Absolute Number Of Neutrophils Research Articles

Enhancing the immunomodulatory effects of the Toll-like receptors 3 agonist poly(I:C) by conjugation with polymers

Background : Poly(I:C) is a Toll-like receptors 3 agonist, which induces potent innate immune responses, and as a consequence adaptive immunity. However, the rapid degradation of poly(I:C) influences its half-life and its adjuvant effects in preclinical and clinical uses. Aims : We aimed in this study to conjugate poly(I:C) with polyethylene glycol/poly D, L-actide-co-glycolide (PLGA/PEG) polymers as an approach for better delivery and immunomodulatory effects. Materials and Methods : Female CD1 mice were treated once with PEG/PLGA, poly(I:C)/PLGA/PEG (50 μg), poly(I:C)/ PLGA/PEG (10 μg) or PEG via intraperitoneal injection and mice were sacrificed 1 day later for complete blood count analysis and analysis of the immune cells by flow cytometry. Results : Treatment with PEG/PLGA, poly(I:C)/PLGA/PEG (50 μg), poly(I:C)/PLGA/PEG (10 μg) or PEG administration resulted in significant (P = 0.0197) increases (1.89, 1.76, 1.69, and 1.42-fold, respectively) in the absolute number of neutrophils as compared to naive mice. Conclusion : Conjugation of poly(I:C) with polymers does not hamper its immunomodulatory effects, instead it enhances its effects on increasing the number of immune cells opening an avenue for further studies on the beneficial effects of this conjugate.

CYTOTOXIC AND ANTI-TUMOR EFFECTS OF THE CHEMOTHERAPEUTIC DRUG CYCLOPHOSPHAMIDE IN TUMOR BEARING MICE IS TIME AND DOSE DEPENDENT

Cyclophosphamide (CTX) has been used to precondition recipient hosts before adoptive T cell therapy to tumor. The aim of the present study was to analyze the host cellularity changes resulting from different doses and timing of CTX administration. To address this aim, Ehrlich ascites carcinoma (EAC)-bearing mice (5×105cells/mouse) were intraperitoneal injected with 2 or 4 mg/mouse CTX either on day 1 or 7. The results revealed a significant decrease in the number of the tumor cells at 4 mg/mouse CTX on day 1 with an induction of 99% (11.6×106 cells/µl) death of the EAC cells. On day 1 post-EAC cells inoculation, there was a significant increase in the absolute number of splenocytes in EAC-bearing mice at 2 and 4 mg/mouse CTX. The absolute number of leukocytes, lymphocytes and monocytes in EAC-bearing mice non-significantly increased at 2 mg/mouse CTX on day 7 and at 4 mg/mouse CTX on day 1 and 7. Additionally, the absolute number of neutrophils was significantly increased at 2 mg/mouse CTX on day 7 and non-significantly increased at 4 mg/mouse CTX on day 7. The results indicate that: 1) the anti-tumor effect of CTX is a dose- and time-dependent, 2) the use of high dose of chemotherapy is preferable to anti-tumor adoptive cell therapy since it induces higher anti-tumor effects with less lymphopenia.

Glutathione S-transferase M1 modulates allergen-induced NF-κB activation in asthmatic airway epithelium.

Glutathione S-transferase M1 (GSTM1) is a phase II enzyme and regulator of inflammatory signaling in airway epithelial cells. We have found upregulation of neutrophilic airway inflammation in atopic asthmatics expressing GSTM1 gene (GSTM1+) compared to GSTM1null asthmatics. We hypothesized that GSTM1 modulates NF-κB activation in bronchial epithelium in atopic asthmatics. We determined regulation of allergen-induced NF-κB activation in bronchial epithelium by GSTM1 in human atopic asthmatics in vivo. Endobronchial biopsies and bronchoalveolar lavage fluid samples were collected from 13 GSTM1+ and 12 GSTM1null human atopic asthmatics at baseline and 24h after segmental allergen challenge. A quantitative analysis of NF-κB activation in airway epithelium was accomplished using a polyclonal antibody against the phosphorylated p65 component of NF-κB. Elastase-positive neutrophils in the bronchial wall were quantified. Postallergen neutrophilia in airway subepithelium and epithelial lining fluid was greater in GSTM1+ compared to GSTM1null asthmatics. Airway eosinophilia was similar in GSTM1+ and GSTM1null asthmatics. Allergen-provoked NF-κB induction in bronchial epithelium was significantly greater in GSTM1+ compared to GSTM1null asthmatics. Activation of NF-κB activation in airway epithelial cells correlated with interleukin-8 concentrations and absolute neutrophil numbers in bronchoalveolar lavage fluid in GSTM1+ but not GSTM1null asthmatics. Allergen-induced neutrophilic airway inflammation in GSTM1+ asthmatics is associated with NF-κB activation in airway epithelial cells in vivo. These novel data provide a potential mechanism of the genomic link between GSTM1 polymorphism and airway neutrophilia in atopic asthma.

