Using staffing ratios for workforce planning: evidence on nine allied health professions.
Modern healthcare managers are faced with pressure to deliver effective, efficient services within the context of fixed budget constraints. Managers are required to make decisions regarding the skill mix of the workforce particularly when staffing new services. One measure used to identify numbers and mix of staff in healthcare settings is workforce ratio. The aim of this study was to identify workforce ratios in nine allied health professions and to identify whether these measures are useful for planning allied health workforce requirements. A systematic literature search using relevant MeSH headings of business, medical and allied health databases and relevant grey literature for the period 2000-2008 was undertaken. Twelve articles were identified which described the use of workforce ratios in allied health services. Only one of these was a staffing ratio linked to clinical outcomes. The most comprehensive measures were identified in rehabilitation medicine. The evidence for use of staffing ratios for allied health practitioners is scarce and lags behind the fields of nursing and medicine.


Co-creation to scale up provision of simplified high-quality comprehensive abortion care in East Central and Southern Africa.
Universal access to comprehensive abortion care (CAC) is a reproductive right and is essential to reduce preventable maternal mortality and morbidity. In East Africa, abortion rates are consistently high, and the vast majority of all abortions are unsafe, significantly contributing to unnecessary mortality and morbidity. The current debate article reflects and summarises key action points required to continue to speed the implementation of and expand access to CAC in the East, Central, and Southern African (ECSA) health community. To ensure universal access to quality CAC, a regional platform could facilitate the sharing of best practices and successful examples from the region, which would help to visualise opportunities. Such a platform could also identify innovative ways to secure women's access to quality care within legally restrictive environments and would provide information and capacity building through the sharing of recent scientific evidence, guidelines, and training programmes aimed at increasing women's access to CAC at the lowest effective level in the healthcare system. This type of infrastructure for exchanging information and developing co-creation could be crucial to advancing the Sustainable Development Goals 2030 agenda.


Exploring gene expression signatures for predicting disease free survival after resection of colorectal cancer liver metastases.
This study was designed to identify and validate gene signatures that can predict disease free survival (DFS) in patients undergoing a radical resection for their colorectal liver metastases (CRLM). Tumor gene expression profiles were collected from 119 patients undergoing surgery for their CRLM in the Paul Brousse Hospital (France) and the University Medical Center Utrecht (The Netherlands). Patients were divided into high and low risk groups. A randomly selected training set was used to find predictive gene signatures. The ability of these gene signatures to predict DFS was tested in an independent validation set comprising the remaining patients. Furthermore, 5 known clinical risk scores were tested in our complete patient cohort. No gene signature was found that significantly predicted DFS in the validation set. In contrast, three out of five clinical risk scores were able to predict DFS in our patient cohort. No gene signature was found that could predict DFS in patients undergoing CRLM resection. Three out of five clinical risk scores were able to predict DFS in our patient cohort. These results emphasize the need for validating risk scores in independent patient groups and suggest improved designs for future studies.


Factors Associated With Frequent Opioid Use in Children With Acute Recurrent and Chronic Pancreatitis.
The aim of the study was to understand the association of frequent opioid use with disease phenotype and pain pattern and burden in children and adolescents with acute recurrent (ARP) or chronic pancreatitis (CP). Cross-sectional study of children <19 years with ARP or CP, at enrollment into the INSPPIRE cohort. We categorized patients as opioid "frequent use" (daily/weekly) or "nonfrequent use" (monthly or less, or no opioids), based on patient and parent self-report. Of 427 children with ARP or CP, 17% reported frequent opioid use. More children with CP (65%) reported frequent opioid use than with ARP (41%, P = 0.0002). In multivariate analysis, frequent opioid use was associated with older age at diagnosis (odds ratio [OR] 1.67 per 5 years, 95% confidence interval [CI] 1.13-2.47, P = 0.01), exocrine insufficiency (OR 2.44, 95% CI 1.13-5.24, P = 0.02), constant/severe pain (OR 4.14, 95% CI 2.06-8.34, P < 0.0001), and higher average pain impact score across all 6 functional domains (OR 1.62 per 1-point increase, 95% CI 1.28-2.06, P < 0.0001). Children with frequent opioid use also reported more missed school days, hospitalizations, and emergency room visits in the past year than children with no frequent use (P < 0.0002 for each). Participants in the US West and Midwest accounted for 83% of frequent opioid users but only 56% of the total cohort. In children with CP or ARP, frequent opioid use is associated with constant pain, more healthcare use, and higher levels of pain interference with functioning. Longitudinal and prospective research is needed to identify risk factors for frequent opioid use and to evaluate nonopioid interventions for reducing pain and disability in these children.


Rationale and Design of the Randomized Evaluation of Default Access to Palliative Services (REDAPS) Trial.
The substantial nationwide investment in inpatient palliative care services stems from their great promise to improve patient-centered outcomes and reduce costs. However, robust experimental evidence of these benefits is lacking. The Randomized Evaluation of Default Access to Palliative Services (REDAPS) study is a pragmatic, stepped-wedge, cluster randomized trial designed to test the efficacy and costs of specialized palliative care consultative services for hospitalized patients with advanced chronic obstructive pulmonary disease, dementia, or end-stage renal disease, as well as the overall effectiveness of ordering such services by default. Additional aims are to identify the types of services that are most beneficial and the types of patients most likely to benefit, including comparisons between ward and intensive care unit patients. We hypothesize that patient-centered outcomes can be improved without increasing costs by simply changing the default option for palliative care consultation from opt-in to opt-out for patients with life-limiting illnesses. Patients aged 65 years or older are enrolled at 11 hospitals using an integrated electronic health record. As a pragmatic trial designed to enroll between 12,000 and 15,000 patients, eligibility is determined using a validated, electronic health record-based algorithm, and all outcomes are captured via the electronic health record and billing systems data. The time at which each hospital transitions from control, opt-in palliative care consultation to intervention, opt-out consultation is randomly assigned. The primary outcome is a composite measure of in-hospital mortality and length of stay. Secondary outcomes include palliative care process measures and clinical and economic outcomes. Clinical trial registered with www.clinicaltrials.gov (NCT02505035).


Quantitative fluorescence correlation spectroscopy reveals a 1000-fold increase in lifetime of protein functionality.
We have investigated dilute protein solutions with fluorescence correlation spectroscopy (FCS) and have observed that a rapid loss of proteins occurs from solution. It is commonly assumed that such a loss is the result of protein adsorption to interfaces. A protocol was developed in which this mode of protein loss can be prevented. However, FCS on fluorescent protein (enhanced green fluorescent protein, mCherry, and mStrawberry) solutions enclosed by adsorption-protected interfaces still reveals a decrease of the fluorescent protein concentration, while the diffusion time is stable over long periods of time. We interpret this decay as a loss of protein functionality, probably caused by denaturation of the fluorescent proteins. We show that the typical lifetime of protein functionality in highly dilute, approximately single molecule per femtoliter solutions can be extended more than 1000-fold (typically from a few hours to >40 days) by adding compounds with surfactant behavior. No direct interactions between the surfactant and the fluorescent proteins were observed from the diffusion time measured by FCS. A critical surfactant concentration of more than 23 muM was required to achieve the desired protein stabilization for Triton X-100. The surfactant does not interfere with DNA-protein binding, because similar observations were made using DNA-cutting restriction enzymes. We associate the occurrence of denaturation of proteins with the activity of water at the water-protein interface, which was recently proposed in terms of the "water attack model". Our observations suggest that soluble biomolecules can extend an influence over much larger distances than suggested by their actual volume.


Effect of pravastatin on sympathetic reinnervation in postinfarcted rats.
We assessed whether pravastatin attenuates cardiac sympathetic reinnervation after myocardial infarction through the activation of ATP-sensitive K(+) (K(ATP)) channels. Epidemiological studies have shown that men treated with statins appear to have a lower incidence of sudden death than men without statins. However, the specific factor for this has remained disappointingly elusive. Twenty-four hours after ligation of the anterior descending artery, male Wistar rats were randomized to groups treated with either vehicle, nicorandil (a specific mitochondrial K(ATP) channel agonist), pinacidil (a nonspecific K(ATP) channel agonist), pravastatin, glibenclamide (a K(ATP) channel blocker), or a combination of nicorandil and glibenclamide, pinacidil and glibenclamide, or pravastatin and glibenclamide for 4 wk. Myocardial norepinephrine levels revealed a significant elevation in vehicle-treated rats at the remote zone compared with sham-operated rats (2.54 +/- 0.17 vs. 1.26 +/- 0.36 mug/g protein, P < 0.0001), consistent with excessive sympathetic reinnervation after infarction. Immunohistochemical analysis for tyrosine hydroxylase, growth-associated factor 43, and neurofilament also confirmed the change of myocardial norepinephrine. This was paralleled by a significant upregulation of tyrosine hydroxylase protein expression and mRNA in vehicle-treated rats, which was reduced after the administration of either nicorandil, pinacidil, or pravastatin. Arrhythmic scores during programmed stimulation in vehicle-treated rats were significantly higher than those treated with pravastatin. In contrast, the beneficial effects of pravastatin were reversed by the addition of glibenclamide, implicating K(ATP) channels as the relevant target. The sympathetic reinnervation after infarction is modulated by the activation of K(ATP) channels. Chronic use of pravastatin after infarction, resulting in attenuated sympathetic reinnervation by the activation of K(ATP) channels, may modify the arrhythomogenic response to programmed electrical stimulation.


