Circulating Neutrophil Counts Research Articles

Long-term cardiovascular inflammation and fibrosis in a murine model of vasculitis induced by Lactobacillus casei cell wall extract.

Kawasaki disease (KD), an acute febrile illness and systemic vasculitis, is the leading cause of acquired heart disease in children in industrialized countries. KD leads to the development of coronary artery aneurysms (CAA) in affected children, which may persist for months and even years after the acute phase of the disease. There is an unmet need to characterize the immune and pathological mechanisms of the long-term complications of KD. We examined cardiovascular complications in the Lactobacillus casei cell wall extract (LCWE) mouse model of KD-like vasculitis over 4 months. The long-term immune, pathological, and functional changes occurring in cardiovascular lesions were characterized by histological examination, flow cytometric analysis, immunofluorescent staining of cardiovascular tissues, and transthoracic echocardiogram. CAA and abdominal aorta dilations were detected up to 16 weeks following LCWE injection and initiation of acute vasculitis. We observed alterations in the composition of circulating immune cell profiles, such as increased monocyte frequencies in the acute phase of the disease and higher counts of neutrophils. We determined a positive correlation between circulating neutrophil and inflammatory monocyte counts and the severity of cardiovascular lesions early after LCWE injection. LCWE-induced KD-like vasculitis was associated with myocarditis and myocardial dysfunction, characterized by diminished ejection fraction and left ventricular remodeling, which worsened over time. We observed extensive fibrosis within the inflamed cardiac tissue early in the disease and myocardial fibrosis in later stages. Our findings indicate that increased circulating neutrophil counts in the acute phase are a reliable predictor of cardiovascular inflammation severity in LCWE-injected mice. Furthermore, long-term cardiac complications stemming from inflammatory cell infiltrations in the aortic root and coronary arteries, myocardial dysfunction, and myocardial fibrosis persist over long periods and are still detected up to 16 weeks after LCWE injection.

Neutrophils cultured ex vivo from CD34+ stem cells are immature and genetically tractable

BackgroundNeutrophils are granulocytes with essential antimicrobial effector functions and short lifespans. During infection or sterile inflammation, emergency granulopoiesis leads to release of immature neutrophils from the bone marrow, serving to boost circulating neutrophil counts. Steady state and emergency granulopoiesis are incompletely understood, partly due to a lack of genetically amenable models of neutrophil development.MethodsWe optimised a method for ex vivo production of human neutrophils from CD34+ haematopoietic progenitors. Using flow cytometry, we phenotypically compared cultured neutrophils with native neutrophils from donors experiencing emergency granulopoiesis, and steady state neutrophils from non-challenged donors. We carry out functional and proteomic characterisation of cultured neutrophils and establish genome editing of progenitors.ResultsWe obtain high yields of ex vivo cultured neutrophils, which phenotypically resemble immature neutrophils released into the circulation during emergency granulopoiesis. Cultured neutrophils have similar rates of ROS production and bacterial killing but altered degranulation, cytokine release and antifungal activity compared to mature neutrophils isolated from peripheral blood. These differences are likely due to incomplete synthesis of granule proteins, as demonstrated by proteomic analysis.ConclusionEx vivo cultured neutrophils are genetically tractable via genome editing of precursors and provide a powerful model system for investigating the properties and behaviour of immature neutrophils.

#1265 Circulating neutrophils in diabetic kidney disease—the relationship with long-term cardiovascular outcomes

Abstract Background and Aims Neutrophils play important roles in the pathogenesis of cardiovascular inflammation and affects cardiovascular disease (CVD) outcomes. The relationship of circulating neutrophil counts with cardiovascular disease (CVD) and all-cause mortality in patients with type 2 diabetes mellitus (T2DM) and diabetic kidney disease (DKD) remains unclear. Method We retrieved data from 44,494 participants in the National Health and Nutrition Examination Surveys (NHANES) from 2005 to 2020, and investigated the associations between circulating neutrophil counts, kidney functions and CVD mortality in T2DM and DKD patients. Clinical predictive models and risk scores for long-term mortality were constructed. Results Among the cohort of 44,332 patients (18% with T2DM and 82% without T2DM), a total of 2,220 patients were diagnosed with diabetic kidney disease (DKD) and 775 of them experienced mortality (31.5% related to CVD) during a follow-up of 6.18 (range: 5.94-6.42) years. The weighted mean of peripheral neutrophil count level (×109 /L) in DKD participants was 4.98. DKD patients in the highest quintile of neutrophil count exhibited a positive correlation with urine albumin-creatinine ratio (uACR) and a negative association with estimated glomerular filtration rate (eGFR), when compared to those in the lowest quintile. In the multivariate logistic regression analysis with RCS, circulating neutrophil counts showed a U-shaped association with CVD mortality. Neutrophil counts of 3.68 × 109 /L and 3.69 × 109 /L were associated with the lowest risk of CVD mortality and all-cause mortality respectively in DKD patients. Clinical predictive models showed good diagnostic performance for 5- and 10-year CVD mortality-free survival (all ROC AUC values >0.75). Conclusion In this study, higher peripheral neutrophil count was associated with an increased risk of cardiovascular mortality in DKD individuals. Our clinical predictive models may help identify at-risk patients for more aggressive CVD risk factors control.

