extracellular traps (NETs). Although NETs are bactericidal and contribute to innate host defense, excessive NET formation has been linked to the pathogenesis of autoinflammatory diseases.

  13. Functional characterization of mitochondria in neutrophils: a role restricted to apoptosis

    NARCIS (Netherlands)

    Maianski, N. A.; Geissler, J.; Srinivasula, S. M.; Alnemri, E. S.; Roos, D.; Kuijpers, T. W.

    2004-01-01

    Mitochondria are known to combine life-supporting functions with participation in apoptosis by controlling caspase activity. Here, we report that in human blood neutrophils the mitochondria are different, because they preserve mainly death-mediating abilities. Neutrophil mitochondria hardly

  14. Increased neutrophil priming and sensitization before commencing cardiopulmonary bypass in cardiac surgical patients

    NARCIS (Netherlands)

    Gu, YJ; Schoen, P; Tigchelaar, [No Value; Loef, BG; Ebels, T; Rankin, AJ; van Oeveren, W

    2002-01-01

    Background. Neutrophil activation is implicated in postoperative complications in patients having cardiac surgery with cardiopulmonary bypass (CPB). This study was designed to determine the temporal fluctuations in the primability of neutrophils in the preoperative, intraoperative, and postoperative

  15. Assessment of neutrophil / lymphocyte ratio in patients with myocardial bridge

    Directory of Open Access Journals (Sweden)

    Abdulkadir Yıldız

    2014-03-01

    Full Text Available Objective: Myocardial bridge (MB is a congenital anomaly characterized by systolic narrowing of the epicardial coronary arterial segment while traveling in the myocardium. It is a benign entity but previous studies showed that the proximal portion is prone to an enhanced atherosclerosis. Neutrophil/lymphocyte ratio (NLR is a sensitive marker of systemic inflammation used as a predictor for adverse cardiovascular outcomes in atherosclerotic heart disease. So in this study, we sought to evaluate the association between NLR and myocardial bridging. Methods: A total of 172 patients (mean age: 50.8 ± 11.5 years, 77.3% men with either angiographically proven MB or normal coronary arteries were included in the study. For the entire study population, hematologic parameters were measured using an automatic blood counter. Results: The study population consisted of 71 patients with MB (mean age: 51.4 ± 11.9 years, 80.3% male and 101 patients with normal coronary arteries (mean age: 50.5 ± 11.3 years, 75.2% male. There were no significant differences between groups regarding hemoglobin level, platelet count, glucose and creatinine. Compared to the control group, NLR was significantly higher in patients with MB (2.45 ± 1.19 vs. 1.72 ± 0.48; p 1.82 predicted myocardial bridge presence with 70% sensitivity and 71% specificity (ROC area under curve: 0.733, 95% CI: 0.654-0.811, p < 0.001. Conclusion: Our study findings demonstrated that MB is associated with elevated NLR, which is used to assess inflammatory status of the body. J Clin Exp Invest 2014; 5 (1: 24-28

  16. 18F-fluoro-2-deoxyglucose PET informs neutrophil accumulation and activation in lipopolysaccharide-induced acute lung injury genetic algorithm

    International Nuclear Information System (INIS)

    Rodrigues, Rosana S.; Bozza, Fernando A.; Hanrahan, Christopher J.; Wang, Li-Ming; Wu, Qi; Hoffman, John M.; Zimmerman, Guy A.; Morton, Kathryn A.

    2017-01-01

    Introduction: Molecular imaging of the earliest events related to the development of acute lung injury (ALI)/acute respiratory distress syndrome (ARDS) could facilitate therapeutic development and patient management. We previously reported that 18 F-fluoro-2-deoxyglucose ( 18 F-FDG) PET identifies ALI/ARDS prior to radiographic abnormalities. The purpose of this study was to establish the time courses of 18 F-FDG uptake, edema and neutrophil recruitment in an endotoxin-induced acute lung injury model and to examine molecular events required for 14 C-2DG uptake in activated neutrophils. Methods: Lung uptake of 18 F-FDG was measured by PET in control male Sprague Dawley rats and at 2, 6 and 24 h following the intraperitoneal injection of 10 mg/kg LPS. Lung edema (attenuation) was measured by microCT. Neutrophil influx into the lungs was measured by myeloperoxidase assay. Control and activated human donor neutrophils were compared for uptake of 14 C-2DG, transcription and content of hexokinase and GLUT isoforms and for hexokinase (HK) activity. Results: Significant uptake of 18 F-FDG occurred by 2 h following LPS, and progressively increased to 24 h. Lung uptake of 18 F-FDG preceded increased CT attenuation (lung edema). Myeloperoxidase activity in the lungs, supporting neutrophil influx, paralleled 18 F-FDG uptake. Activation of isolated human neutrophils resulted in increased uptake of 14 C-2DG, expression of GLUT 3 and GLUT 4 and expression and increased HK1 activity. Conclusion: Systemic endotoxin-induced ALI results in very early and progressive uptake of 18 F-FDG, parallels neutrophil accumulation and occurs earlier than lung injury edema. Activated neutrophils show increased uptake of 14 C-2DG, expression of specific GLUT3, GLUT4 and HK1 protein and HK activity. Advances in knowledge and implications for patient care: 18 F-FDG pulmonary uptake is an early biomarker of neutrophil recruitment in ALI and is associated with specific molecular events that mediate 14

  17. CXC-chemokine regulation and neutrophil trafficking in hepatic ischemia-reperfusion injury in P-selectin/ICAM-1 deficient mice

    Directory of Open Access Journals (Sweden)