TREATMENT WITH WHITE BUTTON MUSHROOM EXTRACT ENHANCED THE ANTI-TUMOR EFFECTS AND LOWERED THE TOXICITY OF CHEMOTHERAPY IN A TUMOR MOUSE MODEL

Cyclophosphamide (CTX) is a chemotherapeutic drug used for treatment of certain solid tumors, lymphomas, and some autoimmune diseases. The antitumor effect of CTX is associated with leukopenia and liver dysfunction. There is an interest in exploring agents that can minimize the adverse effect of CTX without ameliorating its antitumor efficacy. Among the potential agents is a recent study on white button mushroom, which has been found to exert anti-toxic effects. This study was designed to assess the beneficial effects of mushroom extract on the antitumor and toxic effects of CTX. Swiss albino mice were inoculated with intraperitoneal injection (i.p.) of 0.25 x 106 (day 0) of Ehrlich ascites carcinoma (EAC) cells. Mice were treated with a single dose of CTX (200mg/kg) on day 1 or day 7 post EAC inoculation. Agaricus bisporus mushroom extract (666 mg/kg) was administrated (i.p.) on days 8-14 post tumor challenge. The numbers of leukocytes as well as liver function were assessed on day 17. Treatment with CTX on day 7 induced 46% and 37% decrease in the absolute number of monocytes and neutrophils. It had no effect on serum alanine amino transferase activity, while it induced 57% decrease in serum alkaline phosphatase activity. It induced 41% decrease in liver GSH as compared with tumor-bearing mice. Co-administration of mushroom extract with CTX induced 86% decrease in the number of EAC cells, 72% increase in the absolute number of monocytes, 37% increase in the absolute numbers of neutrophils, 57% decrease in serum ALP activity, 100% increase in GPX activity and 35% increase in GSH as compared with tumor-bearing mice. These results indicate to the beneficial effects of mushroom to enhance the antitumor effect of CTX coincided with reduction of its toxic effect.

Clinicoimmunopathologic findings in Atlantic bottlenose dolphins Tursiops truncatus with positive Chlamydiaceae antibody titers

Sera from free-ranging Atlantic bottlenose dolphins Tursiops truncatus inhabiting the Indian River Lagoon (IRL), Florida, and coastal waters of Charleston (CHS), South Carolina, USA, were tested for antibodies to Chlamydiaceae as part of a multidisciplinary study of individual and population health. A suite of clinicoimmunopathologic variables was evaluated in Chlamydiaceae-seropositive dolphins (n = 43) and seronegative healthy dolphins (n = 83). Fibrinogen, lactate dehydrogenase, amylase, and absolute numbers of neutrophils, lymphocytes, and basophils were significantly higher, and serum bicarbonate, total alpha globulin, and alpha-2 globulin were significantly lower in dolphins with positive Chlamydiaceae titers compared with seronegative healthy dolphins. Several differences in markers of innate and adaptive immunity were also found. Concanavalin A-induced T lymphocyte proliferation, lipopolysaccharide-induced B lymphocyte proliferation, and granulocytic phagocytosis were significantly lower, and absolute numbers of mature CD 21 B lymphocytes, natural killer cell activity and lysozyme concentration were significantly higher in dolphins with positive Chlamydiaceae antibody titers compared to seronegative healthy dolphins. Additionally, dolphins with positive Chlamydiaceae antibody titers had significant increases in ELISA antibody titers to Erysipelothrix rhusiopathiae. These data suggest that Chlamydiaceae infection may produce subclinical clinicoimmunopathologic perturbations that impact health. Any potential subclinical health impacts are important for the IRL and CHS dolphin populations, as past studies have indicated that both dolphin populations are affected by other complex infectious and neoplastic diseases, often associated with immunologic perturbations and anthropogenic contaminants.

