Deregulation of apoptosis-related genes is associated with PRV1 overexpression and JAK2 V617F allele burden in Essential Thrombocythemia and Myelofibrosis

<p>Abstract</p> <p>Background</p> <p>Essential Thrombocythemia (ET) and Primary Myelofibrosis (PMF) are Chronic Myeloproliferative Neoplasms (MPN) characterized by clonal myeloproliferation/myeloaccumulation without cell maturation impairment. The JAK2 V617F mutation and <it>PRV1 </it>gene overexpression may contribute to MPN physiopathology. We hypothesized that deregulation of the apoptotic machinery may also play a role in the pathogenesis of ET and PMF. In this study we evaluated the apoptosis-related gene and protein expression of BCL2 family members in bone marrow CD34⁺hematopoietic stem cells (HSC) and peripheral blood leukocytes from ET and PMF patients. We also tested whether the gene expression results were correlated with JAK2 V617F allele burden percentage, <it>PRV1 </it>overexpression, and clinical and laboratory parameters.</p> <p>Results</p> <p>By real time PCR assay, we observed that <it>A1, MCL1, BIK and BID</it>, as well as <it>A1, BCLW </it>and <it>BAK </it>gene expression were increased in ET and PMF CD34⁺cells respectively, while pro-apoptotic <it>BAX </it>and anti-apoptotic <it>BCL2 </it>mRNA levels were found to be lower in ET and PMF CD34⁺cells respectively, in relation to controls. In patients' leukocytes, we detected an upregulation of anti-apoptotic genes <it>A1, BCL2, BCL-X_L</it>and <it>BCLW</it>. In contrast, pro-apoptotic <it>BID </it>and <it>BIM_EL</it>expression were downregulated in ET leukocytes. Increased BCL-X_Lprotein expression in PMF leukocytes and decreased BID protein expression in ET leukocytes were observed by Western Blot. In ET leukocytes, we found a correlation between JAK2 V617F allele burden and <it>BAX, BIK and BAD </it>gene expression and between <it>A1, BAX </it>and <it>BIK </it>and <it>PRV1 </it>gene expression. A negative correlation between <it>PRV1 </it>gene expression and platelet count was observed, as well as a positive correlation between <it>PRV1 </it>gene expression and splenomegaly.</p> <p>Conclusions</p> <p>Our results suggest the participation of intrinsic apoptosis pathway in the MPN physiopathology. In addition, <it>PRV1 </it>and JAK2 V617F allele burden were linked to deregulation of the apoptotic machinery.</p

Variation in antibiotic prescription rates in febrile children presenting to emergency departments across Europe (MOFICHE) : A multicentre observational study