    Crockett Elahé T

    2007-05-01

    Full Text Available Abstract Background Neutrophil adhesion and migration are critical in hepatic ischemia and reperfusion injury (I/R. P-selectin and the intercellular adhesion molecule (ICAM-1 can mediate neutrophil-endothelial cell interactions, neutrophil migration, and the interactions of neutrophils with hepatocytes in the liver. Despite very strong preclinical data, recent clinical trials failed to show a protective effect of anti-adhesion therapy in reperfusion injury, indicating that the length of injury might be a critical factor in neutrophil infiltration. Therefore, the aim of this study was to assess the role of P-selectin and ICAM-1 in neutrophil infiltration and liver injury during early and late phases of liver I/R. Methods Adult male wild-type and P-selectin/ICAM-1-deficient (P/I null mice underwent 90 minutes of partial liver ischemia followed by various periods of reperfusion (6, 15 h, and a survival study. Liver injury was assessed by plasma level of alanine aminotransferase (ALT and histopathology. The plasma cytokines, TNF-α, IL-6, MIP-2 and KC, were measured by ELISA. Results Reperfusion caused significant hepatocellular injury in both wild-type and P/I null mice as was determined by plasma ALT levels and liver histopathology. The injury was associated with a marked neutrophil infiltration into the ischemic livers of both wild-type and P/I null mice. Although the levels of ALT and neutrophil infiltration were slightly lower in the P/I null mice compared with the wild-type mice the differences were not statistically significant. The plasma cytokine data of TNF-α and IL-6 followed a similar pattern to ALT data, and no significant difference was found between the wild-type and P/I null groups. In contrast, a significant difference in KC and MIP-2 chemokine levels was observed between the wild-type and P/I null mice. Additionally, the survival study showed a trend towards increased survival in the P/I null group. Conclusion While ICAM-1 and P

  18. Soluble CD40 ligand stimulates CD40-dependent activation of the β2 integrin Mac-1 and protein kinase C zeda (PKCζ in neutrophils: implications for neutrophil-platelet interactions and neutrophil oxidative burst.

    Directory of Open Access Journals (Sweden)

    Rong Jin

    Full Text Available Recent work has revealed an essential involvement of soluble CD40L (sCD40L in inflammation and vascular disease. Activated platelets are the major source of sCD40L, which has been implicated in platelet and leukocyte activation, although its exact functional impact on leukocyte-platelet interactions and the underlying mechanisms remain undefined. We aimed to determine the impact and the mechanisms of sCD40L on neutrophils. We studied neutrophil interactions with activated, surface-adherent platelets as a model for leukocyte recruitment to the sites of injury. Our data show that CD40L contributes to neutrophil firm adhesion to and transmigration across activated surface-adherent platelets, possibly through two potential mechanisms. One involves the direct interaction of ligand-receptor (CD40L-CD40, i.e., platelet surface CD40L interaction with neutrophil CD40; another involves an indirect mechanism, i.e. soluble CD40L stimulates activation of the leukocyte-specific β2 integrin Mac-1 in neutrophils and thereby further promotes neutrophil adhesion and migration. Activation of the integrin Mac-1 is known to be critical for mediating neutrophil adhesion and migration. sCD40L activated Mac-1 in neutrophils and enhanced neutrophil-platelet interactions in wild-type neutrophils, but failed to elicit such responses in CD40-deficient neutrophils. Furthermore, our data show that the protein kinase C zeta (PKCζ is critically required for sCD40L-induced Mac-1 activation and neutrophil adhesive function. sCD40L strongly stimulated the focal clustering of Mac-1 (CD11b and the colocalization of Mac-1 with PKCζ in wild-type neutrophils, but had minimal effect in CD40-deficient neutrophils. Blocking PKCζ completely inhibited sCD40L-induced neutrophil firm adhesion. Moreover, sCD40L strongly stimulates neutrophil oxidative burst via CD40-dependent activation of PI3K/NF-KB, but independent of Mac-1 and PKCζ. These findings may contribute to a better

  19. File list: His.Bld.10.AllAg.Neutrophils [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  20. File list: His.Bld.50.AllAg.Neutrophils [Chip-atlas[Archive

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  2. File list: His.Bld.20.AllAg.Neutrophils [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  5. File list: ALL.Bld.20.AllAg.Neutrophils [Chip-atlas[Archive

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  10. File list: His.Bld.05.AllAg.Neutrophils [Chip-atlas[Archive

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  11. File list: ALL.Bld.50.AllAg.Neutrophils [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  12. Neutrophil CD64 has a high negative predictive value for exclusion ...

    African Journals Online (AJOL)

    [13] Multiple studies agree that neutrophil CD64 has high diagnostic speci city and sensitivity.[13,18-20] In the light of the above information, we prospectively evaluated the usefulness of neutrophil CD64 expression in diagnosing neonatal infection. e main objective of the study was quantitation of neutrophil. CD64 by ow ...

  13. Evidence for a self-enforcing inflammation in neutrophil-mediated chronic diseases

    NARCIS (Netherlands)

    Overbeek, S.A.

    2011-01-01

    In summary, this thesis provides evidence for the self-sustaining role of neutrophils in the inflammatory state in the pathogenesis of COPD and CD. In active disease, neutrophils release proteolytic enzymes that breakdown collagen. One of the collagen fragments can be neutrophilic chemoattractant

  14. File list: His.Bld.20.AllAg.Neutrophils [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  15. Neutrophil extracellular traps in the host defense against sepsis induced by Burkholderia pseudomallei (melioidosis)

    NARCIS (Netherlands)

    de Jong, Hanna K.; Koh, Gavin C. K. W.; Achouiti, Ahmed; van der Meer, Anne J.; Bulder, Ingrid; Stephan, Femke; Roelofs, Joris J. T. H.; Day, Nick P. J.; Peacock, Sharon J.; Zeerleder, Sacha; Wiersinga, W. Joost

    2014-01-01

    Neutrophil extracellular traps (NETs) are a central player in the host response to bacteria: neutrophils release extracellular DNA (nucleosomes) and neutrophil elastase to entrap and kill bacteria. We studied the role of NETs in Burkholderia pseudomallei infection (melioidosis), an important cause

