A group concept mapping and ethnographic study of intensive care rehabilitation culture.

BACKGROUND: Early physical activity and physical rehabilitation are advocated in the critical care unit for patients recovering from critical illness. Despite this, there are still many factors associated with implementation of early physical rehabilitation into routine critical care and practice. One such factor that has been consistently identified is unit culture, yet there is little understanding of how or why the culture of a critical care unit impacts on implementation of early rehabilitation. AIM: To develop a detailed understanding of the cultural barriers and enablers to the promotion and implementation of physical activity and early mobilization in National Health Service (NHS) critical care units in the United Kingdom (UK). STUDY DESIGN: A mixed-methods, two-phase study incorporating online group concept mapping (GCM) and ethnography. GCM will be conducted to provide a multistakeholder co-authored conceptual framework of rehabilitation culture. Ethnographic observations and interviews will be conducted of culture and behaviours in relation to the implementation and promotion of early physical activity and rehabilitation in two NHS critical care units in the North East of England. RESULTS: The results of the Group Concept Mapping and ethnographic observations and interviews will be triangulated to develop a contextual framework of rehabilitation culture in critical care. RELEVANCE TO CLINICAL PRACTICE: This study will provide a detailed understanding of barriers and facilitators in relation to providing a positive rehabilitation culture in the critical care unit.

Phosphoinositide 3-Kinase δ Inhibition Improves Neutrophil Bacterial Killing in Critically Ill Patients at High Risk of Infection.

Acquired neutrophil dysfunction frequently develops during critical illness, independently increasing the risk for intensive care unit-acquired infection. PI3Kδ is implicated in driving neutrophil dysfunction and can potentially be targeted pharmacologically. The aims of this study were to determine whether PI3Kδ inhibition reverses dysfunction in neutrophils from critically ill patients and to describe potential mechanisms. Neutrophils were isolated from blood taken from critically ill patients requiring intubation and mechanical ventilation, renal support, or blood pressure support. In separate validation experiments, neutrophil dysfunction was induced pharmacologically in neutrophils from healthy volunteers. Phagocytosis and bacterial killing assays were performed, and activity of RhoA and protein kinase A (PKA) was assessed. Inhibitors of PI3Kδ, 3-phosphoinositide-dependent protein kinase-1 (PDK1), and PKA were used to determine mechanisms of neutrophil dysfunction. Sixty-six patients were recruited. In the 27 patients (40.9%) with impaired neutrophil function, PI3Kδ inhibition consistently improved function and significantly increased bacterial killing. These findings were validated in neutrophils from healthy volunteers with salbutamol-induced dysfunction and extended to demonstrate that PI3Kδ inhibition restored killing of clinical isolates of nine pathogens commonly associated with intensive care unit-acquired infection. PI3Kδ activation was associated with PDK1 activation, which in turn phosphorylated PKA, which drove phosphorylation and inhibition of the key regulator of neutrophil phagocytosis, RhoA. These data indicate that, in a significant proportion of critically ill patients, PI3Kδ inhibition can improve neutrophil function through PDK1- and PKA-dependent processes, suggesting that therapeutic use of PI3Kδ inhibitors warrants investigation in this setting.

Atovaquone Suppresses Triple-Negative Breast Tumor Growth by Reducing Immune-Suppressive Cells.

A major contributing factor in triple-negative breast cancer progression is its ability to evade immune surveillance. One mechanism for this immunosuppression is through ribosomal protein S19 (RPS19), which facilitates myeloid-derived suppressor cells (MDSCs) recruitment in tumors, which generate cytokines TGF-ß and IL-10 and induce regulatory T cells (Tregs), all of which are immunosuppressive and enhance tumor progression. Hence, enhancing the immune system in breast tumors could be a strategy for anticancer therapeutics. The present study evaluated the immune response of atovaquone, an antiprotozoal drug, in three independent breast-tumor models. Our results demonstrated that oral administration of atovaquone reduced HCC1806, CI66 and 4T1 paclitaxel-resistant (4T1-PR) breast-tumor growth by 45%, 70% and 42%, respectively. MDSCs, TGF-ß, IL-10 and Tregs of blood and tumors were analyzed from all of these in vivo models. Our results demonstrated that atovaquone treatment in mice bearing HCC1806 tumors reduced MDSCs from tumor and blood by 70% and 30%, respectively. We also observed a 25% reduction in tumor MDSCs in atovaquone-treated mice bearing CI66 and 4T1-PR tumors. In addition, a decrease in TGF-ß and IL-10 in tumor lysates was observed in atovaquone-treated mice with a reduction in tumor Tregs. Moreover, a significant reduction in the expression of RPS19 was found in tumors treated with atovaquone.