Hematopoietic and eosinophil-specific LNK(SH2B3) deficiency promotes eosinophilia and arterial thrombosis

AbstractIncreased eosinophil counts are associated with cardiovascular disease and may be an independent predictor of major cardiovascular events. However, the causality and underlying mechanisms are poorly understood. Genome-wide association studies have shown an association of a common LNK variant (R262W, T allele) with eosinophilia and atherothrombotic disorders. LNK(TT) reduces LNK function, and Lnk-deficient mice display accelerated atherosclerosis and thrombosis. This study was undertaken to assess the role of eosinophils in arterial thrombosis in mice with hematopoietic Lnk deficiency. Hematopoietic Lnk deficiency increased circulating and activated eosinophils, JAK/STAT signaling in eosinophils, and carotid arterial thrombosis with increased eosinophil abundance and extracellular trap formation (EETosis) in thrombi. Depletion of eosinophils by anti–Siglec-F antibody or by the ΔdbIGata1 mutation eliminated eosinophils in thrombi and markedly reduced thrombosis in mice with hematopoietic Lnk deficiency but not in control mice. Eosinophil depletion reduced neutrophil abundance and NETosis in thrombi without altering circulating neutrophil counts. To assess the role of Lnk specifically in eosinophils, we crossed Lnkf/f mice with eoCre mice. LnkΔeos mice displayed isolated eosinophilia, increased eosinophil activation, and accelerated arterial thrombosis associated with increased EETosis and NETosis in thrombi. DNase I infusion abolished EETs and neutrophil extracellular traps (NETs) in thrombi and reversed the accelerated thrombosis. Human induced pluripotent stem cell–derived LNK(TT) eosinophils showed increased activation and EETosis relative to isogenic LNK(CC) eosinophils, demonstrating human relevance. These studies show a direct link between eosinophilia, EETosis, and atherothrombosis in hematopoietic Lnk deficiency and an essential role of eosinophil LNK in suppression of arterial thrombosis.

Type 1 diabetes mellitus: Roles of neutrophils in the pathogenesis.

Circulating neutrophil counts are reduced both in healthy autoantibody-positive individuals and in patients with type 1 diabetes, which may be related on cell-specific autoimmunity. This paper was written to give an update on roles of neutrophils in the pathogenesis of type 1 diabetes mellitus. Different research search engines like PubMed Central, Scopus, Web of Science, Researchgate, Google Scholar etc were utilised for writing this paper. A drop in blood neutrophil counts in type 1 diabetes may be caused by decreased neutrophil generation and maturation, tissue maintenance, consumption, or peripheral damage. Neutrophil count variations between studies may be explained by results from various stages of diabetes or by ethnic groups. Neutrophils can induce type 1 diabetes by colonizing pancreatic islets and interacting with other immune cells, according to exciting findings that shed new light on their role in the pathogenesis of the disease. Knowing more about the function of neutrophils in the pathogenesis of type 1 diabetes will help in early diagnosis, treatment, and even prevention of the disease.