  16. Nicotinamide exacerbates hypoxemia in ventilator-induced lung injury independent of neutrophil infiltration.

    Directory of Open Access Journals (Sweden)

    Heather D Jones

    Full Text Available Ventilator-induced lung injury is a form of acute lung injury that develops in critically ill patients on mechanical ventilation and has a high degree of mortality. Nicotinamide phosphoribosyltransferase is an enzyme that is highly upregulated in ventilator-induced lung injury and exacerbates the injury when given exogenously. Nicotinamide (vitamin B3 directly inhibits downstream pathways activated by Nicotinamide phosphoribosyltransferase and is protective in other models of acute lung injury.We administered nicotinamide i.p. to mice undergoing mechanical ventilation with high tidal volumes to study the effects of nicotinamide on ventilator-induced lung injury. Measures of injury included oxygen saturations and bronchoalveolar lavage neutrophil counts, protein, and cytokine levels. We also measured expression of nicotinamide phosophoribosyltransferase, and its downstream effectors Sirt1 and Cebpa, Cebpb, Cebpe. We assessed the effect of nicotinamide on the production of nitric oxide during ventilator-induced lung injury. We also studied the effects of ventilator-induced lung injury in mice deficient in C/EBPε.Nicotinamide treatment significantly inhibited neutrophil infiltration into the lungs during ventilator-induced lung injury, but did not affect protein leakage or cytokine production. Surprisingly, mice treated with nicotinamide developed significantly worse hypoxemia during mechanical ventilation. This effect was not linked to increases in nitric oxide production or alterations in expression of Nicotinamide phosphoribosyl transferase, Sirt1, or Cebpa and Cebpb. Cebpe mRNA levels were decreased with either nicotinamide treatment or mechanical ventilation, but mice lacking C/EBPε developed the same degree of hypoxemia and ventilator-induced lung injury as wild-type mice.Nicotinamide treatment during VILI inhibits neutrophil infiltration of the lungs consistent with a strong anti-inflammatory effect, but paradoxically also leads to the

  17. Enhanced basophil histamine release and neutrophil chemotactic activity predispose grain dust-induced airway obstruction.

    Science.gov (United States)

    Park, H; Jung, K; Kang, K; Nahm, D; Cho, S; Kim, Y

    1999-04-01

    The pathogenic mechanism of grain dust (GD)-induced occupational asthma (OA) remains unclear. To understand further the mechanism of GD-induced OA. Fifteen employees working in a same GD industry, complaining of work-related respiratory symptoms, were enrolled and were divided into two groups according to the GD-bronchoprovocation test (BPT) result: six positive responders were grouped as group III, nine negative responders as group II and five healthy controls as group I. Serum GD-specific immunoglobulin (Ig)E (sIgE), specific IgG (sIgG) and specific IgG4 (sIgG4) antibodies were detected by enzyme-linked immunosorbent assay. Basophil histamine release was measured by the autofluorometric method, and changes of serum neutrophil chemotactic activity were observed by the Boyden chamber method. For clinical parameters such as degree of airway hyperresponsiveness to methacholine, duration of respiratory symptoms, exposure duration, and prevalences of serum sIgE, sIgG and sIgG4 antibodies, there were no significant differences between group II and III (P > 0.05, respectively). Serum neutrophil chemotactic activity increased significantly at 30 min and decreased at 240 min after the GD-BPT in group III subjects (P 0.05). Basophil histamine release induced by GD was significantly higher in group III than those of group I or group II (P < 0.05, respectively), while minimal release of anti-IgG4 antibodies was noted in all three groups. These results suggest that enhanced basophil histamine release and serum neutrophil chemotactic activity might contribute to the development of GD-induced occupational asthma.

  18. The Beta-2-Adrenoreceptor Agonists, Formoterol and Indacaterol, but Not Salbutamol, Effectively Suppress the Reactivity of Human Neutrophils In Vitro

    Directory of Open Access Journals (Sweden)

    Ronald Anderson

    2014-01-01

    Full Text Available The clinical relevance of the anti-inflammatory properties of beta-2 agonists remains contentious possibly due to differences in their molecular structures and agonist activities. The current study has compared the effects of 3 different categories of β2-agonists, namely, salbutamol (short-acting, formoterol (long-acting and indacaterol (ultra-long-acting, at concentrations of 1–1000 nM, with human blood neutrophils in vitro. Neutrophils were activated with either N-formyl-L-methionyl-L-leucyl-L-phenylalanine (fMLP, 1 µM or platelet-activating factor (PAF, 200 nM in the absence and presence of the β2-agonists followed by measurement of the generation of reactive oxygen species and leukotriene B4, release of elastase, and expression of the β2-integrin, CR3, using a combination of chemiluminescence, ELISA, colorimetric, and flow cytometric procedures respectively. These were correlated with alterations in the concentrations of intracellular cyclic-AMP and cytosolic Ca2+. At the concentrations tested, formoterol and indacaterol caused equivalent, significant (P<0.05 at 1–10 nM dose-related inhibition of all of the pro-inflammatory activities tested, while salbutamol was much less effective (P<0.05 at 100 nM and higher. Suppression of neutrophil reactivity was accompanied by elevations in intracellular cAMP and accelerated clearance of Ca2+ from the cytosol of activated neutrophils. These findings demonstrate that β2-agonists vary with respect to their suppressive effects on activated neutrophils.

  19. Selected mucolytic, anti-inflammatory and cardiovascular drugs change the ability of neutrophils to form extracellular traps (NETs).

