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Bauxite residues (BRs) are highly alkaline wastes generated during alumina production from bauxite ore. Billions of tons have been accumulating worldwide for more than 100 years, they are stored in various forms, and pose environmental and societal issues. At the same time, BRs are promising secondary sources for the production of critical metals including rare earth elements (REEs). However, knowledge on REE speciation is lacking, and is consequently an obstacle to the development of large-scale extraction process. This study analyses the influence of origin of the bauxite ore (lateritic or karstic), the storage conditions and storage time on the properties of ten BR samples, with a particular focus on the speciation of yttrium, which is used as a proxy to identify the behaviour of heavy REE. A multi-scale approach linked yttrium speciation and the origin of the bauxite ore whereas no major variation was observed as a function of storage conditions or ageing of the BRs. Yttrium is mainly found in the form of xenotime phosphate particles in BRs of lateritic origin, while in karstic BRs, the majority of yttrium is probably adsorbed or incorporated into other minerals including iron oxyhydroxide and hydroxyapatite minerals., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

While existing approaches for assessing passenger experience are often limited to surveys of customer satisfaction, societal and technological challenges push the railway industry to adopt a user-centric approach to the design of their service. We used the love and breakup method in a study involving N = 53 passengers making a declaration to their railway company to collect qualitative feedback on the passenger experience. The method allowed to gather personal, emotional, and contextual insights into passengers' experiences that can inform the transportation service design process. We describe 21 factors and 8 needs influencing the passenger experience, thereby consolidating and deepening prior work in the railway context. Using the lens of user experience theories, we argue that the service should be assessed against fulfilling these needs, which can act as guiding principles regarding service improvement. The study also presents valuable insights into the love and breakup method to explore service experiences., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Cancer stem cells (CSCs) serve an essential role in failure of conventional antitumor therapy. In breast cancer, CD24 ‑/low /CD44 + phenotype and high aldehyde dehydrogenase activity are associated with CSC subtypes. Furthermore, CD24 ‑/low /CD44 + pattern is also characteristic of mesenchymal cells generated by epithelial‑mesenchymal transition (EMT). CD24 is a surface marker expressed in numerous types of tumor, however, its biological functions and role in cancer progression and treatment resistance remain poorly documented. Loss of CD24 expression in breast cancer cells is associated with radiation resistance and control of oxidative stress. Reactive oxygen species (ROS) mediate the effects of anticancer drugs as well as ionizing radiation; therefore, the present study investigated if CD24 mediates radiation‑ and chemo‑resistance of breast cancer cells. Using a HMLE breast cancer cell model, CD24 expression has been artificially modulated and it was observed that loss of CD24 expression induced stemness properties associated with acquisition of a hybrid E/M phenotype. CD24 ‑/low cells were more radiation‑ and chemo‑resistant than CD24 + cells. The resistance was associated with lower levels of ROS; CD24 controlled ROS levels via regulation of mitochondrial function independently of antioxidant activity. Together, these results suggested a key role of CD24 in de‑differentiation of breast cancer cells and promoting acquisition of therapeutic resistance properties.

Background: Incorporating the best available evidence into clinical practice is a determining challenge for healthcare professionals and organisations. The role of advanced practice nurses is viewed as a facilitator to adapt guideline recommendations to suit specific contexts and to overcome barriers to implementation. In this study, we evaluate the impact of advanced practice nurses on clinical indicators of hospitalised patients and on adherence to recommendations derived from two clinical practice guidelines (pressure ulcer prevention and treatment and vascular access device management)., Methods: Quasi-experimental study in five intervention (IU) and five control (CU) hospital units at three hospitals in Spain (period 2018-19). Five advanced practice nurses were incorporated into IU, with the intention that would produce attitudinal changes and enhance the skills and knowledge of the nursing team regarding 18 clinical practice recommendations. In this study, 41 indicators were evaluated through direct observation of all patients admitted, at monthly intervals for 1 year. Outcomes were assessed by means of a descriptive, multi-line regression and association analysis., Results: The study population was composed of 3742 inpatients admitted for pressure ulcer assessment and 2631 fitted with vascular access devices. By the end of the study period, all variables had improved in the IU, where average compliance with recommendations was statistically significantly higher (pressure ulcer guidance 7.9 ± 1.9 vs 6.0 ± 1.7. OR 1.86, 95% CI 1.67-2.05; vascular access devices guidance 5.4 ± 1.4 vs 4.4 ± 1,6. OR 1.06, 95% CI 0.95-1.17). The prevalence of pressure lesions and catheter-related adverse events decreased statistically significantly in the IU compared to the CU. The prevalence of pressure ulcers decreases (5.7% in IU vs 8.7% in CU p < 0.005) as well as the prevalence of adverse events related to the catheter (14% In IU vs 21.6% in CU p < 0.005). The unnecessary catheters decressed in IU 10.9% VS CU 15.8% (p < 0.005)., Conclusions: The incorporation of an advanced practice nurse statistically significantly improves clinical indicators related to the prevention and treatment of pressure ulcers and to the management of vascular access devices., Trial Registration: ISRCTN18259923 retrospectively registered on 11/02/2022., (© 2022. The Author(s).)

Yttrium (Y) has gained importance in high tech applications and, together with the other rare earth elements (REEs), is also considered to be an emerging environmental pollutant. The alpine plant Saxifraga paniculata was previously shown to display high metal tolerance and an intriguing REE accumulation potential. In this study, we analysed soil grown commercial and wild specimens of Saxifraga paniculata to assess Y accumulation and shed light on the uptake pathway. Laser ablation inductively coupled plasma mass spectrometry and synchrotron-based micro X-ray fluorescence spectroscopy was used to localise Y within the plant tissues and identify colocalized elements. Y was distributed similarly in commercial and wild specimens. Within the roots, Y was mostly located in the epidermis region. Translocation was low, but wild individuals accumulated significantly more Y than commercial ones. In plants of both origins, we observed consistent colocalization of Al, Fe, Y and Ce in all plant parts except for the hydathodes. This indicates a shared pathway during translocation and could explained by the formation of a stable organic complex with citrate, for example. Our study provides important insights into the uptake pathway of Y in S. paniculata, which can be generalised to other plants., (© 2022. The Author(s).)