Medical records and quality of care in acute coronary syndromes: results from CRUSADE.
Patient medical records are important means of communication among health care providers. Limited evaluation has been performed of the quality of the medical records or its association with health care processes or outcomes. We performed an empirical evaluation of the completeness of medical records from 607 randomly selected patients admitted with non-ST-segment elevation acute coronary syndromes (NSTE ACS) to 219 US hospitals in the CRUSADE National Quality Improvement Initiative. Composite medical records scores were summated and compared by hospital academic status and physician specialty. Correlations between medical records scores, use of evidence-based medicine (EBM), and in-hospital mortality were assessed. Medical records were frequently missing key elements, including cardiac history (23.6%), performance status (64.6%), differential diagnosis (57.8%), and planned use of EBM (44.0%). Evidence-based medicine was more often discussed in medical records from academic medical centers vs nonacademic medical centers (69.7% vs 51.7%) (P < .001) and from cardiologists vs noncardiologists (60.5% vs 48.1%, P = .003). Higher medical records quality scores were associated with greater use of EBM among the medical records quality cohort (P = .006), and a similar trend was observed in CRUSADE overall: adjusted odds ratio, 1.26 (95% confidence interval, 0.92-1.72) for high vs low medical records quality. Higher medical records quality scores were associated with lower in-hospital mortality: adjusted odds ratio, 0.79 (95% confidence interval, 0.65-0.97). Medical records for patients with NSTE ACS often lack key elements of the history and physical examination. Patients treated at hospitals with better medical records quality have significantly lower mortality and may receive more EBM. The relationship between better medical charting and better medical care could lead to new ways to monitor and improve the quality of medical care.


An Integrated Methodology Using Extended Swara and Dea for the Performance Analysis of Wastewater Treatment Plants: Turkey Case.
Public and private companies make significant water infrastructure investments to meet increasing water demand. In this context, investments in wastewater treatment plants (WWTPs), which play an important role in recycling of used water, are also increasing. This study investigates determination of the efficiency scores of WWTPs considering each metropolitan municipality as a decision-making unit (DMU). In this study, a two-step methodology is established to determine efficiency scores of WWTPs. In the first step, the input and output parameters are searched by a literature review for the performance evaluation, and candidate parameters are determined. Then, to determine the most appropriate and related parameters, the importance weights of all candidate inputs and outputs are computed using the extended stepwise weight assessment ratio analysis (SWARA) method. Next, the inputs and outputs are chosen according to their importance weights. In the second step, efficiency scores of WWTPs are calculated using output-oriented data envelopment analysis (DEA) models. Based on the expert opinions, the parameters used as input variables are as follows: Daily Wastewater Amount per Person Discharged in Municipalities, WWTP Capacity, and Number of WWTPs; and the parameters used as output variables are as follows; Amount of Wastewater Treated in WWTPs and Municipal Population Served by WWTPs. The results are presented and discussed by sensitivity analysis. Results show that 14 metropolitan municipalities have total efficiency, 19 metropolitan municipalities have technical efficiency, and 21 metropolitan municipalities have scale efficiency.


Using bioinformatics techniques for gene identification in drug discovery and development.
As more and more evidence has become available, the link between gene and emergent disease has been made including cancer, heart disease and parkinsonism. Analyzing the diseases and designing drugs with respect to the gene and protein level obviously help to find the underlying causes of the diseases, and to improve their rate of cure. The development of modern molecular biology, biochemistry, data collection and analysis techniques provides the scientists with a large amount of gene data. To draw a link between genes and their relation to disease outcomes and drug discovery is a big challenge: how to analyze large datasets and extract useful knowledge? Combining bioinformatics with drug discovery is a promising method to tackle this issue. Most techniques of bioinformatics are used in the first two phases of drug discovery to extract interesting information and find important genes and/or proteins for speeding the process of drug discovery, enhancing the accuracy of analysis and reducing the cost. Gene identification is a very fundamental and important technique among them. In this paper, we have reviewed gene identification algorithms and discussed their usage, relationships and challenges in drug discovery and development.


Stakeholder perspectives on managing the occupational health of UK business drivers: a qualitative approach.
Musculoskeletal disorders are one of the leading causes of work related ill health and sickness absence. Those who drive as part of their job may be at particular risk, with evidence suggesting that prolonged exposure to driving is associated with increased absence from work due to low back pain. Business drivers often work away from a traditional office environment. Such mobile working may pose greater risks to occupational health due to increased ergonomic risks, for example working from the car, longer working hours and a lack of concern amongst drivers about health and safety. It has been suggested that occupational health practices have not adapted to meet the needs of peripatetic workers. The current study explored how occupational health services are delivered to business drivers. Semi-structured interviews were carried out with a sample of 31 stakeholders in 4 organisations. Respondents included, health and safety professionals, occupational health nurses, fleet managers and high mileage business drivers. The interviews were transcribed and analysed using 'Template Analysis'. The data revealed that, within these organisations, the provision of occupational health services was often fragmented and drivers and other key stakeholders were often unaware of the existing systems within their organisations. The peripatetic nature of business drivers meant that they were difficult for occupational health teams to reach. The paper concludes by presenting recommendations for occupational health professionals and researchers engaged with improving the health of peripatetic workers, namely that occupational health policies should be integrated in company strategy and widely disseminated to drivers and those with responsibility for managing their occupational health provision.


Respiratory symptoms, immunological responses, and aeroallergen concentrations at a sawmill.
After identification of a case of extrinsic allergic alveolitis due to exposure to wood dust at a sawmill, all employees at the sawmill where he worked were studied with an occupational, environmental, and symptom questionnaire, spirometry, skin prick tests, and serum specific IgG measurements. Ninety five of current and 14 of 17 ex-sawmill workers were studied. As a basis for comparison, a group of 58 workers from a nearby light engineering factory were also studied. Few women (6) were employed and they were excluded from the analysis. Workers at the sawmill were stratified into high and low exposure groups depending on their place of work. This division was supported both by their subjective assessment of the dustiness of their environment and the results of personal dust samples. There were no significant differences between the three groups in age, height, smoking habits, exposure to other causes of extrinsic allergic alveolitis, forced expiratory volume in one second, forced vital capacity, atopic state, or cutaneous reactivity to moulds. In the high exposure group the prevalence of work related cough and nasal and eye symptoms was higher than in the low exposure and comparison groups. The prevalence of work related wheeze was similar in both the high exposure and comparison groups, but was lower in the low exposure group. The prevalences of chronic bronchitis and symptomatic bronchial hyper-reactivity were similar in the high and low exposure groups but were lower in the comparison group. Serum concentrations of specific IgG against extracts of sawdust and Trichoderma koningii were significantly higher in the high exposure group than in the other two groups. The prevalence of symptoms suggestive of extrinsic allergic alveolitis was 4.4% in the high exposure group, greater than in the low exposure group (0%), and the comparison group (1.9%). In conclusion extrinsic allergic alveolitis probably occurs in British sawmills, and among the exposed population its prevalence may be as high as that reported in Sweden. The allergen responsible is likely to be from mould growing on the wood and may be from Trichoderma koningii.


Breast cancer in systemic lupus.
Objective There is a decreased breast cancer risk in systemic lupus erythematosus (SLE) versus the general population. We assessed a large sample of SLE patients, evaluating demographic and clinical characteristics and breast cancer risk. Methods We performed case-cohort analyses within a multi-center international SLE sample. We calculated the breast cancer hazard ratio (HR) in female SLE patients, relative to demographics, reproductive history, family history of breast cancer, and time-dependent measures of anti-dsDNA positivity, cumulative disease activity, and drugs, adjusted for SLE duration. Results There were 86 SLE breast cancers and 4498 female SLE cancer-free controls. Patients were followed on average for 7.6 years. Versus controls, SLE breast cancer cases tended to be white and older. Breast cancer cases were similar to controls regarding anti-dsDNA positivity, disease activity, and most drug exposures over time. In univariate and multivariate models, the principal factor associated with breast cancers was older age at cohort entry. Conclusions There was little evidence that breast cancer risk in this SLE sample was strongly driven by any of the clinical factors that we studied. Further search for factors that determine the lower risk of breast cancer in SLE may be warranted.


Vitamin D in plants: a review of occurrence, analysis, and biosynthesis.
The major function of vitamin D in vertebrates is maintenance of calcium homeostasis, but vitamin D insufficiency has also been linked to an increased risk of hypertension, autoimmune diseases, diabetes, and cancer. Therefore, there is a growing awareness about vitamin D as a requirement for optimal health. Vitamin D3 is synthesized in the skin by a photochemical conversion of provitamin D3, but the necessary rays are only emitted all year round in places that lie below a 35° latitude. Unfortunately, very few food sources naturally contain vitamin D and the general population as a results fail to meet the requirements. Fish have the highest natural content of vitamin D expected to derive from an accumulation in the food chain originating from microalgae. Microalgae contain both vitamin D3 and provitamin D3, which suggests that vitamin D3 exist in the plant kingdom and vitamin D3 has also been identified in several plant species as a surprise to many. The term vitamin D also includes vitamin D2 that is produced in fungi and yeasts by UVB-exposure of provitamin D2. Small amounts can be found in plants contaminated with fungi and traditionally only vitamin D2 has been considered present in plants. This review summarizes the current knowledge on sterol biosynthesis leading to provitamin D. It also addresses the occurrence of vitamin D and its hydroxylated metabolites in higher plants and in algae and discusses limitations and advantages of analytical methods used in studies of vitamin D and related compounds including recent advances in analytical technologies. Finally, perspectives for a future production of vitamin D biofortified fruits, vegetables, and fish will be presented.


Clear Cell Carcinoma of the Uterine Cervix: A Clinical and Pathological Analysis of 47 Patients Without Intrauterine Diethylstilbestrol Exposure.
The aim of this study was to summarize the clinical and pathological characteristics and to conduct prognosis analysis of patients who were diagnosed with clear cell carcinoma of the uterine cervix (CCCUC) and without a history of exposure to diethylstilbestrol. We performed a retrospective review of all the patients with CCCUC who were diagnosed and treated at Zhejiang Cancer Hospital between 1998 and 2014. Charts were reviewed for clinical and pathological characteristics, and prognosis analysis was conducted. A total of 47 patients were included. Median age was 52 years. No patient had a history of exposure to diethylstilbestrol. The International Federation of Gynecology and Obstetrics stage distribution was 55.3% (n = 26) stage I, 40.4% (n = 19) stage II, 2.1% (n = 1) stage III, and 2.1% (n = 1) stage IV. Forty-two patients (89.4%) underwent radical hysterectomy and pelvic lymphadenectomy. Pathological examination revealed deep cervical stromal invasion (greater than two thirds) in 20 patients (48.4%), pelvic lymph node (PLN) metastasis in 10 patients (23.8%), lymphovascular space involvement in 9 patients (21.4%), and ovarian metastasis in 1 patient (2.4%). Advanced tumor stage (IIB-IV), larger tumor size (>4 cm), and PLN metastasis had negative effects on progression-free survival (PFS) and overall survival (OS) (P < 0.05). Adjuvant radiation therapy alone or concurrent chemoradiation therapy after radical surgery did not affect PFS or OS in patients with risk factors (P > 0.05). International Federation of Gynecology and Obstetrics stage, tumor size, and PLN status were prognostic factors for both PFS and OS in patients with CCCUC. The long-term effects of adjuvant radiation therapy or concurrent chemoradiation therapy may be limited for CCCUC patients with risk factors. Future larger case series or clinical trials are required to confirm these findings.