    Science.gov (United States)

    Zawrotniak, Marcin; Kozik, Andrzej; Rapala-Kozik, Maria

    2015-01-01

    Neutrophils form the first line of host defense against infections that combat pathogens using two major mechanisms, the phagocytosis or the release of neutrophil extracellular traps (NETs). The netosis (NET formation) exerts additional, unfavorable effects on the fitness of host cells and is also involved at the sites of lung infection, increasing the mucus viscosity and in the circulatory system where it can influence the intravascular clot formation. Although molecular mechanisms underlying the netosis are still incompletely understood, a role of NADPH oxidase that activates the production of reactive oxygen species (ROS) during the initiation of NETs has been well documented. Since several commonly used drugs can affects the netosis, our current study was aimed to determine the effects of selected mucolytic, anti-inflammatory and cardiovascular drugs on NET formation, with a special emphasis on ROS production and NADPH oxidase activity. The treatment of neutrophils with N-acetylcysteine, ketoprofen and ethamsylate reduced the production of ROS by these cells in a dose-dependent manner. NET formation was also modulated by selected drugs. N-acetylcysteine inhibited the netosis but in the presence of H2O2 this neutrophil ability was restored, indicating that N-acetylcysteine may influence the NET formation by modulating ROS productivity. The administration of ethamsylate led to a significant reduction in NET formation and this effect was not restored by H2O2 or S. aureus, suggesting the unexpected additional side effects of this drug. Ketoprofen seemed to promote ROS-independent NET release, simultaneously inhibiting ROS production. The results, obtained in this study strongly suggest that the therapeutic strategies applied in many neutrophil-mediated diseases should take into account the NET-associated effects.

  20. On the pharmacology of oxidative burst of neutrophils

    Czech Academy of Sciences Publication Activity Database

    Nosáľ, R.; Drábiková, K.; Harmatha, Juraj; Jančinová, V.; Mačičková, T.; Pečivová, J.; Perečko, T.

    2011-01-01

    Roč. 4, č. 2 (2011), A51-A51 ISSN 1337-6853. [TOXCON 2011. Interdisciplinary Toxicology Conference /16./. 17.05.2011-20.05.2011, Praha] Institutional research plan: CEZ:AV0Z40550506 Keywords : N-feruloyl- serotonin * oxidative burst * inhibition of neutrophil activation Subject RIV: CC - Organic Chemistry

  1. Flavonoids inhibit the respiratory burst of neutrophils in mammals

    Czech Academy of Sciences Publication Activity Database

    Číž, Milan; Denev, P.; Kratchanova, M.; Vašíček, Ondřej; Ambrožová, Gabriela; Lojek, Antonín

    2012-01-01

    Roč. 2012, č. 2012 (2012), ID 181295 ISSN 1942-0900 R&D Projects: GA ČR(CZ) GA524/08/1753 Institutional research plan: CEZ:AV0Z50040702 Keywords : flavonoids * neutrophils * respiratory burst Subject RIV: BO - Biophysics Impact factor: 3.393, year: 2012

  2. Activity of neutrophil elastase reflects the progression of acute pancreatitis

    DEFF Research Database (Denmark)

    Novovic, Srdan; Andersen, Anders M; Nord, Magnus

    2013-01-01

    Abstract Objective. Neutrophil elastase (NE) concentration is associated with progression of acute pancreatitis (AP), but measuring total NE concentration includes biologically inactive NE. This study aims to investigate the relationship between NE activity and the aetiology and severity of AP...... was associated with predicted severity of AP and AP-associated respiratory failure. Specific NE inhibitors may have therapeutic potential in acute pancreatitis....

  3. Neutrophil superoxide-anion generating capacity in chronic smoking ...

    Indian Academy of Sciences (India)

    We investigated whether long-term -tocopherol therapy in chronic smoking affects superoxide generating capacity of neutrophils ex vivo. To this purpose, we randomly assigned 128 male chronic smokers (37 ± 21 pack years of smoking) to treatment with placebo ( = 64) or -tocopherol (400 IU dL--tocopherol daily, ...

  4. Platelet indices and neutrophil to lymphocyte ratio in adults with ...

    African Journals Online (AJOL)

    Background: A study was performed in adults with acute appendicitis and matched controls to assess the utility of the platelet indices and neutrophil to lymphocyte ratio, as a diagnostic adjunct. Methods: Data were retrospectively collected from a complete blood count test of 155 adult patients (72 men and 83 women) with ...

  5. Enhancement by platelets of oxygen radical responses of human neutrophils

    Energy Technology Data Exchange (ETDEWEB)

    McCulloch, K.K.; Powell, J.; Johnson, K.J.; Ward, P.A.

    1986-03-01

    When human blood neutrophils were incubated with immune complexes (consisting of IgG antibody) in the presence of platelets, there was a 2 to 10 fold enhancement in the generation of O-/sub 2/ and H/sub 2/O/sub 2/. This enhancement phenomenon was proportional to the dose of immune complex added and the number of platelets present. The response was not agonist specific since similar enhancement also occurred with the following agonists: phorbol myristate acetate, opsonized zymosan particles and the chemotactic peptide N-formyl-met-leu-phe. The platelet related phenomenon of enhanced O-/sub 2/ generation could not be reproduced by the addition of serotonin, histamine or platelet-derived growth factor and was not affected by prior treatment of platelets with cyclooxygenase inhibitors (indomethacin, piroxicam) or lipoxygenase inhibitors (nafazatrom, BW755C or nordihydroguaiaretic acid). However, activation of platelets by thrombin caused release into the platelet supernatant fluid of a factor that, only in the presence of immune complexes, caused enhanced O-/sub 2/ responses to neutrophils. These data indicate that platelets potentiate oxygen radical responses of human neutrophils and suggest a mechanisms by which platelets may participate in tissue injury which is mediated by oxygen radical products from activated neutrophils.

  6. Yersinia pestis targets neutrophils via complement receptor 3

    Science.gov (United States)

    Merritt, Peter M.; Nero, Thomas; Bohman, Lesley; Felek, Suleyman; Krukonis, Eric S.; Marketon, Melanie M.