Interest in measuring displacement gradients, such as rotation and strain, is growing in many areas of geophysical research. This results in an urgent demand for reliable and field-deployable instruments measuring these quantities. In order to further establish a high-quality standard for rotation and strain measurements in seismology, we organized a comparative sensor test experiment that took place in November 2019 at the Geophysical Observatory of the Ludwig-Maximilians University Munich in Fürstenfeldbruck, Germany. More than 24 different sensors, including three-component and single-component broadband rotational seismometers, six-component strong-motion sensors and Rotaphone systems, as well as the large ring laser gyroscopes ROMY and a Distributed Acoustic Sensing system, were involved in addition to 14 classical broadband seismometers and a 160 channel, 4.5 Hz geophone chain. The experiment consisted of two parts: during the first part, the sensors were co-located in a huddle test recording self-noise and signals from small, nearby explosions. In a second part, the sensors were distributed into the field in various array configurations recording seismic signals that were generated by small amounts of explosive and a Vibroseis truck. This paper presents details on the experimental setup and a first sensor performance comparison focusing on sensor self-noise, signal-to-noise ratios, and waveform similarities for the rotation rate sensors. Most of the sensors show a high level of coherency and waveform similarity within a narrow frequency range between 10 Hz and 20 Hz for recordings from a nearby explosion signal. Sensor as well as experiment design are critically accessed revealing the great need for reliable reference sensors.

Highly sensitive reporter-gene assays have been developed that allow both the direct vascular endothelial growth factor (VEGF) neutralizing activity of bevacizumab and the ability of bevacizumab to activate antibody dependent cellular cytotoxicity (ADCC) to be quantified rapidly and in a highly specific manner. The use of these assays has shown that in 46 patients with ovarian cancer following four cycle of bevacizumab treatment, and in longitudinal samples from the two patients that respond to bevacizumab therapy from a small cohort of patients with glioblastoma, that there is a reasonably good correlation between bevacizumab drug levels determined by ELISA and bevacizumab activity, determined using either the VEGF-responsive reporter gene, or the ADCC assays. One of the two primary non-responders with glioblastoma exhibited high levels of ADCC activity suggesting reduced bevacizumab Fc engagement in vivo in contrast to the other primary non-responder, and the two secondary non-responders with a decreasing bevacizumab PK profile, determined by ELISA that exhibited low to undetectable ADCC activity. Drug levels were consistently higher than bevacizumab activity determined using the reporter gene assay in serial samples from one of the secondary non-responders and lower in some samples from the other secondary non-responder and ADCC activity was markedly lower in all samples from these patients suggesting that bevacizumab activity may be partially neutralized by anti-drug neutralizing antibodies (NAbs). These results suggest that ADCC activity may be correlated with the ability of some patients to respond to treatment with bevacizumab while the use of the VEGF-responsive reporter-gene assay may allow the appearance of anti-bevacizumab NAbs to be used as a surrogate maker of treatment failure prior to the clinical signs of disease progression., (Copyright © 2020 Lallemand, Ferrando-Miguel, Auer, Iglseder, Czech, Gaber-Wagener, Di Pauli, Deisenhammer and Tovey.)

Physical inactivity has become a major public health concern and, consequently, the awareness of striving for a healthy lifestyle has increased. As a result, the popularity of recreational sports, such as running, has increased. Running is known for its low threshold to start and its attractiveness for a heterogeneous group of people. Yet, one can still observe high drop-out rates among (novice) runners. To understand the reasons for drop-out as perceived by runners, we investigate potential reasons to quit running among short distance runners (5 km and 10 km) (n = 898). Data used in this study were drawn from the standardized online Eindhoven Running Survey 2016 (ERS16). Binary logistic regressions were used to investigate the relation between reasons to quit running and different variables like socio-demographic variables, running habits and attitudes, interests, and opinions (AIOs) on running. Our results indicate that, not only people of different gender and age show significant differences in perceived reasons to quit running, also running habits, (e.g., running context and frequency) and AIOs are related to perceived reasons to quit running too. With insights into these related variables, potential drop-out reasons could help health professionals in understanding and lowering drop-out rates among recreational runners.

Among runners, there is a high drop-out rate due to injuries and loss of motivation. These runners often lack personalized guidance and support. While there is much potential for sports apps to act as (e-)coaches to help these runners to avoid injuries, set goals, and maintain good intentions, most available running apps primarily focus on persuasive design features like monitoring, they offer few or no features that support personalized guidance (e.g., personalized training schemes). Therefore, we give a detailed description of the working mechanism of Inspirun e-Coach app and on how this app uses a personalized coaching approach with automatic adaptation of training schemes based on biofeedback and GPS-data. We also share insights into how end-users experience this working mechanism. The primary conclusion of this study is that the working mechanism (if provided with accurate data) automatically adapts training sessions to the runners' physical workload and stimulates runners' goal perception, motivation, and experienced personalization. With this mechanism, we attempted to make optimal use of the potential of wearable technology to support the large group of novice or less experienced runners and that by providing insight in our working mechanisms, it can be applied in other technologies, wearables, and types of sports.

Evidence-based practice (EBP) combined with quality of care improves patient outcomes. However, there are still difficulties for its implementation in daily clinical practice. This project aims to evaluate the impact of the incorporation of the Advanced Practice Nurse (APN) role on the implementation of EBP at three levels: context, nurses' perceptions, and clinical outcomes. Mixed-methods study in two phases is proposed. Phase 1: a quasi-experimental design where five APNs are included in five hospitalization wards that are compared with another five similar wards without APNs. Variables from Practice-Environment-Scale-Nursing-Work-Index, Health-Science-Evidence-Based-Practice-Questionnaire, and Advanced-Practice-Nursing-Competency-Assessment-Instrument are used. Clinical outcomes are followed-up with monthly. A descriptive and exploratory analysis is performed. Phase 2: an exploratory qualitative design through focus groups at the intervention wards after one year of APNs implementation. Explicative data are gathered to explain the progression of change and how actors perceive and attribute triggers, barriers, and facilitators for change. An inductive thematic analysis is performed. The inclusion of APN in hospitalization context is insufficiently studied. It is hoped that these figures provide solutions to the multiple barriers in the development of EBP in these sceneries and contribute to resolve the gap between research results and healthcare practice.

