The role of human CD46 in early xenoislet engraftment in a dual transplant model.

BACKGROUND: Membrane cofactor protein CD46 attenuates the complement cascade by facilitating cleavage of C3b and C4b. In solid organ xenotransplantation, organs expressing CD46 have been shown to resist hyperacute rejection. However, the incremental value of human CD46 expression for islet xenotransplantation remains poorly defined. METHODS: This study attempted to delineate the role of CD46 in early neonatal porcine islet engraftment by comparing Gal-knocked out (GKO) and hCD46-transgenic (GKO/CD46) islets in a dual transplant model. Seven rhesus macaques underwent dual transplant and were sacrificed at 1 hour (n = 4) or 24 hours (n = 3). Both hemilivers were recovered and fixed for immunohistochemistry (CD46, insulin, neutrophil elastase, platelet, IgM, IgG, C3d, C4d, CD68, Caspase 3). Quantitative immunohistochemical analysis was performed using the Aperio Imagescope. RESULTS: Within 1 hour of intraportal infusion of xenografts, no differences were observed between the two types of islets in terms of platelet, antibody, or complement deposition. Cellular infiltration and islet apoptotic activity were also similar at 1 hour. At 24 hours, GKO/CD46 islets demonstrated significantly less platelet deposition (P = 0.01) and neutrophil infiltration (P = 0.01) compared to GKO islets. In contrast, C3d (P = 0.38) and C4d (P = 0.45) deposition was equal between the two genotypes. CONCLUSIONS: Our findings suggest that expression of hCD46 on NPIs potentially provides a measurable incremental survival advantage in vivo by reducing early thrombo-inflammatory events associated with instant blood-mediated inflammatory reaction (IBMIR) following intraportal islet infusion.

The Priority of Administering HIV Postexposure Prophylaxis in Cases of Sexual Assault in an Emergency Department.

INTRODUCTION: Contracting the human immunodeficiency virus (HIV) is a genuine concern for sexually assaulted patients. Emergency departments are a place where sexually assaulted patients seek care, including treatment to prevent HIV. Prompt administration of nonoccupational postexposure prophylaxis is essential because of the time-sensitive nature of the medications. Quality improvement measures at an urban hospital revealed delays in administration of postexposure prophylaxis to these patients. METHODS: A forensic simulation course and checklist was developed for emergency departments to improve care for sexually assaulted patients. Data used for analysis included time of administration of nonoccupational postexposure medication and length of stay before and after intervention with the simulation course and checklist. Points of measurement included student t-test to assess any significant differences and regression analysis to determine associations. RESULTS: When comparing differences between time of nonoccupational postexposure before and after intervention, there was a trend toward improving the time of administration, but it was not found to be significant. Before intervention, an association was found with sexually assaulted patients' lengths of stay and the time that nonoccupational postexposure medication was administered, with a regression equation of R2, 0.76. After intervention, this association was absent, with an R2 of 0.017. DISCUSSION: Implementing a simulation course and checklist for emergency nurses in caring for sexually assaulted patients helps to improve the timeliness of administration of nonoccupational postexposure medications and resolve the association between the length of stay and time of administration of medication.

The Challenge of Emergency Abortion Care Following the Dobbs Ruling.

This Viewpoint discusses how limited or blocked access to legal abortion will affect the provision of emergency and critical care, including negative effects on patient health, legal intrusion into the patient-physician decision-making process, and concerns about legal jeopardy.

A small heat shock protein is essential for thermotolerance and intracellular survival of Leishmania donovani.

Leishmania parasites must survive and proliferate in two vastly different environments - the guts of poikilothermic sandflies and the antigen-presenting cells of homeothermic mammals. The change of temperature during the transmission from sandflies to mammals is both a key trigger for the progression of their life cycle and for elevated synthesis of heat shock proteins, which have been implicated in their survival at higher temperatures. Although the functions of the main heat shock protein families in the Leishmania life cycle have been studied, nothing is known about the roles played by small heat shock proteins. Here, we present the first evidence for the pivotal role played by the Leishmania donovani 23-kDa heat shock protein (which we called HSP23), which is expressed preferentially during the mammalian stage where it assumes a perinuclear localisation. Loss of HSP23 causes increased sensitivity to chemical stressors and renders L. donovani non-viable at 37°C. Consequently, HSP23-null mutants are non-infectious to primary macrophages in vitro. All phenotypic effects could be abrogated by the introduction of a functional HSP23 transgene into the null mutant, confirming the specificity of the mutant phenotype. Thus, HSP23 expression is a prerequisite for L. donovani survival at mammalian host temperatures and a crucial virulence factor.