Application of multivariate modeling for radiation injury assessment: a proof of concept.

Multivariate radiation injury estimation algorithms were formulated for estimating severe hematopoietic acute radiation syndrome (H-ARS) injury (i.e., response category three or RC3) in a rhesus monkey total-body irradiation (TBI) model. Classical CBC and serum chemistry blood parameters were examined prior to irradiation (d 0) and on d 7, 10, 14, 21, and 25 after irradiation involving 24 nonhuman primates (NHP) (Macaca mulatta) given 6.5-Gy 60Co Υ-rays (0.4 Gy min−1) TBI. A correlation matrix was formulated with the RC3 severity level designated as the “dependent variable” and independent variables down selected based on their radioresponsiveness and relatively low multicollinearity using stepwise-linear regression analyses. Final candidate independent variables included CBC counts (absolute number of neutrophils, lymphocytes, and platelets) in formulating the “CBC” RC3 estimation algorithm. Additionally, the formulation of a diagnostic CBC and serum chemistry “CBC-SCHEM” RC3 algorithm expanded upon the CBC algorithm model with the addition of hematocrit and the serum enzyme levels of aspartate aminotransferase, creatine kinase, and lactate dehydrogenase. Both algorithms estimated RC3 with over 90% predictive power. Only the CBC-SCHEM RC3 algorithm, however, met the critical three assumptions of linear least squares demonstrating slightly greater precision for radiation injury estimation, but with significantly decreased prediction error indicating increased statistical robustness.

Neutrophil Related Data Obtained From Newly Developed Automatic Hematology Analyzer Sysmex XN-2000 Can Provide Useful Informations For The Discrimination Of Sepsis Patients

Background In addition to classical biomarkers of sepsis such as C-reactive protein (CRP), lactic acid, and procalcitonin (PCT), new parameters such as immature granulocytes (IG), which can be obtained from automated hematology analyzers without any additional cost, have been recently proposed as biomarkers for the early detection of sepsis and the evaluation of its severity. Recently launched automatic hematology analyzer Sysmex XN-2000 (Sysmex, Kobe, Japan) can provide 36 routine items as well as 57 items of cell population data (CPD) which reflect detailed status of cells examined. In this study, we evaluated the clinical relevance of CPD items as a biomarker for the diagnosis of sepsis, discrimination of sepsis severity, and prediction of outcome in sepsis patients. Methods A total of 280 normal controls (NCs) and 130 patients diagnosed as bacterial sepsis were enrolled in this study. The sepsis patients were classified into uncomplicated sepsis (N=29) and complicated sepsis (N=101, defined as severe sepsis and and septic shock) depending on the sepsis severity at the time of sampling. A total of 93 items (36 routine and 53 CPD) were obtained from the Sysmex XN-2000 automatic hematology analyzer and the results were compared between NC and sepsis group at first and between uncomplicated and complicated sepsis group later. In items with significant differences between 2 groups, receiver operating characteristics (ROC) analysis was performed to evaluate discrimination power of the item between 2 groups. In addition, the results of 93 items were compared between survivors and non-survivors after 28 days from the time of sampling, and ROC analysis was also performed to compare the prediction power of the 28-day mortality of each item which showed significant differences between 2 groups. Results Among the total 93 items, a total of 21 items showed area under the curve (AUC) more than 0.900 for the discrimination of sepsis patients from NCs. These include routine items such as hematocrit (best cutoff values ≤ 37.2%, AUC 0.964), hemoglobin (≤ 11.7 g/dL AUC 0.961), IG (≥ 0.5%, AUC 0.961), CV of RDW (> 13.7%, AUC 0.951), proportion of lymphocytes (≤ 18.5%, AUC 0.941) and neutrophils (> 72.2%, AUC 0.914). Notably, CPD items NE-SFL (defined as the fluorescent light intensity of the neutrophil area on the WDF scattergram) and NE-WY (defined as the fluorescent light distribution width of the neutrophil area on the WDF scattergram) showed comparative AUC of 0.909 (> 51.6 for NE-SFL) and 0.905 (> 641.0 for NE-WY). However, for the discrimination of sepsis severity, the overall performance of all items was relatively poor and only 3 red blood cell CPD items such as RET-UPP (defined as the count in the upper area of the RET scattergram, ≤ 2.0%, AUC of 0.662), reticulocyte hemoglobin (>33.2 pg, AUC of 0.633), and MacroR (defined as the macrocytic RBC ratio, > 4.60%, AUC of 0.607) and only 1 granulocyte CPD item NE-SFL (> 57.6, AUC of 0.603) showed AUC more than 0.600. The performance of classical sepsis biomarkers was also not satisfactory (AUC of 0.695, 0.543, and 0.537 for lactic acid, CRP, and PCT). For the prediction of 28-day mortality, the overall performance of all items was also not good and only 4 routine items such as CV of RDW (> 15.4%, AUC of 0.766), platelet counts (< 54,000/µL, AUC of 0.677), absolute number of neutrophils (> 12,120/µL, AUC of 0.665), absolute number of reticulocytes (> 2.89 x 106/µL, AUC of 0.650), and 1 CPD item RET-TNC (defined as the count in the TNC area of the RET scattergram, > 125.00, AUC of 0.692) showed AUC of more than 0.650. These results were not superior compared with those from classical sepsis biomarkers (AUC of 0.712, 0.547, and 0.592 for lactic acid, CRP, and PCT). Conclusion IG%, the proportion of neutrophils, and 2 granulocytic CPD items (NE-SFL and NE-WY) could discriminate sepsis patients from normal controls when their values increase (which means neutrophil immaturity or activation). Several CPD items showed similar performance compared with lactic acid and slightly better performance than CRP and PCT for the discrimination of sepsis severity and prediction of 28-day mortality. Therefore, it can be speculated that the CPD items of neutrophil immaturity or activation can contribute partly to the discrimination of sepsis patients from NCs, but these items do not discriminate sepsis severity and predict 28-day mortality in sepsis patients as the classical sepsis biomarkers do not. Disclosures: No relevant conflicts of interest to declare.

