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Hillary J Gross,1,* Crystal Watson2,* 1Health Outcomes Practice, Kantar Health, Princeton, NJ, 2Health Economics and Outcomes Research, Biogen, Cambridge, MA, USA *These authors contributed equally to this work Background: Although most patients with relapsing-remitting multiple sclerosis (RRMS) will develop secondary progressive multiple sclerosis (SPMS), little is known about the burden of multiple sclerosis by disease subtype. This study describes the burden of disease in terms of demographics, disease severity, symptoms, health care resource and disease-modifying therapy (DMT) utilization, work and activity impairment, and physical functioning of SPMS and RRMS patients.Methods: SPMS and RRMS patient responses from the 2012 and 2013 waves of the US National Health and Wellness Survey were evaluated to detect differences in demographics, disease severity, symptoms, and health care resource and DMT utilization. In addition, data from the Work Productivity and Activity Impairment and Short Form-36 questionnaires were analyzed.Results: SPMS patients were older than RRMS patients (mean age 55.7 vs 48.9 years; P

Environmental exposures cause substantial morbidity and mortality in the United States. A major goal of Centers for Disease Control and Prevention Environmental Public Health Tracking program is the development of a national network of health and environmental data with analytic tools for rapid evaluation of specific national or regional environmental health concerns. A six-state collaborative project in the northeast United States was established to assess the feasibility of such a system, assessing the possible association between ambient air quality and adverse birth outcomes. For this regional surveillance project, issues were discussed surrounding the design of a mutually acceptable protocol, obtaining human subjects' protection approvals, obtaining and organizing both the exposure and outcome data, analyzing the data both locally and regionally, and planning subsequent interventions to address identified public health concerns. [ABSTRACT FROM AUTHOR]

Background: Capillary sampling of blood counts is a well-established alternative to venipuncture in paediatrics. However, the sampling method has to be considered when interpreting test results, as measurements differ. Ethical and practical considerations prevent simultaneous venous and capillary sample acquisition in comprehensive paediatric cohorts that span all ages for the purpose of a direct method comparison, resulting in uncertainty regarding the interpretation of capillary test results., Methods: We applied a data mining method to calculate the differences between capillary and venous blood count analytes using laboratory data collected during patient care. We examined 486 401 blood counts performed between 2010 and 2017 in two German paediatric tertiary care centers in children from birth to 18 years analysed on SYSMEX XE-2100 and SYSMEX XE-5000 devices, and analysed the differences between capillary and venous test results in 15 218 paired samples performed within 24 h., Results: We identified the mean systematic differences between capillary and venous (capillary-venous) test results for haemoglobin (+6.5 g/L), haematocrit (+2.38%), platelet count (-7.01 × 10 9 /l), red cell count (+0.18 × 10 12 /L), white cell count (-0.64 × 10 9 /L), mean corpuscular cell volume (+2.07 fl), mean corpuscular haemoglobin (+0.33 pg), mean corpuscular haemoglobin concentration (-4.4 g/L) and red cell distribution width (+0.40%). The effect of age on these mean deltas is negligible, while the levels of test results influence the difference between capillary and venous test results in most analytes., Conclusions: Our results improve guidance regarding the interpretation of capillary test results for children of all ages and in both physiological and pathological ranges., (© 2022 The Authors. International Journal of Laboratory Hematology published by John Wiley & Sons Ltd.)

An association between certain ABO/Rh blood groups and susceptibility to SARS-CoV-2 infection has been proposed for adults, although this remains controversial. In children and adolescents, the relationship is unclear due to a lack of robust data. Here, we investigated the association of ABO/Rh blood groups and SARS-CoV-2 in a multi-center study comprising 163 households with 281 children and 355 adults and at least one SARS-CoV-2 seropositive individual as determined by three independent assays as a proxy for previous infection. In line with previous findings, we found a higher frequency of blood group A (+ 6%) and a lower frequency of blood group O (-6%) among the SARS-CoV-2 seropositive adults compared to the seronegative ones. This trend was not seen in children. In contrast, SARS-CoV-2 seropositive children had a significantly lower frequency of Rh-positive blood groups. ABO compatibility did not seem to play a role in SARS-CoV-2 transmission within the families. A correction for family clusters was performed and estimated fixed effects of the blood group on the risk of SARS-CoV-2 seropositivity and symptomatic infection were determined. Although we found a different distribution of blood groups in seropositive individuals compared to the reference population, the risk of SARS-CoV-2 seropositivity or symptomatic infection was not increased in children or in adults with blood group A or AB versus O or B. Increasing age was the only parameter positively correlating with the risk of SARS-CoV-2 infection. In conclusion, specific ABO/Rh blood groups and ABO compatibility appear not to predispose for SARS-CoV-2 susceptibility in children., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Janda, Engel, Remppis, Enkel, Peter, Hörber, Ganzenmueller, Schober, Weinstock, Jacobsen, Fabricius, Zernickel, Stamminger, Dietz, Groß, Bode, Haddad, Elling, Stich, Tönshoff, Henneke, Debatin, Franz and Renk.)