Plasma metabolomic profiling of patients with transient ischemic attack reveals positive role of neutrophils in ischemic tolerance

Transient ischemic attack (TIA) induces ischemic tolerance that can reduce the subsequent ischemic damage and improve prognosis of patients with stroke. However, the underlying mechanisms remain elusive. Recent advances in plasma metabolomics analysis have made it a powerful tool to investigate human pathophysiological phenotypes and mechanisms of diseases. In this study, we aimed to identify the bioactive metabolites from the plasma of patients with TIA for determination of their prophylactic and therapeutic effects on protection against cerebral ischemic stroke, and the mechanism of TIA-induced ischemic tolerance against subsequent stroke. Metabolomic profiling using liquid chromatography-mass spectrometry was performed to identify the TIA-induced differential bioactive metabolites in the plasma samples of 20 patients at day 1 (time for basal metabolites) and day 7 (time for established chronic ischemic tolerance-associated metabolites) after onset of TIA. Mouse middle cerebral artery occlusion (MCAO)-induced stroke model was used to verify their prophylactic and therapeutic potentials. Transcriptomics changes in circulating neutrophils of patients with TIA were determined by RNA-sequencing. Multivariate statistics and integrative analysis of metabolomics and transcriptomics were performed to elucidate the potential mechanism of TIA-induced ischemic tolerance. Plasma metabolomics analysis identified five differentially upregulated metabolites associated with potentially TIA-induced ischemic tolerance, namely all-trans 13,14 dihydroretinol (atDR), 20-carboxyleukotriene B4, prostaglandin B2, cortisol and 9-KODE. They were associated with the metabolic pathways of retinol, arachidonic acid, and neuroactive ligand-receptor interaction. Prophylactic treatment of MCAO mice with these five metabolites significantly improved neurological functions. Additionally, post-stroke treatment with atDR or 9-KODE significantly reduced the cerebral infarct size and enhanced sensorimotor functions, demonstrating the therapeutic potential of these bioactive metabolites. Mechanistically, we found in patients with TIA that these metabolites were positively correlated with circulating neutrophil counts. Integrative analysis of plasma metabolomics and neutrophil transcriptomics further revealed that TIA-induced metabolites are significantly correlated with specific gene expression in circulating neutrophils which showed prominent enrichment in FoxO signaling pathway and upregulation of the anti-inflammatory cytokine IL-10. Finally, we demonstrated that the protective effect of atDR-pretreatment on MCAO mice was abolished when circulating neutrophils were depleted. TIA-induced potential ischemic tolerance is associated with upregulation of plasma bioactive metabolites which can protect against cerebral ischemic damage and improve neurological functions through a positive role of circulating neutrophils. National Natural Science Foundation of China (81974210), Science and Technology Planning Project of Guangdong Province, China (2020A0505100045), Natural Science Foundation of Guangdong Province (2019A1515010671), Science and Technology Program of Guangzhou, China (2023A03J0577), and Natural Science Foundation of Jiangxi, China（20224BAB216043).

INVESTIGATION INTO P2Y RECEPTOR FUNCTION IN PLATELETS FROM PATIENTS WITH SEPSIS.

Key underlying pathological mechanisms contributing to sepsis are hemostatic dysfunction and overwhelming inflammation. Platelet aggregation is required for hemostasis, and platelets are also separately involved in inflammatory responses that require different functional attributes. Nevertheless, P2Y receptor activation of platelets is required for this dichotomy of function. The aim of this study was to elucidate whether P2YR-dependent hemostatic and inflammatory functions were altered in platelets isolated from sepsis patients, compared with patients with mild sterile inflammation. Platelets from patients undergoing elective cardiac surgery (20 patients, 3 female) or experiencing sepsis after community-acquired pneumonia (10 patients, 4 female) were obtained through the IMMunE dysfunction and Recovery from SEpsis-related critical illness in adults (IMMERSE) Observational Clinical Trial. In vitro aggregation and chemotaxis assays were performed with platelets after stimulation with ADP and compared with platelets isolated from healthy control subjects (7 donors, 5 female). Cardiac surgery and sepsis both induced a robust inflammatory response with increases in circulating neutrophil counts with a trend toward decreased circulating platelet counts being observed. The ability of platelets to aggregate in response to ex vivo ADP stimulation was preserved in all groups. However, platelets isolated from patients with sepsis lost the ability to undergo chemotaxis toward N -formylmethionyl-leucyl-phenylalanine, and this suppression was evident at admission through to and including discharge from hospital. Our results suggest that P2Y 1 -dependent inflammatory function in platelets is lost in patients with sepsis resulting from community-acquired pneumonia. Further studies will need to be undertaken to determine whether this is due to localized recruitment to the lungs of a platelet responsive population or loss of function as a result of dysregulation of the immune response.

Loss-of-function variants in SAT1 cause X-linked childhood-onset systemic lupus erythematosus

ObjectivesFamilies that contain multiple siblings affected with childhood onset of systemic lupus erythematosus (SLE) likely have strong genetic predispositions. We performed whole exome sequencing (WES) to identify familial rare risk...