Background: There is increasing interest in the role that technology can play in improving the vitality of knowledge workers. A promising and widely adopted strategy to attain this goal is to reduce sedentary behavior (SB) and increase physical activity (PA). In this paper, we review the state-of-the-art SB and PA interventions using technology in the office environment. By scoping the existing landscape, we identified current gaps and underexplored possibilities. We discuss opportunities for future development and research on SB and PA interventions using technology. Methods: A systematic search was conducted in the Association for Computing Machinery digital library, the interdisciplinary library Scopus, and the Institute of Electrical and Electronics Engineers Xplore Digital Library to locate peer-reviewed scientific articles detailing SB and PA technology interventions in office environments between 2009 and 2019. Results: The initial search identified 1130 articles, of which 45 studies were included in the analysis. Our scoping review focused on the technologies supporting the interventions, which were coded using a grounded approach. Conclusion: Our findings showed that current SB and PA interventions using technology provide limited possibilities for physically active ways of working as opposed to the common strategy of prompting breaks. Interventions are also often offered as additional systems or services, rather than integrated into existing office infrastructures. With this work, we have mapped different types of interventions and provide an increased understanding of the opportunities for future multidisciplinary development and research of technologies to address sedentary behavior and physical activity in the office context.

Aim: The detection of specific DNA methylation patterns bears great promise as biomarker for personalized management of cancer patients. Co-amplification at lower denaturation temperature-PCR (COLD-PCR) assays are sensitive methods, but have previously only been able to analyze loss of DNA methylation., Materials & Methods: Enhanced (E)-ice-COLD-PCR reactions starting from 2 ng of bisulfite-converted DNA were developed to analyze methylation patterns in two promoters with locked nucleic acid (LNA) probes blocking amplification of unmethylated CpGs. The enrichment of methylated molecules was compared to quantitative (q)PCR and quantified using serial dilutions., Results: E-ice-COLD-PCR allowed the multiplexed enrichment and quantification of methylated DNA. Assays were validated in primary breast cancer specimens and circulating cell-free DNA from cancer patients., Conclusion: E-ice-COLD-PCR could prove a useful tool in the context of DNA methylation analysis for personalized medicine.

Novel ADCC effector cells expressing the V-variant or F-variant of Fc γ RIIIa (CD16a) and firefly luciferase under the control of a chimeric promoter incorporating recognition sequences for the principal transcription factors involved in Fc γ RIIIa signal transduction, together with novel target cells overexpressing a constant high level of the specific antigen recognized by rituximab, trastuzumab, cetuximab, infliximab, adalimumab, or etanercept, confer improved sensitivity, specificity, and dynamic range in an ADCC assay relative to effector cells expressing a NFAT-regulated reporter gene and wild-type target cells. The effector cells also contain a normalization gene rendering ADCC assays independent of cell number or serum matrix effects. The novel effector and target cells in a frozen thaw-and-use format exhibit low vial-to-vial and lot-to-lot variation in their performance characteristics reflected by CVs of 10% or less. Homologous control target cells in which the specific target gene has been invalidated by genome editing providing an ideal control and a means of correcting for nonspecific effects were observed with certain samples of human serum. The novel effector cells and target cells expressing noncleavable membrane-bound TNF α have been used to quantify ADCC activity in serum from patients with Crohn's disease treated with infliximab and to relate ADCC activity to drug levels.

Background: Beyond genetics, epigenetics alterations and especially those related to DNA methylation, play key roles in the pathogenesis of autoimmune diseases such as primary Sjögren's syndrome (pSS) and systemic lupus erythematosus. This study aimed to assess the role of methylation deregulation in pSS pathogeny through a genome-wide methylation approach., Patients and Methods: 26 female patients with pSS and 22 age-matched controls were included in this study. CD4+ T cells and CD19+ B cells were isolated from peripheral blood mononuclear cells by magnetic microbeads and their genome-wide DNA methylation profiles were analysed using Infinium Human Methylation 450 K BeadChips. Probes with a median DNA methylation difference of at least 7% and p<0.01 between patients and controls were considered significantly differentially methylated., Results: Methylation alterations were mainly present in B cells compared with T cells. In B cells, an enrichment of genes with differentially methylated probes in genetic at-risk loci was observed, suggesting involvement of both genetic and epigenetic abnormalities in the same genes. Methylation alterations in B cells were more frequent in some specific pathways including Interferon Regulated Genes, mainly among patients who were autoantibody positive. Moreover, genes with differentially methylated probes were over-represented in B cells from patients with active disease., Conclusions: This study demonstrated more important deregulation of DNA methylation patterns in B cells compared with T cells, emphasising the importance of B cells in the pathogenesis of the disease. Overlap between genes with differentially methylated probes in B lymphocytes and genetic at-risk loci is a new finding highlighting their importance in pSS., (Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://www.bmj.com/company/products-services/rights-and-licensing/)

In child-care facilities, taking into account the cultural diversity of families in migration is essential. Professionals can then review their references and habits to meet families and build a trust relationship. This exchange forms a respectful accompaniment of the values and habits of each, favouring the development of young children.

Aim: To discuss the Foucauldian concept of genealogy as a framework for understanding and transforming nurses' professional identity., Background: The professional identity of nurses has primarily been defined by personal and interpersonal attributes and by the intradisciplinary dimensions of nursing, leading to its conceptualization as a universal, monolithic phenomenon. The Foucauldian genealogical perspective offers a critical lens to examine what constitutes this professional identity; Spanish nursing offers a historical case study of an active effort to impose an identity that fits the monolithic ideal., Data Sources: Five of the 33 professional conduct manuals for nurses' training published from 1956-1976 during the Franco dictatorship in Spain and six interviews with nursing instructors or students at the time were analysed using a theoretical framework drawn from Foucault's writing., Discussion: Foucault's genealogical framework considers practices of normalization and resistance as a means of understanding knowledge continuities and discontinuities, clarifying practices that constitute nurses' professional identity in a particular way in specific contexts and analysing the implications of this theoretical frame., Implications for Nursing: The genealogy concept offers valuable tools to determine how professional identities are constituted, questions assumptions about the profession and its professionals and envisions alternative approaches. This theoretical approach helps both scholars and practitioners understand, question and transform their practices as needed., Conclusion: The genealogical approach prioritizes analysis of the phenomenon over its description and challenges many unknown, forgotten, excluded and/or unquestioned aspects of identity from a position of diversity and complexity., (© 2013 John Wiley & Sons Ltd.)