The quality and persistence of children's humoral immune response following SARS-CoV-2 infection remains largely unknown but will be crucial to guide pediatric SARS-CoV-2 vaccination programs. Here, we examine 548 children and 717 adults within 328 households with at least one member with a previous laboratory-confirmed SARS-CoV-2 infection. We assess serological response at 3-4 months and 11-12 months after infection using a bead-based multiplex immunoassay for 23 human coronavirus antigens including SARS-CoV-2 and its Variants of Concern (VOC) and endemic human coronaviruses (HCoVs), and additionally by three commercial SARS-CoV-2 antibody assays. Neutralization against wild type SARS-CoV-2 and the Delta VOC are analysed in a pseudotyped virus assay. Children, compared to adults, are five times more likely to be asymptomatic, and have higher specific antibody levels which persist longer (96.2% versus 82.9% still seropositive 11-12 months post infection). Of note, symptomatic and asymptomatic infections induce similar humoral responses in all age groups. SARS-CoV-2 infection occurs independent of HCoV serostatus. Neutralization responses of children and adults are similar, although neutralization is reduced for both against the Delta VOC. Overall, the long-term humoral immune response to SARS-CoV-2 infection in children is of longer duration than in adults even after asymptomatic infection., (© 2022. The Author(s).)

Reference intervals for laboratory test results have to be appropriate for the population in which they are used to be clinically useful. While sex and age are established partitioning criteria, patients' origin also influences laboratory test results, but is not commonly considered when creating or applying reference intervals. In the German population, stratification for ethnicity is rarely performed, and no ethnicity-specific hematology reference intervals have been reported yet. In this retrospective study, we investigated whether specific reference intervals are warranted for the numerically largest group of non-German descent, individuals originating from Turkey. To this end, we analyzed 1,314,754 test results from 167,294 patients from six German centers. Using a name-based algorithm, 1.9% of patients were identified as originating from Turkey, in line with census data and the algorithm's sensitivity. Reference intervals and their confidence intervals were calculated using an indirect data mining approach, and Turkish and non-Turkish reference limits overlapped completely or partially in nearly all analytes, regardless of age and sex, and only 5/144 (3.5%) subgroups' reference limits showed no overlap. We therefore conclude that the current practice of using common reference intervals is appropriate and allows correct clinical decision-making in patients originating from Turkey., (© 2021. The Author(s).)

Highly sensitive and specific platforms for the detection of anti-severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) antibodies are becoming increasingly important for evaluating potential SARS-CoV-2 convalescent plasma donors, studying the spread of SARS-CoV-2 infections, and identifying individuals with seroconversion. This study provides a comparative validation of 4 anti-SARS-CoV-2 platforms. A unique feature of the study is the use of a representative cohort of convalescent patients with coronavirus disease 2019 and a mild to moderate disease course. All platforms showed significant correlations with a SARS-CoV-2 plaque reduction neutralization test, with highest sensitivities for the Euroimmun and the Roche platforms, suggesting their preferential use for screening persons at increased risk of SARS-CoV-2 infections., (© The Author(s) 2020. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail: journals.permissions@oup.com.)

Objectives: Assessment of children's laboratory test results requires consideration of the extensive changes that occur during physiological development and result in pronounced sex- and age-specific dynamics in many biochemical analytes. Pediatric reference intervals have to account for these dynamics, but ethical and practical challenges limit the availability of appropriate pediatric reference intervals that cover children from birth to adulthood. We have therefore initiated the multi-center data-driven PEDREF project (Next-Generation Pediatric Reference Intervals) to create pediatric reference intervals using data from laboratory information systems., Methods: We analyzed laboratory test results from 638,683 patients (217,883-982,548 samples per analyte, a median of 603,745 test results per analyte, and 10,298,067 test results in total) performed during patient care in 13 German centers. Test results from children with repeat measurements were discarded, and we estimated the distribution of physiological test results using a validated statistical approach ( kosmic )., Results: We report continuous pediatric reference intervals and percentile charts for alanine transaminase, aspartate transaminase, lactate dehydrogenase, alkaline phosphatase, γ-glutamyl-transferase, total protein, albumin, creatinine, urea, sodium, potassium, calcium, chloride, anorganic phosphate, and magnesium. Reference intervals are provided as tables and fractional polynomial functions (i.e., mathematical equations) that can be integrated into laboratory information systems. Additionally, Z -scores and percentiles enable the normalization of test results by age and sex to facilitate their interpretation across age groups., Conclusions: The provided reference intervals and percentile charts enable precise assessment of laboratory test results in children from birth to adulthood. Our findings highlight the pronounced dynamics in many biochemical analytes in neonates, which require particular consideration in reference intervals to support clinical decision making most effectively., (© 2021 Walter de Gruyter GmbH, Berlin/Boston.)