Cigarette smoke aggravates asthma by inducing memory-like type 3 innate lymphoid cells

Although cigarette smoking is known to exacerbate asthma, only a few clinical asthma studies have been conducted involving smokers. Here we show, by comparing paired sputum and blood samples from smoking and non-smoking patients with asthma, that smoking associates with significantly higher frequencies of pro-inflammatory, natural-cytotoxicity-receptor-non-expressing type 3 innate lymphoid cells (ILC3) in the sputum and memory-like, CD45RO-expressing ILC3s in the blood. These ILC3 frequencies positively correlate with circulating neutrophil counts and M1 alveolar macrophage frequencies, which are known to increase in uncontrolled severe asthma, yet do not correlate with circulating eosinophil frequencies that characterize allergic asthma. In vitro exposure of ILCs to cigarette smoke extract induces expression of the memory marker CD45RO in ILC3s. Cigarette smoke extract also impairs the barrier function of airway epithelial cells and increases their production of IL-1β, which is a known activating factor for ILC3s. Thus, our study suggests that cigarette smoking increases local and circulating frequencies of activated ILC3 cells, plays a role in their activation, thereby aggravating non-allergic inflammation and the severity of asthma.

Shift of Neutrophils From Blood to Bone Marrow Upon Extensive Experimental Trauma Surgery.

IntroductionExtensive trauma surgery evokes an immediate cellular immune response including altered circulatory neutrophil numbers. The concurrent bone marrow (BM) response however is currently unclear. We hypothesize that these BM changes include (1) a relative reduction of the bone marrow neutrophil fraction and (2) increasing heterogeneity of the bone marrow neutrophil pool due to (3) the appearance of aged/returning neutrophils from circulation into the BM-compartment.Materials and MethodsEight pigs were included in a standardized extensive trauma surgery model. Blood and bone marrow samples were collected at baseline and after 3 hours of ongoing trauma surgery. Leukocyte and subtype counts and cell surface receptor expression levels were studied by flow cytometry.ResultsAll animals survived the interventions. A significant drop in circulating neutrophil counts from 9.3 to 3.2x106 cells/ml (P=0.001) occurred after intervention, whereas circulatory neutrophil cell surface expression of CD11b increased. The concurrent bone marrow response included an increase of the BM neutrophil fraction from 63 ± 3 to 71 ± 3 percent (P<0.05). Simultaneously, the BM neutrophil pool became increasingly mature with a relative increase of a CXCR4high-neutrophil subtype that was virtually absent at baseline.ConclusionThe current study shows a shift in composition of the BM neutrophil pool during extensive trauma surgery that was associated with a relatively circulatory neutropenia. More specifically, under these conditions BM neutrophils were more mature than under homeostatic conditions and a CXCR4high-neutrophil subset became overrepresented possibly reflecting remigration of aged neutrophils to the BM. These findings may contribute to the development of novel interventions aimed to modify the trauma-induced immune response in the BM.

Abstract WP236: The Gut Microbiome Is Associated With Early Brain Injury After Experimental Subarachnoid Hemorrhage Via Regulation Of Neutrophils

Background: It is known that early brain injury (EBI), which occurs early after the onset of subarachnoid hemorrhage (SAH), largely determines the prognosis of SAH; however, an effective treatment for EBI has not been developed. The outcome of various diseases including ischemic stroke is influenced by the composition of the gut microbiome (GM) and we have previously shown that the GM is also associated with the rupture of cerebral aneurysms. Here, we studied whether prior alteration of the GM can prevent EBI after SAH. Methods: We altered the GM of 7-week-old male rats by administering antibiotics-containing water for 2 weeks or performing fecal microbiome transplantation after antibiotics treatment. The composition of the GM was profiled using 16S rRNA. SAH was induced by injection of blood in the subarachnoid space of control rats and rats with altered GM. We evaluated EBI indicators such as the neurological score, brain water content, Evans blue extravasation, and neuronal apoptosis. We also studied inflammatory cells using immunohistochemistry, immunocytochemistry, quantitative PCR and flow cytometry. Results: There were significant differences in alpha and beta diversity between the control and antibiotics-treated rats. Compared with the control rats, the antibiotics-treated rats had significantly reduced neurological symptoms, brain edema, blood-brain barrier disruption and apoptosis. Among inflammatory cells, neutrophil infiltration into the brain was significantly reduced in the antibiotics-treated rats. Interestingly, the preventive effect on neutrophil infiltration was already observed immediately after SAH. Next, the immune status of each group before SAH onset was examined. There was no difference in circulating neutrophil counts, but flow cytometry showed a significant decrease in aged neutrophils which exhibit a highly reactive phenotype in the antibiotics-treated rats. Finally, the neuroprotective effect of antibiotics against EBI disappeared when the GM was recolonized. Conclusions: Our findings suggest that the GM affects the severity of EBI after SAH by regulating neutrophils. In the future, adjustment of the GM to prevent EBI may become a new strategy in patients who are being followed for cerebral aneurysms.