Effect of Stress Ulcer Prophylaxis With Proton Pump Inhibitors vs Histamine-2 Receptor Blockers on In-Hospital Mortality Among ICU Patients Receiving Invasive Mechanical Ventilation: The PEPTIC Randomized Clinical Trial.

Importance: Proton pump inhibitors (PPIs) or histamine-2 receptor blockers (H2RBs) are often prescribed for patients as stress ulcer prophylaxis drugs in the intensive care unit (ICU). The comparative effect of these drugs on mortality is unknown. Objective: To compare in-hospital mortality rates using PPIs vs H2RBs for stress ulcer prophylaxis. Design, Setting, and Participants: Cluster crossover randomized clinical trial conducted at 50 ICUs in 5 countries between August 2016 and January 2019. Patients requiring invasive mechanical ventilation within 24 hours of ICU admission were followed up for 90 days at the hospital. Interventions: Two stress ulcer prophylaxis strategies were compared (preferential use with PPIs vs preferential use with H2RBs). Each ICU used each strategy sequentially for 6 months in random order; 25 ICUs were randomized to the sequence with use of PPIs and then use of H2RBs and 25 ICUs were randomized to the sequence with use of H2RBs and then use of PPIs (13 436 patients randomized by site to PPIs and 13 392 randomized by site to H2RBs). Main Outcomes and Measures: The primary outcome was all-cause mortality within 90 days during index hospitalization. Secondary outcomes were clinically important upper gastrointestinal bleeding, Clostridioides difficile infection, and ICU and hospital lengths of stay. Results: Among 26 982 patients who were randomized, 154 opted out, and 26 828 were analyzed (mean [SD] age, 58 [17.0] years; 9691 [36.1%] were women). There were 26 771 patients (99.2%) included in the mortality analysis; 2459 of 13 415 patients (18.3%) in the PPI group died at the hospital by day 90 and 2333 of 13 356 patients (17.5%) in the H2RB group died at the hospital by day 90 (risk ratio, 1.05 [95% CI, 1.00 to 1.10]; absolute risk difference, 0.93 percentage points [95% CI, -0.01 to 1.88] percentage points; P = .054). An estimated 4.1% of patients randomized by ICU site to PPIs actually received H2RBs and an estimated 20.1% of patients randomized by ICU site to H2RBs actually received PPIs. Clinically important upper gastrointestinal bleeding occurred in 1.3% of the PPI group and 1.8% of the H2RB group (risk ratio, 0.73 [95% CI, 0.57 to 0.92]; absolute risk difference, -0.51 percentage points [95% CI, -0.90 to -0.12 percentage points]; P = .009). Rates of Clostridioides difficile infection and ICU and hospital lengths of stay were not significantly different by treatment group. One adverse event (an allergic reaction) was reported in 1 patient in the PPI group. Conclusions and Relevance: Among ICU patients requiring mechanical ventilation, a strategy of stress ulcer prophylaxis with use of proton pump inhibitors vs histamine-2 receptor blockers resulted in hospital mortality rates of 18.3% vs 17.5%, respectively, a difference that did not reach the significance threshold. However, study interpretation may be limited by crossover in the use of the assigned medication. Trial Registration: anzctr.org.au Identifier: ACTRN12616000481471.

Excess Mucin Impairs Subglottic Epithelial Host Defense in Mechanically Ventilated Patients.

