Pathogenicity of cationized albumin in the dog: renal and extrarenal effects.
The effects of 21 cationized serum albumin samples of various degrees of cationization on renal function were studied in the dog. The samples were perfused intra- aortically to obtain preferential perfusion of the left kidney in 25 dogs. Standard clearance techniques were used, associated in six dogs with sieving studies of 125I-labelled polyvinylpyrrolidone (125I-PVP) and with an extensive morphological study in 15 dogs. Renal effects were observed. (a) Renal effects in left kidneys. The perfusion with weakly cationized albumin (group 1) produced moderate proteinuria associated with the deposition of cationized albumin on the anionic sites of the basement membrane. Glomerular filtration rate (GFR) was unaltered. Perfusion with highly cationized samples (group 2) produced more severe proteinuria and a significant decrease in GFR. Glomerular permeability to 125I-PVP increased. Perfusion with the four samples of highest pI (group 3) was followed by anuria. (b) Renal effects in right kidneys. A retarded mild proteinuria appeared only in group 2 and group 3 animals without alteration of GFR. All the kidneys (group 1 included), with the exception of two (group 3), showed deposition of the protein in the anionic sites. The following extrarenal effects were observed essentially in group 2 and group 3 animals: erythrocyte agglutination and haemolysis, platelet aggregation and thrombocytopenia, and a decrease in plasma fibrogen level due to fibrinogen precipitation. These effects produced progressive obstruction in the glomerular capillaries, thus explaining the occurrence of anuria. The structural damage in group 2 and group 3 left kidneys bears remarkable resemblance to that observed in the fulminant form of the human so-called 'haemolytic-uraemic syndrome'. The neutralization alone of the fixed negative charges in the glomerular wall appears to produce only mild proteinuria, whereas the various extrarenal effects combine to produce more severe proteinuria associated with functional alteration and vascular obstruction.


Azadioxatriangulenium (ADOTA+): A long fluorescence lifetime fluorophore for large biomolecule binding assay.
Of the many optical bioassays available, sensing by fluorescence anisotropy have great advantages as it provides a sensitive, instrumentally simple, ratiometric method of detection. However, it is hampered by a severe limitation as the emission lifetime of the label needs to be comparable to the correlation lifetime (tumbling time) of the biomolecule which is labelled. For proteins of moderate size this is in the order of 20-200 ns, which due to practical issues currently limits the choice of labels to the dansyl-type dyes and certain aromatics dyes. These have the significant drawback of UV/blue absorption and emission as well as an often significant solvent sensitivity. Here, we report the synthesis and characterization of a new fluorescent label for high molecular weight biomolecules assay based on the azadioxatriangulenium motif. The NHS ester of the long fluorescence lifetime, red emitting fluorophore: azadioxatriangulenium (ADOTA-NHS) was conjugated to anti-rabbit Immunoglobulin G (antiIgG). The long fluorescence lifetime was exploited to determine the correlation time of the high molecular weight antibody and its complex with rabbit Immuniglobulin G (IgG) with steady-state fluorescence anisotropy and time-resolved methods: solution phase immuno-assay was performed following either steady-state or time-resolved fluorescence anisotropy. By performing a variable temperature experiment it was determined that the binding of the ligand resulted in an increase in correlation time by more than 75 %, and a change in the steady-state anisotropy increase of 18%. The results show that the triangulenium class of dyes can be used in anisotropy assay for detecting binding events involving biomolecules of far larger size than what is possible with the other red emitting organic dyes.


Interaction between Burkholderia pseudomallei and Acanthamoeba species results in coiling phagocytosis, endamebic bacterial survival, and escape.
Burkholderia pseudomallei causes melioidosis, a potentially fatal disease whose clinical outcomes include rapid-onset septicemia and relapsing and delayed-onset infections. Like other facultative intracellular bacterial pathogens, B. pseudomallei is capable of survival in human phagocytic cells, but unlike mycobacteria, Listeria monocytogenes, and Salmonella serovar Typhimurium, the species has not been reported to survive as an endosymbiont in free-living amebae. We investigated the consequences of exposing Acanthamoeba astronyxis, A. castellani, and A. polyphaga to B. pseudomallei NCTC 10276 in a series of coculture experiments. Bacterial endocytosis was observed in all three Acanthamoeba species. A more extensive range of cellular interactions including bacterial adhesion, incorporation into amebic vacuoles, and separation was observed with A. astronyxis in timed coculture experiments. Amebic trophozoites containing motile intravacuolar bacilli were found throughout 72 h of coculture. Confocal microscopy was used to confirm the intracellular location of endamebic B. pseudomallei cells. Transmission electron microscopy of coculture preparations revealed clusters of intact bacilli in membrane-lined vesicles inside the trophozoite cytoplasm; 5 x 10(2) CFU of bacteria per ml were recovered from lysed amebic trophozoites after 60 min of coculture. Demonstration of an interaction between B. pseudomallei and free-living acanthamebae in vitro raises the possibility that a similar interaction in vivo might affect environmental survival of B. pseudomallei and subsequent human exposure. Endamebic passage of B. pseudomallei warrants further investigation as a potential in vitro model of intracellular B. pseudomallei infection.


Mental health expenditures for services for people with severe mental illnesses.
To provide comprehensive information on expenditures for mental health and substance abuse services for a large number of people with severe mental illnesses, this study examined use of major types of clinical-medical mental health and psychiatric rehabilitation services over a one-year period. Data were obtained for 1,890 clients in ten public county-based nonmetropolitan mental health systems in Wisconsin. Expenditures were for services provided with public funding, including local sources of funding, Medicaid, and Medicare. Data about services and expenditures were obtained from county records and unduplicated Medicaid claims for 12 months in 1989 and 1990. Expenditures per client averaged $10,995 for one year ($13,992 in 1994 dollars), with a maximum of $95,093. Expenditures for community-based outpatient services, including residential care and vocational services, represented 53.5 percent of all expenditures; residential care accounted for 12.4 percent and vocational services for 5.7 percent. Overall, 46.5 percent was spent for institutional care, with inpatient hospital care accounting for 12.6 percent. Approximately 40.6 percent of total expenditures were for services not typically covered under managed care plans. Expenditures for community-based care accounted for more than half of total expenditures. Expenditure patterns revealed the important role of social and rehabilitation services, a role that must be continued in managed care arrangements if they are to provide adequate services for people with severe mental illnesses.


Early dyadic patterns of mother-infant interactions and outcomes of prematurity at 18 months.
With the increased survival of very preterm infants, there is a growing concern for their developmental and socioemotional outcomes. The quality of the early mother-infant relationship has been noted as 1 of the factors that may exacerbate or soften the potentially adverse impact of preterm birth, particularly concerning the infant's later competencies and development. The first purpose of the study was to identify at 6 months of corrected age whether there were specific dyadic mother-infant patterns of interaction in preterm as compared with term mother-infant dyads. The second purpose was to examine the potential impact of these dyadic patterns on the infant's behavioral and developmental outcomes at 18 months of corrected age. During a 12-month period (January-December 1998), all preterm infants who were <34 weeks of gestational age and hospitalized at the NICU of the Lausanne University Hospital were considered for inclusion in this longitudinal prospective follow-up study. Control healthy term infants were recruited during the same period from the maternity ward of our hospital. Mother-infant dyads with preterm infants (n = 47) and term infants (n = 25) were assessed at 6 months of corrected age during a mother-infant play interaction and coded according to the Care Index. This instrument evaluates the mother's interactional behavior according to 3 scales (sensitivity, control, and unresponsiveness) and the child's interactional behavior according to 4 scales (cooperation, compliance, difficult, and passivity). At 18 months, behavioral outcomes of the children were assessed on the basis of a semistructured interview of the mother, the Symptom Check List. The Symptom Check List explores 4 groups of behavioral symptoms: sleeping problems, eating problems, psychosomatic symptoms, and behavioral and emotional disorders. At the same age, developmental outcomes were evaluated using the Griffiths Developmental Scales. Five areas were evaluated: locomotor, personal-social, hearing and speech, eye-hand coordination, and performance. Among the possible dyadic patterns of interaction, 2 patterns emerge recurrently in mother-infant preterm dyads: a "cooperative pattern" with a sensitive mother and a cooperative-responsive infant (28%) and a "controlling pattern" with a controlling mother and a compulsive-compliant infant (28%). The remaining 44% form a heterogeneous group that gathers all of the other preterm dyads and is composed of 1 sensitive mother-passive infant; 10 controlling mothers with a cooperative, difficult, or passive infant; and 10 unresponsive mothers with a cooperative, difficult, or passive infant. Among the term control subjects, 68% of the dyads are categorized as cooperative pattern dyads, 12% as controlling pattern dyads, and the 20% remaining as heterogeneous dyads. At 18 months, preterm infants of cooperative pattern dyads have similar outcomes as the term control infants. Preterm infants of controlling pattern dyads have significantly fewer positive outcomes as compared with preterm infants of cooperative pattern dyads, as well as compared with term control infants. They display significantly more behavioral symptoms than term infants, including more eating problems than term infants as well as infants from cooperative preterm dyads. Infants of the controlling preterm dyads do not differ significantly for the total development quotient but have worse personal-social development than term infants and worse hearing-speech development than infants from cooperative preterm dyads. The preterm infants of the heterogeneous group have outcomes that can be considered as intermediate with no significant differences compared with preterm infants from the cooperative pattern or the controlling pattern dyads. Among mother-preterm infant dyads, we identified 2 specific patterns of interaction that could play either a protective (cooperative pattern) or a risk-precipitating (controlling pattern) role on developmental and behavioral outcome, independent of perinatal risk factors and of the family's socioeconomic background. The controlling pattern is much more prevalent among preterm than term dyads and is related to a less favorable infant outcome. However, the cooperative pattern still represents almost 30% of the preterm dyads, with infants' outcome comparable to the ones of term infants. These results point out the impact of the quality of mother-infant relationship on the infant's outcome. The most important clinical implication should be to support a healthy parent-infant relationship already in the NICU but also in the first months of the infant's life. Early individualized family-based interventions during neonatal hospitalization and transition to home have been shown to reduce maternal stress and depression and increase maternal self-esteem and to improve positive early parent-preterm infant interactions.


