A semi-automated method for unbiased alveolar morphometry: Validation in a bronchopulmonary dysplasia model.
Reproducible and unbiased methods to quantify alveolar structure are important for research on many lung diseases. However, manually estimating alveolar structure through stereology is time consuming and inter-observer variability is high. The objective of this work was to develop and validate a fast, reproducible and accurate (semi-)automatic alternative. A FIJI-macro was designed that automatically segments lung images to binary masks, and counts the number of test points falling on tissue and the number of intersections of the air-tissue interface with a set of test lines. Manual selection remains necessary for the recognition of non-parenchymal tissue and alveolar exudates. Volume density of alveolar septa ([Formula: see text]) and mean linear intercept of the airspaces (Lm) as measured by the macro were compared to theoretical values for 11 artificial test images and to manually counted values for 17 lungs slides using linear regression and Bland-Altman plots. Inter-observer agreement between 3 observers, measuring 8 lungs both manually and automatically, was assessed using intraclass correlation coefficients (ICC). [Formula: see text] and Lm measured by the macro closely approached theoretical values for artificial test images (R2 of 0.9750 and 0.9573 and bias of 0.34% and 8.7%). The macro data in lungs were slightly higher for [Formula: see text] and slightly lower for Lm in comparison to manually counted values (R2 of 0.8262 and 0.8288 and bias of -6.0% and 12.1%). Visually, semi-automatic segmentation was accurate. Most importantly, manually counted [Formula: see text] and Lm had only moderate to good inter-observer agreement (ICC 0.859 and 0.643), but agreements were excellent for semi-automatically counted values (ICC 0.956 and 0.900). This semi-automatic method provides accurate and highly reproducible alveolar morphometry results. Future efforts should focus on refining methods for automatic detection of non-parenchymal tissue or exudates, and for assessment of lung structure on 3D reconstructions of lungs scanned with microCT.


Osteomyelitis and infective endocarditis.
Osteomyelitis is thought to occur as a complication of infectious endocarditis in as many as 6% of cases of endocarditis. We describe this association in three patients. Osteomyelitis may be difficult to diagnose in patients with endocarditis because symptoms such as fever, bone pain and stiffness are common to both illnesses, therefore physicians need to have a high index of suspicion to avoid missing this important complication. We recommend that patients with endocarditis and persistent or localized musculoskeletal symptoms should be investigated to exclude osteomyelitis. Plain radiographs can be normal in 50% of cases of osteomyelitis in the early stages or show only minor abnormalities, but bone scans are highly sensitive. We suggest that a bone scan is performed if radiography is unhelpful, since a diagnosis of osteomyelitis can effectively be excluded if the bone scan is normal. We advocate close follow-up of these patients with prolonged antibiotic treatment consisting of at least 6 weeks of intravenous therapy, and 3 months or longer of oral therapy.


Ultrastructural study of the granule cell domain of the cochlear nucleus in rats: mossy fiber endings and their targets.
The principal projection neurons of the cochlear nucleus receive the bulk of their input from the auditory nerve. These projection neurons reside in the core of the nucleus and are surrounded by an external shell, which is called the granule cell domain. Interneurons of the cochlear granule cell domain are the target for nonprimary auditory inputs, including projections from the superior olivary complex, inferior colliculus, and auditory cortex. The granule cell domain also receives projections from the cuneate and trigeminal nuclei, which are first-order nuclei of the somatosensory system. The cellular targets of the nonprimary projections are mostly unknown due to a lack of information regarding postsynaptic profiles in the granule cell areas. In the present paper, we examined the synaptic relationships between a heterogeneous class of large synaptic terminals called mossy fibers and their targets within subdivisions of the granule cell domain known as the lamina and superficial layer. By using light and electron microscopic methods in these subdivisions, we provide evidence for three different neuron classes that receive input from the mossy fibers: granule cells, unipolar brush cells, and a previously undescribed class called chestnut cells. The distinct synaptic relations between mossy fibers and members of each neuron class further imply fundamentally separate roles for processing acoustic signals.


Mutations in CBL occur frequently in juvenile myelomonocytic leukemia.
Juvenile myelomonocytic leukemia is an aggressive myeloproliferative disorder characterized by malignant transformation in the hematopoietic stem cell compartment with proliferation of differentiated progeny. Seventy-five percent of patients harbor mutations in the NF1, NRAS, KRAS, or PTPN11 genes, which encode components of Ras signaling networks. Using single nucleotide polymorphism arrays, we identified a region of 11q isodisomy that contains the CBL gene in several JMML samples, and subsequently identified CBL mutations in 27 of 159 JMML samples. Thirteen of these mutations alter codon Y371. In this report, we also demonstrate that CBL and RAS/PTPN11 mutations were mutually exclusive in these patients. Moreover, the exclusivity of CBL mutations with respect to other Ras pathway-associated mutations indicates that CBL may have a role in deregulating this key pathway in JMML.


Utility of routine immunofluorescence staining for C4d in cardiac transplant recipients.
Immunofluorescence staining of endomyocardial biopsy (EMB) specimens to detect the complement fragment C4d is used to diagnose antibody-mediated rejection. However, data are limited regarding the utility of routine staining for C4d in clinical care. This study retrospectively reviewed the clinical course of adult cardiac transplant recipients who underwent > or = 2 EMBs with immunofluorescence C4d staining at the University of Texas Southwestern Medical Center since September 2006. C4d staining was performed by the immunohistochemistry laboratory and interpreted by the members of the surgical pathology department, in conjunction with interpretation of the routine hematoxylin and eosin staining. Donor-specific antibodies (DSA) were routinely assessed at the time of clinical rejection. Of 67 patients, specimens were positive for C4d (C4d+) in 14 and negative for C4d (C4d-) in 53. The frequency of acute cellular rejection (ACR) in these 2 groups was 57% (8 of 14, designated C4d+/ACR+) vs 11% (6 of 53, designated C4d-/ACR+; p < 0.001). Significantly more patients with a positive C4d specimen had a positive retrospective donor specific crossmatch, presence of DSA after transplantation, and depressed graft function (p < 0.01 for each). Positive C4d immunofluorescence staining on EMB specimens was associated with ACR, reduced allograft function, a positive retrospective crossmatch, and the presence of DSA after transplantation. The latter 2 observations support the contention that C4d deposition is a marker of antibody-mediated rejection. Routine evaluation of C4d staining is feasible in the clinical setting and may identify variable patterns of rejection.


Clinical and economic outcomes of Staphylococcus aureus septicemia in ESRD patients receiving hemodialysis.
Serious infections are a common problem in patients with end-stage renal disease (ESRD). The purpose of this study is to identify clinical and economic consequences of hospitalizations for septicemia caused by Staphylococcus aureus in hemodialysis patients with ESRD. We conducted a retrospective analysis of data obtained from the US Renal Data System to determine lengths of stay and Medicare paid costs for index hospitalizations and episodes of care for patients with ESRD hospitalized with septicemia caused by S aureus. Factors associated with hospital length of stay and Medicare paid costs were examined in multivariate analysis. A total of 11,572 patient admissions with septicemia caused by S aureus were included; 20.7% of patients developed 1 or more complications. Average length of stay for the index admission was 13.0 days, and 11.8% of patients were readmitted within 12 weeks for care related to S aureus. Average Medicare cost for the index admission was 17,307 dollars. Average episodic cost of care, including the index hospitalization, outpatient visits, and readmissions related to S aureus during the subsequent 12 weeks, was 20,067 dollars. S aureus--related complications were associated with greater episodic costs of care: no complications, 18,476 dollars; one complication, 25,804 dollars (P < 0.05 versus no complications); and 2 or more complications, 32,102 dollars (P < 0.05 versus no complications). In multivariate analysis, complications resulted in increased mean lengths of stay of 4 to 7 days, and complications were among the strongest predictors of total episodic costs. Patients with septicemia caused by S aureus had costly and lengthy hospitalizations, which frequently were associated with clinically and economically important complications, including hospital readmissions.


A hospitalization from hell: a patient's perspective on quality.
Patients usually cannot assess the technical quality of their care; however, examining a hospitalization through the patients' eyes can reveal important information about the quality of care. Patients are the best source of information about a hospital system's communication, education, and pain-management processes, and they are the only source of information about whether they were treated with dignity and respect. Their experiences often reveal how well a hospital system is operating and can stimulate important insights into the kinds of changes that are needed to close the chasm between the care provided and the care that should be provided. This article examines the case of a patient admitted for ankle arthrodesis due to severe hemophilia-related arthritis. The surgery was successful, but the hospital stay was marked by inefficiency and inconveniences, as well as events that reveal fundamental problems with the hospital's organization and teamwork. These problems could seriously compromise the quality of clinical care. Unfortunately, most of these events occur regularly in U.S. hospitals. Relatively easy and inexpensive ways to avoid many of these problems are discussed, such as reducing variability in non-urgent procedures and routinely asking patients about their experiences and suggestions for improvement.


Regulation of the innate immune response by fibronectin: synergism between the III-1 and EDA domains.
Fibronectin is a critical component of the extracellular matrix and alterations to its structure will influence cellular behavior. Matrix fibronectin is subjected to both mechanical and biochemical regulation. The Type III domains of fibronectin can be unfolded in response to increased cellular contractility, included or excluded from the molecule by alternative splicing mechanisms, or released from the matrix by proteolysis. Using Inflammatory Cytokine microarrays we found that the alternatively spliced fibronectin Type III domain, FnEDA, and the partially unfolded III-1 domain, FnIII-1c, induced the expression of a multitude of pro-inflammatory cytokines in human dermal fibroblasts, most notably CXCL1-3, IL-8 and TNF-α. FnIII-1c, a peptide representing an unfolded intermediate structure of the first Type III domain has been shown to initiate the toll-like receptor-4 (TLR4)-NFκB-dependent release of cytokines from human dermal fibroblasts (You, et al., J. Biol. Chem., 2010). Here we demonstrate that FnIII-1c and the alternatively spliced FnEDA domain induce a TLR4 dependent activation of p38 MAP kinase and its downstream effector, MAPKAP Kinase-2 (MK-2), to regulate cytokine expression in fibroblasts. RT-qPCR analysis indicated that the p38-MK-2 pathway regulates IL-8 mRNA stability. Interestingly, addition of FnIII-1c and FnEDA synergistically enhanced TLR4-dependent IL-8 release. These data indicate that Fn contains two Type III domains which can activate TLR signaling to induce an inflammatory response in fibroblasts. Furthermore, our data identifies the NF-κB and p38/MK2 signaling pathways as transducers of signals initiated in response to structural changes in fibronectin.