RATIONALE: Aspiration of infective subglottic secretions causes ventilator-associated pneumonia (VAP) in mechanically ventilated patients. Mechanisms underlying subglottic colonization in critical illness have not been defined, limiting strategies for targeted prevention of VAP. OBJECTIVES: To characterize subglottic host defense dysfunction in mechanically ventilated patients in the ICU; to determine whether subglottic mucin contributes to neutrophil phagocytic impairment and bacterial growth. METHODS: Prospective subglottic sampling in mechanically ventilated patients (intubated for four or more days), and newly intubated control patients (intubated for less than 30 min); isolation and culture of primary subglottic epithelial cells from control patients; laboratory analysis of host innate immune defenses. MEASUREMENTS AND MAIN RESULTS: Twenty-four patients in the ICU and 27 newly intubated control patients were studied. Subglottic ICU samples had significantly reduced microbiological diversity and contained potential respiratory pathogens. The subglottic microenvironment in the ICU was characterized by neutrophilic inflammation, significantly increased proinflammatory cytokines and neutrophil proteases, and altered physical properties of subglottic secretions, including accumulation of mucins. Subglottic mucin from ICU patients impaired the capacity of neutrophils to phagocytose and kill bacteria. Phagocytic impairment was reversible on treatment with a mucolytic agent. Subglottic mucus promoted growth and invasion of bacterial pathogens in a novel air-liquid interface model of primary human subglottic epithelium. CONCLUSIONS: Mechanical ventilation in the ICU is characterized by substantial mucin secretion and neutrophilic inflammation. Mucin impairs neutrophil function and promotes bacterial growth. Mucolytic agents reverse mucin-mediated neutrophil dysfunction. Enhanced mucus disruption and removal has potential to augment preventive benefits of subglottic drainage.

Intensive versus standard physical rehabilitation therapy in the critically ill (EPICC): a multicentre, parallel-group, randomised controlled trial.

BACKGROUND: Early physical rehabilitation in the intensive care unit (ICU) has been shown to improve short-term clinical outcomes but long-term benefit has not been proven and the optimum intensity of rehabilitation is not known. METHODS: We conducted a randomised, parallel-group, allocation-concealed, assessor-blinded, controlled trial in patients who had received at least 48 hours of invasive or non-invasive ventilation. Participants were randomised in a 1:1 ratio, stratified by admitting ICU, admission type and level of independence. The intervention group had a target of 90 min physical rehabilitation per day, the control group a target of 30 min per day (both Monday to Friday). The primary outcome was the Physical Component Summary (PCS) measure of SF-36 at 6 months. RESULTS: We recruited 308 participants over 34 months: 150 assigned to the intervention and 158 to the control group. The intervention group received a median (IQR) of 161 (67-273) min of physical rehabilitation on ICU compared with 86 (31-139) min in the control group. At 6 months, 62 participants in the intervention group and 54 participants in the control group contributed primary outcome data. In the intervention group, 43 had died, 11 had withdrawn and 34 were lost to follow-up, while in the control group, 56 had died, 5 had withdrawn and 43 were lost to follow-up. There was no difference in the primary outcome at 6 months, mean (SD) PCS 37 (12.2) in the intervention group and 37 (11.3) in the control group. CONCLUSIONS: In this study, ICU-based physical rehabilitation did not appear to improve physical outcomes at 6 months compared with standard physical rehabilitation. TRIAL REGISTRATION NUMBER: ISRCTN 20436833.

Randomised controlled trial of GM-CSF in critically ill patients with impaired neutrophil phagocytosis.

BACKGROUND: Critically ill patients with impaired neutrophil phagocytosis have significantly increased risk of nosocomial infection. Granulocyte-macrophage colony-stimulating factor (GM-CSF) improves phagocytosis by neutrophils ex vivo. This study tested the hypothesis that GM-CSF improves neutrophil phagocytosis in critically ill patients in whom phagocytosis is known to be impaired. METHODS: This was a multicentre, phase IIa randomised, placebo-controlled clinical trial. Using a personalised medicine approach, only critically ill patients with impaired neutrophil phagocytosis were included. Patients were randomised 1:1 to subcutaneous GM-CSF (3 µg/kg/day) or placebo, once daily for 4 days. The primary outcome measure was neutrophil phagocytosis 2 days after initiation of GM-CSF. Secondary outcomes included neutrophil phagocytosis over time, neutrophil functions other than phagocytosis, monocyte HLA-DR expression and safety. RESULTS: Thirty-eight patients were recruited from five intensive care units (17 randomised to GM-CSF). Mean neutrophil phagocytosis at day 2 was 57.2% (SD 13.2%) in the GM-CSF group and 49.8% (13.4%) in the placebo group, p=0.73. The proportion of patients with neutrophil phagocytosis≥50% at day 2, and monocyte HLA-DR, appeared significantly higher in the GM-CSF group. Neutrophil functions other than phagocytosis did not appear significantly different between the groups. The most common adverse event associated with GM-CSF was fever. CONCLUSIONS: GM-CSF did not improve mean neutrophil phagocytosis at day 2, but was safe and appeared to increase the proportion of patients with adequate phagocytosis. The study suggests proof of principle for a pharmacological effect on neutrophil function in a subset of critically ill patients.