MicroLet-7b Regulates Neutrophil Function and Dampens Neutrophilic Inflammation by Suppressing the Canonical TLR4/NF-κB Pathway.

Sepsis is a heterogeneous syndrome caused by a dysregulated host response during the process of infection. Neutrophils are involved in the development of sepsis due to their essential role in host defense. COVID-19 is a viral sepsis. Disfunction of neutrophils in sepsis has been described in previous studies, however, little is known about the role of microRNA-let-7b (miR-let-7b), toll-like receptor 4 (TLR4), and nuclear factor kappa B (NF-κB) activity in neutrophils and how they participate in the development of sepsis. In this study, we investigated the regulatory pathway of miR-let-7b/TLR4/NF-κB in neutrophils. We also explored the downstream cytokines released by neutrophils following miR-let-7b treatment and its therapeutic effects in cecal ligation and puncture (CLP)-induced septic mice. Six-to-eight-week-old male C57BL/6 mice underwent CLP following treatment with miR-let-7b agomir. Survival (n=10), changes in liver and lungs histopathology (n=4), circulating neutrophil counts (n=4), the liver-body weight ratio (n=4–7), and the lung wet-to-dry ratio (n=5–6) were recorded. We found that overexpression of miR-let-7b could significantly down-regulate the expression of human-derived neutrophilic TLR4 at a post-transcriptional level, a decreased level of proinflammatory factors including interleukin-6 (IL-6), IL-8, tumor necrosis factor α (TNF-α), and an upregulation of anti-inflammatory factor IL-10 in vitro. After miR-let-7b agomir treatment in vivo, neutrophil recruitment was inhibited and thus the injuries of liver and lungs in CLP-induced septic mice were alleviated (p=0.01 and p=0.04, respectively), less weight loss was reduced, and survival in septic mice was also significantly improved (p=0.013). Our study suggested that miR-let-7b could be a potential target of sepsis.

Neutrophils promote tumor resistance to radiation therapy

Nearly two-thirds of cancer patients are treated with radiation therapy (RT), often with the intent to achieve complete and permanent tumor regression (local control). RT is the primary treatment modality used to achieve local control for many malignancies, including locally advanced cervical cancer, head and neck cancer, and lung cancer. The addition of concurrent platinum-based radiosensitizing chemotherapy improves local control and patient survival. Enhanced outcomes with concurrent chemoradiotherapy may result from increased direct killing of tumor cells and effects on nontumor cell populations. Many patients treated with concurrent chemoradiotherapy exhibit a decline in neutrophil count, but the effects of neutrophils on radiation therapy are controversial. To investigate the clinical significance of neutrophils in the response to RT, we examined patient outcomes and circulating neutrophil counts in cervical cancer patients treated with definitive chemoradiation. Although pretreatment neutrophil count did not correlate with outcome, lower absolute neutrophil count after starting concurrent chemoradiotherapy was associated with higher rates of local control, metastasis-free survival, and overall survival. To define the role of neutrophils in tumor response to RT, we used genetic and pharmacological approaches to deplete neutrophils in an autochthonous mouse model of soft tissue sarcoma. Neutrophil depletion prior to image-guided focal irradiation improved tumor response to RT. Our results indicate that neutrophils promote resistance to radiation therapy. The efficacy of chemoradiotherapy may depend on the impact of treatment on peripheral neutrophil count, which has the potential to serve as an inexpensive and widely available biomarker.

Neutrophils and neutrophil extracellular traps enhance venous thrombosis in mice bearing human pancreatic tumors.