Cancer of the respiratory tract in nickel sinter plant workers: effect of removal from sinter plant exposure.
The risk of death due to cancer of the nose or lung was studied in nickel sinter plant workers during the period after they left the sintering operations. It was found that the excess risk of death from both diseases continued for many years after leaving the sinter plant. No effect of age at first exposure could be found. Possible explanations for the continued risk include an irreversible cellular change or the persistence of carcinogenic nickel compounds in the mucosa.


Endothelial Cell Proteomic Response to Rickettsia conorii Infection Reveals Activation of the Janus Kinase (JAK)-Signal Transducer and Activator of Transcription (STAT)-Inferferon Stimulated Gene (ISG)15 Pathway and Reprogramming Plasma Membrane Integrin/Cadherin Signaling.
Rickettsia conorii is the etiologic agent of Mediterranean spotted fever, a re-emerging infectious disease with significant mortality. This Gram-negative, obligately intracellular pathogen is transmitted via tick bites, resulting in disseminated vascular endothelial cell infection with vascular leakage. In the infected human, Rickettsia conorii infects endothelial cells, stimulating expression of cytokines and pro-coagulant factors. However, the integrated proteomic response of human endothelial cells to R. conorii infection is not known. In this study, we performed quantitative proteomic profiling of primary human umbilical vein endothelial cells (HUVECs) with established R conorii infection versus those stimulated with endotoxin (LPS) alone. We observed differential expression of 55 proteins in HUVEC whole cell lysates. Of these, we observed induction of signal transducer and activator of transcription (STAT)1, MX dynamin-like GTPase (MX1), and ISG15 ubiquitin-like modifier, indicating activation of the JAK-STAT signaling pathway occurs in R. conorii-infected HUVECs. The down-regulated proteins included those involved in the pyrimidine and arginine biosynthetic pathways. A highly specific biotinylated cross-linking enrichment protocol was performed to identify dysregulation of 11 integral plasma membrane proteins that included up-regulated expression of a sodium/potassium transporter and down-regulation of α-actin 1. Analysis of Golgi and soluble Golgi fractions identified up-regulated proteins involved in platelet-endothelial adhesion, phospholipase activity, and IFN activity. Thirty four rickettsial proteins were identified with high confidence in the Golgi, plasma membrane, or secreted protein fractions. The host proteins associated with rickettsial infections indicate activation of interferon-STAT signaling pathways; the disruption of cellular adhesion and alteration of antigen presentation pathways in response to rickettsial infections are distinct from those produced by nonspecific LPS stimulation. These patterns of differentially expressed proteins suggest mechanisms of pathogenesis as well as methods for diagnosis and monitoring Rickettsia infections.


Malaria epidemics and interventions, Kenya, Burundi, southern Sudan, and Ethiopia, 1999-2004.
Quantitative data on the onset and evolution of malaria epidemics are scarce. We review case studies from recent African Plasmodium falciparum epidemics (Kisii and Gucha Districts, Kenya, 1999; Kayanza Province, Burundi, 2000-2001; Aweil East, southern Sudan, 2003; Gutten and Damot Gale, Ethiopia, 2003-2004). We highlight possible epidemic risk factors and review delays in epidemic detection and response (up to 20 weeks), essentially due to poor case reporting and analysis or low use of public facilities. Epidemics lasted 15-36 weeks, and patients' age profiles suggested departures from classical notions of epidemic malaria everywhere but Burundi. Although emergency interventions were mounted to expand inpatient and outpatient treatment access, we believe their effects were lessened because of delays, insufficient evaluation of disease burden, lack of evidence on how to increase treatment coverage in emergencies, and use of ineffective drugs.


Health status after transcatheter aortic valve replacement in patients at extreme surgical risk: results from the CoreValve U.S. trial.
The purpose of this study was to characterize health status outcomes after transcatheter aortic valve replacement (TAVR) with a self-expanding bioprosthesis among patients at extreme surgical risk and to identify pre-procedural patient characteristics associated with a poor outcome. For many patients considering TAVR, improvement in quality of life may be of even greater importance than prolonged survival. Patients with severe, symptomatic aortic stenosis who were considered to be at prohibitive risk for surgical aortic valve replacement were enrolled in the single-arm CoreValve U.S. Extreme Risk Study. Health status was assessed at baseline and at 1, 6, and 12 months after TAVR using the Kansas City Cardiomyopathy Questionnaire (KCCQ), the Short Form-12, and the EuroQol-5D. The overall summary scale of the KCCQ (range 0 to 100; higher scores = better health) was the primary health status outcome. A poor outcome after TAVR was defined as death, a KCCQ overall summary score (OS) <45, or a decline in KCCQ-OS of 10 points at 6-month follow-up. A total of 471 patients underwent TAVR via the transfemoral approach, of whom 436 (93%) completed the baseline health status survey. All health status measures demonstrated considerable impairment at baseline. After TAVR, there was substantial improvement in both disease-specific and generic health status measures, with an increase in the KCCQ-OS of 23.9 points (95% confidence interval [CI]: 20.3 to 27.5 points) at 1 month, 27.4 points (95% CI: 24.2 to 30.6 points) at 6 months, 27.4 points (95% CI: 24.1 to 30.8 points) at 12 months, along with substantial increases in Short Form-12 scores and EuroQol-5D utilities (all p < 0.003 compared with baseline). Nonetheless, 39% of patients had a poor outcome after TAVR. Baseline factors independently associated with poor outcome included wheelchair dependency, lower mean aortic valve gradient, prior coronary artery bypass grafting, oxygen dependency, very high predicted mortality with surgical aortic valve replacement, and low serum albumin. Among patients with severe aortic stenosis, TAVR with a self-expanding bioprosthesis resulted in substantial improvements in both disease-specific and generic health-related quality of life, but there remained a large minority of patients who died or had very poor quality of life despite TAVR. Predictive models based on a combination of clinical factors as well as disability and frailty may provide insight into the optimal patient population for whom TAVR is beneficial. (Safety and Efficacy Study of the Medtronic CoreValve® System in the Treatment of Symptomatic Severe Aortic Stenosis in High Risk and Very High Risk Subjects Who Need Aortic Valve Replacement; NCT01240902).


Epidemiology, medical outcomes and costs of catheter-related bloodstream infections in intensive care units of four European countries: literature- and registry-based estimates.
Despite high incidence rates, little information is available on the burden of illness of catheter-related bloodstream infections (CRBSIs) in Europe. A review of the available data was performed to estimate the clinical outcomes and costs associated with CRBSIs during intensive care unit (ICU) stays in four European countries (France, Germany, Italy and the UK). Based on these data we have estimated the CRBSI-related mortality and the annual costs associated with CRBSIs in the aforementioned countries. Results show large variation between countries: 1.12-4.2 CRBSI per 1000 catheter days, 8400-14,400 CRBSIs episodes per year, 1000-1584 deaths per year, 15,960-201,600 ICU days caused by CRBSIs and euro35.9 to euro163.9 million associated costs. Discrepancies are mainly explained by the heterogeneous quality of epidemiological studies, as well as the variety of national clinical practices.


Recovery After Genocide: Understanding the Dimensions of Recovery Capital Among Incarcerated Genocide Perpetrators in Rwanda.
Utilizing survey data from 302 men and women incarcerated in the Rwandan correctional system for the crime of genocide, and structured interviews with 75 prisoners, this mixed methods study draws on the concept of recovery capital to understand how individuals convicted of genocide navigate post-genocide healing. Genocide smashes physical and human capital and perverts social and cultural capital. Experiencing high levels of posttraumatic stress symptoms with more than two-thirds of the sample scoring above typical civilian cut-off levels, raised levels of depression, and high levels of anxiety, and failing physical health, the genocide perpetrators require multiple sources of recovery capital to foster internal resilience as they look forward to rebuilding their own lives.


Human-specific organization of primary visual cortex: alternating compartments of dense Cat-301 and calbindin immunoreactivity in layer 4A.
There is evidence that the cortical anatomy of the magnocellular (M) visual pathway, which carries information about motion and luminance contrast, was modified in human evolution. Recent results indicate that layer 4A of humans contains a meshwork of tissue bands that stain densely for nonphosphorylated neurofilament (NPNF), a protein that is preferentially expressed in elements of the M pathway, whereas apes and monkeys lack a comparable pattern. Here we examined the distribution of staining for Cat-301 -- a monoclonal antibody well established to stain M-related structures preferentially -- in area V1 of humans, apes (chimpanzees, orangutan), Old World monkeys (macaques) and New World monkeys (spider monkeys, squirrel monkeys). Single-staining experiments, using a peroxidase-tetramethylbenzidine (TMB) reaction, revealed alternating zones of dark and light staining for Cat-301 in layer 4A of humans, similar to those observed with NPNF. Double-staining studies in humans revealed that Cat-301-immunoreactive somas and neuropil were localized within the same tissue bands that stained strongly for NPNF and, furthermore, that these bands alternated with irregularly shaped territories that stained very strongly for calbindin. Nonhuman primates, by contrast to humans, displayed weak Cat-301 and calbindin staining in layer 4A. The co-localization of Cat-301 and NPNF in human layer 4A, and the weak staining for these molecules in layer 4A of other primates, suggests that the cortical representation of the M channel was modified in recent human evolution. The calbindin-rich compartments in human layer 4A cannot be related to a particular geniculostriate pathway on neurochemical grounds; they may constitute an interneuronal population that increased in human evolution.


Induction of IgE synthesis in normal human B cells. Sequential requirements for activation by an alloreactive T cell clone and IgE-potentiating factors.
Two human alloreactive T cell clones were established from a one-way mixed lymphocyte culture involving two nonatopic donors, and were assessed for their capacity to induce IgE synthesis by B cells obtained from the original stimulator. The two alloreactive T cell clones studied induced IgG but not IgE synthesis in normal B cells. However, one of the two clones, clone 2H6, induced IgE synthesis in the presence of supernatants from T cell lines derived from patients with the hyper-IgE syndrome (HIE), and enriched for T cells bearing receptors for IgE. These supernatants by themselves caused no IgE synthesis in nonatopic B cells. The potentiating factors in these supernatants were shown to bind to IgE. Time sequence experiments indicated that interaction of the B cells with the alloreactive clone 2H6 renders them responsive to the action of the IgE-potentiating factors. These results indicate that induction of IgE synthesis in normal B cells involves at least two sequential T cell derived signals. Furthermore, T cell clones are heterogenous in their capacity to provide these signals.