Impact of the translocational equilibrium of HIV-1 reverse transcriptase on the efficiency of mismatch extensions and the excision of mispaired nucleotides.
The reverse transcriptase of the human immunodeficiency virus type 1 (HIV-1 RT) does not possess an exonucleolytic proofreading activity; however, previous studies have shown that this enzyme can excise incorporated chain-terminators in the presence of pyrophosphate or ATP. This type of reaction provides a plausible mechanism for HIV-1 resistance to several nucleoside analogue inhibitors. Here we studied the efficiency of pyrophosphorolysis in the context of mismatched nucleotides, and found that the removal of dCMP and dTMP opposite T is literally blocked. Thus, pyrophosphorolysis may not provide an alternative, universal proofreading mechanism, although excision of dGMP and the correct dAMP opposite T can occur with considerable efficiency. Site-specific footprinting experiments revealed that the 3' end of C:T- and T:T-mispaired primer strands is predominantly found in a post-translocational configuration, which prevents the removal of terminal nucleotides. In contrast, complexes containing G:T and A:T base pairs can exist in both post- and pre-translocational stages. Excision can only occur in the latter, which helps to explain the observed selectivity of the reaction. The efficiency of mismatch extensions does not appear to depend on pre-existing changes of the translocational equilibrium. However, footprints of complexes containing 3' penultimate mismatches suggest that the incorporation of the first nucleotide following the mispair can force the enzyme to slide backwards, which can inhibit ensuing polymerization events. The fact that misincorporated nucleotides can affect the precise positioning of RT provides a rational for the development of novel nucleoside analogue inhibitors that contain modifications in the base moiety.


Correlates of anxiety symptoms in physically disabled older women.
The authors describe characteristics that are associated with chronic anxiety symptoms and examine the use of anxiolytic and antidepressant medications in physically disabled women with and without symptoms of anxiety. Participants were 791 physically disabled women age 65+ years who participated in the Women's Health and Aging Study for 2-3 years. Anxiety symptoms were measured with four questions from the Hopkins Symptom Checklist, and women were categorized as having no anxiety, intermittent anxiety, and chronic anxiety symptoms. Health-related characteristics, medications, physical functioning, physical activity, and psychosocial variables were also measured. Forty-nine percent of women reported no anxiety symptoms; 41% reported intermittent symptoms; and 10% reported chronic symptoms of anxiety. Depressive symptoms and lack of emotional support were significant correlates of intermittent anxiety symptoms, whereas depressive symptoms, negative life events, and lack of emotional support were significant correlates of chronic anxiety symptoms. Over the course of 3 years, 20.3% of women with no anxiety, 33.0% of women with intermittent anxiety, and 48.7% of women with chronic anxiety symptoms took anxiolytic and/or antidepressant medications. Anxiety symptoms are common among disabled older women. Psychosocial variables were significantly different in women with intermittent or chronic anxiety symptoms, versus women without anxiety.


High isoniazid resistance rates in rifampicin susceptible Mycobacterium tuberculosis pulmonary isolates from Pakistan.
Rapid new diagnostic methods (including Xpert MTB/RIF assay) use rifampicin resistance as a surrogate marker for multidrug resistant tuberculosis. Patients infected with rifampicin susceptible strains are prescribed first line anti-tuberculosis therapy. The roll out of such methods raises a concern that strains with resistance to other first line anti-tuberculosis drugs including isoniazid will be missed and inappropriate treatment given. To evaluate implications of using such methods review of resistance data from high burden settings such as ours is essential. To determine resistance to first line anti-tuberculosis drugs amongst rifampicin susceptible pulmonary Mycobacterium tuberculosis (MTB) isolates from Pakistan. Data of pulmonary Mycobacterium tuberculosis strains isolated in Aga Khan University Hospital (AKUH) laboratory (2009-2011) was retrospectively analyzed. Antimicrobial susceptibility profile of rifampicin susceptible isolates was evaluated for resistance to isoniazid, pyrazinamide, ethambutol, and streptomycin. Pulmonary specimens submitted to AKUH from 2009 to 2011 yielded 7738 strains of Mycobacterium tuberculosis. These included 54% (n 4183) rifampicin susceptible and 46% (n: 3555) rifampicin resistant strains. Analysis of rifampicin susceptible strains showed resistance to at least one of the first line drugs in 27% (n:1133) of isolates. Overall isoniazid resistance was 15.5% (n: 649), with an isoniazid mono-resistance rate of 4% (n: 174). Combined resistance to isoniazid, pyrazinamide, and ethambutol was noted in 1% (n: 40), while resistance to isoniazid, pyrazinamide, ethambutol, and streptomycin was observed in 1.7% (n: 70) of strains. Our data suggests that techniques (including Xpert MTB/RIF assay) relying on rifampicin susceptibility as an indicator for initiating first line therapy will not detect patients infected with MTB strains resistant to other first line drugs (including isoniazid). The roll out of these techniques must therefore be accompanied by strict monitoring ensuring early resistance detection to increase chances of improved patient outcomes.


What can we learn about patient safety from information sources within an acute hospital: a step on the ladder of integrated risk management?
To assess the utility of data already existing within hospitals for monitoring patient safety. An acute hospital in southern England. Mapping of data sources proposed by staff as potentially able to identify patient safety issues followed by an in-depth analysis of the content of seven key sources. Data source analysis: For each data source: scope and depth of content in relation to patient safety, number and type of patient safety incidents identified, degree of overlap with incidents identified by different sources, levels of patient harm associated with incidents. A wide range of data sources existing within the hospital setting have the potential to provide information about patient safety incidents. Poor quality of coding, delays in reports reaching databases, the narrow focus of some data sources, limited data-collection periods and lack of central collation of findings were some of the barriers to making the best use of routine data sources for monitoring patient safety. An in-depth analysis of seven key data sources (Clinical Incident database, Health and Safety Incident database, Complaints database, Claims database and Inquest database, the Patient Administration System and case notes) indicated that case notes have the potential to identify the largest number of incidents and provide the richest source of information on such incidents. The seven data sources identified different types of incidents with differing levels of patient harm. There was little overlap between the incidents identified by different sources. Despite issues related to the quality of coding, depth of information available and accessibility, triangulating information from more than one source can identify a broader range of incidents and provide additional information related to professional groups involved, types of patients affected and important contributory factors. Such an approach can provide a focus for further work and ultimately contributes to the identification of appropriate interventions that improve patient safety.


Readability and content of patient education material related to implantable cardioverter defibrillators.
Implantable cardioverter defibrillators (ICDs) are increasingly offered to patients for primary prevention of sudden cardiac death. Candidates for ICD receive ICD-related patient education material when they make decisions to consent or decline a primary prevention ICD. Printed patient education material directed at ICD candidates has not been the focus of direct appraisal. We evaluated the readability and content of ICD-related print education materials made available to patients who were enrolled in a study involving patient decision making for ICD from 3 ICD sites in southern Ontario, Canada. All ICD print materials referred to during interviews and/or that were available in ICD site waiting rooms were collected for analysis. Readability testing was conducted using the "simple measurement of gobbledygook" and Fry methods. The material was evaluated according to selected plain-language criteria, thematic content analysis, and rhetoric analysis. Twenty-one print materials were identified and analyzed. Documents were authored by device manufacturers, tertiary care hospitals, and cardiac support organizations. Although many documents adhered to plain-language recommendations, text-reading levels were higher than recommended. Twelve major content themes were identified. Content focused heavily on the positive aspects of living with the device to the exclusion of other possible information that could be relevant to the decisions that patients made. Print-based patient education materials for ICD candidates are geared to a highly literate population. The focus on positive information to the exclusion of potentially negative aspects of the ICD, or alternatives to accepting 1, could influence and/or confuse patients about the purpose and implications of this medical device. Development of print materials is indicated that includes information about possible problems and that would be relevant for the multicultural and debilitated population who may require ICDs. The findings are highly relevant for nurses who care for primary prevention ICD candidates.