A translational approach to test anti-inflammatory drugs in a LPS induced acute lung inflammation model in the common marmoset (Callithrix jacchus)

The rising incidence of chronic-obstructive pulmonary diseases is linked to the development of new human-specific anti-inflammatory therapeutics. In many cases classic rodent models fail to predict reliable data of safety and efficacy, which contributes to a growing demand of models that represent the human situation in more detail e. g., anatomy and immune-response. Similar to the human setting of segmental lipopolysaccharide (LPS) challenge, we established a LPS induced acute lung-inflammation model in the common marmoset monkey, a small non-human primate. LPS induced inflammation mimics pro-inflammatory aspects of chronic airway diseases associated with neutrophile aggregation. Healthy animals underwent a first BAL to obtain baseline data. Marmosets were than pre-treated orally with roflumilast (7 µg/kg bw), a recently approved PDE-4-inhibitor, or dexamethasone (2 mg/kg bw) as a treatment control on five consecutive days. Sham treated animals served as positive control. Under general anaesthesia 500 ng LPS were instilled into the left half of the lung of each animal, followed by ipsilateral BAL of the LPS challenged lung 18 h post-provocation. BAL fluid was processed and analysed for cellular and cytokine levels. A whole blood assay performed on day 0 and 5 served as an additional readout. As expected, LPS induced significant influx of total cells into the airways (p<0.001) mainly through neutrophil aggregation. Pre-treatment with roflumilast resulted in a statistically significant decrease of neutrophil numbers in BAL fluid (p=0.047). Dexamethasone pre-treatment showed a significant reduction in relative cell numbers (p=0.047), but not in absolute neutrophil numbers (p=0.076). TNF-α levels were significantly increased after LPS provocation and significantly suppressed in roflumilast (p=0.048) and dexamethasone (p=0.036) pre-treated animals. In the WBA only pre-treatment with dexamethasone resulted in a reduced TNF-α release after ex vivo LPS provocation. Results from this new translational nonhuman primate model indicate the common marmoset monkey as a promising species to test new anti-inflammatory drugs.