Alternating vector immunizations encoding pre-erythrocytic malaria antigens enhance memory responses in a malaria endemic area.
A heterologous prime-boost strategy has been developed to potently induce T cell responses to pre-erythrocytic malaria antigens. Efficacy in the field is likely to depend on both peak immunogenicity and the durability of responses. To improve both immunogenicity and durability of responses, 54 adult males from a malaria endemic area were immunized with different vaccination regimens, systematically varying antigenic insert and the number and sequence of component vaccinations. The component vaccinations were recombinant attenuated viruses, either fowlpox (FP) 9 or modified vaccinia virus Ankara (MVA). These were recombinant for either of two pre-erythrocytic malaria antigens (multiple epitope-thrombospondin-related adhesion protein, ME-TRAP, or circumsporozoite antigen (CS). ELISPOT assays were used to measure the effector and resting memory T cell responses. Sequence, antigen insert and number of vaccinations influenced immunogenicity, but the novel alternating vector immunizations generated the largest resting memory T cell populations. Effector responses were maintained at 84% of the peak response after 270 days. This durability of response is unprecedented. Classical prime-boost vaccination responses were at 5% of the peak after 270 days. Vaccines administered by heterologous prime-boost regimes are being developed for diverse pathogens and cancer. These data suggest these vaccines should also be administered by alternating vector regimens in clinical development.


Exploring the Burden of Mealtime Insulin Dosing in Adults and Children With Type 1 Diabetes.
Timely and accurate mealtime insulin dosing can be an ongoing challenge for people with type 1 diabetes. This multinational, online study aimed to explore attitudes and behaviors around mealtime insulin dosing and the impact of mealtime dose timing, particularly with regard to premeal dosing (15-20 minutes before a meal). Although the majority of surveyed participants (96%) recognized the importance of accurate mealtime bolus insulin dosing, only a small proportion (35%) reported being "very confident" in accurate bolus insulin estimation. Given the choice, the majority of participants would prefer to administer insulin immediately before or after a meal, as this timing would improve their quality of life.


Planning elderly and palliative care in Montenegro.
Montenegro, a newly independent Balkan state with a population of 650,000, has a health care reform programme supported by the World Bank. This paper describes planning for integrated elderly and palliative care. The current service is provided only through a single long-stay hospital, which has institutionalised patients and limited facilities. Broad estimates were made of current financial expenditures on elderly care. A consultation was undertaken with stakeholders to propose an integrated system linking primary and secondary health care with social care; supporting people to live, and die well, at home; developing local nursing homes for people with higher dependency; creating specialised elderly-care services within hospitals; and providing good end-of-life care for all who need it. Effectiveness may be measured by monitoring patient and carers' perceptions of the care experience. Changes in provision of elderly care may be achieved through redirection of existing resources, but the health and social care services also need to enhance elderly care budgets. The challenges for implementation include management skills, engaging professionals and political commitment. Middle-income countries such as Montenegro can develop elderly and palliative care services through redirection of existing finance if accompanied by new service objectives, staff skills and integrated management.


A randomized study of contingency management and spirometric lung age for motivating smoking cessation among injection drug users.
Even after quitting illicit drugs, tobacco abuse remains a major cause of morbidity and mortality in former injection drug users. An important unmet need in this population is to have effective interventions that can be used in the context of community based care. Contingency management, where a patient receives a monetary incentive for healthy behavior choices, and incorporation of individual counseling regarding spirometric "lung age" (the age of an average healthy individual with similar spirometry) have been shown to improve cessation rates in some populations. The efficacy of these interventions on improving smoking cessation rates has not been studied among current and former injection drug users. In a randomized, factorial design study, we recruited 100 active smokers from an ongoing cohort study of current and former injection drug users to assess the impact of contingency management and spirometric lung age on smoking cessation. The primary outcome was 6-month biologically-confirmed smoking cessation comparing contingency management, spirometric lung age or both to usual care. Secondary outcomes included differences in self-reported and biologically-confirmed cessation at interim visits, number of visits attended and quit attempts, smoking rates at interim visits, and changes in Fagerstrom score and self-efficacy. Six-month biologically-confirmed smoking cessations rates were 4% usual care, 0% lung age, 14% contingency management and 0% for combined lung age and contingency management (p = 0.13). There were no differences in secondary endpoints comparing the four interventions or when pooling the lung age groups. Comparing contingency management to non-contingency management, 6-month cessation rates were not different (7% vs. 2%; p = 0.36), but total number of visits with exhaled carbon monoxide-confirmed abstinence were higher for contingency management than non-contingency management participants (0.38 vs. 0.06; p = 0.03), and more contingency management participants showed reduction in their Fagerstrom score from baseline to follow-up (39% vs. 18%; p = 0.03). While lung age appeared ineffective, contingency management was associated with more short-term abstinence and lowered nicotine addiction. Contingency management may be a useful tool in development of effective tobacco cessation strategies among current and former injection drug users. Clinicaltrials.gov NCT01334736 (April 12, 2011).


On the role of keratinised mucosa at dental implants: a 5-year prospective single-cohort study.
The purpose of this study was to evaluate the role of keratinised mucosa on the long-term success of dental implants. Thirty-two edentulous patients had one of their dental arches rehabilitated with a provisional screw-retained resin reinforced cross-arch fixed prosthesis supported by four immediately loaded implants. The two central straight implants were randomly allocated in two equal groups, according to a parallel-group design, to receive or not intermediate abutments (Multi-Unit Abutment, MUA). However, for the purpose of the present publication, the study was considered as a prospective single cohort study. To be immediately loaded, implants had to be inserted with a minimum torque of 30 Ncm, which was achieved by all implants. Provisional prostheses were delivered within 24 hours and were replaced, after 4 months, by definitive screw-retained metal-ceramic prostheses. Patients were followed up to 5 years after loading. Prostheses were removed every 8 months to facilitate professionally delivered maintenance. Outcome measures were prosthesis and implant failures, complications, peri-implant marginal bone level (MBL) changes, bleeding on probing (BoP) and keratinised mucosa height (KMH). The absence of vestibular or lingual keratinised mucosa was put in relation with peri-implant bone loss and BoP. Five-years after loading two patients dropped out and no implant or prosthetic failure occurred. Four patients were affected by prosthetic complications. At delivery of definitive prostheses, 32 (25%) implants had at least one vestibular or lingual site with no KMH and 96 (75%) implants had both sites with some KMH. Multi-level models did not show any statistically significant association between KMH at the time of delivery of the definitive prosthesis and changes in MBL and BoP at 5 years (estimate = 0.02; 95% CI: -0.02 to 0.05; P = 0.3393; and 1.02; 95% CI: 0.79 to 1.33; P = 0.8695, respectively). When KMH was analysed as dichotomous variable, implants with presence of KMH at delivery of definitive prosthesis at both vestibular and lingual aspects showed a trend of less BoP (estimate = -0.8; 95% CI: -1.69 to 0.08; P = 0.0741) but a statistically significant greater MBL loss compared to implants where KMH was only present at one site (estimate = 0.18; 95% CI: -0.1 to 0.3; P = 0.0041). The 5-year after loading outcome of immediately loaded screw-retained cross-arch prostheses supported by four implants is excellent in both the maxilla and mandible. Although the height of the keratinised mucosa did not seem to alter the clinical outcomes, its presence both at vestibular and lingual sites was associated with an increased marginal bone loss when compared to implants having at least one side without keratinised mucosa.


Fast ultrasound beam prediction for linear and regular two-dimensional arrays.
Real-time beam predictions are highly desirable for the patient-specific computations required in ultrasound therapy guidance and treatment planning. To address the longstanding issue of the computational burden associated with calculating the acoustic field in large volumes, we use graphics processing unit (GPU) computing to accelerate the computation of monochromatic pressure fields for therapeutic ultrasound arrays. In our strategy, we start with acceleration of field computations for single rectangular pistons, and then we explore fast calculations for arrays of rectangular pistons. For single-piston calculations, we employ the fast near-field method (FNM) to accurately and efficiently estimate the complex near-field wave patterns for rectangular pistons in homogeneous media. The FNM is compared with the Rayleigh-Sommerfeld method (RSM) for the number of abscissas required in the respective numerical integrations to achieve 1%, 0.1%, and 0.01% accuracy in the field calculations. Next, algorithms are described for accelerated computation of beam patterns for two different ultrasound transducer arrays: regular 1-D linear arrays and regular 2-D linear arrays. For the array types considered, the algorithm is split into two parts: 1) the computation of the field from one piston, and 2) the computation of a piston-array beam pattern based on a pre-computed field from one piston. It is shown that the process of calculating an array beam pattern is equivalent to the convolution of the single-piston field with the complex weights associated with an array of pistons. Our results show that the algorithms for computing monochromatic fields from linear and regularly spaced arrays can benefit greatly from GPU computing hardware, exceeding the performance of an expensive CPU by more than 100 times using an inexpensive GPU board. For a single rectangular piston, the FNM method facilitates volumetric computations with 0.01% accuracy at rates better than 30 ns per field point. Furthermore, we demonstrate array calculation speeds of up to 11.5 X 10(9) field-points per piston per second (0.087 ns per field point per piston) for a 512-piston linear array. Beam volumes containing 256(3) field points are calculated within 1 s for 1-D and 2-D arrays containing 512 and 20(2) pistons, respectively, thus facilitating future real-time thermal dose predictions.