Sugar-sweetened beverages, serum uric acid, and blood pressure in adolescents.
To evaluate whether sugar-sweetened beverage consumption, a significant source of dietary fructose, is associated with higher serum uric acid levels and blood pressure in adolescents. We analyzed cross-sectional data from 4867 adolescents aged 12 to 18 years in the National Health and Nutrition Examination Survey, 1999-2004. Dietary data were assessed from 24-hour dietary recall interviews. Sugar-sweetened beverages included fruit drinks, sports drinks, soda, and sweetened coffee or tea. We used multivariate linear regression to evaluate the association of sugar-sweetened beverage consumption with serum uric acid and with blood pressure. Adolescents who drank more sugar-sweetened beverages tended to be older and male. In the adjusted model, serum uric acid increased by 0.18 mg/dL and systolic blood pressure z-score increased by 0.17 from the lowest to the highest category of sugar-sweetened beverage consumption (P for trend, .01 and .03, respectively). These results from a nationally representative sample of US adolescents indicate that higher sugar-sweetened beverage consumption is associated with higher serum uric acid levels and systolic blood pressure, which may lead to downstream adverse health outcomes.


Additive manufacturing techniques for smart prosthetic liners.
Elastomeric liners are commonly worn between the prosthetic socket and the limb. A number of improvements to the state of the art of liner technology are required to address outstanding problems. A liner that conforms to the residuum more accurately, may improve the skin health at the stump-socket interface. Previous work has shown that for effective thermal management of the socket environment, an active heat removal system is required, yet this is not available. Volume tracking of the stump could be used as a diagnostic tool for looking at the changes that occur across the day for all users, which depend on activity level, position, and the interaction forces of the prosthetic socket with the limb. We believe that it would be advantageous to embed these devices into a smart liner, which could be replaced and repaired more easily than the highly costly and labour-intensive custom-made socket. This paper presents the work to develop these capabilities in soft material technology, with: the development of a printable nanocomposite stretch sensor system; a low-cost digital method for casting bespoke prosthetic liners; a liner with an embedded stretch sensor for growth / volume tracking; a model liner with an embedded active cooling system.


Crystallization and X-ray structure of full-length recombinant human butyrylcholinesterase.
Human butyrylcholinesterase (BChE) has been shown to function as an endogenous scavenger of diverse poisons. BChE is a 340 kDa tetrameric glycoprotein that is present in human serum at a concentration of 5 mg l(-1). The well documented therapeutic effects of BChE on cocaine toxicity and organophosphorus agent poisoning has increased the need for effective methods of producing recombinant therapeutic BChE. In order to be therapeutically useful, BChE must have a long circulatory residence time or associate as tetramers. Full-length recombinant BChE produced in Chinese hamster ovary (CHO) cells or human embryonic kidney cells has been shown to associate as monomers, with a shorter circulatory residence time than the naturally occurring tetrameric serum protein. Based on the preceding observation as well as the need to develop novel methodologies to facilitate the mass production of therapeutic recombinant BChE, studies have been initiated to determine the structural basis of tetramer formation. Towards these ends, full-length monomeric recombinant BChE has been crystallized for the first time. A 2.8 A X-ray structure was solved in space group P42(1)2, with unit-cell parameters a = b = 156, c = 146 A.


Adsorption of perfluoroalkyl substances on microplastics under environmental conditions.
Plastic debris has become an environmental problem during recent years. Among the plastic debris, microplastics (<5 mm; MPLs) imply an extra problem due to their capacity to enter into the fauna through ingestion. In this work, we study the capacity of three MPLs, that include high-density polyethylene (HDPE), polystyrene (PS) and polystyrene carboxylate (PS-COOH), to sorb 18 perfluoroalkyl substances (PFASs; including carboxylic acids, sulphonates and one sulphonamide) from the surrounding waters (freshwater and seawater). Conclusions drawn from the results are that perfluoro sulphonates and sulphonamides have more tendency to be sorbed onto MPLs. In addition, PS and PS-COOH have more affinity for PFASs than HDPE. Finally, the increment of conductivity and pH of the water decreases the exposure time that is necessary to reach equilibrium. However, the presence of salts decreases the tendency of PFASs to be sorbed onto plastic surfaces. These results highlight the problem associated with the presence of MPLs in inland and marine waters since toxic compounds can be sorbed onto surrounding plastics that could be ingested by aquatic fauna.


C1 inhibitor deficiency: consensus document.
We present a consensus document on the diagnosis and management of C1 inhibitor deficiency, a syndrome characterized clinically by recurrent episodes of angio-oedema. In hereditary angio-oedema, a rare autosomal dominant condition, C1 inhibitor function is reduced due to impaired transcription or production of non-functional protein. The diagnosis is confirmed by the presence of a low serum C4 and absent or greatly reduced C1 inhibitor level or function. The condition can cause fatal laryngeal oedema and features indistinguishable from gastrointestinal tract obstruction. Attacks can be precipitated by trauma, infection and other stimulants. Treatment is graded according to response and the clinical site of swelling. Acute treatment for severe attack is by infusion of C1 inhibitor concentrate and for minor attack attenuated androgens and/or tranexamic acid. Prophylactic treatment is by attenuated androgens and/or tranexamic acid. There are a number of new products in trial, including genetically engineered C1 esterase inhibitor, kallikrein inhibitor and bradykinin B2 receptor antagonist. Individual sections provide special advice with respect to diagnosis, management (prophylaxis and emergency care), special situations (childhood, pregnancy, contraception, travel and dental care) and service specification.


Combining two-dimensional diffusion-ordered nuclear magnetic resonance spectroscopy, imaging desorption electrospray ionization mass spectrometry, and direct analysis in real-time mass spectrometry for the integral investigation of counterfeit pharmaceuticals.
During the past decade, there has been a marked increase in the number of reported cases involving counterfeit medicines in developing and developed countries. Particularly, artesunate-based antimalarial drugs have been targeted, because of their high demand and cost. Counterfeit antimalarials can cause death and can contribute to the growing problem of drug resistance, particularly in southeast Asia. In this study, the complementarity of two-dimensional diffusion-ordered (1)H nuclear magnetic resonance spectroscopy (2D DOSY (1)H NMR) with direct analysis in real-time mass spectrometry (DART MS) and desorption electrospray ionization mass spectrometry (DESI MS) was assessed for pharmaceutical forensic purposes. Fourteen different artesunate tablets, representative of what can be purchased from informal sources in southeast Asia, were investigated with these techniques. The expected active pharmaceutical ingredient was detected in only five formulations via both nuclear magnetic resonance (NMR) and mass spectrometry (MS) methods. Common organic excipients such as sucrose, lactose, stearate, dextrin, and starch were also detected. The graphical representation of DOSY (1)H NMR results proved very useful for establishing similarities among groups of samples, enabling counterfeit drug "chemotyping". In addition to bulk- and surface-average analyses, spatially resolved information on the surface composition of counterfeit and genuine antimalarial formulations was obtained using DESI MS that was performed in the imaging mode, which enabled one to visualize the homogeneity of both genuine and counterfeit drug samples. Overall, this study suggests that 2D DOSY (1)H NMR, combined with ambient MS, comprises a powerful suite of instrumental analysis methodologies for the integral characterization of counterfeit antimalarials.


GLP-1 metabolite GLP-1(9-36) is a systemic inhibitor of mouse and human pancreatic islet glucagon secretion.
Diabetes mellitus is associated with impaired insulin secretion, often aggravated by oversecretion of glucagon. Therapeutic interventions should ideally correct both defects. Glucagon-like peptide 1 (GLP-1) has this capability but exactly how it exerts its glucagonostatic effect remains obscure. Following its release GLP-1 is rapidly degraded from GLP-1(7-36) to GLP-1(9-36). We hypothesised that the metabolite GLP-1(9-36) (previously believed to be biologically inactive) exerts a direct inhibitory effect on glucagon secretion and that this mechanism becomes impaired in diabetes. We used a combination of glucagon secretion measurements in mouse and human islets (including islets from donors with type 2 diabetes), total internal reflection fluorescence microscopy imaging of secretory granule dynamics, recordings of cytoplasmic Ca2+ and measurements of protein kinase A activity, immunocytochemistry, in vivo physiology and GTP-binding protein dissociation studies to explore how GLP-1 exerts its inhibitory effect on glucagon secretion and the role of the metabolite GLP-1(9-36). GLP-1(7-36) inhibited glucagon secretion in isolated islets with an IC50 of 2.5 pmol/l. The effect was particularly strong at low glucose concentrations. The degradation product GLP-1(9-36) shared this capacity. GLP-1(9-36) retained its glucagonostatic effects after genetic/pharmacological inactivation of the GLP-1 receptor. GLP-1(9-36) also potently inhibited glucagon secretion evoked by β-adrenergic stimulation, amino acids and membrane depolarisation. In islet alpha cells, GLP-1(9-36) led to inhibition of Ca2+ entry via voltage-gated Ca2+ channels sensitive to ω-agatoxin, with consequential pertussis-toxin-sensitive depletion of the docked pool of secretory granules, effects that were prevented by the glucagon receptor antagonists REMD2.59 and L-168049. The capacity of GLP-1(9-36) to inhibit glucagon secretion and reduce the number of docked granules was lost in alpha cells from human donors with type 2 diabetes. In vivo, high exogenous concentrations of GLP-1(9-36) (>100 pmol/l) resulted in a small (30%) lowering of circulating glucagon during insulin-induced hypoglycaemia. This effect was abolished by REMD2.59, which promptly increased circulating glucagon by >225% (adjusted for the change in plasma glucose) without affecting pancreatic glucagon content. We conclude that the GLP-1 metabolite GLP-1(9-36) is a systemic inhibitor of glucagon secretion. We propose that the increase in circulating glucagon observed following genetic/pharmacological inactivation of glucagon signalling in mice and in people with type 2 diabetes reflects the removal of GLP-1(9-36)'s glucagonostatic action.