What Is Known and Objective: Delayed cerebral ischaemia is an important cause of morbidity and mortality after aneurysmal subarachnoid haemorrhage (aSAH). Nimodipine is the only drug approved by the FDA for improving outcome after aSAH. Clinically, however, there are no specific values of this drug in cerebrospinal fluid (CSF) during aSAH treatment that could be associated to outcome improvement. Furthermore, the neurotransmitter glutamate acts as a secondary marker for brain injury. The aim was to establish a method to measure nimodipine and glutamate concentrations simultaneously in CSF of patients after aSAH., Methods: From June 2017 to June 2018, we prospectively collected clinical data of patients with aSAH admitted to our neurointensive care unit. All included patients received nimodipine orally (60 mg every 4 hours). Patients, who developed clinical vasospasm during their in-hospital stay, underwent intra-arterial application of nimodipine (IAN), followed by angiographic control. A method using high-performance liquid chromatography coupled with mass spectrometric analysis (LC-MS/MS) was established for quantification of both analytes in CSF., Results and Discussion: In 15 (60%) of 25 patients, nimodipine and glutamate concentrations were measured. After IAN for treatment of vasospasms, CSF nimodipine concentrations were slightly higher than in patients who received nimodipine only orally (0.60 ± 0.27 ng/mL vs 0.48 ± 0.18 ng/mL). Patients developing vasospasm exhibited higher glutamate concentrations than patients without vasospasm (188.84 ng/mL vs136.07 ng/mL)., What Is New and Conclusion: The developed method allowed the simultaneous quantification of nimodipine and glutamate in CSF. Furthermore, we demonstrated that IAN resulted in higher concentrations in CSF, when compared to oral application only., (© 2019 John Wiley & Sons Ltd.)

Background Interpreting hematology analytes in children is challenging due to the extensive changes in hematopoiesis that accompany physiological development and lead to pronounced sex- and age-specific dynamics. Continuous percentile charts from birth to adulthood allow accurate consideration of these dynamics. However, the ethical and practical challenges unique to pediatric reference intervals have restricted the creation of such percentile charts, and limitations in current approaches to laboratory test result displays restrict their use when guiding clinical decisions. Methods We employed an improved data-driven approach to create percentile charts from laboratory data collected during patient care in 10 German centers (9,576,910 samples from 358,292 patients, 412,905-1,278,987 samples per analyte). We demonstrate visualization of hematology test results using percentile charts and z-scores (www.pedref.org/hematology) and assess the potential of percentiles and z-scores to support diagnosis of different hematological diseases. Results We created percentile charts for hemoglobin, hematocrit, red cell indices, red cell count, red cell distribution width, white cell count and platelet count in girls and boys from birth to 18 years of age. Comparison of pediatricians evaluating complex clinical scenarios using percentile charts versus conventional/tabular representations shows that percentile charts can enhance physician assessment in selected example cases. Age-specific percentiles and z-scores, compared with absolute test results, improve the identification of children with blood count abnormalities and the discrimination between different hematological diseases. Conclusions The provided reference intervals enable precise assessment of pediatric hematology test results. Representation of test results using percentiles and z-scores facilitates their interpretation and demonstrates the potential of digital approaches to improve clinical decision-making.

Background: Interpretation of alkaline phosphatase activity in children is challenging due to extensive changes with growth and puberty leading to distinct sex- and age-specific dynamics. Continuous percentile charts from birth to adulthood allow accurate consideration of these dynamics and seem reasonable for an analyte as closely linked to growth as alkaline phosphatase. However, the ethical and practical challenges unique to pediatric reference intervals have restricted the creation of such percentile charts, resulting in limitations when clinical decisions are based on alkaline phosphatase activity., Methods: We applied an indirect method to generate percentile charts for alkaline phosphatase activity using clinical laboratory data collected during the clinical care of patients. A total of 361,405 samples from 124,440 patients from six German tertiary care centers and one German laboratory service provider measured between January 2004 and June 2015 were analyzed. Measurement of alkaline phosphatase activity was performed on Roche Cobas analyzers using the IFCC's photometric method., Results: We created percentile charts for alkaline phosphatase activity in girls and boys from birth to 18 years which can be used as reference intervals. Additionally, data tables of age- and sex-specific percentile values allow the incorporation of these results into laboratory information systems., Conclusions: The percentile charts provided enable the appropriate differential diagnosis of changes in alkaline phosphatase activity due to disease and changes due to physiological development. After local validation, integration of the provided percentile charts into result reporting facilitates precise assessment of alkaline phosphatase dynamics in pediatrics.