Case-control investigation of occupational exposure to chlorinated solvents and non-Hodgkin's lymphoma.
Although many studies have investigated the association between trichloroethylene (TCE) exposure and non-Hodgkin's lymphoma (NHL), less is known about other chlorinated solvents. We extended our previous analysis of occupational TCE exposure in a multicentre population-based case-control study of NHL to investigate associations with five additional chlorinated solvents: 1,1,1,-trichloroethane, carbon tetrachloride, chloroform, methylene chloride and perchloroethylene. Cases (n=1189) and controls (n=982) provided detailed information on their occupational histories and workplace exposure to chlorinated solvents for selected occupations using job-specific interview modules. An industrial hygienist used this information and a review of the literature to assess occupational exposure to chlorinated solvents. We computed ORs and 95% CIs for different exposure metrics, with the unexposed group as the referent. We also computed ORs by NHL subtype. High cumulative hours exposed to carbon tetrachloride was associated with NHL (>520 hours: OR 1.9; 95% CI 1.0 to 3.6; Ptrend=0.04). This association remained after restricting to jobs with high-intensity exposure (OR 2.0; 95% CI 1.1 to 3.8; P=0.03) and ≥90% exposure probability (OR 2.1; 95% CI 1.0 to 4.3; P=0.03), adjusting for TCE (OR 2.1; 95% CI 1.0- to 4.1; P=0.04) and incorporating a 15-year lag (OR 1.9; 95% CI 1.0 to 3.6; P=0.06). The other evaluated chlorinated solvents were not associated with NHL. This is the first study using high-quality quantitative exposure assessment methods to identify a statistically significant elevated association between occupational exposure to carbon tetrachloride and NHL. Our findings, although limited by a small number of exposed cases, offer evidence that carbon tetrachloride may be a lymphomagen.


Bats at risk? Bat activity and insecticide residue analysis of food items in an apple orchard.
Although bats are reported as being threatened by pesticides, they are currently not considered in European Union pesticide risk assessments. The reason for that contradiction is probably related to the scarcity of information on bat activity in pesticide-treated fields and the pesticide residues on their food items. The authors recorded bat activity and measured pesticide residues on bat-specific food items following applications of two insecticides in an apple orchard. High activity levels of the common pipistrelle bat, a foraging habitat generalist, were detected. Airborne foragers and bats that take part of their food by gleaning arthropods from the vegetation were recorded frequently. The initial value and the decline of pesticide residues were found to depend on the arthropod type, their surface to volume ratio, their mobility, and the mode of action of the applied pesticide. The highest initial residue values were measured on foliage-dwelling arthropods. By following the toxicity-exposure ratio approaches of the current pesticide risk assessment, no acute dietary risk was found for all recorded bat species. However, a potential reproductive risk for bat species that include foliage-dwelling arthropods in their diet was indicated. The results emphasize the importance of adequately evaluating the risks of pesticides to bats, which, compared to other mammals, are potentially more sensitive due to their ecological traits.


Alpha 2-macroglobulin binds CpG oligodeoxynucleotides and enhances their immunostimulatory properties by a receptor-dependent mechanism.
CpG oligodeoxynucleotides (ODN) stimulate the immune system and are under evaluation as treatments and vaccine adjuvants for infectious diseases, cancer, and immune system disorders. Although they have shown promising results in numerous clinical trials, the ultimate use of CpG ODN-based therapeutics may hinge on improved pharmacokinetics and reduced systemic side-effects. CpG ODN efficacy and potency might be enhanced greatly by packaging them into particles that protect them from degradation and specifically target them for uptake by immune-competent cells. The plasma proteinase inhibitor alpha 2-macroglobulin (alpha 2M) binds numerous biologically active macromolecules, including cytokines, chemokines, and growth factors, and can modulate their activity. Molecules bound to alpha 2M are protected from interactions with neighboring macromolecules and are targeted for receptor-mediated uptake by immune-competent cells. Here, we report that activated alpha 2M (alpha 2M*) binds CpG ODN and enhances their immunostimulatory properties significantly. Murine macrophages treated with alpha 2M*-ODN complexes respond more rapidly and produce a greater cytokine response than induced by free CpG ODN. Using human PBMC, alpha 2M*-ODN complexes exhibit fourfold enhanced potency and 15-fold greater efficacy for stimulating production of inflammatory cytokines. alpha 2M* targets delivery of CpG ODN specifically to immune-competent cells, which endocytose the complexes sixfold more rapidly than free CpG ODN. CpG ODN bound to alpha 2M* are also protected from degradation by nucleases. This novel targeting technology may improve CpG ODN-based therapeutics by increasing efficacy at reduced doses, thus reducing side-effects and cost.


Epithelial β1 integrin is required for lung branching morphogenesis and alveolarization.
Integrin-dependent interactions between cells and extracellular matrix regulate lung development; however, specific roles for β1-containing integrins in individual cell types, including epithelial cells, remain incompletely understood. In this study, the functional importance of β1 integrin in lung epithelium during mouse lung development was investigated by deleting the integrin from E10.5 onwards using surfactant protein C promoter-driven Cre. These mutant mice appeared normal at birth but failed to gain weight appropriately and died by 4 months of age with severe hypoxemia. Defects in airway branching morphogenesis in association with impaired epithelial cell adhesion and migration, as well as alveolarization defects and persistent macrophage-mediated inflammation were identified. Using an inducible system to delete β1 integrin after completion of airway branching, we showed that alveolarization defects, characterized by disrupted secondary septation, abnormal alveolar epithelial cell differentiation, excessive collagen I and elastin deposition, and hypercellularity of the mesenchyme occurred independently of airway branching defects. By depleting macrophages using liposomal clodronate, we found that alveolarization defects were secondary to persistent alveolar inflammation. β1 integrin-deficient alveolar epithelial cells produced excessive monocyte chemoattractant protein 1 and reactive oxygen species, suggesting a direct role for β1 integrin in regulating alveolar homeostasis. Taken together, these studies define distinct functions of epithelial β1 integrin during both early and late lung development that affect airway branching morphogenesis, epithelial cell differentiation, alveolar septation and regulation of alveolar homeostasis.


Larval competition reduces body condition in the female seed beetle, Callosobruchus maculatus.
Early body condition may be important for adult behavior and fitness, and is impacted by a number of environmental conditions and biotic interactions. Reduced fecundity of adult females exposed to larval competition may be caused by reduced body condition or shifts in relative body composition, yet these mechanisms have not been well researched. Here, body mass, body size, scaled body mass index, and two body components (water content and lean dry mass) of adult Callosobruchus maculatus (Fabricius) (Coleoptera: Chrysomelidae: Bruchinae) females exposed to larval competition or reared alone were examined. Experimental females emerged at significantly smaller body mass and body size than control females. Additionally, scaled body mass index and water content, but not lean dry mass, were significantly reduced in experimental females. To our knowledge, these are the first results that demonstrate a potential mechanism for previously documented direct effects of competition on fecundity in female bruchine beetles.


Cytokines and growth factors in wound drainage fluid from patients undergoing incisional hernia repair.
Knowing the dynamics of growth factor and cytokine secretion within the site of a surgical operation is important, as they play a crucial role in the pathophysiology of wound healing and are a target for modifying the repair response. The aim of this study was to evaluate the production of several cytokines and growth factors in the drainage wound fluid from patients undergoing incisional hernia repair: namely, interleukin (IL)-6, IL-10, IL-1alpha, IL-1 ra, interferon-gamma, vascular endothelial growth factors and basic fibroblast growth factor. Ten female patients with abdominal midline incisional hernia undergoing surgical repair were included in this study. In all cases, a closed-suction drain was inserted in the wound below the fascia and removed on postoperative day 4. Wound fluid was collected on postoperative days 1-4 and the amount was recorded each time. Growth factors and cytokines production was evaluated as the whole amount produced over a 24-hour period. In all patients, the amount of drain fluid from surgical wounds was more copious the first day after surgery, it decreased significantly afterward. The presence of all cytokines was highest on postoperative day 1, decreasing over the following days. More specifically, the production of IL-1 ra, IL-6, IL-1alpha, and IL-10 on postoperative day 1 fell sharply on postoperative days 3 and 4, whereas, after an initial reduction, interferon-gamma showed an increase from day 2 onward. Vascular endothelial-derived growth factor production increased progressively after the operation reaching statistical significance only on day 4. As for basic fibroblast growth factor, it showed an opposite pattern: it was higher on postoperative day 1 decreasing thereafter. This analysis of cytokine and growth factor production in the drain fluid will lead us to a better evaluation of the events that follow a surgical wound and to a better understanding of the healing process.


Prospective study of toenail selenium levels and cancer among women.
Inverse associations between selenium status and cancer risk have been observed in animal studies, ecologic studies, and some case-control and prospective studies. Whereas results of some prospective studies have suggested an overall inverse relationship between selenium levels and cancer, other prospective studies have failed to confirm this finding. Prospective data on women are particularly limited because fewer women than men have been studied prospectively. The aim of this study was to prospectively examine the relationship between selenium levels in toenails (previously shown to reflect selenium intake) and incidence of cancer among women. The Nurses' Health Study cohort began in 1976 with 121,700 female nurses aged 30-55 years living in 11 U.S. states. In 1982, we requested toenail clippings from the members of the cohort, and 62,641 participants with no history of cancer returned these clippings. During 41 months of follow-up, 503 cases of cancer other than breast cancer (results previously reported) or nonmelanoma skin cancer were analyzed. For each case patient, a control subject was chosen from women who remained free of diagnosed cancer, matched by age and by date of nail return. No inverse association was observed between selenium levels in toenails and cancer risk. The age- and smoking-adjusted relative risk (RR), comparing the highest with the lowest quintile of toenail selenium level, was 1.44 (95% confidence interval [CI] = 0.97-2.13), and the trend across quintiles was marginally significant (two-sided P = .06). Comparing the highest with the lowest decile, the RR (age- and smoking-adjusted) was 1.77 (95% CI = 1.04-3.02). When these data were combined with the data from 434 breast cancer case patients and their matched control subjects identified in parallel from this same cohort, the RR comparing the highest with the lowest quintile was 1.24 (95% CI = 0.93-1.65). Toenail selenium level was not inversely associated with cancer at any major site, including uterine cancer, colorectal cancer, melanoma, ovarian cancer, or lung cancer (after adjusting for smoking); in fact, nonsignificant positive associations were observed at several sites. Toenail selenium levels were not inversely associated with cancer risk in this study. These data, in conjunction with previous findings of no association between toenail selenium status and breast cancer risk, strongly suggest that higher selenium intake within the range consumed by most U.S. women (as reflected by toenail selenium levels) is not protective against overall cancer incidence in women.