    2015-01-01

    Yersinia species display a tropism for lymphoid tissues during infection, and the bacteria select innate immune cells for delivery of cytotoxic effectors by the type III secretion system. Yet the mechanism for target cell selection remains a mystery. Here we investigate the interaction of Yersinia pestis with murine splenocytes to identify factors that participate in the targeting process. We find that interactions with primary immune cells rely on multiple factors. First, the bacterial adhesin Ail is required for efficient targeting of neutrophils in vivo. However, Ail does not appear to directly mediate binding to a specific cell type. Instead, we find that host serum factors direct Y. pestis to specific innate immune cells, particularly neutrophils. Importantly, specificity towards neutrophils was increased in the absence of bacterial adhesins due to reduced targeting of other cell types, but this phenotype was only visible in the presence of mouse serum. Addition of antibodies against complement receptor 3 and CD14 blocked target cell selection, suggesting that a combination of host factors participate in steering bacteria toward neutrophils during plague infection. PMID:25359083

  7. Circulating neutrophil transcriptome may reveal intracranial aneurysm signature.

    Directory of Open Access Journals (Sweden)

    Vincent M Tutino

    Full Text Available Unruptured intracranial aneurysms (IAs are typically asymptomatic and undetected except for incidental discovery on imaging. Blood-based diagnostic biomarkers could lead to improvements in IA management. This exploratory study examined circulating neutrophils to determine whether they carry RNA expression signatures of IAs.Blood samples were collected from patients receiving cerebral angiography. Eleven samples were collected from patients with IAs and 11 from patients without IAs as controls. Samples from the two groups were paired based on demographics and comorbidities. RNA was extracted from isolated neutrophils and subjected to next-generation RNA sequencing to obtain differential expressions for identification of an IA-associated signature. Bioinformatics analyses, including gene set enrichment analysis and Ingenuity Pathway Analysis, were used to investigate the biological function of all differentially expressed transcripts.Transcriptome profiling identified 258 differentially expressed transcripts in patients with and without IAs. Expression differences were consistent with peripheral neutrophil activation. An IA-associated RNA expression signature was identified in 82 transcripts (p<0.05, fold-change ≥2. This signature was able to separate patients with and without IAs on hierarchical clustering. Furthermore, in an independent, unpaired, replication cohort of patients with IAs (n = 5 and controls (n = 5, the 82 transcripts separated 9 of 10 patients into their respective groups.Preliminary findings show that RNA expression from circulating neutrophils carries an IA-associated signature. These findings highlight a potential to use predictive biomarkers from peripheral blood samples to identify patients with IAs.

  8. Enhancement by platelets of oxygen radical responses of human neutrophils

    International Nuclear Information System (INIS)

    McCulloch, K.K.; Powell, J.; Johnson, K.J.; Ward, P.A.

    1986-01-01

    When human blood neutrophils were incubated with immune complexes (consisting of IgG antibody) in the presence of platelets, there was a 2 to 10 fold enhancement in the generation of O- 2 and H 2 O 2 . This enhancement phenomenon was proportional to the dose of immune complex added and the number of platelets present. The response was not agonist specific since similar enhancement also occurred with the following agonists: phorbol myristate acetate, opsonized zymosan particles and the chemotactic peptide N-formyl-met-leu-phe. The platelet related phenomenon of enhanced O- 2 generation could not be reproduced by the addition of serotonin, histamine or platelet-derived growth factor and was not affected by prior treatment of platelets with cyclooxygenase inhibitors (indomethacin, piroxicam) or lipoxygenase inhibitors (nafazatrom, BW755C or nordihydroguaiaretic acid). However, activation of platelets by thrombin caused release into the platelet supernatant fluid of a factor that, only in the presence of immune complexes, caused enhanced O- 2 responses to neutrophils. These data indicate that platelets potentiate oxygen radical responses of human neutrophils and suggest a mechanisms by which platelets may participate in tissue injury which is mediated by oxygen radical products from activated neutrophils

  9. Modulation of neutrophil oxidative burst via histamine receptors

    Czech Academy of Sciences Publication Activity Database

    Číž, Milan; Lojek, Antonín

    2013-01-01

    Roč. 170, č. 1 (2013), s. 17-22 ISSN 0007-1188 R&D Projects: GA MŠk(CZ) LD11010 Institutional support: RVO:68081707 Keywords : neutrophil * oxidative burst * reactive oxygen species Subject RIV: BO - Biophysics Impact factor: 4.990, year: 2013

  10. On the Pharmacology of Oxidative Burst of Human Neutrophils

    Czech Academy of Sciences Publication Activity Database

    Nosáľ, R.; Drábiková, K.; Jančinová, V.; Mačičková, T.; Pečivová, J.; Perečko, T.; Harmatha, Juraj; Šmidrkal, J.

    2015-01-01

    Roč. 64, Suppl 4 (2015), S445-S452 ISSN 0862-8408 Institutional support: RVO:61388963 Keywords : human neutrophils * oxidative burst * chemiluminescence * protein kinase C * apoptosis Subject RIV: FR - Pharmacology ; Medidal Chemistry Impact factor: 1.643, year: 2015 http://www.biomed.cas.cz/physiolres/pdf/64/64_S445.pdf

  11. ADAM9 Is a Novel Product of Polymorphonuclear Neutrophils

    DEFF Research Database (Denmark)

    Roychaudhuri, Robin; Hergrueter, Anja H; Polverino, Francesca

    2014-01-01

    A disintegrin and a metalloproteinase domain (ADAM) 9 is known to be expressed by monocytes and macrophages. In this study, we report that ADAM9 is also a product of human and murine polymorphonuclear neutrophils (PMNs). ADAM9 is not synthesized de novo by circulating PMNs. Rather, ADAM9 protein...