Genetic differentiation, structure, and a transition zone among populations of the pitcher plant moth Exyra semicrocea: implications for conservation.
Pitcher plant bogs, or carnivorous plant wetlands, have experienced extensive habitat loss and fragmentation throughout the southeastern United States Coastal Plain, resulting in an estimated reduction to <3% of their former range. This situation has lead to increased management attention of these habitats and their carnivorous plant species. However, conservation priorities focus primarily on the plants since little information currently exists on other community members, such as their endemic arthropod biota. Here, we investigated the population structure of one of these, the obligate pitcher plant moth Exyra semicrocea (Lepidoptera: Noctuidae), using mitochondrial cytochrome c oxidase subunit I (COI) gene sequences. Examination of 221 individuals from 11 populations across eight southeastern US states identified 51 unique haplotypes. These haplotypes belonged to one of two divergent (∼1.9-3.0%) lineages separated by the Mississippi alluvial plain. Populations of the West Gulf Coastal Plain exhibited significant genetic structure, contrasting with similarly distanced populations east of the Mississippi alluvial plain. In the eastern portion of the Coastal Plain, an apparent transition zone exists between two regionally distinct population groups, with a well-established genetic discontinuity for other organisms coinciding with this zone. The structure of E. semicrocea appears to have been influenced by patchy pitcher plant bog habitats in the West Gulf Coastal Plain as well as impacts of Pleistocene interglacials on the Apalachicola-Chattahoochee-Flint River Basin. These findings, along with potential extirpation of E. semicrocea at four visited, but isolated, sites highlight the need to consider other endemic or associated community members when managing and restoring pitcher plant bog habitats.


Chromatin remodeling mediated by the FOXA1/A2 transcription factors activates CFTR expression in intestinal epithelial cells.
The forkhead box A transcription factors, FOXA1 and FOXA2, function as pioneer factors to open condensed chromatin and facilitate binding of other proteins. We showed previously that these factors are key components of a transcriptional network that drives enhancer function at the cystic fibrosis transmembrane conductance regulator (CFTR) locus in intestinal epithelial cells. The CFTR promoter apparently lacks tissue-specific regulatory elements and expression of the gene is controlled by multiple cis-acting elements, which coordinate gene expression in different cell types. Here we show that concurrent depletion of FOXA1 and FOXA2 represses CFTR expression and alters the three-dimensional architecture of the active locus by diminishing interactions between the promoter and intronic cis-acting elements. Reduction of FOXA1/A2 also modifies the enrichment profile of the active enhancer marks H3K27ac and H3K4me2 across the CFTR locus and alters chromatin accessibility at individual cis-elements. Moreover, loss of FOXA1/A2 suppresses the recruitment of other members of the transcriptional network including HNF1 and CDX2, to multiple cis-elements. These data reveal a complex molecular mechanism underlying the role of FOXA1/A2 in achieving high levels of CFTR expression in intestinal epithelial cells.


Examination of sleep health dimensions and their associations with perceived stress and health in a UK sample.
Sleep health is a relatively new multidimensional concept, however, there is no consensus on its underlying dimensions. A previous study examined potential indicators of sleep health using an aggregated sleep health measure. However, the psychometric properties are yet to be determined. The primary aim was to assess the factor structure, reliability and validity of this measure. A secondary aim was to explore the relationships with perceived stress, and physical and mental health. A cross-sectional online survey was conducted with 257 adults from the UK aged 18-65 (78.4% female, mean age = 29.39 [SD = 11.37]). Participants completed 13 Sleep health items, the Pittsburgh Sleep Quality Inventory, Insomnia Severity Scale, Epworth Sleepiness Scale, Perceived Stress Scale and SF-12 Health Survey. The measure exhibited good internal consistency (α = 0.785) and construct validity as determined by associations with existing measures. Principle components analysis produced four factors e; sleep quality (α = 0.818), sleep adaptability (α = 0.917), sleep wellness (α = 0.621) and daytime functioning (α = 0.582). Adaptability (β = -241) was strongest predictor of perceived stress, and daytime functioning was strongest predictor of physical (β = 0.322) and mental health (β = 0.312). Sleep health is a multidimensional construct comprising four distinct but related dimensions. The importance of sleep health in terms of perceived stress and mental and physical health is highlighted.


Human cytomegalovirus infection of caco-2 cells occurs at the basolateral membrane and is differentiation state dependent.
Epithelial cells are known to be a major target for human cytomegalovirus (HCMV) infection; however, the analysis of virus-cell interactions has been difficult to approach due to the lack of in vitro models. In this study, we established a polarized epithelial cell model using a colon epithelial cell-derived cell line (Caco-2) that is susceptible to HCMV infection at early stages of cellular differentiation. Infection of polarized cells was restricted to the basolateral surface whereas virus was released apically, which was consistent with the apical and not basolateral surface localization of two essential viral glycoproteins, gB and gH. HCMV infection resulted in the development of a cytopathology characteristic of HCMV infection of colon epithelium in vivo, and infection did not spread from cell to cell. The inability of HCMV to infect Caco-2 cells at late stages of differentiation was due to a restriction at the level of viral entry and was consistent with the sequestration of a cellular receptor for HCMV. These observations provide the first evidence that restriction of HCMV replication in epithelial cells is due to a receptor-mediated phenomenon.


Development of a Clinician-Rated Drop Vertical Jump Scale for Patients Undergoing Rehabilitation After Anterior Cruciate Ligament Reconstruction: A Delphi Approach.
Study Design Delphi panel study. Background Biomechanical parameters measured during a drop vertical jump task are risk factors for anterior cruciate ligament (ACL) injury and are targeted during rehabilitation after ACL reconstruction. A clinically feasible tool that quantifies observed performance on the drop vertical jump would help inform treatment efforts. The content and scoring of such a tool should be deliberated on by a group of experts throughout its development. Objectives To establish consensus on the content and scoring of a clinician-rated drop vertical jump scale (DVJS) for use during rehabilitation after ACL reconstruction. Methods Using a modified Delphi process, a panel of experts (researchers and clinicians) on the risk factors, prevention, treatment, and biomechanics of ACL injury anonymously critiqued versions of a DVJS. The DVJS was developed iteratively, based on the feedback from the panel, using Likert scale responses to questions and providing written comments. Three to 5 rounds were planned a priori, with a requirement of 75% agreement on included items after the final round. Results Twenty of the 31 invited experts (65%) participated. Approximately 93% agreement was achieved after the fourth round. Final items on the scale included the rating of knee valgus collapse (no collapse to extreme collapse) and the presence of other undesirable movements, including lateral trunk lean, insufficient knee flexion, and limb-to-limb asymmetry. Conclusion The Delphi process resulted in a beta version of a DVJS. Expert consensus was achieved on its content and scoring to support further clinical testing of the scale. J Orthop Sports Phys Ther 2017;47(8):557-564. Epub 6 Jul 2017. doi:10.2519/jospt.2017.7183.


Laparoscopic versus open repair of perforated peptic ulcer: Improving outcomes utilizing a standardized technique.
The objective of this study was to compare the outcomes of patients who underwent laparoscopic and open repair of perforated peptic ulcers (PPUs) at our institution. This is a retrospective review of a prospectively collected database of patients who underwent emergency laparoscopic or open repair for PPU between December 2010 and February 2014. A total of 131 patients underwent emergency repair for PPU (laparoscopic repair, n=63, 48.1% vs. open repair, n=68, 51.9%). There were no significant differences in baseline characteristics between both groups in terms of age (p=0.434), gender (p=0.305), body mass index (p=0.180), and presence of comorbidities (p=0.214). Both groups were also comparable in their American Society of Anesthesiologists (ASA) scores (p=0.769), Boey scores 0/1 (p=0.311), Mannheim Peritonitis Index > 27 (p=0.528), shock on admission (p<0.99), and the duration of symptoms > 24 hours (p=0.857). There was no significant difference in the operating time between the two groups (p=0.618). Overall, the laparoscopic group had fewer complications compared with the open group (14.3% vs. 36.8%, p=0.005). When reviewing specific complications, only the incidence of surgical site infection was statistically significant (laparoscopic 0.0% vs. open 13.2%, p=0.003). The other parameters were not statistically significant. The laparoscopic group did have a significantly shorter mean postoperative stay (p=0.008) and lower pain scores in the immediate postoperative period (p<0.05). Mortality was similar in both groups (open, 1.6% vs. laparoscopic, 2.9%, p < 0.99). Laparoscopic repair resulted in reduced wound infection rates, shorter hospitalization, and reduced postoperative pain. Our single institution series and standardized technique demonstrated lower morbidity rates in the laparoscopic group.


Effects of three low-volume, high-intensity exercise conditions on affective valence.
A common barrier to exercise is "lack of time". Accordingly, interest in low-volume, high-intensity training has grown exponentially since this activity is considered time-efficient. However, the high-intensity nature of this exercise may frequently result in feelings of displeasure creating another barrier for many people. The purpose of this study was to compare affective (pleasure-displeasure) responses to three low-volume, high-intensity exercise conditions, including a novel shortened-sprint protocol. Using a within-subjects, randomised crossover experiment, healthy participants (N = 36) undertook a single bout of: 1) traditional reduced-exertion, high-intensity interval training (TREHIT), 2) a novel, shortened-sprint REHIT (SSREHIT) protocol, and 3) sprint continuous training (SCT). Affect and perceived efforts were recorded throughout exercise using the Feeling Scale (FS) and the 15-point Borg Rating of Perceived Exertion (RPE) scale, respectively. Enjoyment was recorded 5 min post-exercise using the Exercise Enjoyment Scale (EES). Differences were found for FS (condition by time interaction: P = 0.01GG, η2 = 0.26), RPE (P = 0.01GG, η2 = 0.23), and enjoyment (P < 0.01) with all outcomes favouring SSREHIT. Shortened-sprint protocols may diminish feelings of displeasure and might be a time-efficient yet tolerable exercise choice to help motivate some people to increase their physical activity and fitness.