Effective monitoring of the development of neutralizing antibodies (NAbs) against IFN-β in multiple sclerosis (MS) patients on IFN-β therapy is important for clinical decision making and disease management. To date, antiviral assays have been the favored approach for NAb determination, but variations in assay conditions between laboratories and the increasing use of novel assays have contributed to the reporting of inconsistent antibody data between laboratories and between products. This study, undertaken at the request of the Committee for Medicinal Products for Human Use (CHMP) of the European Medicines Agency (EMA), is a joint effort by manufacturers of IFN-β products (approved in Europe) towards harmonization of a NAb assay that facilitates generation of comparable NAb data, which, in conjunction with clinical outcomes, should prove useful for clinicians treating MS patients with IFN-β products. This article describes the standardized cellular myxovirus resistance protein A (MxA) protein measurement-based assay for detection of IFN-β NAbs and its use for the validation of assays used for the quantitative determination of such antibodies. Although titers varied between laboratories and the products used, utilization of IFN-β1a rather than IFN-β1b as the challenge antigen produced more consistent results in the NAb assay. Adoption of the standardized assay improves comparability between laboratories circumventing problems that arise when different, nonstandardized assays are employed for immunogenicity assessment. Based on the data, the EMA recommended for standardization purposes, the use of IFN-β1a in NAb assays, independent of the therapeutic product used for therapy and validation of new NAb procedures against the standardized assay described.

Biopharmaceuticals are used extensively for the treatment of a number of chronic debilitating and fatal diseases such as cancer and inflammatory or autoimmune diseases. Although biopharmaceuticals are in general well tolerated, the development of anti-drug antibodies can impair their safety and efficacy. Assessment of immunogenicity is essential for a more effective and rational use of biopharmaceuticals, and is dependent upon the establishment of efficient standardized assays that allow direct comparison of immunogenicity data with clinical outcome. Although regulatory authorities recommend the use of cell-based assays that reflect the mechanism of action of the drug for the detection of neutralizing anti-drug antibodies, conventional cell-based assays are difficult to standardize and often give variable results. A number of strategies have been adopted to improve the performance of cell-based assays, including quantification of drug-induced proteins using either real-time RT-PCR or branched DNA to detect mRNA, or ELISAs to detect protein, bridging assays using immobilized cells and the use of reporter gene assays. The relative merits and limitations of each of these methods is reviewed herein.

Introduction: Participatory action research (PAR) was conducted in an intensive care unit (UCI), with the general purpose of fostering change in clinical practice so as to improve the care offered to families of critically ill patients. As a result of this process, four change-related initiatives were introduced. One specific additional objective was to explore how the unit's background context limited or facilitated change. This paper presents findings based on this objective., Materials and Methods: Qualitative methodology., Design: Participatory-action research (PAR). DATA GATHERING TECHNIQUES: 11 discussion groups incorporating professionals, 5 in-depth interviews with professionals, field diaries kept by the participants, and field diary kept by the lead researcher. Eleven professionals took part in the discussion groups (each one conveyed information made known to them by 3-5 colleagues), 5 professionals were involved in the interviews, and 11 professionals filled in a field diary. A content analysis was performed., Results: Factors limiting change included: 1) Not acknowledging the legitimacy of scientific evidence regarding the families of critically ill patients; 2) Imbalanced power relationships among the members of multi-disciplinary teams; 3) Nurses' lack of involvement in information flow; 4) The organization of time and physical space in the unit. Factors facilitating change: 1) A sense of individual and shared commitment; 2) Leadership in day-to-day matters; 3) A process based on reflection., Conclusions: A process of participatory action research can lead to change in clinical practice, although this is complex and requires substantial input in terms of personal energy. Contextual factors limiting this change are related to the actual structure of the unit, while factors facilitating it are circumstantial ones and are dependent upon individual people. In this sense, professionals working at the bedside are capable of introducing changes to the context in which they work., (Copyright © 2011 Elsevier España, S.L. y SEEIUC. All rights reserved.)

This paper presents the results of an ergonomic intervention conducted within a blast furnace plant. As part of its risk prevention program, the company decided to set up an action plan, in a participatory manner, by setting up working groups to solve health & safety issues. This field mission involved 230 employees, 80 of whom participated actively by being incorporated into working groups. After four months of intervention, a questionnaire survey has been conducted among employees to study the effects of participation on the safety climate. The results seem promising and show that the benefits of participation are numerous: a more positive safety climate associated to safer attitudes and behaviors. However, rather than just participation, it seems to be the employee involvement in the working groups and the satisfaction they derive from their participation that guarantee these positive results. Hence, participatory ergonomics seems to be an effective way to decrease the number of unsafe behaviors at work, provided that the type of participation has been previously well defined and organized according to the specific context of each organization.

The European Prospective Investigation into Cancer and nutrition (EPIC) is a long-term, multi-centric prospective study in Europe investigating the relationships between cancer and nutrition. This study has served as a basis for a number of Genome-Wide Association Studies (GWAS) and other types of genetic analyses. Over a period of 5 years, 52,256 EPIC DNA samples have been extracted using an automated DNA extraction platform. Here we have evaluated the pre-analytical factors affecting DNA yield, including anthropometric, epidemiological and technical factors such as center of subject recruitment, age, gender, body-mass index, disease case or control status, tobacco consumption, number of aliquots of buffy coat used for DNA extraction, extraction machine or procedure, DNA quantification method, degree of haemolysis and variations in the timing of sample processing. We show that the largest significant variations in DNA yield were observed with degree of haemolysis and with center of subject recruitment. Age, gender, body-mass index, cancer case or control status and tobacco consumption also significantly impacted DNA yield. Feedback from laboratories which have analyzed DNA with different SNP genotyping technologies demonstrate that the vast majority of samples (approximately 88%) performed adequately in different types of assays. To our knowledge this study is the largest to date to evaluate the sources of pre-analytical variations in DNA extracted from peripheral leucocytes. The results provide a strong evidence-based rationale for standardized recommendations on blood collection and processing protocols for large-scale genetic studies.