CAR T-Cell Therapy: The Role of Physical Barriers and Immunosuppression in Lymphoma.
Chimeric antigen receptor (CAR) T-cells have shown remarkable results in patients with B-cell leukemia and lymphoma. However, while CAR T-cells have shown complete responses in a majority of patients with acute lymphoblastic leukemia (ALL), lymphomas are more difficult to treat. Different CAR designs and conditioning protocols seem to affect the persistence of patient responses. However, factors that determine if patients receiving the same CARs will respond or not remain obscure. In Sweden, a phase I/IIa trial using third-generation CAR T-cells is ongoing in which we intend to compare tumor biology and immunology, in each patient, to treatment response. CAR T-cell therapy is a powerful tool to add to the treatment options for this patient group but we need to perform the necessary basic research on the multifactorial mechanisms of action to give patients the best possible option of survival. Such studies are also crucial to expand the success of CAR T-cells beyond CD19+ B-cell malignancy. This review will focus on possible barriers of treating lymphoma to define factors that need to be investigated to develop the next generation of CAR T-cell therapy.


ICSH recommendations for identification, diagnostic value, and quantitation of schistocytes.
Schistocytes are fragments of red blood cells (RBCs) produced by extrinsic mechanical damage within the circulation. The detection of schistocytes is an important morphological clue to the diagnosis of thrombotic microangiopathic anemia (TMA). Reporting criteria between different laboratories, however, are not uniform, owing to variability of shape and nature of fragments, as well as subjectivity and heterogeneity in their morphological assessment. Lack of standardization may lead to inconsistency or misdiagnosis, thereby affecting treatment and clinical outcome. The Schistocyte Working Group of the International Council for Standardization in Haematology (ICSH) has prepared specific recommendations to standardize schistocyte identification, enumeration, and reporting. They deal with the type of smear, method of counting, morphological description based on positive criteria (helmet cells, small, irregular triangular, or crescent-shaped cells, pointed projections, and lack of central pallor). A schistocyte count has a definite clinical value for the diagnosis of TMA in the absence of additional severe red cell shape abnormalities, with a confident threshold value of 1%. Automated counting of RBC fragments is also recommended by the ICSH Working Group as a useful complement to the microscope, according to the high predictive value of negative results, but worthy of further research and with limits in quantitation.


Differential treatment response of subtypes of patients with borderline personality organization, as assessed with theory-driven profiles of the Dutch short form of the MMPI: a naturalistic follow-up study.
We investigated the validity of different subtypes of borderline personality organization (BPO) as assessed by theory-driven profiles of the Minnesota Multiphasic Personality Disorder (MMPI; Hathaway & McKinley, 1943 ) Dutch Short Form (DSFM; Eurelings-Bontekoe, Onnink, Williams, & Snellen, 2008 ) in a naturalistic follow-up study among 2,062 psychiatric outpatients who received 6 months of ambulatory treatment. Patients were assessed at intake (T1) and 6 months later (T2). At T2, both patients and therapists rated the level of improvement, using the Global Assessment of Improvement. Patients with the high-level BPO profile showed the largest increase in well-being and the largest decrease in severity of symptomatology, whereas severity of symptomatology and well-being of patients with psychotic BPO profiles did not change over time. Agreement between patients and therapists about improvement was good for the internalizing immature BPO and high-level BPO patients, but poor for the externalizing low-level BPO and narcissistic patients.


Rural-urban differences in health among youth in northern Sweden: an outcome-wide epidemiological approach.
The aim of this research was to contribute knowledge about rural-urban differences in health among young northern Swedish women and men. This study was based on the 2014 "Health on Equal Terms" survey, distributed in the four northernmost counties of Sweden, with complementary information on areas of residence classified as rural, semi-urban and urban from total population registers. The analytical sample included 2,691 individuals who were selected using a probabilistic sampling method. Prevalence ratios were calculated in multivariable log-binomial regression analyses to measure the association between place of residence and nine outcomes covering three health dimensions (general, mental and lifestyle behaviours). The results indicated that daily smoking and being overweight were more common, while feelings of stress and psychological distress were less prevalent, among youths in rural as compared to urban areas. After including covariates, this pattern appeared stronger for young women, although the direction of the results also applied to young men, albeit without revealing significant differences. In conclusion, the findings from this study indicate that for youths - particularly young women - the rural setting may imply an increased risk of poor general health and lifestyle behaviours, while simultaneously playing a partially protective role for mental health.


Alternative strategies for development of a reference transcriptome for quantification of allele specific expression in organisms having sparse genomic resources.
In recent years RNA-Seq technology has been used not only to quantify differences in gene expression but also to understand the underlying mechanisms that lead to these differences. Nucleotide sequence variation arising through evolution may differentially affect the expression profiles of divergent species. RNA-Seq technology, combined with techniques to differentiate parental alleles and quantify their abundance, have recently become popular methods for allele specific gene expression (ASGE) analyses. However, analysis of gene expression within interspecies hybrids may be difficult when one of the two parental genomes represented in the hybrid does not have robust genomic resources or available transcriptome data. Herein, we compare two strategies for analyzing allele specific expression within interspecies hybrids produced from crossing two Xiphophorus fish species. The first strategy relies upon a robust reference transcriptome assembly from one species followed by identification of SNPs and creation of an in silico reference transcriptome for the second species. The second strategy employs de novo assembly of reference transcriptomes for both parental species followed by identification of homologous transcripts prior to mapping hybrid reads to a combined hybrid reference. Our results show that, although both methods are able to achieve balanced allelic distribution upon read mapping of F(1) hybrid fish transcriptomes, the second "de novo" assembly approach is superior for ASGE analyses and leads to results more consistent with those found from quantitative real time PCR assessment of gene expression. In addition, our analysis indicates that indels between the two parental alleles are the major cause of the differences in results observed when employing these two methods.


Analysis of ancient human mitochondrial DNA from the Xiaohe cemetery: insights into prehistoric population movements in the Tarim Basin, China.
The Tarim Basin in western China, known for its amazingly well-preserved mummies, has been for thousands of years an important crossroad between the eastern and western parts of Eurasia. Despite its key position in communications and migration, and highly diverse peoples, languages and cultures, its prehistory is poorly understood. To shed light on the origin of the populations of the Tarim Basin, we analysed mitochondrial DNA polymorphisms in human skeletal remains excavated from the Xiaohe cemetery, used by the local community between 4000 and 3500 years before present, and possibly representing some of the earliest settlers. Xiaohe people carried a wide variety of maternal lineages, including West Eurasian lineages H, K, U5, U7, U2e, T, R*, East Eurasian lineages B, C4, C5, D, G2a and Indian lineage M5. Our results indicate that the people of the Tarim Basin had a diverse maternal ancestry, with origins in Europe, central/eastern Siberia and southern/western Asia. These findings, together with information on the cultural context of the Xiaohe cemetery, can be used to test contrasting hypotheses of route of settlement into the Tarim Basin.


New catalytic approaches towards the enantioselective halogenation of alkenes.
The addition of electrophilic reagents to the carbon-carbon double bond is one of the most fundamental reactions in organic chemistry. Halogen electrophiles constitute probably the most important class of electrophiles and have been widely used to induce electrophilic addition reactions to alkenes like halolactonizations or dihalogenations. Despite their long history and high importance, catalytic, asymmetric variants of these reactions have been underdeveloped until very recently. During the last two years this has changed and many novel approaches have been reported. This review aims to cover these new developments through discussing the common themes as well as the suggested mechanistic scenarios.


A Specific Mutation in Muc2 Determines Early Dysbiosis in Colitis-Prone Winnie Mice.
Inflammatory bowel disease (IBD), including Crohn disease (CD) and ulcerative colitis (UC), is a multifactorial disorder characterized by chronic inflammation and altered gut barrier function. Dysbiosis, a condition defined by dysregulation of the gut microbiome, has been reported in patients with IBD and in experimental models of colitis. Although several factors have been implicated in directly affecting gut microbial composition, the genetic determinants impacting intestinal dysbiosis in IBD remain relatively unknown. We compared the microbiome of normal, uninflamed wild-type (WT) mice with that of a murine model of UC (ie, Winnie strain). Winnie mice possess a missense mutation in Muc2 that manifests in altered mucus production as early as 4 weeks of age, with ensuing colonic inflammation. To better address the potential role of mutant Muc2 in promoting dysbiosis in Winnie mice, we evaluated homozygous mutant mice (Winnie-/-) with their WT littermates that, after weaning from common mothers, were caged separately according to genotype. Histologic and inflammatory status were assessed over time, along with changes in their respective microbiome compositions. Dysbiosis in Winnie mice was already established at 4 weeks of age, before histologic evidence of gut inflammatory changes, in which microbial communities diverged from that derived from their mothers. Furthermore, dysbiosis persisted until 12 weeks of age, with peak differences in microbiome composition observed between Winnie and WT mice at 8 weeks of age. The relative abundance of Bacteroidetes was greater in Winnie compared with WT mice. Verrucomicrobia was detected at the highest relative levels in 4-week-old Winnie mice; in particular, Akkermansia muciniphila was among the most abundant species found at 4 weeks of age. Our results demonstrate that mutant genetic determinants involved in the complex regulation of intestinal homeostasis, such as that observed in Winnie mice, are able to promote early gut dysbiosis that is independent from maternal microbial transfer, including breastfeeding. Our data provide evidence for intestinal dysbiosis attributed to a Muc2-driven mucus defect that leads to colonic inflammation and may represent an important target for the design of future interventional studies.