Chronic condition combinations and health care expenditures and out-of-pocket spending burden among adults, Medical Expenditure Panel Survey, 2009 and 2011.
Little is known about how combinations of chronic conditions in adults affect total health care expenditures. Our objective was to estimate the annual average total expenditures and out-of-pocket spending burden among US adults by combinations of conditions. We conducted a cross-sectional study using 2009 and 2011 data from the Medical Expenditure Panel Survey. The sample consisted of 9,296 adults aged 21 years or older with at least 2 of the following 4 highly prevalent chronic conditions: arthritis, diabetes mellitus, heart disease, and hypertension. Unadjusted and adjusted regression techniques were used to examine the association between chronic condition combinations and log-transformed total expenditures. Logistic regressions were used to analyze the relationship between chronic condition combinations and high out-of-pocket spending burden. Among adults with chronic conditions, adults with all 4 conditions had the highest average total expenditures ($20,016), whereas adults with diabetes/hypertension had the lowest annual total expenditures ($7,116). In adjusted models, adults with diabetes/hypertension and hypertension/arthritis had lower health care expenditures than adults with diabetes/heart disease (P < .001). In adjusted models, adults with all 4 conditions had higher expenditures compared with those with diabetes and heart disease. However, the difference was only marginally significant (P = .04). Among adults with arthritis, diabetes, heart disease, and hypertension, total health care expenditures differed by type of chronic condition combinations. For individuals with multiple chronic conditions, such as heart disease and diabetes, new models of care management are needed to reduce the cost burden on the payers.


Adrenomedullin inhibits hypoxic cell death by upregulation of Bcl-2 in endometrial cancer cells: a possible promotion mechanism for tumour growth.
Regions of hypoxia are a common feature of solid tumours. When tumour cells are exposed to hypoxic stress, transcription of a battery of genes is initiated. The angiogenic factor adrenomedullin (ADM) is a hypoxia regulated gene. ADM is thought to act through the G protein-coupled receptor calcitonin receptor-like receptor (CRLR), with specificity being conferred by the receptor associated modifying protein 2 (RAMP2). Here we report for the first time that ADM treated or stably transfected Ishikawa cells overexpressing ADM show increased resistance to hypoxia induced apoptosis. These cells also show an upregulation of the oncoprotein Bcl-2, which is protective against hypoxic cell death when transiently transfected into Ishikawa cells. Since Ishikawa cells express the putative ADM-receptor CRLR-RAMP2 the production and secretion of ADM with the consecutive upregulation of Bcl-2 could establish an autocrine/paracrine mechanism rescuing malignant cells from hypoxic cell death. These results, taken together with our previous findings that ADM is an angiogenic factor which is upregulated by the nonsteroidal antiestrogen tamoxifen (TAM) in endometrial cells, implicate this peptide as a promoter of tumour growth and a possible target for anticancer strategies.


Variation in heart rate influences the assessment of transient ischemic dilation in myocardial perfusion scintigraphy.
Transient arrhythmias can affect transient ischemic dilation (TID) ratios. This study was initiated to evaluate the frequency and effect of normal heart rate change on TID measures in routine clinical practice. Consecutive patients undergoing stress/rest sestamibi gated myocardial perfusion scintigraphy were studied (N = 407). Heart rate at the time of stress and rest imaging were recorded. TID ratios were analyzed in relation to absolute change in heart rate (stress minus rest) for subjects with normal perfusion and systolic function (Group 1, N = 169) and those with abnormalities in perfusion and/or function (Group 2, N = 238). In Group 1, mean TID ratio was inversely correlated with the change in heart rate (r = -0.47, P < 0.0001). For every increase of 10 BPM in heart rate change, the TID ratio decreased by approximately 0.06 (95% confidence interval 0.04-0.07). In Group 2, multiple linear regression demonstrated that the change in heart rate (beta = -0.25, P < 0.0001) and the summed difference score (beta = 0.36, P < 0.0001) were independent predictors of the TID ratio. Normal variation in heart rate between the stress and rest components of myocardial perfusion scans is common and can influence TID ratios in patients with normal and abnormal cardiac scans.


Community mental health services in Southern Gauteng: An audit using Gauteng District Health Information Systems data.
Community mental health services (CMHS) are a central objective of the National Mental Health Policy Framework and Strategic Plan. Three core components are described: residential facilities, day care and outpatient services. Primary mental health care with specialist support is required according to an intervention pyramid. Staffing norms provide for a minimum mental health service coverage of 2.7% of the population for adults and 1.5% for children and adolescents. The aim of this study was to describe the existing CMHS in Southern Gauteng in terms of the National Mental Health Policy. The CMHS of the City of Johannesburg, Ekurhuleni, Sedibeng and West Rand districts were studied. Information regarding service organisation and staffing was obtained via the Gauteng Directorate of Mental Health. Routinely collected District Health Information Systems data for the 2014/2015 year were analysed. The organisation of services was not consistent with that recommended by the Mental Health Policy, and specialist CMHS were inappropriately situated within primary care. Only 2.23% of clinic visits were for mental health, and 80% of these were at specialist CMHS. Overall mental health coverage was approximately 0.3% of the population for adults and 0.02% for children and adolescents. Staffing, residential facilities and day care were far below the cited norms for minimal cover. Our audit revealed that the CMHS in Southern Gauteng did not meet any of the norms cited by the Mental Health Policy. Barriers to implementation of this aspect of the Mental Health Policy need to be explored.


Error-space estimation method and simplified algorithm for space target tracking.
To improve space target tracking precision and the stability of mobile optoelectronic tracking equipment, an error-space estimation method based on the Kalman filter is discussed, and a simplified algorithm is presented to reduce calculation cost. Based on an available measurement of a space target without sufficient validity and accuracy, the actual position related to the tracking equipment is decomposed to an earlier offline prediction of the kinetic model method and prediction errors. By regarding prediction errors as the motion of a weak maneuver target, the errors can be estimated more accurately in error space. By synthesizing estimation of the errors and offline prediction, the space target position is obtained with higher accuracy to improve tracking performance.


An epidemiologic analysis of clinical practice guidelines for non-arthroplasty treatment of osteoarthritis of the knee.
To analyze the current practice patterns of non-arthroplasty treatment of knee osteoarthritis (OA) and to assess the impact of the American Academy of Orthopaedic Surgeons clinical practice guidelines on the management of OA of the knee, particularly as they relate to the use of arthroscopic treatment. The United Healthcare Database (2004-2009, 11 million patients, 216 million records) was used for the study and was searched using Boolean language for International Classification of Diseases, Ninth Edition, Clinical Modification and Current Procedural Terminology, fourth revision codes. A reference group was defined as patients treated with knee arthroplasty in 2009 and diagnosed with knee OA in the same record. Clinical practice patterns in the 5 years preceding arthroplasty were analyzed in this group. The reference group consisted of 12,806 patients undergoing total knee arthroplasty in 2009 with a documented diagnosis of OA at the time of surgery, with prior nonoperative treatment strategies analyzed during the preceding 5 years (2004-2009); 10.0% of patients were prescribed physical therapy specific to OA, 2.6% received an unloader brace, 0.52% underwent acupuncture, 43.5% were administered intra-articular corticosteroids, and 15.4% received viscosupplementation injections. During the 5 years before arthroplasty, 2,505 patients (19.6%) underwent arthroscopy and debridement/lavage, 35% of whom did not have a diagnosis code for mechanical pathology. Within 1 year of knee arthroplasty, 2,028 of the 2,505 knee arthroscopies (80.9%) were performed. The findings show that significant gaps do exist between the evidence-based American Academy of Orthopaedic Surgeons recommendations and actual practice patterns in the United States between 2004 and 2009. Level IV, diagnostic study.


Growth inhibitory concentrations of EGF induce p21 (WAF1/Cip1) and alter cell cycle control in squamous carcinoma cells.
Previous studies have reported inhibition of A431 squamous carcinoma cell growth by nanomolar concentrations of epidermal growth factor (EGF), a potent mitogen for cells of epithelial origin. In this study, we examined potential mechanisms through which inhibition of keratinocyte growth mediated by EGF might occur by analysing components of the cell cycle regulatory machinery in A431, HN6 and HN30 keratinocytes in the presence of growth inhibitory or growth stimulatory doses of EGF. Treatment of cells with 25 pM EGF produced an increase in [3H]thymidine incorporation in A431, HN6 and HN30 cells, with respect to control cultures. Exposure to 2.5 nM EGF reduced [3H]thymidine incorporation in A431 cells and HN6 cells to 11% and 70% of control levels, respectively, whereas HN30 cells continued to proliferate in the presence of EGF. [3H]thymidine incorporation assays carried out over 24 h revealed repression of DNA synthesis in A431 cells after 12 h exposure to 2.5 nM EGF compared to untreated cells. Flow cytometry studies demonstrated accumulation of cells in G0/G1 after addition of 2.5 nM, but not 25 pM EGF. Western blot analysis revealed elevation of p21 (WAF1/CIP1/SDI1) protein levels in A431 and HN6 cells under growth-inhibitory conditions. Stimulatory doses of EGF did not induce p21 in these cells. Northern blot hybridization demonstrated elevated levels of p21 mRNA within 4 h of exposure of A431 cells to 2.5 nM EGF, which remained elevated above basal levels at 24 h. In vitro kinase assays demonstrated temporal differences in CDK2 and CDK6 activities which were related to EGF concentration. Immunocomplex Western blotting demonstrated increased association of p21 with CDK2 and CDK6 in A431 cells treated with 2.5 nm EGF. Furthermore, temporal alterations in the association of PCNA with p21 and with CDK6 were observed. The data indicate that p21 is a likely mediator of EGF-induced growth-inhibition, probably through mechanisms involving sequestration of PCNA and inhibition of CDK activity.