A cell-based assay has been developed for the quantification of the activity of TNFα antagonists based on human erythroleukemic K562 cells transfected with a NFκB regulated firefly luciferase reporter-gene construct. Both drug activity and anti-drug neutralizing antibodies can be quantified with a high degree of precision within 2h, and without interference from cytokines and other factors known to activate NFκB. The assay cells also contain the Renilla luciferase reporter gene under the control of a constitutive promoter that allows TNFα-induced firefly luciferase activity to be normalized relative to Renilla luciferase expression. Thus, results are independent of cell number or differences in cell viability, resulting in intra and inter assay coefficients of variation of 10% or less. Normalization of results relative to the expression of an internal standard also provides a means for correcting for serum matrix effects and allows residual drug levels or anti-drug neutralizing antibodies to be quantified even in serum samples with a relatively high degree of cytotoxicity., (Copyright © 2011 Elsevier B.V. All rights reserved.)

The safety and efficacy of biopharmaceuticals can be severely impaired by their immunogenicity. A risk-based strategy should be used to assess immunogenicity on a case-by-case basis using standardized methods to correlate anti-drug antibody levels with clinical outcome. In silico and in vitro techniques allow putative T-cell epitopes to be identified and eliminated in candidate molecules while maintaining structure and function. Putative T-cell epitopes can be studied in the context of the HLA allotypes representative of the target population in vitro and in transgenic mice that express human HLA genes. Mice immune tolerant to human proteins allow the study of the effect of factors such as aggregation on the loss of immune tolerance. However, significant challenges remain in order to be able predict the immunogenicity of a therapeutic protein in a particular individual.

Biopharmaceuticals are used widely for the treatment of cancer, chronic viral hepatitis, inflammatory, and autoimmune diseases. Biopharmaceuticals such as interferons are well tolerated for the most part with the most common adverse events observed being 'flu-like' symptoms that resolve rapidly after initial treatment. Prolonged treatment is associated, however, with more serious adverse events including leucopenia, thrombocytopenia, and neuropsychiatric effects, which may necessitate dose reduction or even cessation of treatment in some patients. Recombinant growth factors, such as erythropoietin (EPO), granulocyte colony-stimulating factor, or granulocyte macrophage colony-stimulating factor, are for the most part well tolerated, although severe complications have been reported in patients with cancer or chronic kidney disease treated with EPO. Similarly, treatment of patients with cancer with high doses of interleukin-2 is associated with significant toxicity. Treatment of chronic inflammatory diseases, such as rheumatoid arthritis, psoriasis, and Crohn's disease, with antitumor necrosis factor-alpha monoclonal antibodies is associated with an increased risk of granulomatous infections and, in particular, tuberculosis. The monoclonal antibody, natalizumab, that targets alpha4 integrins is effective in the treatment of multiple sclerosis but is associated with the activation of JC virus and development of progressive multifocal leukoencephalopathy. Repeated administration of recombinant proteins can cause a break in immune tolerance in some patients resulting in the production of a polyclonal antibody response that can adversely affect pharmacokinetics and clinical response. In addition, neutralizing antibodies that cross react with nonredundant essential proteins such as EPO can cause severe autoimmune reactions.

Hoxa1 belongs to the Hox family of homeodomain transcription factors involved in patterning embryonic territories and governing organogenetic processes. In addition to its developmental functions, Hoxa1 has been shown to be an oncogene and to be overexpressed in the mammary gland in response to a deregulation of the autocrine growth hormone. It has therefore been suggested that Hoxa1 plays a pivotal role in the process linking autocrine growth hormone misregulation and mammary carcinogenesis. Like most Hox proteins, Hoxa1 can interact with Pbx proteins. This interaction relies on a Hox hexapeptidic sequence centred on conserved Tryptophan and Methionine residues. To address the importance of the Hox-Pbx interaction for the oncogenic activity of Hoxa1, we characterized here the properties of a Hoxa1 variant with substituted residues in the hexapeptide and demonstrate that the Hoxa1 mutant lost its ability to stimulate cell proliferation, anchorage-independent cell growth, and loss of contact inhibition. Therefore, the hexapeptide motif of Hoxa1 is required to confer its oncogenic activity, supporting the view that this activity relies on the ability of Hoxa1 to interact with Pbx.

Assessment of immunogenicity is an important part of biopharmaceutical drug safety evaluation and a prerequisite for the development of less immunogenic and safer biopharmaceuticals since anti-drug antibodies can impair the activity and compromise the safety of biopharmaceuticals. Although regulatory authorities recommend cell-based assays for detection of neutralizing antibodies (NAbs), such assays are difficult to standardize, and ill adapted to high-throughput analysis. These limitations have been overcome by the development of a unique one-step cell-based assay that allows both drug activity and drug NAbs to be quantified rapidly and with a high degree of precision simply be adding reporter cells to a sample. The reporter cells have been engineered to express firefly luciferase (FL) under the control of a drug-responsive promoter, and to express the drug of interest, the production of which is normalized relative to the expression of Renilla luciferase (RL) transcribed from a common doxycycline-inducible promoter. Residual drug levels present in a sample are first quantified by determination of FL expression, autocrine drug synthesis is then induced, and NAb activity is quantified from the difference in the ratio of FL/RL expression in the presence or absence of the sample. Since assay results are normalized relative to the expression of an internal standard, results are independent of cell number or differences in cell viability thus affording a high degree of assay precision and reducing serum matrix effects to a minimum. This unique assay platform is ideally suited for high-throughput analysis, is applicable to most biopharmaceuticals, and will facilitate standardization and comparison of immunogenicity data. The performance of the one-step assay is illustrated for interferon alpha2 (IFNalpha2) used widely to treated chronic hepatitis C (HCV) infection and neoplastic disease., (2010 Elsevier B.V. All rights reserved.)