Pancreatic cancer is associated with a high incidence of venous thromboembolism. Neutrophils have been shown to contribute to thrombosis in part by releasing neutrophil extracellular traps (NET). A recent study showed that increased plasma levels of the NET biomarker, citrullinated histone H3 (H3Cit), are associated with venous thromboembolism in patients with pancreatic and lung cancer but not in those with other types of cancer, including breast cancer. In this study, we examined the contribution of neutrophils and NET to venous thrombosis in nude mice bearing human pancreatic tumors. We found that tumor-bearing mice had increased circulating neutrophil counts and levels of granulocyte-colony stimulating factor, neutrophil elastase, H3Cit and cell-free DNA compared with controls. In addition, thrombi from tumor-bearing mice contained increased levels of the neutrophil marker Ly6G, as well as higher levels of H3Cit and cell-free DNA. Thrombi from tumor-bearing mice also had denser fibrin with thinner fibers consistent with increased thrombin generation. Importantly, either neutrophil depletion or administration of DNase I reduced the thrombus size in tumor-bearing but not in control mice. Our results, together with clinical data, suggest that neutrophils and NET contribute to venous thrombosis in patients with pancreatic cancer.

Population Pharmacokinetic (PK)/Pharmacodynamic (PD) Modeling of Myelosuppression in Patients with Hematologic Malignancies for CPX-351 and Standard-of-Care 7+3 Therapy

CPX-351 (Vyxeos®), a liposomal encapsulation of cytarabine and daunorubicin at a synergistic ratio, has demonstrated a significant survival benefit vs standard 7+3 in patients (pts) with high-risk/secondary AML. A population PK/PD analysis assessed the correlation between cytarabine and daunorubicin plasma concentrations and myelosuppressive effects (neutropenia, thrombocytopenia) of CPX-351 and 7+3.The PK/PD population for model development included pts with advanced hematologic malignancies from 3 clinical studies. For CPX-351 and 7+3, respectively, 129 and 79 pts were included in the final neutropenia PK/PD analysis and 137 and 86 pts were included in the final thrombocytopenia PK/PD analysis. For the neutropenia model, median age and body weight were 67 y (range: 23-81) and 78.7 kg (39.5-156.5) for CPX-351 and 68 y (60-75) and 83.0 kg (53.9-136.0) for 7+3.PK/PD analyses were conducted using nonlinear mixed-effects modeling in NONMEM. Pt-specific PK profiles were simulated using previously developed population PK models for CPX-351 and 7+3. Blood cell dynamics were described by transit-compartment models with proliferating, maturating, and circulating neutrophils or platelets. The effects of CPX-351 or 7+3 were applied to the proliferation phases of the compartment models by a molar composite PK driver (plasma cytarabine + daunorubicin). Inhibition of proliferation of blood cells by CPX-351 and 7+3 is assumed to be similar, via a sigmoidal Imax function. Co-medication of granulocyte colony stimulating factor (GCSF) or platelet infusion was accounted for during model development. Covariates (eg, demographics, clinical laboratory measures, disease status) were evaluated. Model evaluation and selection were assessed using a standard model discrimination process that included statistical criteria (eg, objective function value) and graphical representations of goodness-of-fit.In the final neutrophil PK/PD models, baseline circulating neutrophil counts were similar for CPX-351 (3.55 × 109/L) and 7+3 (3.76 × 109/L). Mean transit times (MTT) between maturation compartments were estimated at values of 113 h for CPX-351 and 88 h for 7+3. Effects of GCSF on neutrophil production were assumed to be similar for CPX-351 and 7+3. Both treatments had similar maximum inhibition on neutrophil proliferation, with Imax values around 1. However, estimated IC50 values were very different: 24.9 µM for CPX-351 and 0.0286 µM for 7+3.In the final platelet PK/PD models, baseline circulating platelet counts were the same (98.1 × 109/L) for both CPX-351 and 7+3. The MTTs between each compartment of the maturation processes were 91.2 h for CPX-351 and 120 h for 7+3. Drug-specific parameters for CPX-351 and 7+3, respectively, were as follows: Imax, 0.316 and 1; IC50, 0.324 and 0.0982 µM.To better understand the behavior of the models and parameter estimates, simulations were conducted to evaluate the temporal events of myelosuppression. Model simulations were conducted for 200 pts with characteristics similar to the PK/PD model population. During simulations, no platelet transfusion or GCSF was administered. Pts received CPX-351 100 units/m2 (cytarabine 100 mg/m2 + daunorubicin 44 mg/m2) as a 90-min IV infusion on Days 1, 3 and 5 or 7+3 (cytarabine 100 mg/m2/day IV for 7 days continuously + daunorubicin 60 mg/m2 IV on Days 1-3).Median time to initially observe a blood neutrophil count <0.5 × 109/L was longer following CPX-351 (8.3 d) vs 7+3 (7.4 d) treatment. The median duration with neutrophil counts <0.5 × 109/L was longer with CPX-351 (23 d) vs 7+3 (14 d). The median lowest neutrophil counts were well below 0.2 × 109/L for both CPX-351 (0.007 × 109/L) and 7+3 (0.026 × 109/L).Median time to initially observe a platelet count <50 × 109/L was 6.4 d after CPX-351 and 5.8 d after 7+3, while the median time to an observed platelet count <20 × 109/L was 10.8 d and 8.9 d, respectively. The median duration with platelet counts <20 × 109/L was longer with CPX-351 (18 d) vs 7+3 (8 d), and the median duration of platelet counts <50 × 109/L was 22 d and 15 d, respectively. The median lowest platelet counts were 11.3 × 109/L with CPX-351 and 4.7 × 109/L with 7+3.In summary, the median duration of myelosuppressive effects was longer with CPX-351 than 7+3, and the median time for initial detection of myelosuppression with CPX-351 was 1 to 2 days later than with 7+3, which might affect the clinical monitoring scheme. DisclosuresWang:Jazz Pharmaceuticals: Employment, Equity Ownership. Cook:Jazz Pharmaceuticals: Consultancy. Van Wart:Jazz Pharmaceuticals: Consultancy. Mager:Jazz Pharmaceuticals: Consultancy. Faderl:Jazz Pharmaceuticals: Employment, Equity Ownership.