Direct-Acting Antivirals in Chronic Hepatitis C Genotype 4 Infection in Community Care Setting.
Limited data exists comparing the safety, tolerability, and efficacy of direct-acting antivirals (DAAs) in patients with chronic hepatitis C genotype 4 (HCV GT-4) in the community practice setting. We aim to evaluate the treatment response of DAAs in these patients. All the HCV GT-4 patients treated with DAAs between January 2014 and October 2017 in a community clinic setting were retrospectively analyzed. Pretreatment baseline patient characteristics, treatment efficacy with sustained virologic response (SVR) at 12 weeks post treatment (SVR12), and adverse reactions were assessed. Fifty-two patients of Middle Eastern (primarily Egyptian) descent were included in the study. Thirty-two patients were treated with ledipasvir/sofosbuvir (Harvoni®) ± ribavirin, 12 patients were treated with ombitasvir/paritaprevir/ritonavir/dasabuvir (ViekiraPak®) ± ribavirin, and eight patients were treated with sofosbuvir/Velpatasvir (Epclusa®). Ten patients (19.2%) had compensated cirrhosis. Overall, SVR at 12 weeks was achieved in 94% in patients who received one of the three DAA regimens (93.8% in Harvoni® group, 91.7 % in ViekiraPak® group and 100% in Epclusa® group). Prior treatment status and type of regimen used in the presence of compensated cirrhosis had no statistical significance on overall SVR achievement (P value = 0.442 and P value = 0.091, respectively). The most common adverse effect was fatigue (27%). In the real-world setting, DAAs are effective and well tolerated in patients with chronic HCV GT-4 infection with a high overall SVR rate of 94%. Large-scale studies are needed to further assess this SVR in these groups.


Evaluating portable infrared spectrometers for measuring the silica content of coal dust.
Miners face a variety of respiratory hazards while on the job, including exposure to silica dust which can lead to silicosis, a potentially fatal lung disease. Currently, field-collected filter samples of silica are sent for laboratory analysis and the results take weeks to be reported. Since the mining workplace is constantly moving into new and often different geological strata with changing silica levels, more timely data on silica levels in mining workplaces could help reduce exposures. Improvements in infrared (IR) spectroscopy open the prospect for end-of-shift silica measurements at mine sites. Two field-portable IR spectrometers were evaluated for their ability to quantify the mass of silica on filter samples loaded with known amounts of either silica or silica-bearing coal dust (silica content ranging from 10-200 μg/filter). Analyses included a scheme to correct for the presence of kaolin, which is a confounder for IR analysis of silica. IR measurements of the samples were compared to parallel measurements derived using the laboratory-based U.S. Mine Safety and Health Administration P7 analytical method. Linear correlations between Fourier transform infrared (FTIR) and P7 data yielded slopes in the range of 0.90-0.97 with minimal bias. Data from a variable filter array spectrometer did not correlate as well, mainly due to poor wavelength resolution compared to the FTIR instrument. This work has shown that FTIR spectrometry has the potential to reasonably estimate the silica exposure of miners if employed in an end-of-shift method.


The origin and evolution of human glutaminases and their atypical C-terminal ankyrin repeats.
On the basis of tissue-specific enzyme activity and inhibition by catalytic products, Hans Krebs first demonstrated the existence of multiple glutaminases in mammals. Currently, two human genes are known to encode at least four glutaminase isoforms. However, the phylogeny of these medically relevant enzymes remains unclear, prompting us to investigate their origin and evolution. Using prokaryotic and eukaryotic glutaminase sequences, we built a phylogenetic tree whose topology suggested that the multidomain architecture was inherited from bacterial ancestors, probably simultaneously with the hosting of the proto-mitochondrion endosymbiont. We propose an evolutionary model wherein the appearance of the most active enzyme isoform, glutaminase C (GAC), which is expressed in many cancers, was a late retrotransposition event that occurred in fishes from the Chondrichthyes class. The ankyrin (ANK) repeats in the glutaminases were acquired early in their evolution. To obtain information on ANK folding, we solved two high-resolution structures of the ANK repeat-containing C termini of both kidney-type glutaminase (KGA) and GLS2 isoforms (glutaminase B and liver-type glutaminase). We found that the glutaminase ANK repeats form unique intramolecular contacts through two highly conserved motifs; curiously, this arrangement occludes a region usually involved in ANK-mediated protein-protein interactions. We also solved the crystal structure of full-length KGA and present a small-angle X-ray scattering model for full-length GLS2. These structures explain these proteins' compromised ability to assemble into catalytically active supra-tetrameric filaments, as previously shown for GAC. Collectively, these results provide information about glutaminases that may aid in the design of isoform-specific glutaminase inhibitors.


Interaction of methylation-related genetic variants with circulating fatty acids on plasma lipids: a meta-analysis of 7 studies and methylation analysis of 3 studies in the Cohorts for Heart and Aging Research in Genomic Epidemiology consortium.
DNA methylation is influenced by diet and single nucleotide polymorphisms (SNPs), and methylation modulates gene expression. We aimed to explore whether the gene-by-diet interactions on blood lipids act through DNA methylation. We selected 7 SNPs on the basis of predicted relations in fatty acids, methylation, and lipids. We conducted a meta-analysis and a methylation and mediation analysis with the use of data from the CHARGE (Cohorts for Heart and Aging Research in Genomic Epidemiology) consortium and the ENCODE (Encyclopedia of DNA Elements) consortium. On the basis of the meta-analysis of 7 cohorts in the CHARGE consortium, higher plasma HDL cholesterol was associated with fewer C alleles at ATP-binding cassette subfamily A member 1 (ABCA1) rs2246293 (β = -0.6 mg/dL, P = 0.015) and higher circulating eicosapentaenoic acid (EPA) (β = 3.87 mg/dL, P = 5.62 × 10(21)). The difference in HDL cholesterol associated with higher circulating EPA was dependent on genotypes at rs2246293, and it was greater for each additional C allele (β = 1.69 mg/dL, P = 0.006). In the GOLDN (Genetics of Lipid Lowering Drugs and Diet Network) study, higher ABCA1 promoter cg14019050 methylation was associated with more C alleles at rs2246293 (β = 8.84%, P = 3.51 × 10(18)) and lower circulating EPA (β = -1.46%, P = 0.009), and the mean difference in methylation of cg14019050 that was associated with higher EPA was smaller with each additional C allele of rs2246293 (β = -2.83%, P = 0.007). Higher ABCA1 cg14019050 methylation was correlated with lower ABCA1 expression (r = -0.61, P = 0.009) in the ENCODE consortium and lower plasma HDL cholesterol in the GOLDN study (r = -0.12, P = 0.0002). An additional mediation analysis was meta-analyzed across the GOLDN study, Cardiovascular Health Study, and the Multi-Ethnic Study of Atherosclerosis. Compared with the model without the adjustment of cg14019050 methylation, the model with such adjustment provided smaller estimates of the mean plasma HDL cholesterol concentration in association with both the rs2246293 C allele and EPA and a smaller difference by rs2246293 genotypes in the EPA-associated HDL cholesterol. However, the differences between 2 nested models were NS (P > 0.05). We obtained little evidence that the gene-by-fatty acid interactions on blood lipids act through DNA methylation.


Caregiver burden assessed in dementia care networks in Germany: findings from the DemNet-D study baseline.
This paper aimed to describe the burden experienced by informal caregivers supporting a person with dementia (PwD) who lives at home and utilizes a dementia care network (DCN), to investigate the factors that are associated with caregiver burden, and to identify possible differences in caregiver burden among different types of DCNs. This study was part of a multi-center, interdisciplinary evaluation of DCNs in Germany (DemNet-D). Cross-sectional data were collected in face-to-face interviews with people with dementia (PwDs) and their caregivers, and 13 DCNs were represented. Standardized questionnaires were used to assess caregiver burden, challenging PwD behaviors, functional competence and caregiver health status. Based on qualitative data, four DCN governance types were used in a multivariate analysis of burden categories. There were 560 PwD-caregiver dyads enrolled in the study. Informal caregivers (n = 536) reported a low-to-moderate burden associated with PwD characteristics (instrumental activities of daily living, challenging behaviors) and caregiver characteristics (gender) as well as the relationship between the caregivers and PwDs. Women felt more burdened but also showed higher levels of personal development. No differences were observed among the different DCN governance types. DCNs might contribute toward moderate to low caregiver burden. Indicators of positive caregiving aspects can be used by DCNs to advance support structures for informal caregivers drawing upon interventions already established for other community settings. Particular interest should be paid to female and spousal caregivers who might be in in need of greater and/or different kinds of support.