Interferons (IFNs) are class II cytokines that are key components of the innate immune response to virus infection. Three IFN sub-families, type I, II, and III IFNs have been identified in man, Recombinant analogues of type I IFNs, in particular IFNα2 and IFNβ1, have found wide application for the treatment of chronic viral hepatitis and remitting relapsing multiple sclerosis respectively. Type II IFN, or IFN gamma, is used principally for the treatment of chronic granulomatous disease, while the recently discovered type III IFNs, also known as IFN lambda or IL-28/29, are currently being evaluated for the treatment of chronic viral hepatitis. IFNs are in general well tolerated and the most common adverse events observed with IFNα or IFNβ therapy are "flu-like" symptoms such as fever, headache, chills, and myalgia. Prolonged treatment is associated with more serious adverse events including leucopenia, thrombocytopenia, increased hepatic transaminases, and neuropsychiatric effects. Type I IFNs bind to high-affinity cell surface receptors, composed of two transmembrane polypeptides IFNAR1 and IFNAR2, resulting in activation of the Janus kinases Jak1 and Tyk2, phosphorylation and activation of the latent cytoplasmic signal transducers and activators of transcription (STAT1) and STAT2, formation of a transcription complex together with IRF9, and activation of a specific set of genes that encode the effector molecules responsible for mediating the biological activities of type I IFNs. Systemic administration of type I IFN results in activation of IFN receptors present on essentially all types of nucleated cells, including neurons and hematopoietic stem cells, in addition to target cells. This may well explain the wide spectrum of IFN associated toxicities. Recent reports suggest that certain polymorphisms in type I IFN signaling molecules are associated with IFN-induced neutropenia and thrombocytopenia in patients with chronic hepatitis C. IFNγ binds to a cell-surface receptor composed of two transmembrane polypeptides IFGR1 and IFGR2 resulting in activation of the Janus kinases Jak1 and Jak2, phosphorylation of STAT1, formation of STAT1 homodimers, and activation of a specific set of genes that encode the effector molecules responsible for mediating its biological activity. In common with type I IFNs, IFNγ receptors are ubiquitous and a number of the genes activated by IFNγ are also activated by type I IFNs that may well account for a spectrum of toxicities similar to that associated with type I IFNs including "flu-like" symptoms, neutropenia, thrombocytopenia, and increased hepatic transaminases. Although type III IFNs share the major components of the signal transduction pathway and activate a similar set of IFN-stimulated genes (ISGs) as type I IFNs, distribution of the IFNλ receptor is restricted to certain cell types suggesting that IFNλ therapy may be associated with a reduced spectrum of toxicities relative to type I or type II IFNs. Repeated administration of recombinant IFNs can cause in a break in immune tolerance to self-antigens in some patients resulting in the production of neutralizing antibodies (NABs) to the recombinant protein homologue. Appearance of NABs is associated with reduced pharmacokinetics, pharmacodynamics, and a reduced clinical response. The lack of cross-neutralization of IFNβ by anti-IFNα NABs and vice versa, undoubtedly accounts for the apparent lack of toxicity associated with the presence of anti-IFN NABs with the exception of relatively mild infusion/injection reactions.

Aim: A participatory action-research was conducted in an intensive care unit (ICU). A consensus was reached on four proposals for change regarding how to deal with the relatives of the critically ill patients, extending their visiting hours being one of these proposals. This study has aimed to ascertain the contributions of the unit professionals to a new proposal regarding visiting hours., Participants and Methods: The sample included 93 ICU professionals out of 205. A qualitative methodology study that included a two-item open survey was used: 1. Yes, I agree with the proposal (explain how you would improve it); 2. No, I do not agree with the proposal (explain why and how you would improve it). Content analysis and triangulation of researchers and participants were performed. The theoretical saturation point was reached., Results: Five categories emerged: 1) Modify the initial proposal; 2) Agreement with the proposal; 3) Doubts; 4) Barriers to implementation; 5) Flexibilization., Conclusions: ICU professionals are reluctant to open their units, but accept the extension of current visiting hours. The physical structure of the unit is perceived as a major barrier., (Copyright (c) 2009 Elsevier España, S.L. y SEEIUC. All rights reserved.)

The activity of several potent adjuvants, including incomplete Freund's adjuvant, CpG oligodeoxynucleotides, and alum, has been shown to be due at least in part to the induction of cytokines, including type I interferons (IFNs), IFN-gamma, interleukin-2 (IL-2), and IL-12, that play key roles in the regulation of innate and adaptive immunity. The relatively short half-life of recombinant homologues of cytokines has limited their use as vaccine adjuvants. These difficulties have been overcome by encapsulation into liposomes and the use of cytokine expression vectors co-administered with DNA vaccines. Although a number of cytokines including IFN-alpha, IFN-gamma, IL-2, IL-12, IL-15, IL-18, IL-21, GM-CSF, and Flt-3 ligand have been shown to potentiate the immune response to vaccination in various experimental models, the full potential of cytokines as vaccine adjuvants remains to be established.

Introduction: Within the context of participatory action research (PAR), a 4-stage process was established with the general aim of promoting improvements in the care offered to families of patients in the Intensive Care Unit (ICU). The 4 stages consisted of a situational diagnosis, proposals for change, the design and implementation of the proposals, and an evaluation. This paper presents the first 2 stages., Objectives: To define the attention given to families of patients in the Intensive Care Unit. To reach a consensus on areas for action/intervention in the unit., Materials and Method: A qualitative methodology., Design: PAR. Data-collection technique: 4 focus groups made up of 10 professionals, and consensus with support groups made up of 30 members. Content analysis was performed. The theoretical saturation point was reached., Results: Two documents were drawn up: 1) A situational diagnosis, describing the current situation of the attention given to families and 2) 10 proposals for change, 5 of which were given priority. They were: a session to familiarize professionals with evidence concerning families of patients in the ICU and the handing out of informative leaflets, the improvement of (nurse-family and intra-team) informative procedures, more privacy for patients and a reduction in environmental noise, a training plan for professionals, and change in the visiting policy., Conclusion: By using consensus-based methodologies, strategies for change can be prioritized and designed, adapted to the context in which they will be applied., (Copyright 2009 Elsevier España, S.L. y SEEIUC. All rights reserved.)