Funding Information: This project has received funding from the European Union?s Horizon 2020 research and innovation programme under grant agreement No. 668303. The Research was supported by the National Institute for Health Research Biomedical Research Centres at Imperial College London, Newcastle Hospitals NHS Foundation Trust and Newcastle University. The views expressed are those of the author(s) and not necessarily those of the NHS, the NIHR or the Department of Health. For the remaining authors no sources of funding were declared. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. We acknowledge all research nurses for their help in collecting data, and Anda Nagle (Riga) and the Institute of Microbiology at University Medical Centre Ljubljana for their help in collecting data on antimicrobial resistance. Members of the PERFORM consortium are listed in S11 Text. Publisher Copyright: Copyright: © 2020 Hagedoorn et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Copyright: Copyright 2020 Elsevier B.V., All rights reserved.Background The prescription rate of antibiotics is high for febrile children visiting the emergency department (ED), contributing to antimicrobial resistance. Large studies at European EDs covering diversity in antibiotic and broad-spectrum prescriptions in all febrile children are lacking. A better understanding of variability in antibiotic prescriptions in EDs and its relation with viral or bacterial disease is essential for the development and implementation of interventions to optimise antibiotic use. As part of the PERFORM (Personalised Risk assessment in Febrile illness to Optimise Real-life Management across the European Union) project, the MOFICHE (Management and Outcome of Fever in Children in Europe) study aims to investigate variation and appropriateness of antibiotic prescription in febrile children visiting EDs in Europe. Methods and findings Between January 2017 and April 2018, data were prospectively collected on febrile children aged 0–18 years presenting to 12 EDs in 8 European countries (Austria, Germany, Greece, Latvia, the Netherlands [n = 3], Spain, Slovenia, United Kingdom [n = 3]). These EDs were based in university hospitals (n = 9) or large teaching hospitals (n = 3). Main outcomes were (1) antibiotic prescription rate; (2) the proportion of antibiotics that were broad-spectrum antibiotics; (3) the proportion of antibiotics of appropriate indication (presumed bacterial), inappropriate indication (presumed viral), or inconclusive indication (unknown bacterial/viral or other); (4) the proportion of oral antibiotics of inappropriate duration; and (5) the proportion of antibiotics that were guideline-concordant in uncomplicated urinary and upper and lower respiratory tract infections (RTIs). We determined variation of antibiotic prescription and broad-spectrum prescription by calculating standardised prescription rates using multilevel logistic regression and adjusted for general characteristics (e.g., age, sex, comorbidity, referral), disease severity (e.g., triage level, fever duration, presence of alarming signs), use and result of diagnostics, and focus and cause of infection. In this analysis of 35,650 children (median age 2.8 years, 55% male), overall antibiotic prescription rate was 31.9% (range across EDs: 22.4%–41.6%), and among those prescriptions, the broad-spectrum antibiotic prescription rate was 52.1% (range across EDs: 33.0%–90.3%). After standardisation, differences in antibiotic prescriptions ranged from 0.8 to 1.4, and the ratio between broad-spectrum and narrow-spectrum prescriptions ranged from 0.7 to 1.8 across EDs. Standardised antibiotic prescription rates varied for presumed bacterial infections (0.9 to 1.1), presumed viral infections (0.1 to 3.3), and infections of unknown cause (0.1 to 1.8). In all febrile children, antibiotic prescriptions were appropriate in 65.0% of prescriptions, inappropriate in 12.5% (range across EDs: 0.6%–29.3%), and inconclusive in 22.5% (range across EDs: 0.4%–60.8%). Prescriptions were of inappropriate duration in 20% of oral prescriptions (range across EDs: 4.4%–59.0%). Oral prescriptions were not concordant with the local guideline in 22.3% (range across EDs: 11.8%–47.3%) of prescriptions in uncomplicated RTIs and in 45.1% (range across EDs: 11.1%–100%) of prescriptions in uncomplicated urinary tract infections. A limitation of our study is that the included EDs are not representative of all febrile children attending EDs in that country. Conclusions In this study, we observed wide variation between European EDs in prescriptions of antibiotics and broad-spectrum antibiotics in febrile children. Overall, one-third of prescriptions were inappropriate or inconclusive, with marked variation between EDs. Until better diagnostics are available to accurately differentiate between bacterial and viral aetiologies, implementation of antimicrobial stewardship guidelines across Europe is necessary to limit antimicrobial resistance.publishersversionPeer reviewe

Association of kidney disease measures with risk of renal function worsening in patients with type 1 diabetes

Background: Albuminuria has been classically considered a marker of kidney damage progression in diabetic patients and it is routinely assessed to monitor kidney function. However, the role of a mild GFR reduction on the development of stage 653 CKD has been less explored in type 1 diabetes mellitus (T1DM) patients. Aim of the present study was to evaluate the prognostic role of kidney disease measures, namely albuminuria and reduced GFR, on the development of stage 653 CKD in a large cohort of patients affected by T1DM. Methods: A total of 4284 patients affected by T1DM followed-up at 76 diabetes centers participating to the Italian Association of Clinical Diabetologists (Associazione Medici Diabetologi, AMD) initiative constitutes the study population. Urinary albumin excretion (ACR) and estimated GFR (eGFR) were retrieved and analyzed. The incidence of stage 653 CKD (eGFR &lt; 60 mL/min/1.73 m2) or eGFR reduction &gt; 30% from baseline was evaluated. Results: The mean estimated GFR was 98 \ub1 17 mL/min/1.73m2 and the proportion of patients with albuminuria was 15.3% (n = 654) at baseline. About 8% (n = 337) of patients developed one of the two renal endpoints during the 4-year follow-up period. Age, albuminuria (micro or macro) and baseline eGFR &lt; 90 ml/min/m2 were independent risk factors for stage 653 CKD and renal function worsening. When compared to patients with eGFR &gt; 90 ml/min/1.73m2 and normoalbuminuria, those with albuminuria at baseline had a 1.69 greater risk of reaching stage 3 CKD, while patients with mild eGFR reduction (i.e. eGFR between 90 and 60 mL/min/1.73 m2) show a 3.81 greater risk that rose to 8.24 for those patients with albuminuria and mild eGFR reduction at baseline. Conclusions: Albuminuria and eGFR reduction represent independent risk factors for incident stage 653 CKD in T1DM patients. The simultaneous occurrence of reduced eGFR and albuminuria have a synergistic effect on renal function worsening