  12. Nucleases from Prevotella intermedia can degrade neutrophil extracellular traps.

    Science.gov (United States)

    Doke, M; Fukamachi, H; Morisaki, H; Arimoto, T; Kataoka, H; Kuwata, H

    2017-08-01

    Periodontitis is an inflammatory disease caused by periodontal bacteria in subgingival plaque. These bacteria are able to colonize the periodontal region by evading the host immune response. Neutrophils, the host's first line of defense against infection, use various strategies to kill invading pathogens, including neutrophil extracellular traps (NETs). These are extracellular net-like fibers comprising DNA and antimicrobial components such as histones, LL-37, defensins, myeloperoxidase, and neutrophil elastase from neutrophils that disarm and kill bacteria extracellularly. Bacterial nuclease degrades the NETs to escape NET killing. It has now been shown that extracellular nucleases enable bacteria to evade this host antimicrobial mechanism, leading to increased pathogenicity. Here, we compared the DNA degradation activity of major Gram-negative periodontopathogenic bacteria, Porphyromonas gingivalis, Prevotella intermedia, Fusobacterium nucleatum, and Aggregatibacter actinomycetemcomitans. We found that Pr. intermedia showed the highest DNA degradation activity. A genome search of Pr. intermedia revealed the presence of two genes, nucA and nucD, putatively encoding secreted nucleases, although their enzymatic and biological activities are unknown. We cloned nucA- and nucD-encoding nucleases from Pr. intermedia ATCC 25611 and characterized their gene products. Recombinant NucA and NucD digested DNA and RNA, which required both Mg 2+ and Ca 2+ for optimal activity. In addition, NucA and NucD were able to degrade the DNA matrix comprising NETs. © 2016 The Authors Molecular Oral Microbiology Published by John Wiley & Sons Ltd.

  13. Neutrophil elastase-mediated increase in airway temperature during inflammation

    DEFF Research Database (Denmark)

    Schmidt, Annika; Belaaouaj, Azzaq; Bissinger, Rosi

    2014-01-01

    in the exhaled air of cystic fibrosis (CF) patients. To further test our hypothesis, a pouch inflammatory model using neutrophil elastase-deficient mice was employed. Next, the impact of temperature changes on the dominant CF pathogen Pseudomonas aeruginosa growth was tested by plating method and RNAseq. Results...

  14. Neutrophil extracellular traps in patients with pulmonary tuberculosis

    NARCIS (Netherlands)

    van der Meer, Anne Jan; Zeerleder, Sacha; Blok, Dana C.; Kager, Liesbeth M.; Lede, Ivar O.; Rahman, Wahid; Afroz, Rumana; Ghose, Aniruddha; Visser, Caroline E.; Zahed, Abu Shahed Md; Husain, Md Anwar; Alam, Khan Mashrequl; Barua, Pravat Chandra; Hassan, Mahtabuddin; Tayab, Md Abu; Dondorp, Arjen M.; van der Poll, Tom

    2017-01-01

    Tuberculosis is a devastating infectious disease causing many deaths worldwide. Recent investigations have implicated neutrophil extracellular traps (NETs) in the host response to tuberculosis. The aim of the current study was to obtain evidence for NETs release in the circulation during human

  15. Increased neutrophil expression of pattern recognition receptors during COPD exacerbations

    NARCIS (Netherlands)

    Pouwels, Simon D.; Van Geffen, Wouter H.; Jonker, Marnix R.; Kerstjens, Huib A. M.; Nawijn, Martijn C.; Heijink, Irene H.

    Previously, we observed increased serum levels of damage-associated molecular patterns (DAMPs) during COPD exacerbations. Here, gene expression of DAMP receptors was measured in peripheral blood neutrophils of COPD patients during stable disease and severe acute exacerbation. The expression of

  16. The strength of integrin binding between neutrophils and endothelial cells

    Czech Academy of Sciences Publication Activity Database

    Labrador, V.; Říha, Pavel; Muller, S.; Dumas, D.; Wang, X.; Stoltz, J. F.

    2003-01-01

    Roč. 32, č. 8 (2003), s. 684-688 ISSN 0175-7571 R&D Projects: GA ČR GA305/01/1605 Institutional research plan: CEZ:AV0Z2060917 Keywords : endothelium * integrins * neutrophil adhesion * scanning microscopy Subject RIV: BO - Biophysics Impact factor: 1.769, year: 2003

  17. Pharmacological intervention with oxidative burst in human neutrophils

    Czech Academy of Sciences Publication Activity Database

    Nosál, R.; Drábiková, K.; Jančinová, V.; Mačičková, T.; Pečivová, J.; Perečko, T.; Harmatha, Juraj

    2017-01-01

    Roč. 10, č. 2 (2017), s. 56-60 ISSN 1337-6853 Institutional support: RVO:61388963 Keywords : human neutrophils * oxidative burst * tharapeutical drugs * natural antioxidants Subject RIV: FR - Pharmacology ; Medidal Chemistry OBOR OECD: Pharmacology and pharmacy https://www.degruyter.com/downloadpdf/j/intox.2017.10.issue-2/intox-2017-0009/intox-2017-0009.pdf

  18. Circulating neutrophil transcriptome may reveal intracranial aneurysm signature

    Science.gov (United States)

    Tutino, Vincent M.; Poppenberg, Kerry E.; Jiang, Kaiyu; Jarvis, James N.; Sun, Yijun; Sonig, Ashish; Siddiqui, Adnan H.; Snyder, Kenneth V.; Levy, Elad I.; Kolega, John