A randomized double-blind placebo-controlled study was conducted to determine the effect of sublingual administration of IFNalpha on the immune response to influenza vaccination in elderly institutionalized individuals. Sublingual administration of 10 million IU of IFNalpha immediately prior to vaccination, reduced the geometric mean haemagglutination inhibitory (HAI) and IgG2 circulating antibody titers, and the secretory IgA (sIgA) response in saliva, to the New York strain of influenza A virus, 21 days post-vaccination, without detectable drug-related local or systemic toxicity. IFN treatment did not inhibit the immune response to the other components of the vaccine; the New Caledonia strain of influenza A virus, or the Jiangsu strain of influenza B virus. At the dose tested sublingual administration of IFNalpha reduces the immune response to influenza vaccination in elderly institutionalized individuals.

Type I interferons (IFNs) produced primarily by plasmacytoid dendritic cells (pDCs) as part of the innate immune response to infectious agents induce the maturation of myeloid DCs and enhance antigen presentation. Type I IFNs also enhance apoptosis of virus-infected cells, stimulate cross priming and enhanced presentation of viral peptides. Type I IFNs are powerful polyclonal B-cell activators that induce a strong primary humoral immune response characterized by isotype switching and protection against virus challenge. Type I IFNs stimulate an IgG2a antibody response characteristic of Th1 immunity when ad-mixed with influenza virus vaccine and injected intramuscurarly (i.m.) or administered intranasally. The adjuvant activity of type I IFNs has been shown to involve direct effects of IFN on B-cells, effects on T-cells, as well as effects on antigen presentation. Oromucosal administration of type I IFNs concomitantly with i.m. injection of vaccine alone can also enhance the antibody response to influenza vaccination by enhancing trafficking of antigen-presenting cells towards the site of vaccination. Recombinant IFNs are potent adjuvants that may find application in both parenterally and mucosally administered vaccines.

In this article, the authors analyze the role the "Sant Vicente de Paül" Sisters of Charity had in treating the ill in Majorca during the 19th Century. The authors carried out their research on two primary resources: The Rules of Life and a record of members of the order called the Book of the Curia. The authors tried to identify the methods the sisters followed in their work inside the Rules, searching for those aspects indicating their possible influence on the health of the population to whom they served as well as searching for references to their dedication as teachers, nurses and/or managers. In the Book of the Curia, the authors analyze the social data registered for the first 320 members of the order; evaluating their age upon entrance, their length of stay in the order, their job, and the cause of their death in order to compare these according to whether a sister was a teacher or a nurse.

Recent data showed that p53 stimulates the expression of genes encoding not only pro- but also antioxidant enzymes. It was suggested that antioxidant genes could be induced under physiologic levels of stress while the prooxidant ones respond to higher level of stress. Results presented in this article illustrate an additional degree of complexity. We show that the expression of Haeme-oxygenase 1 (HO-1), a stress-inducible gene that codes for an enzyme having antioxidant properties, is stimulated in a p53-dependent manner in the thymus and spleen of irradiated mice. We prove that HO-1 is a direct p53 target gene by showing that the p53RE identified within human and mouse genes is specifically bound by p53. The threshold of irradiation dose required to induce a significant response of HO-1 in the lymphoid organs of the irradiated mice is higher than that for Waf1/p21 that encodes an universal inhibitor of cell cycle. Moreover, induction of HO-1 occurs later than that of Waf1/p21. Finally, the higher stimulation of HO-1 is reached when Waf1/p21 stimulation starts to decrease.

Interferon alpha (IFNalpha), produced primarily by plasmacytoid dendritic cells as part of the innate immune response to infectious agents, is a powerful polyclonal B-cell activator that induces a strong primary humoral immune response characterized by isotype switching and protection against virus challenge. IFNalpha has also been shown to stimulate an IgG2a antibody response characteristic of Th1 immunity when ad-mixed with influenza virus vaccine. The use of transgenic mice expressing a green fluorescent protein reporter gene regulated by an IFN responsive element has shown that IFN-activated cells are present in the peripheral circulation of influenza vaccinated mice as early as 4 h after initiation of IFNalpha treatment and that the principal cell populations activated by IFN treatment included B220 (-) Ly6c (-), CD11c (high), CD11b (high), CD8 (+) cells, and B220 (high), Ly6c (high), CD11c (low), CD11b (-), CD4 (+), CD19 (-) cells. The effect of IFNalpha on the antibody response to influenza vaccination was shown to be dependent upon the route of administration. Differential display analysis showed that numerous IFN responsive genes were induced in the lymphoid tissue of IFN treated animals together with a number of genes not previously shown to be induced by IFNalpha including Crg2 and other chemokines, proteases associated with antigen processing, and genes involved in lymphocyte activation, apoptosis, and protein degradation. Together these results may explain in part the mechanism(s) of the adjuvant activity of IFNalpha.

Replicative senescence appears after a finite number of cell divisions. After proliferation has ceased, senescent cells remain viable for long periods and metabolic modifications are observed such as lipofuscin accumulation. In order to understand this phenomenon, we examined the emergence of subcellular modifications corresponding to autophagy in MRC5 normal human fibroblasts. An increase of monodansylcadaverine fluorescence, a specific marker of autophagy, in aging compared to young fibroblasts was observed (p<0.0001). The increase of autophagic vacuoles in aging fibroblasts was confirmed by electron microscopy. We compared young versus senescent fibroblasts and showed that autophagic vacuoles, already present in young cells, became larger in senescent fibroblasts with a significant relative increase of inclusion area with respect to measured cell area (p=0.0041). However, autophagy-associated-gene expression remained stable in senescent compared to young fibroblasts, suggesting that the autophagy process per se is not enhanced. In parallel, transmission electron microscopy analysis showed that beta-galactosidase activity distribution was modified by aging: beta-galactosidase (an enzyme linked to lysosome) was scattered in young fibroblasts, but clustered at the level of autophagic vacuoles in senescent fibroblasts, suggesting a predominance of autolysosomes at this stage. These results support the hypothesis that, during fibroblast aging, the increase of autophagic vacuoles, as well as that of beta-galactosidase activity, may be associated to an increase of lysosomal mass and to an accumulation of degradative autolysosomes with lipofuscin. This phenomenon could be involved in the death of senescent fibroblasts.