Sulforaphane induces oxidative stress and death by p53-independent mechanism: implication of impaired glutathione recycling

Sulforaphane (SFN) is a naturally-occurring isothiocyanate best known for its role as an indirect antioxidant. Notwithstanding, in different cancer cell lines, SFN may promote the accumulation of reactive oxygen species (ROS) and cause cell death e.g. by apoptosis. Osteosarcoma often becomes chemoresistant, and new molecular targets to prevent drug resistance are needed. Here, we aimed to determine the effect of SFN on ROS levels and to identify key biomarkers leading to ROS unbalance and apoptosis in the p53-null MG-63 osteosarcoma cell line. MG-63 cells were exposed to SFN for up to 48 h. At 10 μM concentration or higher, SFN decreased cell viability, increased the%early apoptotic cells and increased caspase 3 activity. At these higher doses, SFN increased ROS levels, which correlated with apoptotic endpoints and cell viability decline. In exposed cells, gene expression analysis revealed only partial induction of phase-2 detoxification genes. More importantly, SFN inhibited ROS-scavenging enzymes and impaired glutathione recycling, as evidenced by inhibition of glutathione reductase (GR) activity and combined inhibition of glutathione peroxidase (GPx) gene expression and enzyme activity. In conclusion, SFN induced oxidative stress and apoptosis via a p53-independent mechanism. GPx expression and activity were found associated with ROS accumulation in MG-63 cells and are potential biomarkers for the efficacy of ROS-inducing agents e.g. as co-adjuvant drugs in osteosarcoma

Impact of Epstein-Barr virus in the clinical evolution of patients with classical Hodgkin's Lymphoma in Brazil

Introduction: Classical Hodgkin's Lymphoma (cHL) has been frequently associated with Epstein Barr virus (EBV), which can be found in a latent pattern in Reed-Sternberg (RS) cells. However, the impact of the presence of EBV in RS cells and its prognosis are still controversial. We analysed the presence of EBV in RS cells and its influence in the clinical evolution of patients with cHL treated in two public hospitals in the city of São Paulo, Brazil.Materials and Methods: We selected 97 patients with cHL from 1994 to 2004. Patients were only included in this study if they had (1) >18 years, (2) negative HIV serology, (3) undergone similar chemotherapy protocols, (4) paraffin blocks available with enough material for systematic review and histological reclassification and for detection of EBV in RS cells by in situ hybridization and immunohistochemistry and (5) clinical, epidemiological and laboratorial parameters available after a thorough chart review.Results: EBV was identified in 52.5% of the cases. Mixed cellularity (MC) subtype was more common in EBV-related tumours (25.5%) (p =0.005). There was no difference on age, gender, stage and the presence of B symptoms between the two groups. the presence of EBV did not influence event free survival (EFS) (p =0.38) or overall survival (OS) (p = 0.80) with a median follow-up of 80 months.Conclusion: We demonstrate that the prevalence of EBV-related cHL in this Brazilian population is 52.5% and, that, the presence of EBV does not change the clinical evolution and OS of patients treated with similar chemotherapy protocols. Copyright (C) 2010 John Wiley & Sons, Ltd.Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Universidade Federal de São Paulo, Disciplina Hematol & Hemoterapia, Hematol & Transfus Serv, BR-04023060 São Paulo, BrazilBrigadeiro State Hosp, São Paulo, BrazilUniversidade Federal de São Paulo, Disciplina Hematol & Hemoterapia, Hematol & Transfus Serv, BR-04023060 São Paulo, BrazilFAPESP: 02/10290-1FAPESP: 02/09994-4Web of Scienc