Distinct neutrophil counts and functions in newly diagnosed type 1 diabetes, latent autoimmune diabetes in adults, and type 2 diabetes

Recent discoveries from animal models demonstrated that neutrophils can induce type 1 diabetes (T1D) through infiltrating into the islets. However, the evidence of their actions in T1D patients is relatively rare, and the change trend of neutrophil numbers and functions in different subtypes of diabetes has not been investigated. Patients with newly diagnosed T1D (n=189), latent autoimmune diabetes in adults (LADA) (n=86), T2D (n=235), and healthy controls (n=709) were enrolled. Circulating neutrophil counts were measured, and their correlations with clinical parameters were analysed. Neutrophils were isolated by density gradient centrifugation and magnetic bead cell sorting method. Neutrophil migration rate and chemokine levels in the blood were explored by trans-well and ELISA, respectively. Neutrophil phagocytosis rate, adhesion molecules and chemokine receptors expression were investigated by flow cytometry. Compared with controls, neutrophil counts decreased in T1D patients but increased in T2D patients, with no change in LADA patients. The numbers showed a gradual increase trend from T1D, LADA to T2D. In autoimmune diabetes, neutrophil counts were associated with the number and titre of positive autoantibodies against β-cell antigens. No difference was found in neutrophil phagocytosis rate, but neutrophil migration in T1D patients was impaired and associated with CD62L expression, which was related closely to the titre of autoantibody. Neutrophil numbers and migration abilities displayed distinct levels in different types of diabetes. In T1D, CD62L seems to play an important role in the migration of neutrophils and β-cell autoimmunity.

Circulating Neutrophil Counts Decrease in Response to Mitigated Air Quality in Stable COPD Patients

This three-year study, based on the Guangzhou Institute of Respiratory Disease (GRID), chronic obstructive pulmonary disease (COPD) Biobank, was conducted in 36 COPD patients to estimate whether changes in levels of leukocytes, erythrocytes, hemoglobin, and platelets were related to changes in air pollutant concentration. Daily NO2 levels exhibited significant differences between baseline years and the 2010 Asian Game period. We observed significant reductions in leukocyte and neutrophils counts levels, by 15.51% and 23.01%, from pre-Asian Games to during-Asian Games, respectively. In the post-Asian Game period, most pollutants approximated pre-Asian Game period levels, and similar effects were demonstrated in leukocyte and neutrophil counts. For both items, we identified significant increases resulting from elevated NO2 at lag days 0-2/5-6. We concluded that reductions in pollutants during the intervention period were associated with inactivation of hematological events in COPD.