Introduction and aim. Nursing professionals in intensive care units (ICU) face situations of uncertainty and anxiety, above all in patients' relatives. In the literature, we find that the way in which the family and their needs are valued by professionals in the unit is not very systematical, even though there are instruments that have been validated for this purpose. Therefore, we decided to value what perspective ICU nurses have of the relationship they establish with patients' relatives, in order to find possible explanations for this situation. Material and methods. A qualitative study with a critical-social orientation. This was carred out in three ICUs in three state hospitals in Mallorca. Research techniques: 8 observations of the visit of relatives to the unit and 6 semi-structured interviews with nurses from the three hospitals. The transcripts of the interviews were analysed and codified in significance units. Sources and methods were triangulated.Results. Categories of the relationship process: 1) who the social actors are; 2) what the actions of the nurses with respect to the relatives are; 3) what the relationships are like between both; and 4) expectations of the nurses regarding this relationship.Conclusions. a) It is necessary to establish quality standards for attending the relatives of critical patients. b) Continuous nursing training is necessary in order to provide emotional support to families and to protect nurses from the added stress involved in providing this emotional support.

The mechanisms that regulate the transcription of the tumour suppressor genes p53 and IRF-1 are poorly understood. We have characterized a 68-kDa transcription factor, GAAP-1 (gatekeeper of apoptosis activating proteins), encoded by an alternative splice product of the PRDII-BF1 gene, that recognizes a novel regulatory element within the p53 and IRF-1 promoters. Transfection of U937 cells with GAAP-1 activates p53 and IRF-1 expression and leads to apoptosis, whereas over-expression of GAAP-1 in K562 cells that lack p53 and IRF-1 induces cell differentiation. Alterations in the 6p24 locus containing the GAAP-1 gene are frequent in acute myelogenous leukemia (AML), and AML-derived cell lines display reduced GAAP-1 mRNA levels. Together, these results suggest that GAAP-1 acts as a gatekeeper at a critical point in the tumour suppressor gene pathway.

Constitutive expression of IFN-alpha5 and IFN-beta was detected in different lymphoid cells including peripheral blood mononuclear cells from normal individuals following amplification of IFN mRNA by reverse transcriptase-polymerase chain reaction and direct sequencing of the amplified product. The activated form of the interferon-induced transcription factor complex ISGF3 was also detected in nuclear extracts from uninduced cells. Culture supernatants from uninduced U937 cells were also found to activate an ISRE cloned upstream of the luciferase reporter gene, indicating the presence of endogenous IFN activity equivalent to approximately 0.3 to 0.5 IU/mL. This endogenous IFN was also shown to play a role in maintaining the basal level of expression of the major histocompatibility class I genes in lymphoid cells. These results suggest that IFN-alpha5 and IFN-beta are produced at low levels in normal tissues and play an important role in the regulation of cell function and in the maintenance of homeostasis.

To determine the cellular functions which are modified when interferon-beta (IFN-beta) gene expression is inhibited, a plasmid allowing the constitutive expression of RNA complementary to IFN-beta mRNA was constructed and stably introduced into L929 cells. Some of the selected clones expressing this antisense IFN-beta mRNA, named L-ASI, were unable to produce IFN-beta and lost the ability to arrest in the G0 phase of the cell cycle. Indeed, the usual transrepression of the c-fos gene observed in quiescent cells was blocked in IFN-beta antisense L-ASI clones and the c-fos gene was permanently stimulated. This overexpression of c-fos was not modified in response to protein kinase C agonists such as phorbol esters, but increased in response to the adenylate cyclase activator forskolin. In addition, the ability to induce major histocompatibility class I genes following recombinant IFN-beta treatment was impaired in antisense IFN-beta L-ASI clones, suggesting an important alteration of this cell with regard to the interferon system. Unexpectedly, the tumorigenicity of the clones was significantly diminished. We postulate that IFN-beta antisense RNA blocks the repression of the c-fos gene and thus prevents the arrest of cells in the G0 phase of the cycle.

A revertant cell line was selected from Moloney sarcoma virus-transformed BALB/c cells after long-term treatment with type I interferon. Despite an actively transcribed and transfectable v-mos gene, these revertant cells were nontumorigenic in nude mice. The functionality of the mos protein was investigated, focusing on the alpha 2(1) collagen promoter regulation, which is known to be affected by mos-induced trans-acting factors. Both in transient expression assays and after stable integration into the cellular genome, the transfected alpha 2(1) collagen promoter fused to the cat reporter gene was activated in the revertant while being downregulated in the original transformed cells. In retransformation assays of the revertant by Moloney sarcoma virus strains homologous to the original transforming virus, no detectable change was noted in the in vitro phenotype or in tumorigenicity. These results reveal that the mos-directed factors were no longer effective on their specific targets. Thus, the R.MSVIF cell could be either an oncoprotein-deficient or a target-related revertant. Attempts at retransformation with unrelated sarcoma viruses bearing v-sis, v-fms, or v-fos oncogenes were also negative. In contrast, tumorigenicity was obtained with the unrelated Kirsten sarcoma virus without any change in the revertant morphology or collagen expression. These findings showed that the common pathway blocked by the reversion and shared by v-mos and v-ras was not required for ras-induced tumorigenicity.

Serum lipoproteins were explored on the first day of life (D1) and on the sixth day (D6) in blood drawn by peripheral venipuncture from 43 normal term newborns, 22 breast-fed and 21 formula-fed, and were compared to those of a control group of 28 young adults. With the exception of apolipoprotein E (Apo E), values of lipoprotein components obtained at D1 were similar, although generally slightly lower than those previously reported for cord blood serum. Total Apo E concentration at D1 (71 +/- 25 mg/l) was much higher than that obtained for the adult group (30 +/- 7 mg/l). Apo E distribution within the lipoprotein spectrum confirmed the presence of an already known Apo E-rich high density lipoprotein subfraction, which was responsible for the high total Apo E level at birth. The rise of lipoproteins of low density from D1 to D6, as evidenced by the increase of very low density lipoprotein + low density lipoprotein cholesterol and low density lipoprotein apolipoprotein B concentrations, was shown to be diet dependent. It was significantly less important in newborns fed a standard formula moderately enriched in unsaturated fatty acids than in breast-fed newborns.