    2018-01-01

    Background Unruptured intracranial aneurysms (IAs) are typically asymptomatic and undetected except for incidental discovery on imaging. Blood-based diagnostic biomarkers could lead to improvements in IA management. This exploratory study examined circulating neutrophils to determine whether they carry RNA expression signatures of IAs. Methods Blood samples were collected from patients receiving cerebral angiography. Eleven samples were collected from patients with IAs and 11 from patients without IAs as controls. Samples from the two groups were paired based on demographics and comorbidities. RNA was extracted from isolated neutrophils and subjected to next-generation RNA sequencing to obtain differential expressions for identification of an IA-associated signature. Bioinformatics analyses, including gene set enrichment analysis and Ingenuity Pathway Analysis, were used to investigate the biological function of all differentially expressed transcripts. Results Transcriptome profiling identified 258 differentially expressed transcripts in patients with and without IAs. Expression differences were consistent with peripheral neutrophil activation. An IA-associated RNA expression signature was identified in 82 transcripts (pIAs on hierarchical clustering. Furthermore, in an independent, unpaired, replication cohort of patients with IAs (n = 5) and controls (n = 5), the 82 transcripts separated 9 of 10 patients into their respective groups. Conclusion Preliminary findings show that RNA expression from circulating neutrophils carries an IA-associated signature. These findings highlight a potential to use predictive biomarkers from peripheral blood samples to identify patients with IAs. PMID:29342213

  19. Changes in neutrophil count, creatine kinases and muscle soreness ...

    African Journals Online (AJOL)

    Objective. A primary objective was to examine circulating neutrophil count after repeated bouts of downhill running. An additional aim was to determine creatine kinase (CK) levels during the initial 12 hours, after repeated DHRs. Design. Eleven healthy, untrained Caucasian males performed 2 x 60 min bouts of DHR ...

  20. Activated human neutrophils release hepatocyte growth factor/scatter factor.

    LENUS (Irish Health Repository)

    McCourt, M

    2012-02-03

    BACKGROUND: Hepatocyte growth factor or scatter factor (HGF\\/SF) is a pleiotropic cytokine that has potent angiogenic properties. We have previously demonstrated that neutrophils (PMN) are directly angiogenic by releasing vascular endothelial growth factor (VEGF). We hypothesized that the acute inflammatory response can stimulate PMN to release HGF. AIMS: To examine the effects of inflammatory mediators on PMN HGF release and the effect of recombinant human HGF (rhHGF) on PMN adhesion receptor expression and PMN VEGF release. METHODS: In the first experiment, PMN were isolated from healthy volunteers and stimulated with tumour necrosis factor-alpha (TNF-alpha), lipopolysaccharide (LPS), interleukin-8 (IL-8), and formyl methionyl-leucyl-phenylalanine (fMLP). Culture supernatants were assayed for HGF using ELISA. In the second experiment, PMN were lysed to measure total HGF release and HGF expression in the PMN was detected by Western immunoblotting. Finally, PMN were stimulated with rhHGF. PMN CD 11a, CD 11b, and CD 18 receptor expression and VEGF release was measured using flow cytometry and ELISA respectively. RESULTS: TNF-alpha, LPS and fMLP stimulation resulted in significantly increased release of PMN HGF (755+\\/-216, 484+\\/-221 and 565+\\/-278 pg\\/ml, respectively) compared to controls (118+\\/-42 pg\\/ml). IL-8 had no effect. Total HGF release following cell lysis and Western blot suggests that HGF is released from intracellular stores. Recombinant human HGF did not alter PMN adhesion receptor expression and had no effect on PMN VEGF release. CONCLUSIONS: This study demonstrates that pro-inflammatory mediators can stimulate HGF release from a PMN intracellular store and that activated PMN in addition to secreting VEGF have further angiogenic potential by releasing HGF.

  1. Influence of neutrophil defects on Burkholderia cepacia complex pathogenesis

    Directory of Open Access Journals (Sweden)

    Laura A. Porter

    2011-11-01

    Full Text Available The Burkholderia cepacia complex (Bcc is a group of Gram-negative bacteria that are ubiquitous in the environment and have emerged as opportunistic pathogens in immunocompromised patients. The primary patient populations infected with Bcc include individuals with cystic fibrosis (CF, as well as those with chronic granulomatous disease (CGD. While Bcc infection in CF is better characterized than in CGD, these two genetic diseases are not obviously similar and it is currently unknown if there is any commonality in host immune defects that is responsible for the susceptibility to Bcc. CF is caused by mutations in the CF transmembrane conductance regulator, resulting in manifestations in various organ systems, however the major cause of morbidity and mortality is currently due to bacterial respiratory infections. CGD, on the other hand, is a genetic disorder that is caused by defects in phagocyte NADPH oxidase. Because of the defect in CGD, phagocytes in these patients are unable to produce reactive oxygen species, which results in increased susceptibility to bacterial and fungal infections. Despite this significant defect in microbial clearance, the spectrum of pathogens frequently implicated in infections in CGD is relatively narrow and includes some bacterial species that are considered almost pathognomonic for this disorder. Very little is known about the cause of the specific susceptibility to Bcc over other potential pathogens more prevalent in the environment, and a better understanding of specific mechanisms required for bacterial virulence has become a high priority. This review will summarize both the current knowledge and future directions related to Bcc virulence in immunocompromised individuals with a focus on the roles of bacterial factors and neutrophil defects in pathogenesis.