P-168 Familial myelodysplastic syndrome/acute leukemia in adult setting – Experience from a single Brazilian center

Introduction: Myelodysplastic syndromes (MDS) are a group of heterogenous clonal stem cell disorders characterized by inefficient haematopoiesis with one or more cytopenias and increased risk of transformation to acute leukemia. Purpose: The current diagnosis includes peripheral blood morphology, bone marrow biopsy and cytogenetic exam. Clonal cytogenetic abnormalities are described in 40-60% of cases. In 1997, the established System of International Prognostic Score was based on the number of cytopenias on the peripheral blood, precentage bone marrow blasts and karyotype abnomalities. In this system are described a limited number of cytogenetic abnormalities and the importance of their forecast is underestimated. This score classify patients into four risk groups, allow overall survival and estimate the risk of progression to acute leukemia. Materials and Methods: In September 2012 the Revised International Prognostic Scoring System (R-IPSS) was developed in the framework of a project of the International Working Group for Prognosis in MDS (IWG-PM) peak. For scoring according to IPSS-R, it was taken into account: severity of cytopenias (Hb level, the number of platelets, the absolute number of neutrophils), the percentage of bone marrow blasts and cytogenetic exam. It was found that this system prognostic allows a better classification of patients with MDS into risk groups. We analyzed 25 patients diagnosed with MDS, according to theWHO classification 2001, with low risk and intermediate-1 during the period 2008-2012. These patients have been hospitalized in three centers of Hematology Departments from Romania. Results: The lot of patients was evaluated according to R-IPSS (2012) and it was found that the new system allows a better prognosis estimation and more accurate patient risk group classification. Conclusions: Although according to IPSS (1997), all patients included in this studywere low risk (0-1) but applying the new scoring system has allowed a better classification of patients, six of them were with intermediate and high risk (3,5-6).

Association of high levels of α-defensins and S100A proteins with Candida mannan detection in bronchoalveolar lavage fluid of preterm neonates

Candida mannan (Mn) detection in bronchoalveolar lavage fluid (BALF) was shown to be useful for earlier identification and preemptive therapy targeting in preterm infants at high risk of invasive Candida infection. We investigated whether early detection of Candida Mn in BALF is associated with the presence of some neutrophilic products, as markers of prenatal infection/inflammation. BALF specimens were collected during the first 48 h of life from mechanically ventilated preterm newborns. Samples were analyzed by high-performance liquid chromatography-electrospray ionization-mass spectrometry. The relative amounts of α-defensins 1-4 and S100A proteins were measured by extracted ion current peak area. Absolute and differential white cell counts in BALF were obtained. Mn antigen concentrations were determined by the Platelia Candida antigen kit. Twenty-five studied neonates were divided into two groups: Mn-positive group and Mn-negative group. Levels of α-defensins 1-4 and S100A12 were significantly higher in the Mn-positive group than in the Mn-negative group. Moreover, positive significant correlations between the absolute number of neutrophils and the levels of α-defensins 1-4 and S100A8 were observed. The detection of Mn antigen in BALF of preterm infants is consistent with evidence of an innate immune response in their lungs as demonstrated by higher levels of α-defensins and S100A proteins.

How to approach neutropenia.

Neutropenia is defined as the reduction in the absolute number of neutrophils in the blood circulation. Acute neutropenia is a relatively frequent finding, whereas disorders of production of neutrophils are quite rare. Acute neutropenia is often well tolerated and normalizes rapidly. Neutropenia arising as a result of underlying hematologic disorders is far more significant. Such a patient may be at risk for infectious complications and will likely require a thorough investigation. Acute neutropenia evolves over a few days and occurs when neutrophil use is rapid and production is impaired. Chronic neutropenia may last for 3 months or longer and is a result of reduced production, increased destruction, or excessive splenic sequestration of neutrophils. Neutropenia may be classified by whether it arises secondarily to causes extrinsic to BM myeloid cells, which is common; as an acquired disorder of myeloid progenitor cells, which is less frequent; or as an intrinsic defect arising from impaired proliferation and maturation of myeloid progenitor cells in the BM, which is rare. Severe neutropenia with absolute neutrophil counts below 500/μL increases susceptibility to bacterial or fungal infections. Multiple disorders of severe congenital neutropenia have been found by the discovery of genetic defects affecting differentiation, adhesion, and apoptosis of neutrophil precursors. Elucidation of the multiple genetic defects have provided insight into the biology of the cell involving membrane structures, secretory vesicles, mitochondrial metabolism, ribosome biogenesis, transcriptional regulation, and cytoskeletal dynamics, as well as the risk for myelodysplasia and acute myeloid leukemia.