Why don't academic physicians seek needed professional help for psychological distress?
Suicidal thoughts, burnout and other signs of psychological distress are prevalent among physicians. There are no studies concerning help-seeking for psychological distress among university hospital physicians, who face a particularly challenging, competitive work environment. We compare psychologically-distressed university hospital physicians who have not sought needed help with those who have sought such help. We thereby aim to identify factors that may hinder help-seeking and factors that may trigger seeking help. Analysis was performed among university hospital physicians reporting recent suicidal thoughts and/or showing other indications of current psychological ill-health. These distressed physicians were a subgroup (42.7%) from the cross-sectional phase I HOUPE study (Health and Organization among University Hospital Physicians in Europe): 366 from Sweden and 150 from Italy. Having sought professional help for depression or burnout was the outcome variable. Multiple logistic regression was performed with socio-demographic factors as covariates. Altogether 404 (78.3%) of these distressed physicians had never sought professional help for depression/burnout. Physicians who were currently involved in medical research, taking night call, surgical specialists, male, or Italian were least likely to have sought help. Physicians who faced harassment at work or who self-diagnosed and self-treated were more likely to have sought help. Very few of these university hospital physicians with signs of psychological distress sought help from a mental-health professional. This has implications for physicians themselves and for patient care, clinical research, and education of future physicians. More study, preferably of interventional design, is warranted concerning help-seeking among these physicians in need.


Dam and Dcm methylations prevent gene transfer into Clostridium pasteurianum NRRL B-598: development of methods for electrotransformation, conjugation, and sonoporation.
Butanol is currently one of the most discussed biofuels. Its use provides many benefits in comparison to bio-ethanol, but the price of its fermentative production is still high. Genetic improvements could help solve many problems associated with butanol production during ABE fermentation, such as its toxicity, low concentration achievable in the cultivation medium, the need for a relatively expensive substrate, and many more. Clostridium pasteurianum NRRL B-598 is non-type strain producing butanol, acetone, and a negligible amount of ethanol. Its main benefits are high oxygen tolerance, utilization of a wide range of carbon and nitrogen sources, and the availability of its whole genome sequence. However, there is no established method for the transfer of foreign DNA into this strain; this is the next step necessary for progress in its use for butanol production. We have described functional protocols for conjugation and transformation of the bio-butanol producer C. pasteurianum NRRL B-598 by foreign plasmid DNA. We show that the use of unmethylated plasmid DNA is necessary for efficient transformation or successful conjugation. Genes encoding DNA methylation and those for restriction-modification systems and antibiotic resistance were searched for in the whole genome sequence and their homologies with other clostridial bacteria were determined. Furthermore, activity of described novel type I restriction system was proved experimentally. The described electrotransformation protocol achieved an efficiency 1.2 × 10(2) cfu/μg DNA after step-by-step optimization and an efficiency of 1.6 × 10(2) cfu/μg DNA was achieved by the sonoporation technique using a standard laboratory ultrasound bath. The highest transformation efficiency was achieved using a combination of these approaches; sono/electroporation led to an increase in transformation efficiency, to 5.3 × 10(2) cfu/μg DNA. Both Dam and Dcm methylations are detrimental for transformation of C. pasteurianum NRRL B-598. Methods for conjugation, electroporation, sonoporation, and a combined method for sono/electroporation were established for this strain. The methods described could be used for genetic improvement of this strain, which is suitable for bio-butanol production.


Interaction Between Breast Cancer Cells and Adipose Tissue Cells Derived from Fat Grafting.
Adipose tissue transplantation has the benefit of providing both regenerative and aesthetic outcomes in breast cancer treatment. However, the transplanted tissue can stimulate the growth of residual cancer cells. The aim of this study is to identify the interactions between adipose tissue cell subpopulations and human cancer cell lines. Intact adipose tissue from lipofilling procedures as well as fibroblasts derived from adipose tissue, were cocultured in the presence of MDA-MB-231, MCF-7 e ZR-75-1 breast cancer cell lines. The influence on cancer cell lines of fibroblasts, induced to differentiate into specific adipocytes, was also assayed. All cancer cell lines displayed a significant increase in proliferation rate when cocultured in the presence of either intact adipose tissue or induced adipocytes. To a lesser extent, uninduced fibroblasts stimulate breast cancer cell proliferation. Recent studies have shown that the microenvironment surrounding breast cancer cells may stimulate growth and promote progression of residual cancer cells when surgery is performed on the main tumor mass. Accordingly, the graft of adipose tissue could potentially promote or accelerate the development of a subclinical tumor or support its locoregional recurrence. Our data suggest that adipocytes have a remarkable influence on the proliferation of cancer cell lines. The oncological safety of the lipofilling procedure outcome is still debated; thus, further studies and consistent follow-up examination are needed.


State hospital rate-setting revisited.
In an attempt to control rapid growth in hospital costs, beginning in the mid-1970s several states implemented rate-setting programs to regulate hospital payments. In seven states, rate-setting was in effect for a substantial period of time (14 years or more). While most of these programs were discontinued by the mid-1990s, two are still active. In five of the seven states, the rates of increase in hospital costs were lower than the corresponding national rates during the periods in which the regulation programs were in place. Four of the states--Maryland, Massachusetts, New York, and New Jersey--had some of the lowest rates of hospital cost increases among all the states. This indicates that hospital rate regulation may be a useful approach in managing a major component of health care spending.


Parents' Beliefs About Physical Activity for Their Children With Visual Impairments.
Despite having the desire to become physically active as a family, parents of children with visual impairments often lack the skills and resources needed to provide appropriate physical activities (PAs) for their children. The purpose of this study was to explore the intentions of parents of children with visual impairments toward including their children in PAs after participating in a PA program. In this descriptive qualitative study, the participants were 10 parents of children with visual impairments. A series of workshops were designed to provide parents with the skills and resources needed to promote PA for their family. Upon completion of the workshops, parents took part in one-on-one semistructured interviews that were subsequently transcribed and analyzed using a thematic line-by-line process. Two interdependent themes emerged from the data analyses: (a) eye-opening experiences and (b) transformed, more hopeful, and optimistic outlook. The results revealed that through the PA intervention, parents learned teaching strategies that were intended to increase their PA opportunities and garnered resources that allowed them to teach their children to participate in PA.


The relationship between non-communicable disease occurrence and poverty-evidence from demographic surveillance in Matlab, Bangladesh.
In low-income countries, a growing proportion of the disease burden is attributable to non-communicable diseases (NCDs). There is little knowledge, however, of their impact on wealth, human capital, economic growth or household poverty. This article estimates the risk of being poor after an NCD death in the rural, low-income area of Matlab, Bangladesh. In a matched cohort study, we estimated the 2-year relative risk (RR) of being poor in Matlab households with an NCD death in 2010. Three separate measures of household economic status were used as outcomes: an asset-based index, self-rated household economic condition and total household landholding. Several estimation methods were used including contingency tables, log-binomial regression and regression standardization and machine learning. Households with an NCD death had a large and significant risk of being poor. The unadjusted RR of being poor after death was 1.19, 1.14 and 1.10 for the asset quintile, self-rated condition and landholding outcomes. Adjusting for household and individual level independent variables with log-binomial regression gave RRs of 1.19 [standard error (SE) 0.09], 1.16 (SE 0.07) and 1.14 (SE 0.06), which were found to be exactly the same using regression standardization (SE: 0.09, 0.05, 0.03). Machine learning-based standardization produced slightly smaller RRs though still in the same order of magnitude. The findings show that efforts to address the burden of NCD may also combat household poverty and provide a return beyond improved health. Future work should attempt to disentangle the mechanisms through which economic impacts from an NCD death occur.


Effects of biliopanceratic diversion on type 2 diabetes in patients with BMI 25 to 35.
Biliopancreatic diversion (BPD) resolves type 2 diabetes in near totality of morbidly obeses [BMI (body mass index) ≥35 kg/m]. However, studies of BPD effect in BMI range 25.0 to 34.9 kg/m, including about 90% of diabetic patients, are lacking. If BPD effects are independent of weight changes, they should be maintained in patients who, being mildly obese or overweight, will lose little or no weight after operation. Thirty type 2 diabetic patients with BMI 25 to 34.9 were submitted to BPD and monitored 12 months. Thirty-eight diabetic patients selected from a large database, kept 1 year on medical therapy, served as controls. Nineteen male and 11 female. Mean age 56.4 ± 7.4 years, weight 84.8 ± 11.1 kg, BMI 30.6 ± 2.9 kg/m, waist circumference 104 ± 9.4 cm, diabetes duration 11.2 ± 6.9 years, HbA1c 9.3±1.5. Twelve patients on insulin. Fifteen (2 F) with BMI < 30 (mean: 28.1). No mortality or major adverse events occurred. BMI progressively decreased, stabilizing around 25 since the fourth month, without excessive weight loss. One year after BPD, mean HbA1c was 6.3%±0.8, with 25 patients (83%) controlled (HbA1c≤7%) on free diet, without antidiabetics, and the remaining improved. Acute insulin response to intravenous glucose had increased from 1.2 ± 2.9 to 4.2 ± 4.4 μIU/mL. Diabetes resolution correlated positively with BMI. HbA1c decreased at 1 year in the control group, along with an overall increased amount of antidiabetic therapy. BPD improves or resolves diabetes in BMI 25 to 35 without causing excessive weight loss, its action being on insulin sensitivity and beta-cell function. The strikingly different response between morbidly obese and low BMI patients might depend on different beta-cell defect. ClinicalTrials.gov Identifier: NCT00996294.


Polycythemia vera in young patients: a study on the long-term risk of thrombosis, myelofibrosis and leukemia.
Polycythemia vera (PV) is unusual in young patients, so that little information is available on long-term clinical evolution in this particular group. The aim of this study was to define the long-term risk of thrombosis, acute leukemia (AL) and myelofibrosis with myeloid metaplasia (MMM) in young PV patients. From 1975 to 2000, 70 PV patients aged less than 50 years were followed for a median time of 14 years (range 2-26). About three quarters were treated with pipobroman. The Kaplan-Meier method and Cox regression were used for survival analysis. The standardized mortality ratio (SMR) was calculated using Italian age/sex specific mortality rates. The risk of thrombosis increased during the observation period, reaching a plateau of 14% at 10 years and was markedly higher in individuals with a previous history of thrombosis (p=0.0023). No patient had progression into AL or MMM before the 9th year of follow-up. Subsequently, five patients (7%) developed AL and five (7%) MMM, with a 20-year cumulative risk of 15% and 10%, respectively. Overall survival at 20 years was 62%, with nine patients dying of progression into AL or MMM, four of vascular events, one of lung cancer, and four of non-PV-related causes. The SMR was 5.3, indicating a mortality significantly higher than that of the general population (p<0.000). This long-term retrospective cohort study shows that although the median survival of young patients with PV exceeds 23 years, their life expectancy is markedly lower than that of the general population because of disease evolution into AL or MMM.