What Is Known and Objectives: Venous thromboembolism (VTE), comprising deep vein thrombosis (DVT) and pulmonary embolism (PE), is a serious, life-threatening condition that often complicates treatment of individuals who are already ill and increases in risk with age. The comorbidity burden of VTE can complicate treatment; therefore, treatment should be influenced by the presence of comorbidities (Kearon 2012). The prevalence of common conditions in the VTE population is, therefore, an important subject of research. Prevalence of two common comorbid burdens, prior myocardial infarction (MI) and upper gastrointestinal (GI) conditions, was studied among survey respondents who reported DVT or PE., Methods: Responses to the 2010 wave of the National Health and Wellness Survey (NHWS), a self-administered, internet-based questionnaire from a nationwide, demographically representative sample of adults, were evaluated., Results and Discussion: Among the 814 participants reporting a history of VTE, 9·7% (n = 60) of the DVT subpopulation and 13·2% (n = 39) of the PE subpopulation also reported prior MI. In respondents with prior MI, cardiovascular, urological, and pain comorbidities were each reported as additional comorbidities by at least two thirds of respondents in both the PE and DVT subpopulations, with cardiovascular and urological conditions reported significantly (P < 0·05) more often than among respondents with no prior MI. Among the respondents reporting VTE, 48·9% (n = 302) of the subpopulation reporting DVT and 52·2% (n = 154) of those reporting PE also reported upper GI comorbidities. Cardiovascular and pain conditions in the respondents reporting upper GI comorbidities were each reported by more than three quarters of VTE patients in both the DVT and PE subpopulations and were significantly more common (P < 0·05) than among their counterparts without upper GI comorbidities., What Is New and Conclusion: The results of the NHWS indicate that VTE patients who have either of two common comorbid burdens, prior MI and concomitant upper GI conditions, also showed high levels of additional, concurrent comorbidities and generally poor health status. Clinicians must be aware of the total comorbidity profile of their patients who have experienced VTE in order to best manage them and prescribe appropriate therapy., (© 2014 John Wiley & Sons Ltd.)

Objectives: The aims of this study were to examine the impact of peripheral joint OA across five large European countries and how people with OA use pharmacotherapies., Methods: People with self-reported peripheral joint OA were selected from the 2011 five European countries (5EU) National Health and Wellness Survey (NHWS), which included 57 512 respondents from France, Germany, Italy, Spain and the UK. Information was recorded on symptoms, health status, health care utilization, work productivity and medication usage. All variables were analysed descriptively for the total population and individual countries., Results: A total of 3750 respondents met the inclusion criteria: 1635 (43.6%) UK, 961 (25.6%) France, 570 (15.2%) Germany, 316 (8.4%) Spain and 268 (7.1%) Italy. The majority were ages 55-74 years and most were overweight or obese. Health status [12-item Short Form version 2 (SF12v2)] was similar across all countries, with a mean (s.d.) of 40.53 (10.99); 21.5% self-reported experiencing depression. Most had visited a health care provider in the previous 6 months (n = 3537; 94.3%). One third were employed: 7% reported absenteeism and 24% presenteeism. The use of prescription medication for OA was reported by 46.9% of patients, over-the-counter (OTC) medication by 26.5%, and both by 9.4%. Medication use increased with pain severity. NSAIDs were the most commonly used medication. Opioid use varied from 1.8% in Italy to 54.5% in France. Fifty per cent reported full adherence (4-point Morisky Medication Adherence Scale), but only 30% reported satisfaction with their OA medication. Most used medication for half the days of the month., Conclusion: Despite some wide variations in pharmacotherapy for OA treatment, the impact of OA on health status and work productivity is substantial and looks largely similar across major European countries.

In the first decade of the 20th century, a horse named Hans drew worldwide attention in Berlin as the first and most famous "speaking" and thinking animal. Hans solved calculations by tapping numbers or letters with his hoof in order to answer questions. Later on, it turned out that the horse was able to give the correct answer by reading the microscopic signals in the face of the questioning person. This observation caused a revolution and as a consequence, experimenters avoided strictly any face-to-face contact in studies about cognitive abilities of animals-a fundamental lesson that is still not applied rigorously.

Pseudothrombocytopenia remains a challenge in the haematological laboratory. The pre-analytical problem that platelets tend to easily aggregate in vitro, giving rise to lower platelet counts, has been known since ethylenediamine-tetra acetic acid EDTA and automated platelet counting procedures were introduced in the haematological laboratory. Different approaches to avoid the time and temperature dependent in vitro aggregation of platelets in the presence of EDTA were tested, but none of them proved optimal for routine purposes. Patients with unexpectedly low platelet counts or flagged for suspected aggregates, were selected and smears were examined for platelet aggregates. In these cases patients were asked to consent to the drawing of an additional sample of blood anti-coagulated with a magnesium additive. Magnesium was used in the beginning of the last century as anticoagulant for microscopic platelet counts. Using this approach, we documented 44 patients with pseudothrombocytopenia. In all cases, platelet counts were markedly higher in samples anti-coagulated with the magnesium containing anticoagulant when compared to EDTA-anticoagulated blood samples. We conclude that in patients with known or suspected pseudothrombocytopenia the magnesium-anticoagulant blood samples may be recommended for platelet counting., (© 2013 John Wiley & Sons Ltd.)