How to approach neutropenia

AbstractNeutropenia is defined as the reduction in the absolute number of neutrophils in the blood circulation. Acute neutropenia is a relatively frequent finding, whereas disorders of production of neutrophils are quite rare. Acute neutropenia is often well tolerated and normalizes rapidly. Neutropenia arising as a result of underlying hematologic disorders is far more significant. Such a patient may be at risk for infectious complications and will likely require a thorough investigation. Acute neutropenia evolves over a few days and occurs when neutrophil use is rapid and production is impaired. Chronic neutropenia may last for 3 months or longer and is a result of reduced production, increased destruction, or excessive splenic sequestration of neutrophils. Neutropenia may be classified by whether it arises secondarily to causes extrinsic to BM myeloid cells, which is common; as an acquired disorder of myeloid progenitor cells, which is less frequent; or as an intrinsic defect arising from impaired proliferation and maturation of myeloid progenitor cells in the BM, which is rare. Severe neutropenia with absolute neutrophil counts below 500/μL increases susceptibility to bacterial or fungal infections. Multiple disorders of severe congenital neutropenia have been found by the discovery of genetic defects affecting differentiation, adhesion, and apoptosis of neutrophil precursors. Elucidation of the multiple genetic defects have provided insight into the biology of the cell involving membrane structures, secretory vesicles, mitochondrial metabolism, ribosome biogenesis, transcriptional regulation, and cytoskeletal dynamics, as well as the risk for myelodysplasia and acute myeloid leukemia.

Cytokine profile of bronchoalveolar lavage fluid in patients with combined pulmonary fibrosis and emphysema

Combined pulmonary fibrosis and emphysema (CPFE) is characterized by upper lobe emphysema together with lower lobe fibrosis. The aim of this study was to examine whether cytokine levels in the alveolar space are associated with emphysematous changes superimposed on pulmonary fibrosis. Consecutive patients (n = 102), diagnosed with pulmonary fibrosis were retrospectively evaluated. Cytokine levels and differential cell counts in bronchoalveolar lavage (BAL) fluid, pulmonary function, computed tomography (CT) scores and levels of serum markers were compared between patients with or without emphysema. Among the 102 patients (14 females, mean age 68 years), 38 (37%) had evidence of upper lobe emphysema on computed tomography (CT). Levels of epithelial neutrophil activating peptide 78 (ENA-78/CXCL5) and interleukin (IL)-8/CXCL8 in BAL fluid were significantly higher in patients with emphysema. Vital capacity (VC, % predicted) was greater, and ratio of forced expiratory volume in 1 s/forced vital capacity and diffusing capacity of carbon monoxide (DL(CO))/alveolar volume (V(A)) were lower in patients with emphysema. CXCL8 and CXCL5 levels were associated with percentage or absolute numbers of neutrophils in BAL fluid. In addition, CXCL8 levels were inversely correlated with VC and DL(CO)/V(A), and positively correlated with composite physiological index (CPI) and the extent of areas of low attenuation on CT. Increased CXC chemokine levels in the airspaces may be associated with emphysematous lung changes in patients with pulmonary fibrosis.

Inflammatory Marker sTREM-1 Reflects the Clinical Stage and Respiratory Tract Obstruction in Allergic Asthma Bronchiale Patients and Correlates with Number of Neutrophils

The knowledge that asthma is an inflammatory disorder has prompted us to investigate the plasma levels of a new inflammatory marker sTREM-1 that is released from the surfaces of activated neutrophils and monocytes. The plasma levels of sTREM-1 were analysed by a sandwich ELISA test in the cohort of 76 patients with allergic asthma bronchiale and 39 healthy controls. Our results revealed more than 3.5 times higher levels of sTREM-1 in AB patients (92.3 pg/mL ± 125.6) compared with healthy subjects (25.7 pg/mL ± 9.2; P = 0.0001). Higher levels of sTREM-1 were found also in patients with exacerbated AB (170.5 pg/mL ± 78.2) compared with nonexacerbated AB patients (59.1 ± 78.2; P < 0.0001), patients with respiratory tract obstruction (176.4 pg/mL ± 177.8), than those without obstruction (51.99 pg/mL ± 64.0; P < 0.0001) and patients with anti-IgE therapy (P < 0.0001). Levels of sTREM-1 correlated with number of leucocytes (P = 0.002), and absolute number of neutrophils (P = 0.001). Elevated plasma levels of sTREM-1 reflect the severity, state of exacerbation, presence of respiratory tract obstruction in AB patients and together with increased number of neutrophils point to the role of neutrophils in inflammation accompanying AB.