The Effect of Perioperative Cimetidine Administration on Time to Colorectal Cancer Recurrence.

BACKGROUND: Studies have reported that the perioperative use of cimetidine, a histamine type 2 receptor antagonist, in addition to chemotherapy in patients with lymph node-positive colorectal cancer (CRC) improves the survival. STUDY QUESTION: To determine if time to CRC recurrence could be prolonged with cimetidine. STUDY DESIGN: Cimetidine was prescribed to American Joint on Cancer Committee (AJCC) stage III CRC patients perioperatively. Tumor recurrence was defined as the time (in days) between tumor resection and CRC recurrence. Medical charts of patients diagnosed with CRC between 1996 and 2006 were reviewed. Inclusion criteria were patients with (a) AJCC stage III CRC, (b) who had undergone surgical resection of the tumor, and (c) who received chemotherapy (5-fluorouracil). MEASURES AND OUTCOMES: AJCC stage III CRC patients who did and did not receive cimetidine as part of the treatment regimen were compared with respect to their clinical outcomes using univariate analysis and Kaplan-Meier modeling. RESULTS: Between 1996 and 2006, 38 patients met our inclusion criteria. Twenty-six percent (10/38) received perioperative cimetidine (mean daily dose, 750 mg; mean duration, 369 days; mean total cumulative cimetidine dose, 274,070 mg/d) in addition to chemotherapy. Time to recurrence and cancer deaths were prolonged in the chemotherapy plus cimetidine group compared with the group that received chemotherapy alone (mean ± SD: 1078 ± 290 vs. 446 ± 62; P = 0.03). In addition, we found a significant positive relationship between the duration of cimetidine therapy (days) and survival duration (correlation coefficient = 0.94, P = 0.02) and time until cancer recurrence (correlation coefficient = 0.99, P = 0.04). Moreover, there was a significant positive relationship between the total cumulative cimetidine dose and survival duration (correlation coefficient = 0.92, P = 0.03). CONCLUSIONS: Prolonged duration of cimetidine may be superior to shorter courses in prolonging recurrence of CRC and thus survival.

Reducing the incidence of pressure ulcers in critical care units: a 4-year quality improvement.

QUALITY PROBLEM: Critical care patients often have several risk factors for pressure ulceration and implementing prevention interventions have been shown to decrease risk. INITIAL ASSESSMENT: We identified a high incidence of pressure ulcers in the four adult critical care units in our organization. Therefore, avoiding pressure ulceration was an important quality priority. CHOICE OF SOLUTION: We undertook a quality improvement programme aimed at reducing the incidence of pressure ulceration using an evidence-based bundle approach. IMPLEMENTATION: A bundle of technical and non-technical interventions were implemented supported by clinical leadership on each unit. Important components were evidence appraisals; changes to mattresses; focussed risk assessment alongside mandating patients at very high risk to be repositioned two hourly; and staff training to increase awareness of how to prevent pressure ulcers. EVALUATION: Pressure ulcer numbers, incidence and categories were collected continuously and monitored monthly by unit staff. Pressure ulcer rates reduced significantly from 8.08/100 patient admissions to 2.97/100 patient admissions, an overall relative rate reduction of 63% over 4 years. The greatest reduction was seen in the most severe category of pressure ulceration. The average estimated cost saving was £2.6 million (range £2.1-£3.1). LESSONS LEARNED: A quality improvement programme including technical and non-technical interventions, data feedback to staff and clinical leadership was associated with a sustained reduction in the incidence of pressure ulceration in the critically ill. Strategies used in this programme may be transferable to other critical care units to bring more widespread patient benefit.

Exchange protein directly activated by cyclic AMP (EPAC) activation reverses neutrophil dysfunction induced by ß2-agonists, corticosteroids, and critical illness.