Biofilms growing on copper and stainless steel substrata in natural drinking water were investigated. A modular pilot-scale distribution facility was installed at four waterworks using different raw waters and disinfection regimes. Three-month-old biofilms were analysed using molecular biology and microscopy methods. High total cell numbers, low counts of actively respiring cells and low numbers of cultivable bacteria indicated the high abundance of viable but not cultivable bacteria in the biofilms. The expression of the recA SOS responsive gene was detected and underlined the presence of transcriptionally active bacteria within the biofilms. This effect was most evident after UV disinfection, UV oxidation and UV disinfection with increased turbidity at waterworks compared to chemically treated and non-disinfected systems. Furthermore, live/dead staining techniques and environmental scanning electron microscopy imaging revealed the presence of living and intact bacteria in biofilms on copper substrata. Cluster analyses of DGGE profiles demonstrated differences in the composition of biofilms on copper and steel materials.

Objective: This study examined comorbidity prevalence and general medication use among individuals with atrial fibrillation in the United States to convey a more comprehensive picture of their total disease burden., Methods: This was a retrospective, observational evaluation of responses to the 2009 wave of the annual Internet-based National Health and Wellness survey, which collects health data including epidemiologic data and information on medical treatment from a representative nationwide sample of adults in the United States. Responses were assessed to determine three measures of comorbidity: mean number of comorbidities, CHADS2 score reflecting stroke risk (0-6 points; low risk: 0; moderate risk: 1; high risk: ≥ 2), and scores on the Charlson Comorbidity Index, which is a measure of general comorbidity reflecting presence of a wide range of comorbidities., Results: Of the overall sample, 1297 participants reported having been diagnosed with atrial fibrillation. Almost all (98%) of the predominantly male (65.1%) and older (≥ 65 years of age, 65.7%) population with atrial fibrillation had at least one additional comorbidity, and 90% had cardiovascular comorbidities. On the Charlson Comorbidity Index, 44.9% of the respondents had scores of 1-2 and 20.5% had scores of 3 or higher. High risk for stroke, demonstrated by a CHADS2 score of at least 2, was present in 45% and moderate risk (CHADS2 score 1) in 36%. Of the respondents with atrial fibrillation, 71% reported current use of medication to manage the condition, but only 48% of individuals at high risk for stroke reported use of anticoagulation therapy. Of those who reported having common risk factors for stroke, the majority reported receiving prescription therapy for these conditions., Conclusions: The health burden carried by patients often extends far beyond atrial fibrillation. Physicians should carefully consider comorbidities and concomitant medications when managing patients with atrial fibrillation.

Polymorphonuclear granulocytes (PMNs) have been attributed a primarily deleterious role in the pathogenesis of acute lung injury (ALI). However, evidence exists that PMNs might also act beneficially in certain types of ALI. In this regard, we investigated the role of activated neutrophils in the pathophysiology of lung contusion-induced ALI. We used the model of blunt chest trauma accompanied by PMN-depletion in male C3H/HeN mice. Animals received 25 μg/g body weight PMN-depleting antibody Gr-1 intravenously 48 h before trauma. Bronchoalveolar lavage (BAL) and lung tissue interleukin 6 (IL-6) were similarly elevated in PMN-depleted and control animals after trauma, whereas macrophage inflammatory protein 2 and monocyte chemoattractant protein 1 in BAL and lungs, IL-10 in BAL, and lung keratinocyte chemoattractant (KC) were even further increased in the absence of PMNs. Plasma IL-6 and KC were also increased in response to the insult and even further in the absence of PMNs. Chest trauma induced an enhanced release of IL-6, tumor necrosis factor α, macrophage inflammatory protein 2, monocyte chemoattractant protein 1, and IL-10 from isolated KU, which was blunted in the absence of PMNs. In the presence of PMNs, BAL protein was further increased at 30 h when compared with the 3-h time point, which was not the case in the absence of PMNs. Taken together, in response to lung trauma, activated neutrophils control inflammation including mediator release from distant immune cells but simultaneously mediate pulmonary tissue damage. Thus, keeping in mind potential inflammatory adverse effects, modulation of neutrophil activation or trafficking might be a reasonable therapeutic approach in chest trauma-induced lung injury.

Precise recognition of small object numbers without counting is a widespread phenomenon. It is well documented for humans and for a series of non-human vertebrates. Recently this has been confirmed for an invertebrate, the honeybee.(1) This type of inborn numerical competence has been named "subitizing", from the Latin subito = suddenly, immediately. It differs from the classical, sequential counting which has to be trained, starting with the help of our fingers. For humans it had been established since 1871 by Jevons(2) that only up to four objects are precisely recognized and memorized. Under conditions which do not allow sequential counting, mistakes start to occur in case of more than four objects. This result has been confirmed whenever the range of visual attention has been carefully tested under a variety of rigorous conditions. It provides the basis for a novel hypothesis about the evolution of counting and numbering systems in ancient civilizations.(3)