Danger in the reef: Proteome, toxicity, and neutralization of the venom of the olive sea snake, Aipysurus laevis.
Four specimens of the olive sea snake, Aipysurus laevis, were collected off the coast of Western Australia, and the venom proteome was characterized and quantitatively estimated by RP-HPLC, SDS-PAGE, and MALDI-TOF-TOF analyses. A. laevis venom is remarkably simple and consists of phospholipases A2 (71.2%), three-finger toxins (3FTx; 25.3%), cysteine-rich secretory proteins (CRISP; 2.5%), and traces of a complement control module protein (CCM; 0.2%). Using a Toxicity Score, the most lethal components were determined to be short neurotoxins. Whole venom had an intravenous LD50 of 0.07 mg/kg in mice and showed a high phospholipase A2 activity, but no proteinase activity in vitro. Preclinical assessment of neutralization and ELISA immunoprofiling showed that BioCSL Sea Snake Antivenom was effective in cross-neutralizing A. laevis venom with an ED50 of 821 μg venom per mL antivenom, with a binding preference towards short neurotoxins, due to the high degree of conservation between short neurotoxins from A. laevis and Enhydrina schistosa venom. Our results point towards the possibility of developing recombinant antibodies or synthetic inhibitors against A. laevis venom due to its simplicity.


Experimental optic neuritis induced by the microinjection of lipopolysaccharide into the optic nerve.
Optic neuritis (ON) is a condition involving primary inflammation, demyelination, and axonal injury in the optic nerve which leads to retinal ganglion cell (RGC) loss, and visual dysfunction. We investigated the ability of a single microinjection of bacterial lipopolysaccharide (LPS) directly into the optic nerve to induce functional and structural alterations compatible with ON. For this purpose, optic nerves from male Wistar rats remained intact or were injected with vehicle or LPS. The effect of LPS was evaluated at several time points post-injection in terms of: i) visual pathway and retinal function (visual evoked potentials (VEPs) and electroretinograms, (ERGs), respectively), ii) anterograde transport from the retina to its projection areas, iii) consensual pupil light reflex (PLR), iv) optic nerve histology, v) microglia/macrophage reactivity (by Iba-1- and ED1-immunostaining), vi) astrocyte reactivity (by glial fibrillary acid protein-immunostaining), vii) axon number (by toluidine blue staining), vii) demyelination (by myelin basic protein immunoreactivity and luxol fast blue staining), viii) optic nerve ultrastructure, and ix) RGC number (by Brn3a immunoreactivity). LPS induced a significant and persistent decrease in VEP amplitude and PLR, without changes in the ERG. In addition, LPS induced a deficit in anterograde transport, and an early inflammatory response consisting in an increased cellularity, and Iba-1 and ED1-immunoreactivity in the optic nerve, which were followed by changes in axonal density, astrocytosis, demyelination, and axon and RGC loss. These results suggest that the microinjection of LPS into the optic nerve may serve as a new experimental model of primary ON.


A method to measure sound transmission via the malleus-incus complex.
The malleus-incus complex (MIC) plays a crucial role in the hearing process as it transforms and transmits acoustically-induced motion of the tympanic membrane, through the stapes, into the inner-ear. However, the transfer function of the MIC under physiologically-relevant acoustic stimulation is still under debate, especially due to insufficient quantitative data of the vibrational behavior of the MIC. This study focuses on the investigation of the sound transformation through the MIC, based on measurements of three-dimensional motions of the malleus and incus with a full six degrees of freedom (6 DOF). The motion of the MIC was measured in two cadaveric human temporal bones with intact middle-ear structures excited via a loudspeaker embedded in an artificial ear canal, in the frequency range of 0.5-5 kHz. Three-dimensional (3D) shapes of the middle-ear ossicles were obtained by sequent micro-CT imaging, and an intrinsic frame based on the middle-ear anatomy was defined. All data were registered into the intrinsic frame, and rigid body motions of the malleus and incus were calculated with full six degrees of freedom. Then, the transfer function of the MIC, defined as velocity of the incus lenticular process relative to velocity of the malleus umbo, was obtained and analyzed. Based on the transfer function of the MIC, the motion of the lenticularis relative to the umbo reduces with frequency, particularly in the 2-5 kHz range. Analysis of the individual motion components of the transfer function indicates a predominant medial-lateral component at frequencies below 1 kHz, with low but considerable anterior-posterior and superior-inferior components that become prominent in the 2-5 kHz range. The transfer function of the human MIC, based on motion of the umbo and lenticularis, has been visualized and analyzed. While the magnitude of the transfer function decreases with frequency, its spatio-temporal complexity increases significantly.


Ionotropic glutamate receptor binding and subunit mRNA expression in thalamic nuclei in schizophrenia.
Both thalamic and glutamatergic dysfunction have been implicated in the pathophysiology of schizophrenia. The authors examined ionotropic glutamate receptor expression in postmortem samples from patients with schizophrenia and comparison subjects, using the hypothesis that glutamate receptor expression differs in limbic nuclei of the thalamus in schizophrenia. N-Methyl-D-aspartate (NMDA), AMPA, and kainate receptor expression was determined in six thalamic nuclei from 12 subjects with DSM-III-R diagnoses of schizophrenia and eight psychiatrically normal individuals. The authors used in situ hybridization to determine NMDAR1, NMDAR2A-NMDAR2D, gluR1-gluR7, KA1, and KA2 subunit mRNA levels and receptor autoradiography to determine binding to glutamate binding sites of the three receptor subtypes and to the glycine, polyamine, and ion channel binding sites of the NMDA receptor. Glutamate receptor expression was lower at both transcriptional (NMDAR1, NMDAR2B, NMDAR2C, gluR1, gluR3, and KA2 subunit mRNAs) and posttranscriptional ([(3)H]ifenprodil and [(3)H]MDL105,519 binding to polyamine and glycine sites of the NMDA receptor) levels in the thalamus in patients with schizophrenia than in comparison subjects, but differences were most prominent in nuclei with reciprocal projections to limbic regions. Abnormalities in NMDA, AMPA, and kainate receptor expression in limbic thalamus are suggestive of the NMDA receptor hypoactivity hypothesis of schizophrenia and are consistent with diminished glutamatergic activity in the thalamus in schizophrenia. Alternatively, these results could suggest abnormal glutamatergic innervation in afferent and/or efferent regions, which are limbic structures that have been implicated in this illness. These results may provide a neurochemical anatomical substrate for antipsychotic therapies targeting ionotropic glutamate receptors.


Use of Personal Protective Equipment Among Healthcare Workers During the First and the Second Wave of the COVID-19 Pandemic.
To treat and properly care for COVID-19 patients it is vital to have healthy healthcare workers to ensure the continued function of the healthcare system and to prevent transmission of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) to patients, co-workers, and the community. Personal protective equipment (PPE) can prevent healthcare workers from being infected with and transmitting SARS-CoV-2. Experience and training are pivotal to ensure optimal protection. This study aims to examine the use and failure of PPE and compliance with PPE guidelines during the first and the second wave of the COVID-19 pandemic among Danish healthcare workers. Healthcare workers from the Central Denmark Region and the Capital Region of Denmark were invited to participate April-June 2020 during the first wave and November 2020-April 2021 during the second wave. Day-by-day, participants reported work procedures, use and failure of PPE, and compliance with PPE guidelines. Register-based information on sex, age, department, and profession was available for all participants. In total, 21 684 and 10 097 healthcare workers participated during the first and the second wave, respectively. During the first wave, 1.7% used filtering face piece-2 or -3 (FFP2 or FFP3) respirators and 8.2% used face masks [fluid resistant (type IIR) masks, masks with visor (typically type IIR), and other unspecified face masks] during physical contact with patients. During the second wave, the corresponding figures increased to 17.8% and 80.7%. During respiratory procedures, the use of FFP2 or FFP3 respirators increased from 5.6 to 24.3%, and the use of face masks from 14.7 to 77.8%. The no PPE use decreased from 21.3% during the first wave to 0.4% in the second wave, during respiratory procedures. Total PPE failures decreased from 0.7 to 0.4% from the first to second wave. The proportion not complying with PPE guidelines declined from 3.6 to 2.2% during physical contact with patients and from 6.5 to 4.6% during respiratory procedures. PPE failure and non-compliance varied by age, sex and type of department. Frequent reasons for non-compliance were forgetfulness and lack of time, and during the first but not during the second wave, limited availability of PPE. We found a substantial increase in the use of PPE and a substantial decrease in PPE failures from the first to the second wave of COVID-19 in Denmark. However, there is still a need for continuous focus on compliance in use of PPE among healthcare workers.


Copper Resistance Mediates Long-Term Survival of Cupriavidus metallidurans in Wet Contact With Metallic Copper.
Metallic copper to combat bacterial proliferation in drinking water systems is being investigated as an attractive alternative to existing strategies. A potential obstacle to this approach is the induction of metal resistance mechanisms in contaminating bacteria, that could severely impact inactivation efficacy. Thus far, the role of these resistance mechanisms has not been studied in conditions relevant to drinking water systems. Therefore, we evaluated the inactivation kinetics of Cupriavidus metallidurans CH34 in contact with metallic copper in drinking water. Viability and membrane permeability were examined for 9 days through viable counts and flow cytometry. After an initial drop in viable count, a significant recovery was observed starting after 48 h. This behavior could be explained by either a recovery from an injured/viable-but-non-culturable state or regrowth of surviving cells metabolizing lysed cells. Either hypothesis would necessitate an induction of copper resistance mechanisms, since no recovery was seen in a CH34 mutant strain lacking metal resistance mechanisms, while being more pronounced when copper resistance mechanisms were pre-induced. Interestingly, no biofilms were formed on the copper surface, while extensive biofilm formation was observed on the stainless steel control plates. When CH34 cells in water were supplied with CuSO4, a similar initial decrease in viable counts was observed, but cells recovered fully after 7 days. In conclusion, we have shown that long-term bacterial survival in the presence of a copper surface is possible upon the induction of metal resistance mechanisms. This observation may have important consequences in the context of the increasing use of copper as an antimicrobial surface, especially in light of potential co-selection for metal and antimicrobial resistance.