BACKGROUND: Neutrophils play a role in the pathogenesis of asthma, chronic obstructive pulmonary disease, and pulmonary infection. Impaired neutrophil phagocytosis predicts hospital-acquired infection. Despite this, remarkably few neutrophil-specific treatments exist. OBJECTIVES: We sought to identify novel pathways for the restoration of effective neutrophil phagocytosis and to activate such pathways effectively in neutrophils from patients with impaired neutrophil phagocytosis. METHODS: Blood neutrophils were isolated from healthy volunteers and patients with impaired neutrophil function. In healthy neutrophils phagocytic impairment was induced experimentally by using ß2-agonists. Inhibitors and activators of cyclic AMP (cAMP)-dependent pathways were used to assess the influence on neutrophil phagocytosis in vitro. RESULTS: ß2-Agonists and corticosteroids inhibited neutrophil phagocytosis. Impairment of neutrophil phagocytosis by ß2-agonists was associated with significantly reduced RhoA activity. Inhibition of protein kinase A (PKA) restored phagocytosis and RhoA activity, suggesting that cAMP signals through PKA to drive phagocytic impairment. However, cAMP can signal through effectors other than PKA, such as exchange protein directly activated by cyclic AMP (EPAC). An EPAC-activating analog of cAMP (8CPT-2Me-cAMP) reversed neutrophil dysfunction induced by ß2-agonists or corticosteroids but did not increase RhoA activity. 8CPT-2Me-cAMP reversed phagocytic impairment induced by Rho kinase inhibition but was ineffective in the presence of Rap-1 GTPase inhibitors. 8CPT-2Me-cAMP restored function to neutrophils from patients with known acquired impairment of neutrophil phagocytosis. CONCLUSIONS: EPAC activation consistently reverses clinical and experimental impairment of neutrophil phagocytosis. EPAC signals through Rap-1 and bypasses RhoA. EPAC activation represents a novel potential means by which to reverse impaired neutrophil phagocytosis.

Phenethyl isothiocyanate: a comprehensive review of anti-cancer mechanisms.

The epidemiological evidence suggests a strong inverse relationship between dietary intake of cruciferous vegetables and the incidence of cancer. Among other constituents of cruciferous vegetables, isothiocyanates (ITC) are the main bioactive chemicals present. Phenethyl isothiocyanate (PEITC) is present as gluconasturtiin in many cruciferous vegetables with remarkable anti-cancer effects. PEITC is known to not only prevent the initiation phase of carcinogenesis process but also to inhibit the progression of tumorigenesis. PEITC targets multiple proteins to suppress various cancer-promoting mechanisms such as cell proliferation, progression and metastasis. Pre-clinical evidence suggests that combination of PEITC with conventional anti-cancer agents is also highly effective in improving overall efficacy. Based on accumulating evidence, PEITC appears to be a promising agent for cancer therapy and is already under clinical trials for leukemia and lung cancer. This is the first review which provides a comprehensive analysis of known targets and mechanisms along with a critical evaluation of PEITC as a future anti-cancer agent.

Diagnostic accuracy of pulmonary host inflammatory mediators in the exclusion of ventilator-acquired pneumonia.

BACKGROUND: Excessive use of empirical antibiotics is common in critically ill patients. Rapid biomarker-based exclusion of infection may improve antibiotic stewardship in ventilator-acquired pneumonia (VAP). However, successful validation of the usefulness of potential markers in this setting is exceptionally rare. OBJECTIVES: We sought to validate the capacity for specific host inflammatory mediators to exclude pneumonia in patients with suspected VAP. METHODS: A prospective, multicentre, validation study of patients with suspected VAP was conducted in 12 intensive care units. VAP was confirmed following bronchoscopy by culture of a potential pathogen in bronchoalveolar lavage fluid (BALF) at >10(4) colony forming units per millilitre (cfu/mL). Interleukin-1 beta (IL-1ß), IL-8, matrix metalloproteinase-8 (MMP-8), MMP-9 and human neutrophil elastase (HNE) were quantified in BALF. Diagnostic utility was determined for biomarkers individually and in combination. RESULTS: Paired BALF culture and biomarker results were available for 150 patients. 53 patients (35%) had VAP and 97 (65%) patients formed the non-VAP group. All biomarkers were significantly higher in the VAP group (p<0.001). The area under the receiver operator characteristic curve for IL-1ß was 0.81; IL-8, 0.74; MMP-8, 0.76; MMP-9, 0.79 and HNE, 0.78. A combination of IL-1ß and IL-8, at the optimal cut-point, excluded VAP with a sensitivity of 100%, a specificity of 44.3% and a post-test probability of 0% (95% CI 0% to 9.2%). CONCLUSIONS: Low BALF IL-1ß in combination with IL-8 confidently excludes VAP and could form a rapid biomarker-based rule-out test, with the potential to improve antibiotic stewardship.

Pheromone responsiveness is regulated by components of the Gpr1p-mediated glucose sensing pathway in Saccharomyces cerevisiae.