Enhanced neutrophil longevity and recruitment contribute to the severity of oviduct pathology during Chlamydia muridarum infection.

Our previous studies revealed that intravaginal infection of mice with a plasmid-deficient strain of Chlamydia muridarum, CM3.1, does not induce the development of oviduct pathology. In this study, we determined that infection with CM3.1 resulted in a significantly reduced frequency and absolute number of neutrophils in the oviducts during acute infection. This reduction in neutrophils was associated with significantly lower levels of neutrophil chemokines in the oviducts and decreased production of neutrophil chemokines by oviduct epithelial cells infected with CM3.1 in vitro. Infection with CM3.1 also resulted in an increased frequency of late apoptotic/dead neutrophils in the oviduct. Examination of the ability of Chlamydia trachomatis to prevent neutrophil apoptosis in vitro revealed that C. trachomatis strain D/UW-3/Cx exhibited an enhanced ability to prevent neutrophil apoptosis compared to plasmid-deficient CTD153, and this effect was dependent on the presence of CD14(high) monocytes. The presence of monocytes also resulted in enhanced neutrophil cytokine production and increased production of tissue-damaging molecules in response to D/UW-3/Cx relative to results with CTD153. Attempts to use antibody-mediated depletion to discern the specific role of neutrophils in infection control and pathology in vivo revealed that although Ly6G(high) neutrophils were eliminated from the blood and oviducts with this treatment, immature neutrophils and high levels of tissue-damaging molecules were still detectable in the upper genital tract. These data support the role of neutrophils in chlamydia-induced pathology and reveal that novel methods of depletion must be developed before their role can be specifically determined in vivo.

Reconstitution and functional analyses of neutrophils and distinct subsets of monocytes after allogeneic stem cell transplantation

The aim of the study was to investigate the recovery of the innate immune system within the first 100days after allogeneic peripheral blood stem cell transplantation (PBSCT) and to elucidate a potential correlation with such important events as severe infectious complications or graft-versus-host disease (GvHD). In 30 consecutive patients who underwent allogeneic PBSCT, absolute numbers of neutrophils and monocytes were determined and different functional analyses performed at different time points (day +30, +60 and +90, respectively). The capacity to phagocyte Escherichia coli (E. coli) as well as the induction of oxidative burst after incubation with different stimuli (Phorbol-12-myristate-13-acetate; PMA, the chemotactic peptide N-formyl-Met-Leu-Phe; f-MLP or opsonized E. coli) were analysed after engraftment. There was a rapid reconstitution concerning the capability of both neutrophils and monocytes to phagocyte E. coli without a significant increase between day +30 and +90. In contrast, a twofold increase of monocyte oxidative burst after incubation with PMA at day +90 was observed (P=0.017). Furthermore, the ability of neutrophils to induce oxidative burst after ingestion with E. coli was impaired on day +30 with a significant functional reconstitution on day +60 (P=0.01). The oxidative burst activity following incubation with f-MLP did not show significant changes after stem cell engraftment. Analysis of numeric reconstitution of CD14+CD16+ monocytes demonstrated a potential correlation with a decreased incidence of chronic GvHD. The functional recovery of neutrophils and monocytes in the early period after allogeneic PBSCT differs not only concerning phagocytosis and oxidative burst but also with respect to the stimulus and the cell population that was analysed for oxidative burst activity. The subset of CD16+CD14+ monocytes might be a predictor for a reduced risk of chronic GvHD.