RNA-containing viruses represent a global threat to the health and wellbeing of humans and animals. Hence, the discovery of new approaches for the design of novel vaccines and antiviral compounds attains high attention. Here we describe the potential of artificial ribonucleases (aRNases), low molecular weight compounds capable to cleave phosphodiester bonds in RNA under mild conditions, to act as antiviral compounds via destroying the genome of non-enveloped RNA viruses, and the potential of utilizing honey bee larvae and adult bees (Apis mellifera) as a novel experimental system for the screening of new antiviral compounds. Pre-incubation of an Acute bee paralysis virus (ABPV) suspension with aRNases D3-12, K-D-1 or Dp12F6 in a concentration-dependent manner increased the survival rate of bee larvae and adult bees subsequently infected with these preparations, whereas incubation of the virus with aRNases ABL3C3 or L2-3 had no effect at all. The results of RT-PCR analysis of viral RNA isolated from aRNase-treated virus particles confirmed that virus inactivation occurs via degradation of viral genomic RNA: dose-dependent inactivation of ABPV correlates well with the cleavage of viral RNA. Electron microscopy analysis revealed that the morphology of ABPV particles inactivated by aRNases remains unaffected as compared to control virus preparations. Altogether the obtained results clearly demonstrate the potential of aRNases as a new virus inactivation agents and bee larvae/ABPV as a new in vivo system for the screening of antiviral compounds., (Copyright © 2011 Elsevier B.V. All rights reserved.)

Human inborn numerical competence means our ability to recognize object numbers precisely under circumstances which do not allow sequential counting. This archaic process has been called "subitizing," from the Latin "subito" = suddenly, immediately, indicating that the objects in question are presented to test persons only for a fraction of a second in order to prevent counting. In contrast, however, sequential counting, an outstanding cultural achievement of mankind, means to count "1, 2, 3, 4, 5, 6, 7, 8…" without a limit. The following essay will explain how the limit of numerical competence, i.e., the recognition of object numbers without counting, has been determined for humans and how this has been achieved for the first time in case of an invertebrate, the honeybee. Finally, a hypothesis explaining the influence of our limited, inborn numerical competence on counting in our times, e.g., in the Russian language, will be presented. Subitizing versus counting by young Down syndrome infants and autistics and the Savant syndrome will be discussed.

Congenital dyserythropoietic anaemias (CDAs) are heterogeneous, hereditary disorders hallmarked by ineffective erythropoiesis and tissue iron overload. Growth differentiation factor 15 (GDF15) was suggested to mediate iron overload in iron-loading anaemias, such as the thalassaemias and CDAI by suppressing hepcidin, the key regulator of iron absorption. Here, we show that serum GDF15 concentrations are elevated in subjects with CDAI and CDAII. Despite similar disease characteristics, CDAI patients present with significantly higher GDF15 concentrations compared to CDAII patients. Hepcidin concentrations are inappropriately low in CDAII patients considering the severe hepatic iron overload associated with this disorder. GDF15 significantly correlates with the degree of anaemia (Hb), the response of erythropoiesis (reticulocyte index) as well as with iron availability in the serum (transferrin saturation). The observation that GDF15 is elevated in CDAII patients is consistent with the proposal that GDF15 is among the erythroid factors down-regulating hepcidin and contributing to iron overload in conditions of dyserythropoiesis.

Precise recognition of small numbers of objects without counting is an archaic, inborn ability of humans. Since almost 140 years it is known that we can recognize precisely only up to four objects if sequential counting is prevented. Vertebrates and invertebrates such as honeybees can recognize and remember three and up to four objects, respectively. A synopsis of counting systems in ancient civilizations reveals that our limited ability to recognize only four objects without counting influenced our counting and numbering systems and enforced the need for new symbols for numbers beyond four.

Although the numerical abilities of many vertebrate species have been investigated in the scientific literature, there are few convincing accounts of invertebrate numerical competence. Honeybees, Apis mellifera, by virtue of their other impressive cognitive feats, are a prime candidate for investigations of this nature. We therefore used the well-established delayed match-to-sample paradigm, to test the limits of honeybees' ability to match two visual patterns solely on the basis of the shared number of elements in the two patterns. Using a y-maze, we found that bees can not only differentiate between patterns containing two and three elements, but can also use this prior knowledge to differentiate three from four, without any additional training. However, bees trained on the two versus three task could not distinguish between higher numbers, such as four versus five, four versus six, or five versus six. Control experiments confirmed that the bees were not using cues such as the colour of the exact configuration of the visual elements, the combined area or edge length of the elements, or illusory contours formed by the elements. To our knowledge, this is the first report of number-based visual generalisation by an invertebrate.

Phagocytosis is an essential event in the complex process of tissue repair. Here we examined the effect of low intensity pulsed ultrasound (US), which promotes fracture and wound healing, on phagocytosis by mouse macrophage cell line J774A.1 and human monocyte-derived macrophages. First, 10 to 40 min low intensity pulsed US increased uptake of serum opsonized E. coli by J774A.1 cells during a 50 min phagocytosis period. In addition, when the E. coli exposure time was varied between 35 to 80 min, the maximum increase in phagocytosis was observed in the first 35 min upon US exposure. In parallel, US induced robust actin polymerization in a time dependent manner in J774A.1 cells, showing the peak effect 30 min after stimulation. Interestingly, a low concentration of cytochalasin D (0.25-0.5 microM) prevented US-induced phagocytosis of E. coli. Furthermore, we demonstrated US enhanced activation of RhoA. Blocking its downstream effector Rho associated kinase (ROCK) with Y27632 abrogated US-induced phagocytosis. We also show that US induced activation of ERK and p38 MAPK. Pretreatment of the cells with the corresponding inhibitors PD98059 and SB203580 reduced US-induced phagocytosis. In addition, activity of tyrosine kinase Src was required for US-induced phagocytosis. Here Src represents an upstream activator of ERK and p38 MAPK. Depolymerization of actin by cytochalasin D prevented US-induced Src, ERK, and p38 activation. Our data provide a new insight into the cellular and molecular mechanisms by which low intensity pulsed US promotes tissue repair.