Many fungi have evolved mechanisms to assess environmental nutrient availability prior to the energy-intensive process of mating. In this study, we examined one such system in Saccharomyces cerevisiae, involving a glucose-sensing pathway mediated by Gpr1p and the pheromone-induced mating pathway. Initially we observed that the mating pathway in MATa cells is sensitive to environmental glucose depletion. This phenomenon can be partially reversed with a high glucose spike, but not with the addition of low levels of glucose. Deletion of the low-affinity glucose receptor, Gpr1p, eliminated this glucose-induced recovery of pheromone responsiveness. We then determined the impact of GPR1 deletion on the mating pathway and observed that, in all end points studied, the mating pathway response to pheromone is reduced in the absence of Gpr1p. Similarly, elimination of the Gα for Gpr1p, Gpa2p, resulted in reduction in pheromone sensitivity in all assays studied. The negative effect of removing Gpr1p on mating pathway activation could be recovered by overexpressing the mating receptor, Ste2p. Furthermore, Ste2p levels are reduced in the absence of glucose and GPR1. These data suggest that activity of the GPCR-mediated mating pathway in S. cerevisiae is modulated by extracellular glucose concentrations through the only other GPCR in MATa cells, Gpr1p.

Tucaresol down-modulation of MUC1-stimulated human mononuclear cells.

Interaction between antigen presenting cells and T lymphocytes, through receptors and co-stimulatory molecules present on the surface of these cells, is one of the key means to modulate the adaptive immune system. Tucaresol, a Schiff-base-forming chemical, can be used as a substitute for the physiological donor of carbonyl groups of antigen presenting cells, which can interact with the amine groups of T lymphocytes to modulate the adaptive immune system. This study was done to determine whether tucaresol can enhance killing of cancer cells in vitro as well as protect non-obese diabetic severe combined immunodeficient mice from tumor development by mucin 1 stimulated human mononuclear cells through the adaptive immune system. The expected hypothesis was not supported. Percent specific lysis of MCF-7 tumor cells by mucin 1 stimulated human mononuclear cells was reduced by tucaresol. Furthermore, tucaresol abolished the protective effect of mucin 1 stimulated human mononuclear cells against MCF-7 breast cancer cell growth in non-obese diabetic severe combined immunodeficient mice. This study implies that tucaresol may be of use as an immunosuppressive agent.

Assessment of the comparative agreement between chest radiographs and CT scans in intensive care units.

PURPOSE: Chest radiographs in critically ill patients can be difficult to interpret due to technical and clinical factors. We sought to determine the agreement of chest radiographs and CT scans, and the inter-observer variation of chest radiograph interpretation, in intensive care units (ICUs). METHODS: Chest radiographs and corresponding thoracic computerised tomography (CT) scans (as reference standard) were collected from 45 ICU patients. All radiographs were analysed by 20 doctors (radiology consultants, radiology trainees, ICU consultants, ICU trainees) from 4 different centres, blinded to CT results. Specificity/sensitivity were determined for pleural effusion, lobar collapse and consolidation/atelectasis. Separately, Fleiss' kappa for multiple raters was used to determine inter-observer variation for chest radiographs. RESULTS: The median sensitivity and specificity of chest radiographs for detecting abnormalities seen on CTs scans were 43.2% and 85.9% respectively. Diagnostic sensitivity for pleural effusion was significantly higher among radiology consultants but no specialty/experience distinctions were observed for specificity. Median inter-observer kappa coefficient among assessors was 0.295 ("fair"). CONCLUSIONS: Chest radiographs commonly miss important radiological features in critically ill patients. Inter-observer agreement in chest radiograph interpretation is only "fair". Consultant radiologists are least likely to miss thoracic radiological abnormalities. The consequences of misdiagnosis by chest radiographs remain to be determined.

Tissue factor expression in monocyte subsets during human immunothrombosis, endotoxemia and sepsis.