Molecular epidemiology of H5N1 avian influenza.
The highly pathogenic Asian H5N1 influenza virus that was first detected in Guangdong in the People's Republic of China (China) in 1996 is unique in having spread to humans and other mammalian species. To date, this virus has not consistently transmitted between any mammalian species but the continued spread and evolution of these viruses in domestic poultry across Eurasia presents a continuing pandemic threat. These viruses have caused devastation in domestic poultry and have killed over 60% of infected humans. The H5N1 viruses are unique in having evolved into multiple clades and subclades by reassortment with other influenza viruses in the epicentre of southern China, and accumulation of point mutations has resulted in antigenic differences between the clades. Three waves of spread have occurred, wave one to East Asia and Southeast Asia, wave two through Qinghai Lake, China, to Europe, India and Africa, and wave three to Southeast Asia again. This paper deals with the molecular epidemiology of the evolution of the multiplicity of H5N1 clades. The continuing evolution of these H5N1 viruses and the possible establishment of secondary epicentres in Indonesia, Egypt and Nigeria present a continuing threat to poultry and people globally.


Consensus guidelines on the testing and clinical management issues associated with HLA and non-HLA antibodies in transplantation.
The introduction of solid-phase immunoassay (SPI) technology for the detection and characterization of human leukocyte antigen (HLA) antibodies in transplantation while providing greater sensitivity than was obtainable by complement-dependent lymphocytotoxicity (CDC) assays has resulted in a new paradigm with respect to the interpretation of donor-specific antibodies (DSA). Although the SPI assay performed on the Luminex instrument (hereafter referred to as the Luminex assay), in particular, has permitted the detection of antibodies not detectable by CDC, the clinical significance of these antibodies is incompletely understood. Nevertheless, the detection of these antibodies has led to changes in the clinical management of sensitized patients. In addition, SPI testing raises technical issues that require resolution and careful consideration when interpreting antibody results. With this background, The Transplantation Society convened a group of laboratory and clinical experts in the field of transplantation to prepare a consensus report and make recommendations on the use of this new technology based on both published evidence and expert opinion. Three working groups were formed to address (a) the technical issues with respect to the use of this technology, (b) the interpretation of pretransplantation antibody testing in the context of various clinical settings and organ transplant types (kidney, heart, lung, liver, pancreas, intestinal, and islet cells), and (c) the application of antibody testing in the posttransplantation setting. The three groups were established in November 2011 and convened for a "Consensus Conference on Antibodies in Transplantation" in Rome, Italy, in May 2012. The deliberations of the three groups meeting independently and then together are the bases for this report. A comprehensive list of recommendations was prepared by each group. A summary of the key recommendations follows. Technical Group: (a) SPI must be used for the detection of pretransplantation HLA antibodies in solid organ transplant recipients and, in particular, the use of the single-antigen bead assay to detect antibodies to HLA loci, such as Cw, DQA, DPA, and DPB, which are not readily detected by other methods. (b) The use of SPI for antibody detection should be supplemented with cell-based assays to examine the correlations between the two types of assays and to establish the likelihood of a positive crossmatch (XM). (c) There must be an awareness of the technical factors that can influence the results and their clinical interpretation when using the Luminex bead technology, such as variation in antigen density and the presence of denatured antigen on the beads. Pretransplantation Group: (a) Risk categories should be established based on the antibody and the XM results obtained. (b) DSA detected by CDC and a positive XM should be avoided due to their strong association with antibody-mediated rejection and graft loss. (c) A renal transplantation can be performed in the absence of a prospective XM if single-antigen bead screening for antibodies to all class I and II HLA loci is negative. This decision, however, needs to be taken in agreement with local clinical programs and the relevant regulatory bodies. (d) The presence of DSA HLA antibodies should be avoided in heart and lung transplantation and considered a risk factor for liver, intestinal, and islet cell transplantation. Posttransplantation Group: (a) High-risk patients (i.e., desensitized or DSA positive/XM negative) should be monitored by measurement of DSA and protocol biopsies in the first 3 months after transplantation. (b) Intermediate-risk patients (history of DSA but currently negative) should be monitored for DSA within the first month. If DSA is present, a biopsy should be performed. (c) Low-risk patients (nonsensitized first transplantation) should be screened for DSA at least once 3 to 12 months after transplantation. If DSA is detected, a biopsy should be performed. In all three categories, the recommendations for subsequent treatment are based on the biopsy results. A comprehensive list of recommendations is provided covering the technical and pretransplantation and posttransplantation monitoring of HLA antibodies in solid organ transplantation. The recommendations are intended to provide state-of-the-art guidance in the use and clinical application of recently developed methods for HLA antibody detection when used in conjunction with traditional methods.


Medicinal chemistry and pharmacology of cyclic ADP-ribose.
Cyclic ADP-ribose (cADPR) is a signaling molecule that has been shown to regulate calcium mobilization from intracellular stores in a wide variety of biological systems. Synthesis of structural analogs of cADPR has provided insights into structure-activity relationships as well as produced pharmacological research tools with useful properties such as, hydrolysis-resistance and cell permeability. The first generation of cADPR analogs was synthesized by a chemo-enzymatic approach that took advantage of the broad substrate specificity of Aplysia ADP-ribosyl cyclase. Analogs synthesized by this approach provided useful structure-activity information, including the importance of the 8-position of the adenine in determining agonistic or antagonistic activity and of the 3'-hydroxyl group of the southern ribose for activity. Hydrolysis resistant analogs were generated by replacing the southern ribose with a carbocyclic structure or by replacing the adenine ring with 7-deaza- or 3-deaza-adenine. Approaches to synthesize cADPR analogs by total chemical approaches have been recently reported. These approaches allow the synthesis of analogs with stable linkages between N1 of adenine and the northern ribose (or surrogate) that are not possible with the enzymatic strategy. This review will focus on the synthesis and properties of analogs that have been shown to have utility in dissecting the role of cADPR in calcium signaling.


The intergenerational transfer of mother-daughter risk for gender-based abuse.
In this 10-year longitudinal study 150 mother-daughter pairs were recruited to participate in a study examining gender-based abuse across three generations. Forms of gender-based abuse included: child sexual abuse, witnessing intimate partner violence against their mothers, and intimate partner violence or dating violence in adolescence or adulthood. Daughters were interviewed when they were on average 9, 14, and 16 years old. Regression analyses revealed that if the grandmother (G1) was abused by her husband, her daughter (G2) was more likely to be sexually molested in childhood and was also more likely to be in an abusive relationship as an adult. If the mother (G2) was sexually abused as a child her daughter (G3) was at increased risk for child sexual abuse. In turn, child sexual abuse for the daughters related to their reports of dating violence in adolescence. Daughters (G3) who were sexually abused expressed more anxiety about romantic relationships, reflecting early attachment conflicts. Both child sexual abuse and anxious romantic attachment style independently predicted adolescent sexual risk-taking as in having multiple sexual partners or dating older men. These findings demonstrate how informative it is to include multiple forms of gender-based abuse in research and practice to better illuminate complex family dynamics. In addition, the findings support previous empirical work showing the importance of attachment behavior in women who are in abusive relationships, which has unique clinical implications.


How Close is too Close? The Effect of a Non-Lethal Electric Shark Deterrent on White Shark Behaviour.
Sharks play a vital role in the health of marine ecosystems, but the potential threat that sharks pose to humans is a reminder of our vulnerability when entering the ocean. Personal shark deterrents are being marketed as the solution to mitigate the threat that sharks pose. However, the effectiveness claims of many personal deterrents are based on our knowledge of shark sensory biology rather than robust testing of the devices themselves, as most have not been subjected to independent scientific studies. Therefore, there is a clear need for thorough testing of commercially available shark deterrents to provide the public with recommendations of their effectiveness. Using a modified stereo-camera system, we quantified behavioural interactions between white sharks (Carcharodon carcharias) and a baited target in the presence of a commercially available, personal electric shark deterrent (Shark Shield Freedom7™). The stereo-camera system enabled an accurate assessment of the behavioural responses of C. carcharias when encountering a non-lethal electric field many times stronger than what they would naturally experience. Upon their first observed encounter, all C. carcharias were repelled at a mean (± std. error) proximity of 131 (± 10.3) cm, which corresponded to a mean voltage gradient of 9.7 (± 0.9) V/m. With each subsequent encounter, their proximity decreased by an average of 11.6 cm, which corresponded to an increase in tolerance to the electric field by an average of 2.6 (± 0.5) V/m per encounter. Despite the increase in tolerance, sharks continued to be deterred from interacting for the duration of each trial when in the presence of an active Shark Shield™. Furthermore, the findings provide no support to the theory that electric deterrents attract sharks. The results of this study provide quantitative evidence of the effectiveness of a non-lethal electric shark deterrent, its influence on the behaviour of C. carcharias, and an accurate method for testing other shark deterrent technologies.