Oxidized low density lipoproteins (oxLDL), macrophages and T-lymphocytes are present in atherosclerotic lesions. We and others have shown that oxLDL is cytotoxic for macrophages, endothelial, smooth muscle and activated T-lymphocytes and induce apoptosis. Here we demonstrate that (i) oxidized LDL (oxLDL), oxidized VLDL (oxVLDL) and hydrogen peroxide (H2O2) induce apoptosis in human T-lymphocytes and (ii) mitogen-activated protein kinases are involved in this process. Apoptosis was monitored by immunofluorescence microscopy and flow cytometry for annexin V binding, Apo 2.7 expression, the TUNEL reaction and caspase 3 activity. In the presence of oxLDL (100 microg/ml), oxVLDL (50 microg/ml) and H2O2 (5 mM), the fraction of apoptotic cells increased within 6 hours to more than 70%. Preincubation of lymphocytes with the MAPKK inhibitor PD-98059 and the p38MAPK inhibitor SB-203580 almost completely abolished these effects. Furthermore, oxLDL and H2O2 but not native LDL strongly enhanced phosphorylation of JNK, p38MAPK and p42/44MAPK. The results suggest that in the resting lymphocyte apoptosis triggered by oxidized lipoproteins and oxidative stress depends on the activation of p44/42MAPK and p38MAPK cascades.

Artificial ribonucleases, conjugates of short oligodeoxyribonucleotides and peptides built of arginine, leucine, proline, and serine, were synthesized and assessed in terms of ribonuclease activity and specificity of RNA cleavage. A specific group of the conjugates was identified that display T1-ribonuclease-like activity and cleave RNA predominantly at G-X sequences. Circular dichroism study of the structures of the most active conjugates, free peptide (LR)4G, and oligonucleotides revealed that conjugation of oligonucleotide to the peptide results in a specific peptide folding that possibly provides ribonuclease activity to the conjugate.

Cleavage of phosphodiester bonds by small ribonuclease mimics within different bulge-loops of RNA was investigated. Bulge-loops of different size (1-7 nt) and sequence composition were formed in a 3' terminal fragment of influenza virus M2 RNA (96 nt) by hybridization of complementary oligodeoxynucleotides. Small bulges (up to 4 nt) were readily formed upon oligonucleotide hybridization, whereas hybridization of the RNA to the oligonucleotides designed to produce larger bulges resulted in formation of several alternative structures. A synthetic ribonuclease mimic displaying Pyr-Pu cleavage specificity cleaved CpA motifs located within bulges faster than similar motifs within the rest of the RNA. In the presence of 10 mM MgCl2, 75% of the cleavage products resulted from the attack of this motif. Thus, selective RNA cleavage at a single target phosphodiester bond was achieved by using bulge forming oligonucleotides and a small ribonuclease A mimic.

Kwashiorkor is a severe edematous form of malnutrition with high prevalence and lethality in many African countries, and repeatedly has been reported to be associated with oxidative stress. The therapy of kwashiorkor is still ineffective. In this pilot study, we tested the hypothesis that oral application of thiol-containing antioxidants increases glutathione status and is beneficial for the clinical recovery of kwashiorkor patients. The longitudinal clinical intervention study was carried out at St Joseph's Hospital, Jirapa, Ghana. Children with severe kwashiorkor were randomly assigned to either a standard treatment (ST) receiving a therapeutic protocol based on the recommendations of the WHO or to one of three study groups receiving in addition 2 x 600 mg reduced glutathione or 2 x 50 mg alpha-lipoic acid or 2 x 100 mg N-acetylcysteine per day. Patients were followed up clinically and biochemically for 20 days and compared with 37 healthy controls. Both glutathione and alpha-lipoic acid supplementation had positive effects on survival. Also, the blood glutathione concentrations correlated positively with survival rates. Furthermore, the initial skin lesions, glutathione and total protein concentrations were found to be strong predictors of survival. The data strongly suggest that a therapy restoring the antioxidative capacity by applying cysteine equivalents in the form of glutathione and/or alpha-lipoic acid is beneficial for biochemical and clinical recovery of kwashiorkor patients.