Fresh cow mastitis monitoring on day 3 postpartum and its relationship to subsequent milk production

The purpose was to determine the association of milk California Mastitis Test (CMT), somatic cell concentration (SCC), and milk differential cell count results on day 3 postcalving with subsequent lactation production and health events. On d 3 postcalving, the CMT was performed and quarter milk samples were collected from 130 dairy cows. Quarter SCC and milk differential cell counts were determined. Microbiology on duplicate quarter milk samples was used to determine the presence of intramammary infection by major or minor pathogens. Production measures obtained using Dairy Herd Improvement Association testing were 150-d standardized and summit milks. Milk culture results on a cow basis included 82 (63.1%) samples with no growth, 31 (23.9%) with major pathogens, and 17 (13.1%) with minor pathogens. Milk culture results comparing cows with no growth to those with any growth (major or minor pathogens) were not associated with statistically significant differences in milk production. Milk culture results comparing cows with major pathogens to those with no growth and minor pathogens combined were associated with statistically significant differences in 150 d milk. Milk production did not differ for cows with CMT results above and below a cut-off of trace, and for SCC results above and below cut-offs of 200,000, 300,000, and 400,000/mL, respectively. Statistically significant differences in milk production were found for cows above and below cut-offs for percentage neutrophils in milk and for absolute neutrophil counts. Associations were found for milk production and number of quarters (0, 1, 2, or 3 and 4 combined) above respective cut-offs for SCC, percentage neutrophils in milk, and absolute numbers of neutrophils in milk, but not for CMT. Milk production differed for cows experiencing any health event versus those with no health event. The most commonly recorded health event was clinical mastitis. Statistically significant associations were detected between health events and milk culture results, SCC, neutrophil percentage, and neutrophil absolute counts. Results of the present investigation indicate that milk monitoring on d 3 of lactation using milk neutrophil percentage or neutrophil absolute counts may be useful as an indication of subsequent milk production.

Effect of storage time on automated cell count and cytological interpretation of body cavity effusions

The effect of 24- and 48-hour storage at room temperature on automated total nucleated cell count (TNCC), differential cell count (DCC) and cell morphology was assessed, and the effect of...

Correlation of Levels of α–Defensins Determined by HPLC-ESI-MS in Bronchoalveolar Lavage Fluid With the Diagnosis of Pneumonia in Premature Neonates

The presence of alpha-defensins in bronchoalveolar lavage fluid (BALF) was investigated in a cohort of preterm newborns with gestational age (GA) < or =30 wk. Specimens were collected during the first week of life from 24 preterm neonates mechanically ventilated. The studied population was divided into two groups: pneumonia group of nine neonates suffering from pulmonary infection (GA: 26.1 +/- 2.1 wk; birth weight: 787.4 +/- 309.9 g), with or without associated bloodstream infection, and nonpneumonia group of 15 neonates (GA: 27.7 +/- 2.0 wk; birth weight: 1019.0 +/- 319.8 g). BALF culture was positive for CONS (n = 5), Staphylococcus aureus (n = 1), and Candida spp (n = 3). BALF samples were analyzed by HPLC-electrospray Ionization-mass spectrometer. The alpha-defensins 1-4 concentration, absolute and differential white cells count were measured. Relative amounts of alpha-defensins 1-4 and the absolute number of neutrophils were found significantly higher in the pneumonia group with respect to the nonpneumonia group (p < 0.05). Moreover, positive significant correlations between the number of neutrophils and the alpha-defensins 1-3 levels were observed. In conclusion, our data show that preterm newborns, also at the lower GA, are able to produce alpha-defensins, underlining that their innate defense system is already active before the at-term delivery date.

Controversial data on simvastatin in asthma: What about the rat model?

The effects of simvastatin on lung inflammation in asthma are controversial. Reduction of inflammation and hyperreactivity has been reported in studies using murine models of asthma. In contrast, a clinical study has not found beneficial effects in patients. The rat model of asthma has some distinct advantages and is still widely used in industrial studies. Therefore, the role of simvastatin was investigated in this rat model using intraperitoneal and intratracheal administration. With both simvastatin administration routes, the relative and absolute numbers of neutrophils, eosinophils, and lymphocytes were only partially reduced after increasing dosages (0.1, 1.0, and 10 mg per animal). The most obvious effect was on CD4 T cell numbers, which were reduced in most treatment groups. The results presented here suggest that treatment with simvastatin differs between species, and that it is too early for extrapolation of these data to humans.