Identification and sequence characterization of novel Theileria genotypes from the waterbuck (Kobus defassa) in a Theileria parva-endemic area in Kenya.
Waterbuck (Kobus defassa), an ungulate species endemic to the Eastern African savannah, is suspected of being a wildlife reservoir for tick-transmitted parasites infective to livestock. Waterbuck is infested by large numbers of Rhipicephalus appendiculatus, the tick vector for Theileria parva, and previous data suggests that the species may be a source of T. parva transmission to cattle. In the present study, a total of 86 cattle and 26 waterbuck blood samples were obtained from Marula, a site in Kenya endemic for East Coast fever (ECF) where the primary wildlife reservoir of T. parva the Cape buffalo (Syncerus caffer) is also common. To investigate for the presence of cattle-infective Theileria parasites, DNA specimens extracted from the blood samples were subjected to two diagnostic assays; a nested PCR based on the p104 gene that is specific for T. parva, and a reverse line blot (RLB) incorporating 13 oligonucleotide probes including all of the Theileria spp. so far described from livestock and wildlife in Kenya. Neither assay provided evidence of T. parva or Theileria sp. (buffalo) infection in the waterbuck DNA samples. By contrast, majority of the cattle samples (67.4%) were positive for T. parva using a nested PCR assay. The RLB assay, including a generic probe for the genus Theileria, indicated that 25/26 (96%) of the waterbuck samples were positive for Theileria, while none of the 11 Theileria species-specific probes hybridized with the waterbuck-derived PCR products. Phylogenetic analysis of 18S ribosomal RNA (18S rRNA) and internal transcribed spacer (ITS) sequences within the RLB-positive waterbuck samples revealed the occurrence of three Theileria genotypes of unknown identity designated A, B and C. Group A clustered with Theileria equi, a pathogenic Theileria species and a causative agent of equine piroplasmosis in domestic equids. However, DNA from this group failed to hybridize with the T. equi oligonucleotide present on the RLB filter probe, suggesting the occurrence of novel taxa in these animals. This was confirmed by DNA sequencing that revealed heterogeneity between the waterbuck isolates and previously reported T. equi genotypes. Group B parasites clustered closely with Theileria luwenshuni, a highly pathogenic parasite of sheep and goats reported from China. Group C was closely related to Theileria ovis, an apparently benign parasite of sheep. Together, these findings provided no evidence that waterbuck plays a role in the transmission of T. parva. However, novel Theileria genotypes detected in this bovid species may be of veterinary importance.


Cell type-specific subunit composition of G protein-gated potassium channels in the cerebellum.
G protein-gated inwardly rectifying potassium (GIRK/Kir3) channels regulate cellular excitability and neurotransmission. In this study, we used biochemical and morphological techniques to analyze the cellular and subcellular distributions of GIRK channel subunits, as well as their interactions, in the mouse cerebellum. We found that GIRK1, GIRK2, and GIRK3 subunits co-precipitated with one another in the cerebellum and that GIRK subunit ablation was correlated with reduced expression levels of residual subunits. Using quantitative RT-PCR and immunohistochemical approaches, we found that GIRK subunits exhibit overlapping but distinct expression patterns in various cerebellar neuron subtypes. GIRK1 and GIRK2 exhibited the most widespread and robust labeling in the cerebellum, with labeling particularly prominent in granule cells. A high degree of molecular diversity in the cerebellar GIRK channel repertoire is suggested by labeling seen in less abundant neuron populations, including Purkinje neurons (GIRK1/GIRK2/GIRK3), basket cells (GIRK1/GIRK3), Golgi cells (GIRK2/GIRK4), stellate cells (GIRK3), and unipolar brush cells (GIRK2/GIRK3). Double-labeling immunofluorescence and electron microscopies showed that GIRK subunits were mainly found at post-synaptic sites. Altogether, our data support the existence of rich GIRK molecular and cellular diversity, and provide a necessary framework for functional studies aimed at delineating the contribution of GIRK channels to synaptic inhibition in the cerebellum.


Humidity- and light-driven actuators based on carbon nanotube-coated paper and polymer composite.
Multi-responsive actuators driven by different stimuli (e.g. light, humidity, electricity) have attracted intense attention recently for the advantages of being used in various environments and show enormous actuation. In this work, we propose humidity- and light-driven actuators based on carbon nanotube (CNT)-coated paper and a biaxially oriented polypropylene (BOPP) composite. The CNT-paper/BOPP actuator shows large bending actuation when driven by humidity change (curvature of 1.2 cm-1) and near infrared (NIR) light irradiation (curvature up to 1.6 cm-1). The great actuation performances outperform most other paper-based actuators. Finally, a smart gripper, of which the initial opening width can be enlarged, is fabricated on the basis of the CNT-paper/BOPP actuators. By utilizing the bidirectional bending motion of the actuator, the opening width of the gripper can increase to a width that is 4 times larger than its initial width, so as to grasp a large object. The gripper is also able to raise and move an object that is 20 times heavier than one actuator of the gripper. We assume that this new type of actuator has great potential in artificial muscle, soft robotics and biomimetic applications.


The role of infectious disease impact in informing decision-making for animal health management in aquaculture systems in Bangladesh.
The aquaculture sector in Bangladesh is an important employer and a significant source of foreign exchange. In addition, it contributes significantly to food security due to the role of fish in peoples' diets, the most important source of protein and micronutrients. However, infectious diseases represent an important barrier to sector development due to economic losses and vulnerability of smallholders. The aim of this study was to gain an overview of the impact of infectious diseases in the aquaculture sector, and to assess the usefulness and use of impact studies in decision making for animal health management and biosecurity governance in Bangladesh. A review of scientific and grey literature on infectious disease impact in different aquaculture systems was conducted and their methodologies and findings summarised. Subsequently, interviews with 28 stakeholders from the private and public sector were conducted to enquire about decision-making structures in animal health management. The data were analysed using the framework method to allow the development of themes, by using the information, experiences and opinions inductively obtained from interviewees, deductively through the reviewed literature. Results showed a substantial socio-economic impact of infectious diseases. The numerous stakeholders involved in the decision-making process explained that key barriers to effective aquaculture health management were insufficient resources to investigate and tackle infectious aquatic animal diseases, a dearth of legislation and capacity for disease surveillance, a reliance on reactive response, and a lack of impact and evidence-based approaches for prioritising problem-solving, commonly based on anecdotal evidence. Furthermore, communication among the multiple stakeholders involved was reported to be weak. This complex situation requires a multi-level response, which should span from strengthening the knowledge of farmers and professionals in the field to the improvement of surveillance and diagnostic systems. Improved systems along with evidence on disease impact could inform the prioritisation of diseases and resource allocation for disease control in Bangladesh. Further, this evidence needs to be used to advise decisions to have a true value, for which establishing and strengthening communication pathways and processes is critical to make systematic use of the information and improve animal health management. In the light of future threats to Bangladesh such as climate change, increasing population density and demand for animal source foods, it is crucial to strengthen animal health management systems to reduce livelihoods vulnerability, food insecurity and the likelihood of disease emergence.


A single-molecule diode.
We have designed and synthesized a molecular rod that consists of two weakly coupled electronic pi -systems with mutually shifted energy levels. The asymmetry thus implied manifests itself in a current-voltage characteristic with pronounced dependence on the sign of the bias voltage, which makes the molecule a prototype for a molecular diode. The individual molecules were immobilized by sulfur-gold bonds between both electrodes of a mechanically controlled break junction, and their electronic transport properties have been investigated. The results indeed show diode-like current-voltage characteristics. In contrast to that, control experiments with symmetric molecular rods consisting of two identical pi-systems did not show significant asymmetries in the transport properties. To investigate the underlying transport mechanism, phenomenological arguments are combined with calculations based on density functional theory. The theoretical analysis suggests that the bias dependence of the polarizability of the molecule feeds back into the current leading to an asymmetric shape of the current-voltage characteristics, similar to the phenomena in a semiconductor diode.


Visualization of mouse pancreas architecture using MR microscopy.
Pancreatic diseases, which include diabetes, pancreatitis, and pancreatic cancer, are often difficult to detect and/or stage, contributing to a reduced quality of life and lifespan for patients. Thus, there is need for a technology that can visualize tissue changes in the pancreas, improve understanding of disease progression, and facilitate earlier detection in the human population. Because of low spatial resolution, current clinical magnetic resonance imaging (MRI) at low field strength has yet to fully visualize the exocrine, endocrine, vascular, and stromal components of the pancreas. We used high field strength magnetic resonance microscopy (μMRI) to image mouse pancreas ex vivo without contrast agents at high spatial resolution. We analyzed the resulting high-resolution images using volume rendering to resolve components in the pancreas, including acini, islets, blood vessels, and extracellular matrix. Locations and dimensions of pancreatic components as seen in three-dimensional μMRI were compared with histological images, and good correspondence was found. Future longitudinal studies could expand on the use of in vivo μMRI in mouse models of pancreatic diseases. Capturing three-dimensional structural changes through μMRI could help to identify early cellular and tissue changes associated with pancreatic disease, serving as a mode of improved detection in the clinic for endocrine and exocrine pathologies.


The Skeletal Muscle Anabolic Response to Plant- versus Animal-Based Protein Consumption.
Clinical and consumer market interest is increasingly directed toward the use of plant-based proteins as dietary components aimed at preserving or increasing skeletal muscle mass. However, recent evidence suggests that the ingestion of the plant-based proteins in soy and wheat results in a lower muscle protein synthetic response when compared with several animal-based proteins. The possible lower anabolic properties of plant-based protein sources may be attributed to the lower digestibility of plant-based sources, in addition to greater splanchnic extraction and subsequent urea synthesis of plant protein-derived amino acids compared with animal-based proteins. The latter may be related to the relative lack of specific essential amino acids in plant- as opposed to animal-based proteins. Furthermore, most plant proteins have a relatively low leucine content, which may further reduce their anabolic properties when compared with animal proteins. However, few studies have actually assessed the postprandial muscle protein synthetic response to the ingestion of plant proteins, with soy and wheat protein being the primary sources studied. Despite the proposed lower anabolic properties of plant vs. animal proteins, various strategies may be applied to augment the anabolic properties of plant proteins. These may include the following: 1) fortification of plant-based protein sources with the amino acids methionine, lysine, and/or leucine; 2) selective breeding of plant sources to improve amino acid profiles; 3) consumption of greater amounts of plant-based protein sources; or 4) ingesting multiple protein sources to provide a more balanced amino acid profile. However, the efficacy of such dietary strategies on postprandial muscle protein synthesis remains to be studied. Future research comparing the anabolic properties of a variety of plant-based proteins should define the preferred protein sources to be used in nutritional interventions to support skeletal muscle mass gain or maintenance in both healthy and clinical populations.