The adsorption equilibrium and kinetics of polyethylene glycol (PEG) in three aqueous systems were examined in this study. Langmuir isotherm was used to satisfactorily predict the adsorption capacity of PEG on activated carbon F-400 and applied to the investigation of adsorption kinetics. The surface diffusion, pore diffusion, and branched pore kinetics models successfully described the adsorption behavior of PEG on F-400 in the completely stirred tank reactor. The pore diffusion coefficients obtained from the pore diffusion model were compared with those computed by the experimental data of the short fixed-bed reactor combined with the assumption of non-hindered pore diffusion. In addition, the effects of initial concentrations of PEG and the relative importance of external and internal mass transfers for the adsorption were also taken into account and discussed in this study.

New artificial ribonucleases, conjugates of short oligodeoxyribonucleotides with peptides containing alternating arginine and leucine, were synthesized and characterized in terms of their catalytic activity and specificity of RNA cleavage. The conjugates efficiently cleave different RNAs within single-stranded regions. Depending on the sequence and length of the oligonucleotide, the conjugates display either G-X>>Pyr-A or Pyr-A>>G-X cleavage specificity. Preferential RNA cleavage at G-X phosphodiester bonds was observed for conjugate NH2-Gly-[ArgLeu]4-CCAAACA. The conjugates function as true catalysts, exhibiting reaction turnover up to 175 for 24 h. Our data show that in the conjugate the oligonucleotide plays the role of a factor which provides an 'active' conformation of the peptide via intramolecular interactions, and that it is the peptide residue itself which is responsible for substrate affinity and catalysis.

Surface expressed negatively charged sialoglycans contribute to the regulation of adhesive cellular interactions and are thus involved in the growth and differentiation of hematopoietic progenitor cells. In particular, the cell surface sialylation state may govern the liberation of CD34+ hematopoietic precursors from bone marrow stroma cells and extracellular matrix. In order to assess the overall surface sialylation of live human CD34+ hematopoietic precursor cells, we applied a previously described flow cytometric enzyme assay. Cells with and without sialidase pretreatment were incubated in the presence of fluorescent CMP-sialic acid and exogenous ST6GalI. Thus sialylation of surface-expressed lactosamine residues was analysed. We demonstrated that surface lactosamines of CD34+ precursors derived from bone marrow and peripheral blood are over 95% sialylated, predominantly in alpha2-6 linkage. These results are in accordance with flow cytometric analysis of surface lectin staining. Sialic acid specific lectins MAA and SNA were strongly bound whereas SBA, VVA, and PNA became reactive only after sialidase pretreatment. CD34+ leukemia cell lines TF1 and KG1a also showed a high degree of surface sialylation, whereas cell line KG1 expressed to the largest extent free lactosamines. In these cell lines, alpha2-6 and alpha2-3 sialylated residues were present in equal amounts. In a variation of the flow cytometric enzyme assay, live cells were incubated without exogenous STGal I to measure the activity of endogenous ecto-sialyltransferase. Ecto sialyltransferase activity was observed in all CD34+ cells which was able to resialylate major surface glycoproteins such as HLA Class I, CD45, CD43, and CD34. The ecto-sialyltransferase may serve to maintain or increase surface sialylation rapidly without de novo synthesis.

Objective: To study the sensitivity and specificity of different staining methods to monitor apoptosis induced by oxidative stress in adherent cells., Methods: Sensitivity and specificity of several common methods for apoptosis determination were evaluated (Apo2.7-expression, Annexin V-binding, TUNEL-reaction, poly-(ADP-ribose)-polymerase-(PARP) cleavage and single-stranded-DNA (ssDNA) staining). Apoptosis was induced by oxidative stress generated by hydrogen peroxide in 3 cultured cells types growing as adherent monolayer (MiaPaCa-2, Hep-G2 and human skin fibroblasts), necrosis was induced by depletion of cellular ATP using sodium azide. Cells positively stained by the respective apoptosis assay were quantified and alterations of cell morphology were monitored by fluorescence microscopy. The date was analyzed by one-way analysis of variance and significance test of correlation coefficient., Results: One hour after apoptosis induction significant cell fractions were positively stained for ssDNA (33% with MiaPaCa-2 cells, 35% with Hep-G2 cells, 56% with human skin fibroblasts). PARP-cleavage was less sensitive compared to the ssDNA-staining. Apo2.7-expression, Annexin V-binding and TUNEL-reaction were not applicable to detect early apoptosis induced by oxidative stress (below 2 hours), but were efficiently monitoring late apoptosis. Specificity of ssDNA-staining was complete with each cell type even 4 hs after induction of necrosis by the highest sodium azide concentration. In contrast, the same experimental conditions resulted in 50% - 90% positively stained necrotic cells by using Apo2.7-expression, TUNEL-reaction or Annexin V-binding. Surprisingly, specificity of PARP-cleavage was highly depending on the respective cell type., Conclusions: Our study prove that among the five methods investigated only ssDNA-staining allowed to completely differentiate apoptosis from necrosis, and is thus suitable to reliably detect early as well as late apoptosis. Therefore, the ssDNA-staining may be used as reference method to clearly identify apoptosis induced by oxidative stress in adherent cells. The TUNEL-reaction, annexin-V-binding and Apo-2.7-expression may be used to quantify the number of apoptotic and necrotic cells especially at later stages but without discrimination of apoptosis and primary or secondary necrosis.