The cytochrome P-450 monooxygenase system of rabbit lung enzyme components, activities, and induction in the nonciliated bronchiolar epithelial (Clara) cell, alveolar type II cell, and alveolar macrophage.
Enzyme components and activities of the cytochrome P-450 monooxygenase system in microsomal preparations from the Clara cell, alveolar type II cell, and alveolar macrophage fractions isolated from lungs of untreated rabbits and rabbits treated with 2,3,7,8-tetrachlorodibenzo-p-dioxin were examined. Results are compared to those obtained with microsomal preparations from whole lung. Concentrations of cytochrome P-450 isozymes 2 and 5 and NADPH-cytochrome P-450 reductase activities were higher in preparations from Clara cell fractions than in preparations from type II cell fractions or whole lung. For the most part, however, differences among these preparations were 2-fold or less. Microsomal preparations from the macrophage fraction contained low or undetectable levels of cytochrome P-450 isozymes but relatively high levels of cytochrome P-450 reductase activity. The concentration of cytochrome P-450 isozyme 6, in contrast to those of isozymes 2 and 5, was found to be highest in microsomal preparations from whole lung. Treatment of rabbits with 2,3,7,8-tetrachlorodibenzo-p-dioxin increased the concentrations of isozyme 6 in preparations from the Clara and type II cell fractions and from whole lung about 20-fold. In contrast, the content of isozyme 6 in preparations from the macrophage fraction increased greater than 90-fold. In all cases, induction of isozyme 6 resulted in substantial increases in the O-deethylation of 7-ethoxyresorufin and only minor increases in the hydroxylation of benzo(a)pyrene. Activities per unit of isozyme 6, following induction, were similar in all preparations, and we estimate that less than 20% of the potential activity of isozyme 6 is expressed with benzo(a)pyrene and greater than 40% with 7-ethoxyresorufin. These similarities exist in spite of significant differences among the preparations from different fractions in the ratios of isozyme 6 to NADPH-cytochrome P-450 reductase.


Boundary element method for surface nonlinear optics of nanoparticles.
We present the frequency-domain boundary element formulation for solving surface second-harmonic generation from nanoparticles of virtually arbitrary shape and material. We use the Rao-Wilton-Glisson basis functions and Galerkin's testing, which leads to very accurate solutions for both near and far fields. This is verified by a comparison to a solution obtained via multipole expansion for the case of a spherical particle. The frequency-domain formulation allows the use of experimentally measured linear and nonlinear material parameters or the use of parameters obtained using ab-initio principles. As an example, the method is applied to a non-centrosymmetric L-shaped gold nanoparticle to illustrate the formation of surface nonlinear polarization and the second-harmonic radiation properties of the particle. This method provides a theoretically well-founded approach for modelling nonlinear optical phenomena in nanoparticles.


Optical forces on metallic nanoparticles induced by a photonic nanojet.
We investigate the optical forces acting on a metallic nanoparticle when the nanoparticle is introduced within a photonic nanojet (PNJ). Optical forces at resonance and off-resonance conditions of the microcylinder or nanoparticle are investigated. Under proper polarization conditions, the whispering gallery mode can be excited in the microcylinder, even at off resonance provided that scattering from the nanoparticle is strong enough. The optical forces are enhanced at resonance either of the single microcylinder or of the nanoparticle with respect to the forces under off-resonant illuminations. We found that the optical forces acting on the nanoparticle depend strongly on the dielectric permittivity of the nanoparticle, as well as on the intensity and the beam width of the PNJ. Hence, metallic sub-wavelength nanoparticle can be efficiently trapped by PNJs. Furthermore, the PNJ's attractive force can be simply changed to a repulsive force by varying the polarization of the incident beam. The changed sign of the force is related to the particle's polarizability and the excitation of localized surface plasmons in the nanoparticle.


Mdm2 binds to Nbs1 at sites of DNA damage and regulates double strand break repair.
Mdm2 directly regulates the p53 tumor suppressor. However, Mdm2 also has p53-independent activities, and the pathways that mediate these functions are unresolved. Here we report the identification of a specific association of Mdm2 with Mre11, Nbs1, and Rad50, a DNA double strand break repair complex. Mdm2 bound to the Mre11-Nbs1-Rad50 complex in primary cells and in cells containing inactivated p53 or p14/p19ARF, a regulator of Mdm2. Further analysis revealed that Mdm2 directly bound to Nbs1 but not to Mre11 or Rad50. Amino acids 198-314 of Mdm2 were required for Mdm2/Nbs1 association, and neither the N terminus forkhead-associated and breast cancer C-terminal domains nor the C terminus Mre11 binding domain of Nbs1 mediated the interaction of Nbs1 with Mdm2. Mdm2 co-localized with Nbs1 to sites of DNA damage following gamma-irradiation. Notably, Mdm2 overexpression inhibited DNA double strand break repair, and this was independent of p53 and ARF, the alternative reading frame of the Ink4alocus. The delay in DNA repair imposed by Mdm2 required the Nbs1 binding domain of Mdm2, but the ubiquitin ligase domain in Mdm2 was dispensable. Therefore, Nbs1 is a novel p53-independent Mdm2 binding protein and links Mdm2 to the Mre11-Nbs1-Rad50-regulated DNA repair response.


Conditional root expansion mutants of Arabidopsis.
Regulation of cell expansion is essential to the formation of plant organs. We have characterized 21 mutations, representing six loci, that cause abnormal cell expansion in the root of Arabidopsis thaliana. The phenotype of these mutants is conditional upon the rate of root growth. Calculation of cell volumes indicated that the mutations resulted in defects in either the orientation or the extent of expansion or in both. Analysis of cortical microtubules in the mutants suggested that a shift in the orientation of cell expansion may not be dependent on a change in the orientation of the microtubules. Double mutant combinations resulted in loss of the conditional phenotype suggesting that the genes may act in a similar pathway or encode partially redundant functions.


Development of a three-scale MMPI: the MMPI-TRI.
A 60-item short form of the MMPI with very high content validity and items that appear on both the MMPI and MMPI-2 was developed and named the MMPI-TRI. It contains three 20-item scales--the Subjective Distress, Acting-Out, and Psychosis scales. These three scales have excellent internal consistency and sufficient independence from each other. An anxiety and depression group of patients, prison inmate group, and a schizophrenic and other psychotic group had the highest mean scores on Subjective Distress, Acting-Out, and Psychosis, respectively. Correlations with the 13 regular scales of the MMPI and MMPI-2, their content and supplementary scales, and four other psychometric instruments provided very strong evidence for validity. Norms are provided.


Whortleberry protects kidney against the cisplatin-induced nephrotoxicity: an experimental study.
This study investigated the antioxidant effects of whortleberry against cisplatin-induced nephrotoxicity in rats. This study included 48 female Sprague-Dawley rats weighing 263.68 ± 8.29 g. The rats were divided into the following six groups, with eight rats in each group: control, ethanol control, whortleberry control, cisplatin control, 16 mg/kg cisplatin +100 mg/kg whortleberry, and 16 mg/kg cisplatin +200 mg/kg whortleberry groups. Biochemical analysis was performed by measuring total oxidant status and total antioxidant status, histopathological analysis was performed by calculating proximal and distal tubule areas (μm2), and immunohistochemical analysis was performed by determining anti-Caspase-3 immunostaining. Differences among the groups were examined using one-way analysis of variance, and p < .05 was considered statistically significant. Cisplatin treatment decreased the total antioxidant status and increased the total oxidant status and Caspase-3 level. Moreover, it resulted in the dilatation, vacuolization and loss of tubular epithelial cells; and glomerular degeneration and edema in the kidney tissues (p < .05). Treatment with 100 and 200 mg whortleberries increased the total antioxidant status; decreased the total oxidant status and Caspase-3 level and ameliorated distal and proximal tubule degeneration, glomerular degeneration and edema in the kidney tissues (p < .05). Our results indicate that the antioxidant effects of the whortleberry decrease cisplatin-associated nephrotoxicity.


Tic32, an essential component in chloroplast biogenesis.
Chloroplast protein import across the inner envelope is facilitated by the translocon of the inner envelope of chloroplasts (Tic). Here we have identified Tic32 as a novel subunit of the Tic complex. Tic32 can be purified from solubilized inner envelope membranes by chromatography on Tic110 containing affinity matrix. Co-immunoprecipitation experiments using either Tic32 or Tic110 antisera indicated a tight association between these polypeptides as well as with other Tic subunits, e.g. Tic40, Tic22, or Tic62, whereas the outer envelope protein Toc75 was not found in this complex. Chemical cross-linking suggests that Tic32 is involved late in the overall translocation process, because both the precursor form as well as the mature form of Rubisco small subunit can be detected. We were unable to isolate Arabidopsis null mutants of the attic32 gene, indicating that Tic32 is essential for viability. Deletion of the attic32 gene resulted in early seed abortion because the embryo was unable to differentiate from the heart stage to the torpedo stage. The homology of Tic32 to short-chain dehydrogenases suggests a dual role of Tic32 in import, one as a regulatory component and one as an important subunit in the assembly of the entire complex.


Barcoded Consortium Infections Resolve Cell Type-Dependent Salmonella enterica Serovar Typhimurium Entry Mechanisms.
Bacterial host cell invasion mechanisms depend on the bacterium's virulence factors and the properties of the target cell. The enteropathogen Salmonella enterica serovar Typhimurium (STm) invades epithelial cell types in the gut mucosa and a variety of immune cell types at later infection stages. The molecular mechanism(s) of host cell entry has, however, been studied predominantly in epithelial cell lines. STm uses a type three secretion system (TTSS-1) to translocate effectors into the host cell cytosol, thereby sparking actin ruffle-dependent entry. The ruffles also fuel cooperative invasion by bystander bacteria. In addition, several TTSS-1-independent entry mechanisms exist, involving alternative STm virulence factors, or the passive uptake of bacteria by phagocytosis. However, it remains ill-defined how STm invasion mechanisms vary between host cells. Here, we developed an internally controlled and scalable method to map STm invasion mechanisms across host cell types and conditions. The method relies on host cell infections with consortia of chromosomally tagged wild-type and mutant STm strains, where the abundance of each strain can be quantified by qPCR or amplicon sequencing. Using this methodology, we quantified cooccurring TTSS-1-dependent, cooperative, and TTSS-1-independent invasion events in epithelial, monocyte, and macrophage cells. We found STm invasion of epithelial cells and monocytes to proceed by a similar MOI-dependent mix of TTSS-1-dependent and cooperative mechanisms. TTSS-1-independent entry was more frequent in macrophages. Still, TTSS-1-dependent invasion dominated during the first minutes of interaction also with this cell type. Finally, the combined action of the SopB/SopE/SopE2 effectors was sufficient to explain TTSS-1-dependent invasion across both epithelial and phagocytic cells.IMPORTANCESalmonella enterica serovar Typhimurium (STm) is a widespread and broad-host-spectrum enteropathogen with the capacity to invade diverse cell types. Still, the molecular basis for the host cell invasion process has largely been inferred from studies of a few selected cell lines. Our work resolves the mechanisms that Salmonellae employ to invade prototypical host cell types, i.e., human epithelial, monocyte, and macrophage cells, at a previously unattainable level of temporal and quantitative precision. This highlights efficient bacterium-driven entry into innate immune cells and uncovers a type III secretion system effector module that dominates active bacterial invasion of not only epithelial cells but also monocytes and macrophages. The results are derived from a generalizable method, where we combine barcoding of the bacterial chromosome with mixed consortium infections of cultured host cells. The application of this methodology across bacterial species and infection models will provide a scalable means to address host-pathogen interactions in diverse contexts.