INTRODUCTION: Tissue factor expression on monocytes is implicated in the pathophysiology of sepsis-induced coagulopathy. How tissue factor is expressed by monocyte subsets (classical, intermediate and non-classical) is unknown. METHODS: Monocytic tissue factor surface expression was investigated during three conditions. Primary human monocytes and microvascular endothelial cell co-cultures were used for in vitro studies. Volunteers received a bolus of lipopolysaccharide (2 ng/kg) to induce endotoxemia. Patients with sepsis, or controls with critical illness unrelated to sepsis, were recruited from four intensive care units. RESULTS: Contact with endothelium and stimulation with lipopolysaccharide reduced the proportion of intermediate monocytes. Lipopolysaccharide increased tissue factor surface expression on classical and non-classical monocytes. Endotoxemia induced profound, transient monocytopenia, along with activation of coagulation pathways. In the remaining circulating monocytes, tissue factor was up-regulated in intermediate monocytes, though approximately 60 % of individuals (responders) up-regulated tissue factor across all monocyte subsets. In critically ill patients, tissue factor expression on intermediate and non-classical monocytes was significantly higher in patients with established sepsis than among non-septic patients. Upon recovery of sepsis, expression of tissue factor increased significantly in classical monocytes. CONCLUSION: Tissue factor expression in monocyte subsets varies significantly during health, endotoxemia and sepsis.

Immunotherapy with IL-10- and IFN-γ-producing CD4 effector cells modulate "Natural" and "Inducible" CD4 TReg cell subpopulation levels: observations in four cases of patients with ovarian cancer.

Adoptive T cell therapy for cancer patients optimally requires participation of CD4 T cells. In this phase I/II study, we assessed the therapeutic effects of adoptively transferred IL-10- and IFN-γ-producing CD4 effector cells in patients with recurrent ovarian cancer. Using MUC1 peptide and IL-2 for ex vivo CD4 effector cell generation, we show that three monthly treatment cycles of autologous T cell restimulation and local intraperitoneal re-infusion-modulated T cell-mediated immune responses that were associated with enhanced patient survival. One patient remains disease-free, another patient experienced prolonged survival for nearly 16 months with recurrent disease, and two patients expired within 3-5 months following final infusion. Prolonged survivors showed elevated levels of systemic CD3(+)CD4(+)CD25(+) and CD3(+)CD4(+)CD25(-) T cells when compared to that of pre-treatment levels and similarly treated short-term survivors. Such cell populations among these patients contained variable levels of "Inducible" Tr1 (CD4(+)CD25(-)FoxP3(-)IL-10(+)) and "Natural" (CD4(+)CD25(+)CD45RO(+)FoxP3(+)) TReg cell numbers and ratios that were associated with prolonged and/or disease-free survival. Moreover, peptide-restimulated T cells from these patients showed an elevation in both IFN-γ production, memory cell phenotype, and select TNF family ligands associated with enhanced T cell survival and apoptosis-inducing activities. This suggests that intraperitoneally administered Th1-like cells, producing elevated levels of IL-10, may require and/or induce differential levels of distinct systemic TReg subpopulations that influence, in part, long-term tumor immunity and enhanced memory/effector CD4-mediated therapeutic potentials. Furthermore, treatment efficacy and enhanced memory cell phenotype did not appear to be dependent on TReg cell numbers but upon ratios of "Inducible" and "Natural" TReg subpopulations.

Blocking epidermal growth factor receptor activation by 3,3'-diindolylmethane suppresses ovarian tumor growth in vitro and in vivo.

Genetic alterations, including the overexpression of epidermal growth factor receptor (EGFR) (in approximately 70% of ovarian tumors), play a crucial role in the signal transduction pathways that regulate key cellular functions, such as cell survival and proliferation, and are responsible for compromising traditional chemotherapy. 3,3'-Diindolylmethane (DIM) is an indole compound present in Brassica vegetables. In our previous studies, we demonstrated that BR-DIM, a formulated version of DIM, suppressed the growth of ovarian cancer cells by causing cell cycle arrest and apoptosis. In the present study, we delineated the mechanism by which DIM suppressed the growth of SKOV-3, OVCAR-3, and TOV-21G human ovarian cancer cells. DIM treatment caused significant down-regulation of the constitutive EGFR protein level as well as phosphorylation of EGFR at Tyr1068, Tyr992, Tyr845, and Tyr1173 in various ovarian cancer cells. To determine whether DIM suppressed the activation of EGFR by activating phosphorylation, cells were treated with epidermal growth factor. Epidermal growth factor treatment significantly blocked the DIM-mediated inhibition of EGFR activation and apoptosis in both SKOV-3 and OVCAR-3 cells. In addition, DIM treatment drastically reduced the phosphorylation of mitogen-activated protein kinase kinase (MEK) and extracellular signal-regulated kinase (ERK), which are downstream to EGFR, without affecting their protein levels. DIM treatment also inhibited the kinase activity of ERK, as observed by the down-regulation of phospho-E twenty-six like transcription factor 1 (p-ELK1) in all three ovarian cancer cell lines. DIM significantly suppressed the growth of ovarian tumors in vivo. Tumor growth suppressive effects of DIM in SKOV-3 tumor xenografts were associated with reduced phosphorylation of EGFR, MEK, and ERK. These results indicate that DIM induces apoptosis in ovarian cancer cells by inhibiting the EGFR-ERK pathway in vitro and in vivo.