Iapetus: unique surface properties and a global color dichotomy from Cassini imaging.
Since 2004, Saturn's moon Iapetus has been observed repeatedly with the Imaging Science Subsystem of the Cassini spacecraft. The images show numerous impact craters down to the resolution limit of approximately 10 meters per pixel. Small, bright craters within the dark hemisphere indicate a dark blanket thickness on the order of meters or less. Dark, equator-facing and bright, poleward-facing crater walls suggest temperature-driven water-ice sublimation as the process responsible for local albedo patterns. Imaging data also reveal a global color dichotomy, wherein both dark and bright materials on the leading side have a substantially redder color than the respective trailing-side materials. This global pattern indicates an exogenic origin for the redder leading-side parts and suggests that the global color dichotomy initiated the thermal formation of the global albedo dichotomy.


Evaluating the cardiovascular safety of sclerostin inhibition using evidence from meta-analysis of clinical trials and human genetics.
Inhibition of sclerostin is a therapeutic approach to lowering fracture risk in patients with osteoporosis. However, data from phase 3 randomized controlled trials (RCTs) of romosozumab, a first-in-class monoclonal antibody that inhibits sclerostin, suggest an imbalance of serious cardiovascular events, and regulatory agencies have issued marketing authorizations with warnings of cardiovascular disease. Here, we meta-analyze published and unpublished cardiovascular outcome trial data of romosozumab and investigate whether genetic variants that mimic therapeutic inhibition of sclerostin are associated with higher risk of cardiovascular disease. Meta-analysis of up to three RCTs indicated a probable higher risk of cardiovascular events with romosozumab. Scaled to the equivalent dose of romosozumab (210 milligrams per month; 0.09 grams per square centimeter of higher bone mineral density), the SOST genetic variants were associated with lower risk of fracture and osteoporosis (commensurate with the therapeutic effect of romosozumab) and with a higher risk of myocardial infarction and/or coronary revascularization and major adverse cardiovascular events. The same variants were also associated with increased risk of type 2 diabetes mellitus and higher systolic blood pressure and central adiposity. Together, our findings indicate that inhibition of sclerostin may elevate cardiovascular risk, warranting a rigorous evaluation of the cardiovascular safety of romosozumab and other sclerostin inhibitors.


Fine-needle aspiration for diagnosis of tuberculous lymphadenitis in children in Bangui, Central African Republic.
Tuberculosis (TB) is a major cause of childhood morbidity and mortality in developing countries. One of the main difficulties is obtaining adequate specimens for bacteriological confirmation of the disease in children.The aim of this study is to evaluate the adequacy of fine-needle aspiration (FNA) for the diagnosis of TB. In a prospective study conducted at the paediatric hospital in Bangui in 2007-2009, we used fine-needle aspiration to obtain samples for diagnosis of TB from 131 children aged 0-17 years with persistent lymphadenitis. Fine-needle aspiration provided samples that could be used for bacteriological confirmation of TB. Ziehl-Neelsen staining for acid-fast bacilli was positive in 42.7% of samples, and culture identified TB in 67.2% of cases. Of 75 samples that were stain-negative, 49 (65.3%) were culture-positive, while 12 stain-positive samples remained culture-negative. Ten of the 12 stain-positive, culture-negative samples were from patients who had received previous antimicrobial therapy. With regard to phenotypic drug susceptibility, 81/88 strains (91.1%) were fully susceptible to isoniazid, rifampicin, ethambutol and streptomycin, six (6.8%) were resistant to one drug, and one multidrug-resistant strain was found. Fine-needle aspiration is simple, cost-effective and non-invasive and can be performed by trained staff. Combined with rapid molecular diagnostic tests, fine-needle aspirates could improve the diagnosis of TB and provide valuable information for appropriate treatment and drug resistance.


Human coronavirus NL63 open reading frame 3 encodes a virion-incorporated N-glycosylated membrane protein.
Human pathogenic coronavirus NL63 (hCoV-NL63) is a group 1 (alpha) coronavirus commonly associated with respiratory tract infections. In addition to known non-structural and structural proteins all coronaviruses have one or more accessory proteins whose functions are mostly unknown. Our study focuses on hCoV-NL63 open reading frame 3 (ORF 3) which is a highly conserved accessory protein among coronaviruses. In-silico analysis of the 225 amino acid sequence of hCoV-NL63 ORF 3 predicted a triple membrane-spanning protein. Expression in infected CaCo-2 and LLC-MK2 cells was confirmed by immunofluorescence and Western blot analysis. The protein was detected within the endoplasmatic reticulum/Golgi intermediate compartment (ERGIC) where coronavirus assembly and budding takes place. Subcellular localization studies using recombinant ORF 3 protein transfected in Huh-7 cells revealed occurrence in ERGIC, Golgi- and lysosomal compartments. By fluorescence microscopy of differently tagged envelope (E), membrane (M) and nucleocapsid (N) proteins it was shown that ORF 3 protein colocalizes extensively with E and M within the ERGIC. Using N-terminally FLAG-tagged ORF 3 protein and an antiserum specific to the C-terminus we verified the proposed topology of an extracellular N-terminus and a cytosolic C-terminus. By in-vitro translation analysis and subsequent endoglycosidase H digestion we showed that ORF 3 protein is N-glycosylated at the N-terminus. Analysis of purified viral particles revealed that ORF 3 protein is incorporated into virions and is therefore an additional structural protein. This study is the first extensive expression analysis of a group 1 hCoV-ORF 3 protein. We give evidence that ORF 3 protein is a structural N-glycosylated and virion-incorporated protein.


Dietary treatment in adults with refractory epilepsy: a review.
We review adjunctive ketogenic diet (KD) and modified Atkins diet (MAD) treatment of refractory epilepsy in adults. Only a few studies have been published, all open-label. Because of the disparate, uncontrolled nature of the studies, we analyzed all studies individually, without a meta-analysis. Across all studies, 32% of KD-treated and 29% of MAD-treated patients achieved ≥ 50% seizure reduction, including 9% and 5%, respectively, of patients with >90% seizure frequency reduction. The effect persists long term, but, unlike in children, may not outlast treatment. The 3:1 and 4:1 [fat]:[carbohydrate + protein] ratio KD variants and MAD are similarly effective. The anticonvulsant effect occurs quickly with both diets, within days to weeks. Side effects of both diets are benign and similar. The most serious, hyperlipidemia, reverses with treatment discontinuation. The most common, weight loss, may be advantageous in patients with obesity. Potential barriers to large-scale use of both diets in adults include low rate of diet acceptance and high rates of diet discontinuation. The eligible screened/enrolled subject ratios ranged from 2.9 to 7.2. Fifty-one percent of KD-treated and 42% of MAD-treated patients stopped the diet before study completion. Refusal to participate was due to diet restrictiveness and complexity, which may be greater for KD than MAD. However, long-term adherence is low for both diets. Most patients eventually stop the diet because of culinary and social restrictions. For treatment of refractory status epilepticus, only 14 adult cases of KD treatment have been published, providing insufficient data to allow evaluation. In summary, KD and MAD treatment show modest efficacy, although in some patients the effect is remarkable. The diets are well-tolerated, but often discontinued because of their restrictiveness. In patients willing to try dietary treatment, the effect is seen quickly, giving patients the option whether to continue the treatment.


Characterisation of an oxidative response inhibitor produced by Streptococcus pneumoniae.
Pneumonia caused by infection with Streptococcus pneumoniae is still a major clinical problem. Reactive oxygen species contribute to the killing of these bacteria by polymorphonuclear leucocytes (PMNs). Defence mechanisms of Str pneumoniae which counter reactive oxygen species are characterised. PMNs were stimulated with phorbol myristate acetate (PMA) in the presence and absence of Str pneumoniae and supernatants from them, and superoxide (O2-) production was measured by the reduction of ferricytochrome c. Streptococcus pneumoniae, but not Klebsiella pneumoniae or Staphylococcus aureus, inhibited PMA stimulated superoxide production by PMNs. Washed PMNs which had been preincubated with Str pneumoniae autolysis phase supernatants also exhibited depressed H2O2 production in response to PMA. The inhibitory activity was not attributable to non-specific cytotoxicity as assessed by release of the cytoplasmic enzyme lactate dehydrogenase, nor did the supernatants inhibit PMA stimulated degranulation of PMNs. Fractionation of the autolysis phase supernatants revealed inhibitory activity in both the fractions greater than and less than 10 kD. Like pneumolysin the inhibitory activity was heat sensitive. However, both a parent and pneumolysin negative mutant Str pneumoniae, and autolysis phase supernatants from them, inhibited PMN superoxide production. Antisera to pneumolysin failed to abrogate the inhibitory effect of intact Str pneumoniae or autolysis phase supernatants from types 1 or 14 Str pneumoniae. The inhibitory effect of Str pneumoniae on the respiratory burst of PMNs is not shared by two other common lung pathogens. The existence of a novel inhibitor of the PMN respiratory burst, distinct from pneumolysin, has been demonstrated. The inhibitor is specific for the respiratory burst and is active both in the logarithmic phase of growth and during autolysis.