Phenotypic Characterization of a Leishmania donovani Cyclophilin 40 Null Mutant.

Protozoan parasites of the genus Leishmania adapt to their arthropod and vertebrate hosts through the development of defined life cycle stages. Stage differentiation is triggered by environmental stress factors and has been linked to parasite chaperone activities. Using a null mutant approach we previously revealed important, nonredundant functions of the cochaperone cyclophilin 40 in L. donovani-infected macrophages. Here, we characterized in more detail the virulence defect of cyp40-/- null mutants. In vitro viability assays, infection tests using macrophages, and mixed infection experiments ruled out a defect of cyp40-/- parasites in resistance to oxidative and hydrolytic stresses encountered inside the host cell phagolysosome. Investigation of the CyP40-dependent proteome by quantitative 2D-DiGE analysis revealed up regulation of various stress proteins in the null mutant, presumably a response to compensate for the lack of CyP40. Applying transmission electron microscopy we showed accumulation of vesicular structures in the flagellar pocket of cyp40-/- parasites that we related to a significant increase in exosome production, a phenomenon previously linked to the parasite stress response. Together these data suggest that cyp40-/- parasites experience important intrinsic homeostatic stress that likely abrogates parasite viability during intracellular infection.

The Leishmania donovani chaperone cyclophilin 40 is essential for intracellular infection independent of its stage-specific phosphorylation status.

During its life cycle, the protozoan pathogen Leishmania donovani is exposed to contrasting environments inside insect vector and vertebrate host, to which the parasite must adapt for extra- and intracellular survival. Combining null mutant analysis with phosphorylation site-specific mutagenesis and functional complementation we genetically tested the requirement of the L. donovani chaperone cyclophilin 40 (LdCyP40) for infection. Targeted replacement of LdCyP40 had no effect on parasite viability, axenic amastigote differentiation, and resistance to various forms of environmental stress in culture, suggesting important functional redundancy to other parasite chaperones. However, ultrastructural analyses and video microscopy of cyp40-/- promastigotes uncovered important defects in cell shape, organization of the subpellicular tubulin network and motility at stationary growth phase. More importantly, cyp40-/- parasites were unable to establish intracellular infection in murine macrophages and were eliminated during the first 24 h post infection. Surprisingly, cyp40-/- infectivity was restored in complemented parasites expressing a CyP40 mutant of the unique S274 phosphorylation site. Together our data reveal non-redundant CyP40 functions in parasite cytoskeletal remodelling relevant for the development of infectious parasites in vitro independent of its phosphorylation status, and provide a framework for the genetic analysis of Leishmania-specific phosphorylation sites and their role in regulating parasite protein function.

Association of RBC Transfusion Thresholds and Outcomes in Medical Patients With Acute Respiratory Failure Supported With Extracorporeal Membrane Oxygenation: A Single-Center Retrospective Cohort Study.

BACKGROUND: The hemoglobin value to trigger RBC transfusion for patients receiving venovenous extracorporeal membrane oxygenation (ECMO) is controversial. Previous guidelines recommended transfusing to a normal hemoglobin level, but recent studies suggest that more RBC transfusions are associated with increased adverse outcomes. RESEARCH QUESTION: Is implementation of different institutional RBC transfusion thresholds for patients receiving venovenous ECMO associated with changes in RBC use and patient outcomes? STUDY DESIGN AND METHODS: This single-center retrospective study of patients receiving venovenous ECMO used segmented regression to test associations between implementation of institutional transfusion thresholds and trends in RBC use. Associations with secondary outcomes, including in-hospital survival, also were assessed. RESULTS: The study included 229 patients: 91 in the no threshold cohort, 48 in the hemoglobin < 8 g/dL cohort, and 90 in the hemoglobin < 7 g/dL cohort. Despite a decrease in number of RBC units transfused per day of ECMO support after implementation of different thresholds (mean ± SD: 0.6 ± 1.0 in the no threshold cohort, 0.3 ± 0.8 in the hemoglobin < 8 g/dL cohort, and 0.3 ± 1.1 in the hemoglobin < 7 g/dL cohort; P < .001), segmented regression showed no association between implementation of transfusion thresholds and changes in trends in number of RBC units per day of ECMO. We observed an increased hazard of death in the no threshold cohort compared with the hemoglobin < 8 g/dL cohort (adjusted hazard ratio [aHR], 2.08; 95% CI, 1.12-3.88) and in the hemoglobin < 7 g/dL cohort compared with the hemoglobin < 8 g/dL cohort (aHR, 1.93; 95% CI, 1.02-3.62). No difference was found in the hazard of death between the no threshold and hemoglobin < 7 g/dL cohorts (aHR, 1.08; 95% CI, 0.69-1.69). INTERPRETATION: We observed a decrease in number of RBC units per day of ECMO over time, but changes were not associated temporally with implementation of transfusion thresholds. A transfusion threshold of hemoglobin < 8 g/dL was associated with a lower hazard of death, but these findings are limited by study methodology. Further research is needed to investigate optimal RBC transfusion practices for patients supported with venovenous ECMO.