  2. Ensemble models of neutrophil trafficking in severe sepsis.

    Directory of Open Access Journals (Sweden)

    Sang Ok Song

    Full Text Available A hallmark of severe sepsis is systemic inflammation which activates leukocytes and can result in their misdirection. This leads to both impaired migration to the locus of infection and increased infiltration into healthy tissues. In order to better understand the pathophysiologic mechanisms involved, we developed a coarse-grained phenomenological model of the acute inflammatory response in CLP (cecal ligation and puncture-induced sepsis in rats. This model incorporates distinct neutrophil kinetic responses to the inflammatory stimulus and the dynamic interactions between components of a compartmentalized inflammatory response. Ensembles of model parameter sets consistent with experimental observations were statistically generated using a Markov-Chain Monte Carlo sampling. Prediction uncertainty in the model states was quantified over the resulting ensemble parameter sets. Forward simulation of the parameter ensembles successfully captured experimental features and predicted that systemically activated circulating neutrophils display impaired migration to the tissue and neutrophil sequestration in the lung, consequently contributing to tissue damage and mortality. Principal component and multiple regression analyses of the parameter ensembles estimated from survivor and non-survivor cohorts provide insight into pathologic mechanisms dictating outcome in sepsis. Furthermore, the model was extended to incorporate hypothetical mechanisms by which immune modulation using extracorporeal blood purification results in improved outcome in septic rats. Simulations identified a sub-population (about 18% of the treated population that benefited from blood purification. Survivors displayed enhanced neutrophil migration to tissue and reduced sequestration of lung neutrophils, contributing to improved outcome. The model ensemble presented herein provides a platform for generating and testing hypotheses in silico, as well as motivating further experimental

  3. d(− Lactic Acid-Induced Adhesion of Bovine Neutrophils onto Endothelial Cells Is Dependent on Neutrophils Extracellular Traps Formation and CD11b Expression

    Directory of Open Access Journals (Sweden)

    Pablo Alarcón

    2017-08-01

    Full Text Available Bovine ruminal acidosis is of economic importance as it contributes to reduced milk and meat production. This phenomenon is mainly attributed to an overload of highly fermentable carbohydrate, resulting in increased d(− lactic acid levels in serum and plasma. Ruminal acidosis correlates with elevated acute phase proteins in blood, along with neutrophil activation and infiltration into various tissues leading to laminitis and aseptic polysynovitis. Previous studies in bovine neutrophils indicated that d(− lactic acid decreased expression of L-selectin and increased expression of CD11b to concentrations higher than 6 mM, suggesting a potential role in neutrophil adhesion onto endothelia. The two aims of this study were to evaluate whether d(− lactic acid influenced neutrophil and endothelial adhesion and to trigger neutrophil extracellular trap (NET production (NETosis in exposed neutrophils. Exposure of bovine neutrophils to 5 mM d(− lactic acid elevated NET release compared to unstimulated neutrophil negative controls. Moreover, this NET contains CD11b and histone H4 citrullinated, the latter was dependent on PAD4 activation, a critical enzyme in DNA decondensation and NETosis. Furthermore, NET formation was dependent on d(− lactic acid plasma membrane transport through monocarboxylate transporter 1 (MCT1. d(− lactic acid enhanced neutrophil adhesion onto endothelial sheets as demonstrated by in vitro neutrophil adhesion assays under continuous physiological flow conditions, indicating that cell adhesion was a NET- and a CD11b/ICAM-1-dependent process. Finally, d(− lactic acid was demonstrated for the first time to trigger NETosis in a PAD4- and MCT1-dependent manner. Thus, d(− lactic acid-mediated neutrophil activation may contribute to neutrophil-derived pro-inflammatory processes, such as aseptic laminitis and/or polysynovitis in animals suffering acute ruminal acidosis.

  4. A novel neutrophil derived inflammatory biomarker of pulmonary exacerbation in cystic fibrosis.

    LENUS (Irish Health Repository)

    2012-02-01

    BACKGROUND: The focus of this study was to characterize a novel biomarker for cystic fibrosis (CF) that could reflect exacerbations of the disease and could be useful for therapeutic stratification of patients, or for testing of potential drug treatments. This study focused exclusively on a protein complex containing alpha-1 antitrypsin and CD16b (AAT:CD16b) which is released into the bloodstream from membranes of pro-inflammatory primed neutrophils. METHODS: Neutrophil membrane expression and extracellular levels of AAT and CD16b were quantified by flow cytometry, Western blot analysis and by 2D-PAGE. Interleukin-8 (IL-8), tumor necrosis factor-alpha (TNF-alpha) and AAT:CD16b complex were quantified in CF plasma (n=38), samples post antibiotic treatment for 14days (n=10), chronic obstructive pulmonary disease (n=10), AAT deficient (n=10) and healthy control (n=14) plasma samples by ELISA. RESULTS: Cell priming with IL-8 and TNF-alpha caused release of the AAT:CD16b complex from the neutrophil cell membrane. Circulating plasma levels of IL-8, TNF-alpha and AAT:CD16b complex were significantly higher in patients with CF than in the other patient groups or healthy controls (P<0.05). Antibiotic treatment of pulmonary exacerbation in patients with CF led to decreased plasma protein concentrations of AAT:CD16b complex with a significant correlation with improved FEV1 (r=0.81, P=0.003). CONCLUSION: The results of this study have shown that levels of AAT:CD16b complex present in plasma correlate to the inflammatory status of patients. The AAT:CD16b biomarker may become a useful addition to the clinical diagnosis of exacerbations in CF.

  5. Malnutrition in Healthy Individuals Results in Increased Mixed Cytokine Profiles, Altered Neutrophil Subsets and Function

    Science.gov (United States)

    Takele, Y.; Adem, E.; Getahun, M.; Tajebe, F.; Kiflie, A.; Hailu, A.; Raynes, J.; Mengesha, B.; Ayele, T. A.; Shkedy, Z.; Lemma, M.; Diro, E.; Toulza, F.; Modolell, M.; Munder, M.; Müller, I.; Kropf, P.

    2016-01-01

    Malnutrition is commonly associated with increased infectious disease susceptibility and severity. Whereas malnutrition might enhance the incidence of disease as well as its severity, active infection can in turn exacerbate malnutrition. Therefore, in a malnourished individual suffering from a severe infection, it is not possible to determine the contribution of the pre-existing malnutrition and/or the infection itself to increased disease severity. In the current study we focussed on two groups of malnourished, but otherwise healthy individuals: moderately malnourished (BMI: 18.4–16.5) and severely malnourished (BMI