Taste transductions in taste receptor cells: basic tastes and moreover.
In the oral cavity, taste receptor cells dedicate to detecting chemical compounds in foodstuffs and transmitting their signals to gustatory nerve fibers. Heretofore, five taste qualities (sweet, umami, bitter, salty and sour) are generally accepted as basic tastes. Each of these may have a specific role in the detection of nutritious and poisonous substances; sweet for carbohydrate sources of calories, umami for protein and amino acid contents, bitter for harmful compounds, salty for minerals and sour for ripeness of fruits and spoiled foods. Recent studies have revealed molecular mechanisms for reception and transduction of these five basic tastes. Sweet, umami and bitter tastes are mediated by G-protein coupled receptors (GPCRs) and second-messenger signaling cascades. Salty and sour tastes are mediated by channel-type receptors. In addition to five basic tastes, taste receptor cells may have the ability to detect fat taste, which is elicited by fatty acids, and calcium taste, which is elicited by calcium. Taste compounds eliciting either fat taste or calcium taste may be detected by specific GPCRs expressed in taste receptor cells. This review will focus on transduction mechanisms and cellular characteristics responsible for each of basic tastes, fat taste and calcium taste.


Biodistribution of Liposome-Encapsulated Bacteriophages and Their Transcytosis During Oral Phage Therapy.
This study sheds light on the biodistribution of orally administered, liposome-encapsulated bacteriophages, and their transcytosis through intestinal cell layers. Fluorochrome-labeled bacteriophages were used together with a non-invasive imaging methodology in the in vivo visualization of bacteriophages in the stomach and intestinal tract of mice. In those studies, phage encapsulation resulted in a significant increase of the labeled phages in the mouse stomach, even 6 h after their oral administration, and without a decrease in their concentration. By contrast, the visualization of encapsulated and non-encapsulated phages in the intestine were similar. Our in vivo observations were corroborated by culture methods and ex vivo experiments, which also showed that the percentage of encapsulated phages in the stomach remained constant (50%) compared to the amount of initially administered product. However, the use of conventional microbiological methods, which employ bile salts to break down liposomes, prevented the detection of encapsulated phages in the intestine. The ex vivo data showed a higher concentration of non-encapsulated than encapsulated phages in liver, kidney, and even muscle up to 6 h post-administration. Encapsulated bacteriophages were able to reach the liver, spleen, and muscle, with values of 38% ± 6.3%, 68% ± 8.6%, and 47% ± 7.4%, respectively, which persisted over the course of the experiment. Confocal laser scanning microscopy of an in vitro co-culture of human Caco-2/HT29/Raji-B cells revealed that Vybrant-Dil-stained liposomes containing labeled bacteriophages were preferably embedded in cell membranes. No transcytosis of encapsulated phages was detected in this in vitro model, whereas SYBR-gold-labeled non-encapsulated bacteriophages were able to cross the membrane. Our work demonstrates the prolonged persistence of liposome-encapsulated phages in the stomach and their adherence to the intestinal membrane. These observations could explain the greater long-term efficacy of phage therapy using liposome-encapsulated phages.


Serum Albumin Domain Structures in Human Blood Serum by Mass Spectrometry and Computational Biology.
Chemical cross-linking combined with mass spectrometry has proven useful for studying protein-protein interactions and protein structure, however the low density of cross-link data has so far precluded its use in determining structures de novo. Cross-linking density has been typically limited by the chemical selectivity of the standard cross-linking reagents that are commonly used for protein cross-linking. We have implemented the use of a heterobifunctional cross-linking reagent, sulfosuccinimidyl 4,4'-azipentanoate (sulfo-SDA), combining a traditional sulfo-N-hydroxysuccinimide (sulfo-NHS) ester and a UV photoactivatable diazirine group. This diazirine yields a highly reactive and promiscuous carbene species, the net result being a greatly increased number of cross-links compared with homobifunctional, NHS-based cross-linkers. We present a novel methodology that combines the use of this high density photo-cross-linking data with conformational space search to investigate the structure of human serum albumin domains, from purified samples, and in its native environment, human blood serum. Our approach is able to determine human serum albumin domain structures with good accuracy: root-mean-square deviation to crystal structure are 2.8/5.6/2.9 Å (purified samples) and 4.5/5.9/4.8Å (serum samples) for domains A/B/C for the first selected structure; 2.5/4.9/2.9 Å (purified samples) and 3.5/5.2/3.8 Å (serum samples) for the best out of top five selected structures. Our proof-of-concept study on human serum albumin demonstrates initial potential of our approach for determining the structures of more proteins in the complex biological contexts in which they function and which they may require for correct folding. Data are available via ProteomeXchange with identifier PXD001692.


Quantitative comparison of analysis methods for spectroscopic optical coherence tomography.
Spectroscopic optical coherence tomography (sOCT) enables the mapping of chromophore concentrations and image contrast enhancement in tissue. Acquisition of depth resolved spectra by sOCT requires analysis methods with optimal spectral/spatial resolution and spectral recovery. In this article, we quantitatively compare the available methods, i.e. the short time Fourier transform (STFT), wavelet transforms, the Wigner-Ville distribution and the dual window method through simulations in tissue-like media. We conclude that all methods suffer from the trade-off in spectral/spatial resolution, and that the STFT is the optimal method for the specific application of the localized quantification of hemoglobin concentration and oxygen saturation.


CT with the hepatocyte-specific contrast medium FP 736-04 in an experimental model of liver steatosis.
To investigate whether liver uptake of the iodinated hepatocyte-specific lipid emulsion FP 736-04 is altered by fatty infiltration of the liver. Fatty infiltration of the liver was induced in female Sprague-Dawley rats by an intraperitoneal injection of L-ethionine preceded by 15 h of food withdrawal. CT of the liver was performed before and 24 h after the administration of L-ethionine and, in addition at the latter time point, after an i.v. injection of 1.0 ml/kg b.w. of FP 736-04. A control group was subjected to the same CT examination protocol. Intraperitoneal administration of L-ethionine caused liver steatosis, as established by liver triglyceride analysis, leading to a significant decrease in the liver attenuation, from 69.2 +/- 2.4 to 27.8 +/- 12.0 HU. The uptake of FP 736-04 by the fatty liver was significantly reduced, yielding a maximum enhancement of 22.3 +/- 7.9 HU as compared to a value of 32.3 +/- 5.0 HU in the control group. Enhancement by FP 736-04 was reduced in the steatotic liver compared with the normal liver. It remains to be established whether this degree of enhancement is sufficient for reliable lesion detection.


Continuous replication of Friend virus complex (spleen focus-forming virus-lymphatic leukemia-inducing virus) in mouse embryo fibroblasts. Retention of leukemogenicity and loss of immunosuppressive properties.
Exposure of NIH Swiss mouse embryo fibroblasts (MEF) to infectious Friend virus (FV) complex [containing defective spleen focus-forming virus (SFFV) and endogenous NB-tropic leukemia-inducing helper virus (LLV-F)] resulted in the productive infection of these cells by both SFFV and LLV-F. Stocks of SFFV derived after extensive growth in this Swiss MEF cell culture system are fully leukemogenic in adult mice for the induction of erythroleukemia and spleen foci. In addition, in vitro-derived LLV-F, when isolated free of SFFV, is fully leukemogenic for the induction of lymphatic leukemia when inoculated into susceptible newborn BALB/c mice. The host range of in vitro-derived FV complex (i.e., FV-TC) for focus formation in vivo is NB-tropic. Unlike in vivo-derived FV complex, FV-TC does not suppress the responsiveness of murine thymocytes to concanavalin A (Con A) in vitro. Rather, FV-TC acts as a mitogen to nonspecifically stimulate the proliferation of BALB/c thymocytes. The mitogenicity of in vitro-derived FV complex is directly associated with the presence of type-C virus particles, is a heat-labile and UV-sensitive property of the virus, and may be primarily due to LLV since equivalent amounts of LLV with or without SFFV present are equally mitogenic. One in vivo passage of FV-TC resulted in the total loss of this mitogenic property with the reappearance of full immunosuppressive properties. This result demonstrates a clear association between in vivo growth of FV and its ability to suppress mouse thymocytes, and suggests that FV complex (SFFV-LLV) is not inherently immunosuppressive for these cells. While the mechanism of this interconversion between immunostimulatory and fully suppressive virus is unknown, both virus markers appear to be dependent upon the presence of infectious FV.


Synergistic Communication between CD4+ T Cells and Monocytes Impacts the Cytokine Environment.
Physiological cytokine environments arise from factors produced by diverse cell types in coordinated concert. Understanding the contributions of each cell type in the context of cell-cell communication is important for effectively designing disease modifying interventions. Here, we present multi-plexed measurement of 48 cytokines from a coculture system of primary human CD4+ T cells and monocytes across a spectrum of stimuli and for a range of relative T cell/monocyte compositions, coupled with corresponding measurements from PBMCs and plasma from the same donors. Computational analysis of the resulting data-sets elucidated communication-independent and communication-dependent contributions, including both positive and negative synergies. We find that cytokines in cell supernatants were uncorrelated to those found in plasma. Additionally, as an example of positive synergy, production levels of CXCR3 cytokines IP-10 and MIG, depend non-linearly on both IFNγ and TNFα levels in cross-talk between T cells and monocytes. Overall, this work demonstrates that communication between cell types can significantly impact the consequent cytokine environment, emphasizing the value of mixed cell population studies.