Importance of duodenal bulb biopsies in children for diagnosis of celiac disease in clinical practice.

BACKGROUND: The patchy nature of villous lesion in celiac disease is increasingly being recognized. Current guidelines recommend four endoscopic duodenal mucosal biopsies from the second or more distal part of the duodenum to confirm the diagnosis of celiac disease. The purpose of the study was to investigate the usefulness of duodenal bulb mucosal biopsies in confirming the diagnosis of celiac disease in everyday clinical practice. METHODS: All patients with a positive tissue-transglutaminase antibody requiring biopsy-confirmation of celiac disease over a two-year period were studied. Two endoscopic biopsies were taken from the duodenal bulb and four biopsies from the second (or distal) part of the duodenum. RESULTS: Thirty-five patients were included, mean age 8.1 (+/- 4.7) years. Thirty-one (88.6%) patients had abnormal distal duodenal biopsies, one had Marsh type 1, one had Marsh type 2 and twenty-nine had Marsh type 3 lesion. All but two patients with abnormal distal duodenal biopsies also had abnormal bulb biopsies. Four (11.4%) patients had normal distal duodenal biopsies but abnormal bulb biopsies. Of these, one patient had Marsh type 2 and three had Marsh type 3 lesion. The distal duodenum was also grossly normal in these four patients. The histological diagnosis of celiac disease would not have been possible in these four cases with distal duodenal biopsies only. CONCLUSION: The lesion in celiac disease in children can be patchy with duodenal bulb mucosa being the only area showing histological changes. The recommendations regarding the site of biopsies should be revised to include biopsies not only from distal duodenum but also from bulb to improve the diagnostic yield.

Electronic cigarettes and cardiovascular health: what do we know so far?

While tobacco cigarette (TC) smoking has continued to drop to all-time lows, the use of electronic cigarettes (ECs), introduced in the US in 2007, has been rising dramatically, especially among youth. In EC emissions, nicotine is the major biologically active element, while levels of carcinogens and harmful combustion products that typify TC smoke are very low or even undetectable. TCs cause cardiovascular harm by activation of inflammatory pathways and oxidative damage, leading to atherogenesis and thrombosis, as well as through sympathetic activation triggering ischemia and arrhythmia. While ECs are generally believed to be safer than TCs, there remain many uncertainties regarding the overall cardiovascular health effects of EC usage. In this review, we discuss the various components of EC smoke and review the potential mechanisms of cardiovascular injury caused by EC use. We also discuss the controversy regarding the increasing epidemic of youth EC use weighed against the use of ECs as a smoking-cessation aid.

Mothers' Perceived Barriers to and Recommendations for Health Care Appointment Keeping for Children Who Have Cerebral Palsy.

Children with cerebral palsy (CP) require ongoing rehabilitation services to address complex health care needs. Attendance at appointments ensures continuity of care and improves health and well-being. The study's aim was to gain insight into mothers' perspectives of the factors associated with nonattendance. A qualitative descriptive design was conducted to identify barriers and recommendations for appointment keeping. Semi-structured interviews were conducted with 15 mothers of children with CP. Data underwent inductive qualitative analysis. Mothers provided rich context regarding barriers confronted for appointment keeping-transportation and travel, competing priorities for the child and family, and health services. Mothers' recommendations for improving the experience of attending appointments included virtual care services, transportation support, multimethod scheduling and appointment reminders, extended service hours, and increased awareness among staff of family barriers to attendance. The results inform services/policy strategies to facilitate appointment keeping, thereby promoting access to ongoing rehabilitation services for children with CP.