A randomised, placebo-controlled trial of anti\u2013interleukin-1 receptor 1 monoclonal antibody MEDI8968 in chronic obstructive pulmonary disease

BackgroundInterleukin-1 receptor 1 (IL-1R1) inhibition is a potential strategy for treating patients with chronic obstructive pulmonary disease (COPD). MEDI8968, a fully human monoclonal antibody, binds selectively to IL-1R1, inhibiting activation by IL-1α and IL-1β. We studied the efficacy and safety/tolerability of MEDI8968 in adults with symptomatic, moderate-to-very severe COPD.MethodsThis was a phase II, randomised, double-blind, placebo-controlled, multicentre, parallel-group study. Subjects aged 45–75 years and receiving standard maintenance therapy with ≥2 exacerbations in the past year were randomised 1:1 to receive placebo or MEDI8968 300 mg (600 mg intravenous loading dose) subcutaneously every 4 weeks, for 52 weeks. The primary endpoint was the moderate/severe acute exacerbations of COPD (AECOPD) rate (week 56 post-randomisation). Secondary endpoints were severe AECOPD rate and St George’s Respiratory Questionnaire-COPD (SGRQ-C) score (week 56 post-randomisation).ResultsOf subjects randomised to placebo (n = 164) and MEDI8968 (n = 160), 79.3% and 75.0%, respectively, completed the study. There were neither statistically significant differences between treatment groups in moderate/severe AECOPD rate ([90% confidence interval]: 0.78 [0.63, 0.96], placebo; 0.71 [0.57, 0.90], MEDI8968), nor in severe AECOPD rate or SGRQ-C scores. Post-hoc analysis of subject subgroups (by baseline neutrophil count or tertiles of circulating neutrophil counts) did not alter the study outcome. The incidence of treatment-emergent adverse events (TEAEs) with placebo and MEDI8968 treatment was similar. The most common TEAE was worsening of COPD.ConclusionsIn this phase II study, MEDI8968 did not produce statistically significant improvements in AECOPD rate, lung function or quality of life.Trial registrationClinicalTrials.gov, NCT01448850, date of registration: 06 October 2011.

Meta-analysis of rare and common exome chip variants identifies S1PR4 and other loci influencing blood cell traits.

Hematologic measures such as hematocrit and white blood cell (WBC) count are heritable and clinically relevant. Erythrocyte and WBC phenotypes were analyzed with Illumina HumanExome BeadChip genotypes in 52,531 individuals (37,775 of European ancestry; 11,589 African Americans; 3,167 Hispanic Americans) from 16 population-based cohorts. We then performed replication analyses of novel discoveries in 18,018 European American women and 5,261 Han Chinese. We identified and replicated four novel erythrocyte trait-locus associations (CEP89, SHROOM3, FADS2, and APOE) and six novel WBC loci for neutrophil count (S1PR4), monocyte count (BTBD8, NLRP12, and IL17RA), eosinophil count (IRF1), and total WBC (MYB). The novel association of a rare missense variant in S1PR4 supports the role of sphingosine-1-phosphate signaling in leukocyte trafficking and circulating neutrophil counts. Loss-of-function experiments of S1pr4 in mouse and zebrafish demonstrated phenotypes consistent with the association observed in humans and altered kinetics of neutrophil recruitment and resolution in response to tissue injury.

A Systemic Inflammatory Endotype of Asthma With More Severe Disease Identified by Unbiased Clustering of the Serum Cytokine Profile.

Asthma is considered as a clinical and molecularly heterogeneous disorder. Systemic inflammation is suggested to play an important role in a group of asthma patients. We hypothesized that there is a subgroup of patients with asthma characterized by systemic inflammation. In this study, we aimed to discriminate asthma subtypes based on circulating biomarkers and to determine whether a systemic inflammatory endotype of asthma could be identified. In the present cross-sectional study, 50 patients with untreated asthma were prospectively recruited from a single academic outpatient clinic, and characterized with respect to clinical, functional, and inflammatory parameters. The expression profiles of 20 serum cytokines were assessed by anti-human cytokine antibody array. Then, hierarchical clustering analysis was performed based on principal component analysis (PCA)-transformed data to classify the clinical groups. PCA showed that 6 independent components accounted for 80.113% of the variance, and PCA-based hierarchical clustering identified 3 endotypes. One of the endotypes was evidenced by elevated systemic inflammation markers such as leptin, vascular endothelial growth factor (VEGF), and reduced levels of soluble receptor for advanced glycation end products (sRAGE), an anti-inflammatory molecule. More female patients were included, with higher circulating neutrophil counts and more severe symptoms. In conclusion, we identified an endotype of asthma characterized by systemic inflammation and severe symptoms. Increased levels of VEGF, leptin and decreased level of sRAGE may contribute to the systemic inflammation of this asthma endotype.