Generating in vivo cloning vectors for parallel cloning of large gene clusters by homologous recombination.
A robust method for the in vivo cloning of large gene clusters was developed based on homologous recombination (HR), requiring only the transformation of PCR products into Escherichia coli cells harboring a receiver plasmid. Positive clones were selected by an acquired antibiotic resistance, which was activated by the recruitment of a short ribosome-binding site plus start codon sequence from the PCR products to the upstream position of a silent antibiotic resistance gene in receiver plasmids. This selection was highly stringent and thus the cloning efficiency of the GFPuv gene (size: 0.7 kb) was comparable to that of the conventional restriction-ligation method, reaching up to 4.3 × 10(4) positive clones per μg of DNA. When we attempted parallel cloning of GFPuv fusion genes (size: 2.0 kb) and carotenoid biosynthesis pathway clusters (sizes: 4 kb, 6 kb, and 10 kb), the cloning efficiency was similarly high regardless of the DNA size, demonstrating that this would be useful for the cloning of large DNA sequences carrying multiple open reading frames. However, restriction analyses of the obtained plasmids showed that the selected cells may contain significant amounts of receiver plasmids without the inserts. To minimize the amount of empty plasmid in the positive selections, the sacB gene encoding a levansucrase was introduced as a counter selection marker in receiver plasmid as it converts sucrose to a toxic levan in the E. coli cells. Consequently, this method yielded completely homogeneous plasmids containing the inserts via the direct transformation of PCR products into E. coli cells.


Uncovering cryptic glycan markers in multiple sclerosis (MS) and experimental autoimmune encephalomyelitis (EAE).
Using an integrated antigen microarray approach, we observed epitope-spreading of autoantibody responses to a variety of antigenic structures in the cerebrospinal fluid (CSF) of patients with multiple sclerosis (MS) and in the serum of mice with experimental autoimmune encephalomyelitis (EAE). These included previously described protein- and lipid-based antigenic targets and newly discovered autoimmunogenic sugar moieties, notably, autoantibodies specific for the oligomannoses in both MS patient CSF and the sera of mice with EAE. These glycans are often masked by other sugar moieties and belong to a class of cryptic autoantigens. We further determined that these targets are highly expressed on multiple cell types in MS and EAE lesions. Co-immunization of SJL/J mice with a Man9-KLH conjugate at the time of EAE induction elicited highly significant levels of anti-Man9-cluster autoantibodies. Nevertheless, this anti-glycan autoantibody response was associated with a significantly reduced clinical severity of EAE. The potential of these cryptic glycan markers and targeting antibodies for diagnostic and therapeutic interventions of neurological disorders has yet to be explored.


Bioactive norditerpenoids and neolignans from the roots of salvia miltiorrhiza.
Two new norditerpenoids, miltiolactones A and B (1a, 1b); seven new neolignans, miltiolignanolides A-G (2a, 2b and 3a, 3b and 4a, 4b, and 5) were obtained from the root extract of Salvia miltiorrhiza. Using HPLC separation with a chiral column, compounds 1-4 were found to exist as four pairs of enantiomers. Compounds 2-5 are novel neolignans with a dibenzocycloheptatriene ring system, which form an unprecedented 6/7/6 carbon skeleton. The structures were established via extensive spectroscopic analysis, and experimental and calculated electronic circular dichroism (ECD) spectra, and compound 1 was also elucidated by X-ray diffraction experiments. Compounds 2a and 2b significantly increased the viability of H9c2 cells from H2O2-induced cell death, and 3a showed significant antioxidant activity in an ABTS assay. In addition, compounds 2a, 2b and 3a, 3b display moderate inhibitory effects on nitric oxide (NO) production in LPS-induced microglial cells.


Chest physician-reported, work-related, long-latency respiratory disease in Great Britain.
Much of the current burden of long-latency respiratory disease (LLRD) in Great Britain is attributed to historical asbestos exposure. However, continuing exposure to other agents, notably silica, also contributes to disease burden. The aim of this study was to investigate the incidence of work-related LLRD reported by chest physicians in Great Britain, including variations by age, gender, occupation and suspected agent.LLRD incidence and incidence rate ratios by occupation were estimated (1996-2014). Mesothelioma cases by occupation were compared with proportional mortality ratios.Cases were predominantly in men (95%) and 92% of all cases were attributed to asbestos. Annual average incidence rates (males) per 100 000 were: benign pleural disease, 7.1 (95% CI 6.0-8.2); mesothelioma, 5.4 (4.8-6.0); pneumoconiosis, 1.9 (1.7-2.2); lung cancer, 0.8 (0.6-1.0); chronic obstructive pulmonary disease (COPD), 0.3 (0.2-0.4). Occupations with a particularly high incidence of LLRD were miners and quarrymen (COPD), plumbers and gas fitters (asbestosis), and shipyard and dock workers (all other categories). There was a clear concordance between cases of SWORD mesothelioma and proportional mortality ratios by occupation.Occupationally caused LLRD continues to contribute to a significant disease burden. Many cases are attributable to past exposure to agents such as asbestos and silica, but the potential for occupational exposures persists.


Influence of heel lifts during standing in children with motor disorders.
Heel wedges may influence standing posture but how and to what extent are unknown. Thirty-two children with motor disorders - 16 with arthrogryposis multiplex congenita (AMC) and 16 with cerebral palsy (CP) - and 19 control children underwent a three-dimensional motion analysis. Unassisted standing during 20s with shoes only and with heel lifts of 10, 20 and 30mm heights was recorded in a randomized order. The more weight-bearing limb or the right limb was chosen for analysis. In both the AMC and CP groups, significant changes were seen between various heel lifts in ankle, knee and pelvis, and in the control group in the ankle only. Between orthosis and non-orthosis users significant differences were seen between different heel lift conditions in ankle, knee and trunk in the AMC group and in the ankle in the CP group. Pelvis position changed toward less anterior tilt with increasing heel height, but led to increasing knee flexion in most of the children, except for the AMC Non-Ort group. Children with AMC and CP represent different motor disorders, but the heel wedges had a similar influence on pelvis, hip and knee positions in all children with CP and in the AMC orthosis users. A challenge is to apply heel heights adequate to each individual's orthopaedic and neurologic conditions to improve biomechanical alignment with respect to all body segments.


[A Comparison of Methods for Modeling Factors Associated with Hospital Admissions in Incident Cases of Heart Failure].
Heart failure (HF) is an important public health problem due to its increasing prevalence, and the decompensation associated with hospital admission represents an increased risk of death. The objective of this study was to compare several methods to model the variable hospitalizations and to determine the effect of factors associated with hospital admissions in incident cases of HF. Study of a retrospective cohort of patients with information extracted from electronic medical records of PC was performed. Patients 24 year and older with at least 1 visit to PC in 2006 were included. Registered hospital admissions of HF incident cases between 2006 and 2010 or until death were analyzed and comparison of Poisson, Negative Binomial (NB), zero-inflated and Hurdle regression models were conducted to identify factors associated con hospitalizations. 3,061 patients were identified in a cohort of 227,984. Regarding the factors associated with hospitalizations and according to the zero inflated NB regression model, patients who presented valvular disease (OR=2.01; CI95% 1.22-3.30), or were being treated with antithrombotics (OR=3.45; CI95%: 1.61-7.42) or diuretics (OR=2.28; CI95% 1.13-4.58) had a lower likelihood of hospitalization. Factors associated with a higher rate of hospital admissions were having valvular disease (IRR=1.37; CI95% 1.03-1.81) or diabetes (IRR=1.38; 1.07-1.78), and being treated with calcium antagonists (IRR=1.35; CI95% 1.05- 1.73) or ACE inhibitors (IRR=1.43; CI95% 1.06- 1.92). Having being referred to a cardiologist had a protective effect (IRR=0.86; CI95% 0.76- 0.97). The regression model that obtained the best adjustment was the zero inflated NB. According to this model, the factors associated with an increase in hospital admissions were valvulopathies, diabetes and treatment with calcium antagonists.


Mortality in SARS-CoV-2 Hospitalized Patients Treated with Remdesivir: A Nationwide, Registry-Based Study in Italy.
Remdesivir is the first drug approved for treatment of COVID-19 but current evidence for recommending its use for the treatment of moderate-to-severe disease is still controversial among clinical guidelines. We performed a nationwide, registry-based study including all Italian hospitalized patients with COVID-19 treated with remdesivir to assess the impact of major confounders on crude 15-day and 29-day mortality. Mortality was calculated using the Kaplan-Meier estimator and the Cox proportional-hazards model was applied to analyze the risks by patient's baseline features. In total, 16,462 patients treated with remdesivir from 29 October 2020 to 17 December 2020 were entered in the study. Crude 15-day and 29-day mortality were 7.1% (95% CI, 6.7-7.5%) and 11.7% (95% CI, 11.2-12.2%), respectively. Being treated within two days of admission reduced the risk of death by about 40% (HR 1.4, 95% CI, 1.2-1.6). Results from the largest cohort of remdesivir-treated patients suggests that mortality in SARS-CoV-2 hospitalized patients is substantially influenced by the days between SARS-CoV-2 diagnosis and drug prescription. Current recommendations and future clinical trials for remdesivir alone or in combination should carefully consider the target population and timing for best efficacy of treatment.


Regulators, Pivotal Clinical Trials, and Drug Regulation in the Age of COVID-19.
Medicine regulators rely on pivotal clinical trials to make decisions about approving a new drug, but little is known about how they judge whether pivotal trials justify the approval of new drugs. We explore this issue by looking at the positions of 3 major regulators: the European Medicines Agency, Food and Drug Administration, and Health Canada. Here we report their views and the implications of those views for the approval process. On various points, the 3 regulators are ambiguous, consistent, and demonstrate flexibility. The range of views may well reflect different regulatory cultures. Although clinical trial information from pivotal trials is becoming more available, regulators are still reluctant to provide detailed information about how that information is interpreted. As medicines and vaccines come up for approval for treatment of COVID-19, transparency in how pivotal trials are interpreted will be critical in determining how these treatments should be used.


Spatial relative risk patterns of autism spectrum disorders in Utah.
Heightened areas of spatial relative risk for autism spectrum disorders (ASD), or ASD hotspots, in Utah were identified using adaptive kernel density functions. Children ages four, six, and eight with ASD from multiple birth cohorts were identified by the Utah Registry of Autism and Developmental Disabilities. Each ASD case was gender-matched to 20 birth cohort controls. Demographic and socioeconomic characteristics of children born inside versus outside ASD hotspots were compared. ASD hotspots were found in the surveillance area for all but one birth cohort and age group sample; maximum relative risk in these hotspots ranged from 1.8 to 3.0. Associations were found between higher socioeconomic status and birth residence in an ASD hotspot in five out of six birth cohort and age group samples.