Leishmania donovani 90 kD Heat Shock Protein - Impact of Phosphosites on Parasite Fitness, Infectivity and Casein Kinase Affinity.

Leishmania parasites are thought to control protein activity at the post-translational level, e.g. by protein phosphorylation. In the pathogenic amastigote, the mammalian stage of Leishmania parasites, heat shock proteins show increased phosphorylation, indicating a role in stage-specific signal transduction. Here we investigate the impact of phosphosites in the L. donovani heat shock protein 90. Using a chemical knock-down/genetic complementation approach, we mutated 11 confirmed or presumed phosphorylation sites and assessed the impact on overall fitness, morphology and in vitro infectivity. Most phosphosite mutations affected the growth and morphology of promastigotes in vitro, but with one exception, none of the phosphorylation site mutants had a selective impact on the in vitro infection of macrophages. Surprisingly, aspartate replacements mimicking the negative charge of phosphorylated serines or threonines had mostly negative impacts on viability and infectivity. HSP90 is a substrate for casein kinase 1.2-catalysed phosphorylation in vitro. While several putative phosphosite mutations abrogated casein kinase 1.2 activity on HSP90, only Ser289 could be identified as casein kinase target by mass spectrometry. In summary, our data show HSP90 as a downstream client of phosphorylation-mediated signalling in an organism that depends on post-transcriptional gene regulation.

Human Trafficking: How Nurses Can Make a Difference.

Human trafficking is a human rights violation and a global health problem. Victims of human trafficking have medical and mental health sequelae requiring specific healthcare interventions. Healthcare professionals may be the initial contact that these victims make outside the world of trafficking. Healthcare professionals are key agents in the identification of human trafficking, which is essential in eliminating this public health problem. Unfortunately, healthcare professionals are not always able to detect signs of human trafficking. Failure to detect results in missed opportunities to assist victims. This is a case report of a victim of human trafficking who presented to an emergency department with medical and mental health issues. Despite numerous encounters with different healthcare professionals, signs and symptoms of human trafficking were not identified. Skilled assessment made by a forensic nurse alerted the healthcare team to clear features of human trafficking associated with this person. Through this case report we illustrate the key role the nurse played in identifying signs of human trafficking. Improvement of human trafficking educational programs is highlighted as a key adjunct to improving detection and facilitating the proper treatment of victims.

Heparan sulfate is a plasma biomarker of acute cellular allograft rejection.

Despite advances in management of immunosuppression, graft rejection remains a significant clinical problem in solid organ transplantation. Non-invasive biomarkers of graft rejection can facilitate earlier diagnosis and treatment of acute rejection. The purpose of this study was to investigate the potential role of heparan sulfate as a novel biomarker for acute cellular rejection. Heparan sulfate is released from the extracellular matrix during T-cell infiltration of graft tissue via the action of the enzyme heparanase. In a murine heart transplant model, serum heparan sulfate is significantly elevated during rejection of cardiac allografts. Moreover, expression of the enzyme heparanase is significantly increased in activated T-cells. In human studies, plasma heparan sulfate is significantly elevated in kidney transplant recipients with biopsy-proven acute cellular rejection compared to healthy controls, recipients with stable graft function, and recipients without acute cellular rejection on biopsy. Taken together, these findings support further investigation of heparan sulfate as a novel biomarker of acute cellular rejection in solid organ transplantation.

T Cell Repertoire Maturation Induced by Persistent and Latent Viral Infection Is Insufficient to Induce Costimulation Blockade Resistant Organ Allograft Rejection in Mice.