Assessment of the magnetic field exposure due to the battery current of digital mobile phones.
Hand-held digital mobile phones generate pulsed magnetic fields associated with the battery current. The peak value and the waveform of the battery current were measured for seven different models of digital mobile phones, and the results were applied to compute approximately the magnetic flux density and induced currents in the phone-user's head. A simple circular loop model was used for the magnetic field source and a homogeneous sphere consisting of average brain tissue equivalent material simulated the head. The broadband magnetic flux density and the maximal induced current density were compared with the guidelines of ICNIRP using two various approaches. In the first approach the relative exposure was determined separately at each frequency and the exposure ratios were summed to obtain the total exposure (multiple-frequency rule). In the second approach the waveform was weighted in the time domain with a simple low-pass RC filter and the peak value was divided by a peak limit, both derived from the guidelines (weighted peak approach). With the maximum transmitting power (2 W) the measured peak current varied from 1 to 2.7 A. The ICNIRP exposure ratio based on the current density varied from 0.04 to 0.14 for the weighted peak approach and from 0.08 to 0.27 for the multiple-frequency rule. The latter values are considerably greater than the corresponding exposure ratios 0.005 (min) to 0.013 (max) obtained by applying the evaluation based on frequency components presented by the new IEEE standard. Hence, the exposure does not seem to exceed the guidelines. The computed peak magnetic flux density exceeded substantially the derived peak reference level of ICNIRP, but it should be noted that in a near-field exposure the external field strengths are not valid indicators of exposure. Currently, no biological data exist to give a reason for concern about the health effects of magnetic field pulses from mobile phones.


Expression pattern of two paralogs encoding cinnamyl alcohol dehydrogenases in Arabidopsis. Isolation and characterization of the corresponding mutants.
Studying Arabidopsis mutants of the phenylpropanoid pathway has unraveled several biosynthetic steps of monolignol synthesis. Most of the genes leading to monolignol synthesis have been characterized recently in this herbaceous plant, except those encoding cinnamyl alcohol dehydrogenase (CAD). We have used the complete sequencing of the Arabidopsis genome to highlight a new view of the complete CAD gene family. Among nine AtCAD genes, we have identified the two distinct paralogs AtCAD-C and AtCAD-D, which share 75% identity and are likely to be involved in lignin biosynthesis in other plants. Northern, semiquantitative restriction fragment-length polymorphism-reverse transcriptase-polymerase chain reaction and western analysis revealed that AtCAD-C and AtCAD-D mRNA and protein ratios were organ dependent. Promoter activities of both genes are high in fibers and in xylem bundles. However, AtCAD-C displayed a larger range of sites of expression than AtCAD-D. Arabidopsis null mutants (Atcad-D and Atcad-C) corresponding to both genes were isolated. CAD activities were drastically reduced in both mutants, with a higher impact on sinapyl alcohol dehydrogenase activity (6% and 38% of residual sinapyl alcohol dehydrogenase activities for Atcad-D and Atcad-C, respectively). Only Atcad-D showed a slight reduction in Klason lignin content and displayed modifications of lignin structure with a significant reduced proportion of conventional S lignin units in both stems and roots, together with the incorporation of sinapaldehyde structures ether linked at Cbeta. These results argue for a substantial role of AtCAD-D in lignification, and more specifically in the biosynthesis of sinapyl alcohol, the precursor of S lignin units.


Direct growth of carbon nanofibers to generate a 3D porous platform on a metal contact to enable an oxygen reduction reaction.
For carbon nanotube-based electronics to achieve their full performance potential, it is imperative to minimize the contact resistance between macroscale metal contacts and the carbon nanotube (CNT) nanoelectrodes. We have developed a three-dimensional electrode platform that consists of carbon nanofibers (CNFs) that are directly grown on a metal contact, such as copper (Cu). Carbon nanofiber morphology can be tailored by adjusting the annealing time of a thin electrochemically deposited nickel catalyst layer on copper. We demonstrate that increasing the annealing time increases the amount of copper infused into the nickel catalyst layer. This reduces the carbon deposition rate, and consequently a more well-defined CNF 3D architecture can be fabricated. This direct growth of CNFs on a Cu substrate yields an excellent electron transfer pathway, with contact resistance between CNFs and Cu being comparable to that of a Cu-Cu interface. Furthermore, the excellent bonding strength between CNFs and Cu can be maintained over prolonged periods of ultrasonication. The porous 3D platform affixed with intertwined CNFs allows facile surface functionalization. Using a simple solution soaking procedure, the CNF surface has been successfully functionalized with iron(II) phthalocyanine (FePc). FePc functionalized CNFs exhibit excellent oxygen reduction capability, equivalent to platinum-carbon electrodes. This result demonstrates the technological promise of this new 3D electrode platform that can be exploited in other applications that include sensing, battery, and supercapacitors.


Control of invasive rats on islands and priorities for future action.
Invasive rats are one of the world's most successful animal groups that cause native species extinctions and ecosystem change, particularly on islands. On large islands, rat eradication is often impossible and population control, defined as the local limitation of rat abundance, is now routinely performed on many of the world's islands as an alternative management tool. However, a synthesis of the motivations, techniques, costs, and outcomes of such rat-control projects is lacking. We reviewed the literature, searched relevant websites, and conducted a survey via a questionnaire to synthesize the available information on rat-control projects in island natural areas worldwide to improve rat management and native species conservation. Data were collected from 136 projects conducted over the last 40 years; most were located in Australasia (46%) and the tropical Pacific (25%) in forest ecosystems (65%) and coastal strands (22%). Most of the projects targeted Rattus rattus and most (82%) were aimed at protecting birds and endangered ecosystems. Poisoning (35%) and a combination of trapping and poisoning (42%) were the most common methods. Poisoning allows for treatment of larger areas, and poison projects generally last longer than trapping projects. Second-generation anticoagulants (mainly brodifacoum and bromadiolone) were used most often. The median annual cost for rat-control projects was US$17,262 or US$227/ha. Median project duration was 4 years. For 58% of the projects, rat population reduction was reported, and 51% of projects showed evidence of positive effects on biodiversity. Our data were from few countries, revealing the need to expand rat-control distribution especially in some biodiversity hotspots. Improvement in control methods is needed as is regular monitoring to assess short- and long-term effectiveness of rat-control.


Dextropropoxyphene induced hepatotoxicity: a report of nine cases.
Nine patients are described with jaundice, upper abdominal pain and malaise attributable to dextropropoxyphene hepatotoxicity. In each case the history was suggestive of large bile duct obstruction. All patients underwent ultrasound examination and percutaneous liver biopsy. Three patients also underwent endoscopic retrograde cholangio pancreatography. The histological features of the biopsies concur with previously reported cases of dextropropoxyphene hepatotoxicity. The histological changes seen on biopsy were remarkably constant, consisting of centrilobular cholestasis, portal tract inflammation and bile duct abnormalities, in all cases mimicking large bile duct obstruction. Fifteen previous patients with probable dextropropoxyphene hepatotoxicity have been described. The occurrence of 9 further cases at one centre, 6 presenting within 12 months, suggests that it is much more common than previously assumed and may be misdiagnosed as large bile duct obstruction.


Redox markers for drought-induced nodule senescence, a process occurring after drought-induced senescence of the lowest leaves in soybean (Glycine max).
Water is an increasingly scarce resource that limits crop productivity in many parts of the world, and the frequency and severity of drought are predicted to increase as a result of climate change. Improving tolerance to drought stress is therefore important for maximizing future crop yields. The aim of this study was to compare the effects of drought on soybean (Glycine max) leaves and nodules in order to define phenotypic markers and changes in cellular redox state that characterize the stress response in different organs, and to characterize the relationships between leaf and nodule senescence during drought. Leaf and crown nodule metabolite pools were measured together with leaf and soil water contents, and leaf chlorophyll, total protein contents and chlorophyll a fluorescence quenching parameters in nodulated soybeans that were grown under either well-watered conditions or deprived of water for up to 21 d. Ureides, ascorbate, protein, chlorophyll and the ratios of variable chlorophyll a fluorescence (Fv') to maximal chlorophyll a fluorescence (Fm') fell to levels below detection in the oldest leaves after 21 d of drought. While these drought-induced responses were not observed in the youngest leaf ranks, the Fv'/Fm' ratios, pyridine nucleotide levels and the reduction state of the ascorbate pool were lower in all leaf ranks after 21 d of drought. In contrast to leaves, total nodule protein, pyridine nucleotides, ureides, ascorbate and glutathione contents increased as a result of the drought treatment. However, the nodule ascorbate pool was significantly less reduced as a result of drought. Higher levels of transcripts encoding two peroxiredoxins were detected in nodules exposed to drought stress but senescence-associated transcripts and other mRNAs encoding redox-related proteins were similar under both conditions. While the physiological impact of the drought was perceived throughout the shoot, stress-induced senescence occurred only in the oldest leaf ranks. At this stage, a number of drought-induced changes in nodule metabolites were observed but no metabolite or transcript markers of senescence could be detected. It is concluded that stress-induced senescence in the lowest leaf ranks precedes nodule senescence, suggesting that leaves of low photosynthetic capacity are sacrificed in favour of nodule nitrogen metabolism.