Maternal and foetal immune responses of cattle following an experimental challenge with Neospora caninum at day 70 of gestation.

The immune responses of pregnant cattle and their foetuses were examined following inoculation on day 70 of gestation either intravenously (iv) (group 1) or subcutaneously (sc) (group 2) with live NC1 strain tachyzoites or with Vero cells (control) (group 3). Peripheral blood mononuclear cell (PBMC) responses to Neospora antigen and foetal viability were assessed throughout the experiment. Two animals from each group were sacrificed at 14, 28, 42 and 56 days post inoculation (pi). At post mortem, maternal lymph nodes, spleen and PBMC and when possible foetal spleen, thymus and PBMC samples were collected for analysis. Inoculation with NC1 (iv and sc) lead to foetal deaths in all group 1 dams (6/6) and in 3/6 group 2 dams from day 28pi; statistically significant (p ≤ 0.05) increases in cell-mediated immune (CMI) responses including antigen-specific cell proliferation and IFN-γ production as well as increased levels of IL-4, IL-10 and IL-12 were observed in challenged dams compared to the group 3 animals. Lymph node samples from the group 2 animals carrying live foetuses showed greater levels of cellular proliferation as well as significantly (p ≤ 0.05) higher levels of IFN-γ compared to the dams in group 2 carrying dead foetuses. Foetal spleen, thymus and PBMC samples demonstrated cellular proliferation as well as IFN-γ, IL-4, IL-10 and IL-12 production following mitogenic stimulation with Con A from day 14pi (day 84 gestation) onwards. This study shows that the generation of robust peripheral and local maternal CMI responses (lymphoproliferation, IFN-γ) may inhibit the vertical transmission of the parasite.

Efficacy and safety of proton pump inhibitors versus histamine-2 receptor blockers in the cardiac surgical population: insights from the PEPTIC trial.

OBJECTIVES: The comparative effectiveness and safety of proton pump inhibitors (PPIs) versus histamine-2 receptor blockers for stress ulcer prophylaxis in the cardiac surgical intensive care unit population is uncertain. Although the Proton Pump Inhibitors versus Histamine-2 Receptor Blockers for Ulcer Prophylaxis Therapy in the Intensive Care Unit (PEPTIC) trial reported a higher risk of mortality in the PPI arm with no difference in gastrointestinal bleeding, detailed information on surgical variables and clinically relevant surgical subgroups was not available. METHODS: The analysis included all Canadian cardiac surgery patients enrolled in the PEPTIC trial. Data were electronically linked using unique patient identifiers to a clinical information system. Outcomes of interest included in-hospital mortality, gastrointestinal bleeding, Clostridium difficile infections, ventilator-associated conditions and length of stay. RESULTS: We studied 823 (50.6%) randomized to PPIs and 805 (49.4%) to histamine-2-receptor blockers. In the intention-to-treat analysis, there were no differences in hospital mortality [PPI: 4.3% vs histamine-2 receptor blockers: 4.8%, adjusted odds ratio (aOR) 0.97, 95% confidence interval (CI) 0.55-1.70], gastrointestinal bleeding (3.9% vs 4.8%, aOR 1.09, 95% CI 0.66-1.81), C. difficile infections (0.9% vs 0.1%, aOR 0.18, 95% CI 0.02-1.59), ventilator-associated conditions (1.6% vs 1.7%, aOR 0.92, 95% CI 0.85-1.00) or median length of stay (9.2 vs 9.8 days, adjusted risk ratio 1.06, 85% CI 0.99-1.13). No significant treatment differences were observed among subgroups of interest or per-protocol populations. CONCLUSIONS: In a secondary analysis of cardiac surgery patients enrolled in the PEPTIC trial in Canada, no differences in effectiveness or safety were observed between use of PPIs and histamine-2 receptor blockers for stress ulcer prophylaxis. CLINICAL TRIAL REGISTRATION NUMBER: anzctr.org.au identifier: ACTRN12616000481471.