CD28:CD80/86 pathway costimulation blockade (CoB) with the CD80/86-specific fusion protein CTLA4-Ig prevents T cell-mediated allograft rejection in mice. However, in humans, transplantation with CoB has been hampered by CoB-resistant rejection (CoBRR). CoBRR has been attributed in part to pathogen-driven T cell repertoire maturation and resultant heterologous alloreactive memory. This has been demonstrated experimentally in mice. However, prior murine models have used viral pathogens, CoB regimens, graft types, and/or antigen systems atypically encountered clinically. We therefore sought to explore whether CoBRR would emerge in a model of virus-induced memory differentiation designed to more closely mimic clinical conditions. Specifically, we examined mouse homologs of clinically prevalent viruses including murine polyomavirus, cytomegalovirus, and gammaherpesvirus 68 in the presence of clinically relevant maintenance CoB regimens using a fully MHC-mismatched, vascularized allograft model. Infected mice developed a significant, sustained increase in effector memory T cells consistent with that seen in humans, but neither developed heterologous alloreactivity nor rejected primarily vascularized heterotopic heart transplants at an increased rate compared with uninfected mice. These results indicate that memory acquisition alone is insufficient to provoke CoBRR and suggest that knowledge of prior latent or persistent viral infection may have limited utility in anticipating heterologous CoB-resistant alloimmunity.

Effect of sex on ethanol consumption and conditioned taste aversion in adolescent and adult rats.

RATIONALE: Vulnerability to alcoholism is determined by many factors, including the balance of pleasurable vs. aversive alcohol-induced sensations: pleasurable sensations increase intake, while aversive sensations decrease it. Female sex and adolescent age are associated with lower sensitivity to intake-reducing effects and more rapid development of alcohol abuse. OBJECTIVES: This study assessed voluntary drinking and the aversive effects of alcohol to determine whether these measures are inversely related across the sexes and development. METHODS: Voluntary drinking of 20 % ethanol in an every-other-day (EOD) availability pattern and the dose-response relationship of ethanol conditioned taste aversion (CTA) were assessed in male and female adolescent and adult rats. RESULTS: CTA was sex specific in adult but not adolescent rats, with adult females exhibiting less aversion. Voluntary ethanol consumption varied according to age and individual differences but was not sex specific. Adolescents initially drank more than adults, exhibited greater day-to-day variation in consumption, were more susceptible to the alcohol deprivation effect, and took longer to establish individual differences in consumption patterns. CONCLUSIONS: These results show that the emergence of intake patterns differs between adolescents and adults. Adolescents as a group initiate drinking at high levels but decrease intake as they mature. A subset of adolescents maintained high drinking levels into adulthood. In contrast, most adults consumed at steady, low levels, but a small subset quickly established and maintained high-consumption patterns. Adolescents also showed marked deprivation-induced increases. Sex differences were not observed in EOD drinking during either adolescence or adulthood.

Psychosis in temporal lobe epilepsy: atypical presentation.

A 55-year-old lady was admitted following a concern raised by family members who had noticed a change in behaviour in terms of declining mood, paranoia with expression of belief that she was being bugged, also reported smelling perfume and after shave lotion. She had a prior diagnosis of bipolar mood disorder and was on lithium but remained no-compliant with her prescribed medication in the 10 weeks before admission. Upon admission, necessary investigations were performed including CT scan and EEG. Her CT was normal but EEG was grossly abnormal. Neurology consultation was sought and a sleep deprived EEG and MRI was ordered. Meanwhile, carbamezipine was commenced in view of the change in diagnosis to that of temporal lobe epilepsy. The patient responded very well to this regime and improved in all spheres. She was finally discharged with follow-up appointments with both neurology and psychiatry clinics.

Inhibition of HSP90 in Trypanosoma cruzi induces a stress response but no stage differentiation.

The 90-kDa heat shock proteins (HSP90) are important in the regulation of numerous intracellular processes in eukaryotic cells. In particular, HSP90 has been shown to be involved in the control of the cellular differentiation of the protozoan parasite Leishmania donovani. We investigated the role of HSP90 in the related parasite Trypanosoma cruzi by inhibiting its function using geldanamycin (GA). GA induced a dose-dependent increase in heat shock protein levels and a dose-dependent arrest of proliferation. Epimastigotes were arrested in G(1) phase of the cell cycle, but no stage differentiation occurred. Blood form trypomastigotes showed conversion towards spheromastigote-like forms when they were cultivated with GA, but differentiation into epimastigotes was permanently blocked. We conclude that, similar to leishmanial HSP90, functional HSP90 is essential for cell division in T. cruzi and serves as a feedback inhibitor in the cellular stress response. In contrast to L. donovani cells, however, T. cruzi cells treated with GA do not begin to differentiate into relevant life cycle stages.