Update on the genetics of the epilepsy-aphasia spectrum and role of GRIN2A mutations.
Formerly idiopathic, focal epilepsies (IFE) are self-limiting, "age-related" diseases that mainly occur during critical developmental periods. Childhood epilepsy with centrotemporal spikes, or Rolandic epilepsy (RE), is the most frequent form of IFE. Together with the Landau-Kleffner syndrome and the epileptic Encephalopathy related to Status Epilepticus during slow Sleep syndrome (ESES), RE is part of a single and continuous spectrum of childhood epilepsies and epileptic encephalopathies with acquired cognitive, behavioral and speech and/or language impairment, known as the epilepsy-aphasia spectrum (EAS). The pathophysiology has long been attributed to an elusive and complex interplay between brain development and maturation processes on the one hand, and susceptibility genes on the other hand. Studies based on the variable combination of molecular cytogenetics, Sanger and next-generation sequencing tools, and functional assays have led to the identification and validation of genetic mutations in the GRIN2A gene that can directly cause various types of EAS disorders. The recent identification of GRIN2A defects in EAS represents a first and major break-through in our understanding of the underlying pathophysiological mechanisms. In this review, we describe the current knowledge on the genetic architecture of IFE.


Synergy between the C2 allele of transferrin and the C282Y allele of the haemochromatosis gene (HFE) as risk factors for developing Alzheimer's disease.
There is evidence that iron may play a role in the pathology of Alzheimer's disease (AD). There may be genetic factors that contribute to iron deposition resulting in tissue damage thus exacerbating AD. We have genotyped 269 healthy elderly controls, 191 cases with definite or probable AD, and 69 with mild cognitive impairment (MCI) from the OPTIMA cohort. We have examined the interaction between the C2 variant of the transferrin (TF) gene and the C282Y allele of the haemochromatosis (HFE) gene as risk factors for developing AD. Our results showed that each of the two variants was associated with an increased risk of AD only in the presence of the other. Neither allele alone had any effect. Carriers of both variants were at 5 times greater risk of AD compared with all others. The interaction was significant by logistic regression (p = 0.014) and by synergy factor analysis (p = 0.015, synergy factor = 5.1). Further, carriers of these two alleles plus apolipoprotein E epsilon4 (APOE4) were at still higher risk of AD: of the 14 tri-carriers of the three variants, identified in this study, 12 had AD and two MCI. We suggest that the combination of TF C2 and HFE C282Y may lead to an excess of redox-active iron and the induction of oxidative stress in neurones, which is exacerbated in carriers of APOE4. Since 4% of Northern Europeans carry the two iron-related variants and since iron overload is a treatable condition, these results merit replication.


Transcription Factor Activity Mapping of a Tissue-Specific in vivo Gene Regulatory Network.
A wealth of physical interaction data between transcription factors (TFs) and DNA has been generated, but these interactions often do not have apparent regulatory consequences. Thus, equating physical interaction data with gene regulatory networks (GRNs) is problematic. Here, we comprehensively assay TF activity, rather than binding, to construct a network of gene regulatory interactions in the C. elegans intestine. By manually observing the in vivo tissue-specific knockdown of 921 TFs on a panel of 19 fluorescent transcriptional reporters, we identified a GRN of 411 interactions between 19 promoters and 177 TFs. This GRN shows only modest overlap with physical interactions, indicating that many regulatory interactions are indirect. We applied nested effects modeling to uncover information flow between TFs in the intestine that converges on a small set of physical TF-promoter interactions. We found numerous cell nonautonomous regulatory interactions, illustrating tissue-to-tissue communication. Altogether, our study illuminates the complexity of gene regulation in the context of a living animal.


Protective effect of an acute oral loading dose of trimetazidine on myocardial injury following percutaneous coronary intervention.
To evaluate the effect of pre-procedural acute oral administration of trimetazidine (TMZ) on percutaneous coronary intervention (PCI)-induced myocardial injury. Single-centre, prospective, randomised evaluation study. Patients with stable angina pectoris and single-vessel disease undergoing PCI. 582 patients were prospectively randomised. Patients who underwent more than one inflation during PCI were excluded, resulting in 266 patients randomly assigned to 2 groups. Patients were randomly assigned to receive or not an acute loading dose of 60 mg of TMZ prior to intervention. The frequency and the increase in the level of cardiac troponin Ic (cTnI) after successful PCI. cTnI levels were measured before and 6, 12, 18 and 24 h after PCI. 136 patients were assigned to the TMZ group and 130 to the control group. Although no statistically significant difference was observed in the frequency of cTnI increase between the two groups, post-procedural cTnI levels were significantly reduced in the TMZ group at all time points (6 h: mean (SD) 4.2 (0.8) vs 1.7 (0.2), p<0.001; 12 h: 5.5 (1.5) vs 2.3 (0.4), p<0.001; 18 h: 9 (2.3) vs 3 (0.5), p<0.001; and 24 h: 3.2 (1.2) vs 1 (0.5), p<0.001). Moreover, the total amount of cTnI released after PCI, as assessed by the area under the curve of serial measurement, was significantly reduced in the TMZ group (p<0.05). Pre-procedural acute oral TMZ administration significantly reduces PCI-induced myocardial infarction.


Implementation of a Stepwise, Multidisciplinary Intervention for Pain and Challenging Behaviour in Dementia (STA OP!): A Process Evaluation.
A stepwise, multidisciplinary and multicomponent intervention (called STA OP!) was implemented in Dutch nursing home units, which included a comprehensive multidisciplinary team training. A cluster-randomised controlled trial showed that the intervention reduced symptoms of pain and challenging behaviour. To describe the experiences around the implementation of the intervention; to examine the extent to which the STA OP! intervention was delivered and implemented as intended (at the level of the team, and the individual resident/professional); and to understand factors influencing the implementation process. A process evaluation was performed using a mixed-methods design encompassing several data sources. Quantitative data (i.e. from the written evaluations by healthcare professionals, management, and the research database) were analysed using descriptive statistics. Qualitative data (i.e. semi-structured interviews, notes, completed intervention forms, and written evaluations) were analysed according to the principles of thematic analysis. The implementation process and the influencing factors were categorised according to the i) organisational level, ii) the team level, and iii) the level of the individual resident/professional. In total, 39.2% of the residents with pain and/or challenging behaviour were treated following the stepwise approach of the STA OP! intervention. The training manual and forms used were found to be relevant and feasible. Factors inhibiting the implementation process at the i) organisational level concerned instability of the organisation and the team (e.g. involvement in multiple projects/new innovations, staff turnover/absence of essential disciplines, and/or high workload). At the team level (ii), we found that presence of a person with a motivational leadership style facilitated the implementation. Also, interdisciplinary cooperation through the design/setting of the multidisciplinary training, securing the intervention by use of clear agreements, and written reporting or transfers facilitated implementation. At the individual level (iii), perceived value of the stepwise working method, and enhanced awareness facilitated the implementation. Although the intervention was not implemented as planned, the intervention empowered healthcare professionals and increased their awareness of the signals of pain and challenging behaviour. Future implementation of the intervention should start on units with a motivational leader, and specific features of the organisation and the team should be considered to facilitate implementation, e.g. stability, support, and shared focus to change.


Sociodemographic characteristics, clinical signs and quality of life in patients with fibromyalgia.
We aimed to evaluate the sociodemographic and clinical characteristics as well as the quality of life of patients with fibromyalgia (FM). A total of 37 female patients diagnosed with FM and 31 healthy females were included into the study. Individuals were asked about their demographic characteristics. The number of sensitive points (NSP), skin fold sensitivity, cutaneous hyperemia and reticular skin changes of patients and healthy controls were evaluated during physical examination. Individuals were evaluated for pain severity using the Visual Analogue Scale (VAS), for psychological states using the Beck Depression Scale (BDS), for quality of life using the Short Form-36 (SF-36) and for functional status using the Fibromyalgia Impact Questionnaire (FIQ). The mean age of FM patients was 39.2±6.5 years versus 39.1±6.2 years in the controls. The most frequently encountered FM symptoms were fatigue (94.6%), sleep disturbances (86.5%) and anxiety (86.5%). Differences between FM patients and controls were statistically significant for NSP (p=0.001), VAS (p=0.001), FIQ (p=0.001), BDS (p=0.001) and SF-36 (p=0.003), and FIQ subgroups were also different between the two groups (p<0.001). In the SF-36 survey, FM patients were different from the control in the physical function (p=0.001), pain (p=0.005), general health (p=0.017), physical fitness (p=0.003), and mental health (p=0.008) portions of the survey. Fibromyalgia has distinct clinical features that lead to low functional capacity and quality of life. Thus, patients' sociodemographic characteristics and the evaluation of their quality of life may be important in the diagnosis and monitoring of treatment progress.


Comparative histomorphological review of rat and human hepatocellular proliferative lesions.
In this comparative review, histomorphological features of common nonneoplastic and neoplastic hepatocyte lesions of rats and humans are examined using H&E-stained slides. The morphological similarities and differences of both neoplastic (hepatocellular carcinoma and hepatocellular adenoma) and presumptive preneoplastic lesions (large and small cell change in humans and foci of cellular alteration in rats) are presented and discussed. There are major similarities in the diagnostic features, growth patterns and behavior of both rat and human hepatocellular proliferative lesions and in the process of hepatocarcinogenesis. Further study of presumptive preneoplastic lesions in humans and rats should help to further define their role in progression to hepatocellular neoplasia in both species.


The DeepTree Exhibit: Visualizing the Tree of Life to Facilitate Informal Learning.
In this paper, we present the DeepTree exhibit, a multi-user, multi-touch interactive visualization of the Tree of Life. We developed DeepTree to facilitate collaborative learning of evolutionary concepts. We will describe an iterative process in which a team of computer scientists, learning scientists, biologists, and museum curators worked together throughout design, development, and evaluation. We present the importance of designing the interactions and the visualization hand-in-hand in order to facilitate active learning. The outcome of this process is a fractal-based tree layout that reduces visual complexity while being able to capture all life on earth; a custom rendering and navigation engine that prioritizes visual appeal and smooth fly-through; and a multi-user interface that encourages collaborative exploration while offering guided discovery. We present an evaluation showing that the large dataset encouraged free exploration, triggers emotional responses, and facilitates visitor engagement and informal learning.