Community hospitals--the place of local service provision in a modernising NHS: an integrative thematic literature review.
Recent developments within the United Kingdom's (UK) health care system have re-awakened interest in community hospitals (CHs) and their role in the provision of health care. This integrative literature review sought to identify and assess the current evidence base for CHs. A range of electronic reference databases were searched from January 1984 to either December 2004 or February 2005: Medline, Embase, Web of Knowledge, BNI, CINAHL, HMIC, ASSIA, PsychInfo, SIGLE, Dissertation Abstracts, Cochrane Library, Kings Fund website, using both keywords and text words. Thematic analysis identified recurrent themes across the literature; narrative analyses were written for each theme, identifying unifying concepts and discrepant issues. The search strategy identified over 16,000 international references. We included papers of any study design focussing on hospitals in which care was led principally by general practitioners or nurses. Papers from developing countries were excluded. A review of titles revealed 641 potentially relevant references; abstract appraisal identified 161 references for review. During data extraction, a further 48 papers were excluded, leaving 113 papers in the final review. The most common methodological approaches were cross-sectional/descriptive studies, commentaries and expert opinion. There were few experimental studies, systematic reviews, economic studies or studies that reported on longer-term outcomes. The key themes identified were origin and location of CHs; their place in the continuum of care; services provided; effectiveness, efficiency and equity of CHs; and views of patients and staff. In general, there was a lack of robust evidence for the role of CHs, which is partly due to the ad hoc nature of their development and lack of clear strategic vision for their future. Evidence for the effectiveness and efficiency of the services provided was limited. Most people admitted to CHs appeared to be older, suggesting that admittance to CHs was age-related rather than condition-related. Overall the literature surveyed was long on opinion and short of robust studies on CHs. While lack of evidence on CHs does not imply lack of effect, there is an urgent need to develop a research agenda that addresses the key issues of health care delivery in the CH setting.


Long-Term Follow-Up of the French Stop Imatinib (STIM1) Study in Patients With Chronic Myeloid Leukemia.
Purpose Imatinib (IM) can safely be discontinued in patients with chronic myeloid leukemia (CML) who have had undetectable minimal residual disease (UMRD) for at least 2 years. We report the final results of the Stop Imatinib (STIM1) study with a long follow-up. Patients and Methods IM was prospectively discontinued in 100 patients with CML with UMRD sustained for at least 2 years. Molecular recurrence (MR) was defined as positivity of BCR-ABL transcript in a quantitative reverse transcriptase polymerase chain reaction assay confirmed by a second analysis point that indicated an increase of one log in relation to the first analysis point at two successive assessments or loss of major molecular response at one point. Results The median molecular follow-up after treatment discontinuation was 77 months (range, 9 to 95 months). Sixty-one patients lost UMRD after a median of 2.5 months (range, 1 to 22 months), and one patient died with UMRD at 10 months. Molecular recurrence-free survival was 43% (95% CI, 33% to 52%) at 6 months and 38% (95% CI, 29% to 47%) at 60 months. Treatment was restarted in 57 of 61 patients with MR, and 55 patients achieved a second UMRD with a median time of 4 months (range, 1 to 16 months). None of the patients experienced a CML progression. Analyses of the characteristics of the study population identified that the Sokal risk score and duration of IM treatment were significantly associated with the probability of MR. Conclusion With a median follow-up of more than 6 years after treatment discontinuation, the STIM1 study demonstrates that IM can safely be discontinued in patients with a sustained deep molecular response with no late MR.


The androgen receptor is a tumor suppressor in estrogen receptor-positive breast cancer.
The role of the androgen receptor (AR) in estrogen receptor (ER)-α-positive breast cancer is controversial, constraining implementation of AR-directed therapies. Using a diverse, clinically relevant panel of cell-line and patient-derived models, we demonstrate that AR activation, not suppression, exerts potent antitumor activity in multiple disease contexts, including resistance to standard-of-care ER and CDK4/6 inhibitors. Notably, AR agonists combined with standard-of-care agents enhanced therapeutic responses. Mechanistically, agonist activation of AR altered the genomic distribution of ER and essential co-activators (p300, SRC-3), resulting in repression of ER-regulated cell cycle genes and upregulation of AR target genes, including known tumor suppressors. A gene signature of AR activity positively predicted disease survival in multiple clinical ER-positive breast cancer cohorts. These findings provide unambiguous evidence that AR has a tumor suppressor role in ER-positive breast cancer and support AR agonism as the optimal AR-directed treatment strategy, revealing a rational therapeutic opportunity.


PALM: a paralleled and integrated framework for phylogenetic inference with automatic likelihood model selectors.
Selecting an appropriate substitution model and deriving a tree topology for a given sequence set are essential in phylogenetic analysis. However, such time consuming, computationally intensive tasks rely on knowledge of substitution model theories and related expertise to run through all possible combinations of several separate programs. To ensure a thorough and efficient analysis and avert tedious manipulations of various programs, this work presents an intuitive framework, the phylogenetic reconstruction with automatic likelihood model selectors (PALM), with convincing, updated algorithms and a best-fit model selection mechanism for seamless phylogenetic analysis. As an integrated framework of ClustalW, PhyML, MODELTEST, ProtTest, and several in-house programs, PALM evaluates the fitness of 56 substitution models for nucleotide sequences and 112 substitution models for protein sequences with scores in various criteria. The input for PALM can be either sequences in FASTA format or a sequence alignment file in PHYLIP format. To accelerate the computing of maximum likelihood and bootstrapping, this work integrates MPICH2/PhyML, PalmMonitor and Palm job controller across several machines with multiple processors and adopts the task parallelism approach. Moreover, an intuitive and interactive web component, PalmTree, is developed for displaying and operating the output tree with options of tree rooting, branches swapping, viewing the branch length values, and viewing bootstrapping score, as well as removing nodes to restart analysis iteratively. The workflow of PALM is straightforward and coherent. Via a succinct, user-friendly interface, researchers unfamiliar with phylogenetic analysis can easily use this server to submit sequences, retrieve the output, and re-submit a job based on a previous result if some sequences are to be deleted or added for phylogenetic reconstruction. PALM results in an inference of phylogenetic relationship not only by vanquishing the computation difficulty of ML methods but also providing statistic methods for model selection and bootstrapping. The proposed approach can reduce calculation time, which is particularly relevant when querying a large data set. PALM can be accessed online at http://palm.iis.sinica.edu.tw.


Rapid detection of Salmonella in milk by combined immunomagnetic separation-polymerase chain reaction assay.
During the past few years, milk has presented a risk of Salmonella contamination; it has been implicated as the cause in several outbreaks of salmonellosis. Because conventional detection methods require 5 to 7 d for completion and involve several subcultivation stages followed by biochemical and serological tests, rapid and sensitive methods have been sought, mainly at the DNA level. Therefore, a study including milk samples was conducted to evaluate the performance of a combination of 2 techniques--immunomagnetic separation and polymerase chain reaction (PCR)--for the detection of Salmonella. The 16-, 14-, 12-, 10-, and 8-h nonselective pre-enrichment steps before immunomagnetic separation and the high-pure DNA preparation method before PCR were used in a combined assay. Milk samples, which were found to be Salmonella-negative by a reference method, were first inoculated with Salmonella Enteritidis. Next, the shortest pre-enrichment time that is required for detection of 1 or 10 cfu of Salmonella/mL by combined immunomagnetic separation-PCR assay was found by using 16-, 14-, 12-, 10-, and 8-h incubation periods. The detection limit using a 16-, 14-, or 12-h nonselective pre-enrichment was 1 to 10 cfu/mL. However, the sensitivity decreased to 10(1) and 10(2) cfu/mL, respectively, when 10- and 8-h pre-enrichments were used. This assay, in conjunction with a 12-h pre-enrichment, proved to be rapid (overall 16 h) and sensitive (1-10 cfu/mL) for the detection of Salmonella in milk samples and promising for routine use in the detection of Salmonella in milk.


Extracting Fundamental Periods to Segment Biomedical Signals.
We address the problem of segmenting nearly periodic time series into period-like segments. We introduce a definition of nearly periodic time series via triplets 〈 basic shape, shape transformation, time scaling 〉 that covers a wide range of time series. To split the time series into periods, we select a pair of principal components of the Hankel matrix. We then cut the trajectory of the selected principal components by its symmetry axis and, thus, obtaining half-periods that are merged into segments. We describe a method of automatic selection of periodic pairs of principal components, corresponding to the fundamental periodicity. We demonstrate the application of the proposed method to the problem of period extraction for accelerometric time series of human gait. We see the automatic segmentation into periods as a problem of major importance for human activity recognition problem, since it allows to obtain interpretable segments: each extracted period can be seen as an ultimate entity of gait. The method we propose is more general compared to the application specific methods and can be used for any nearly periodical time series. We compare its performance to classical mathematical methods of period extraction and find that it is not only comparable to the alternatives, but in some cases performs better.


Common origins of hippocampal Ivy and nitric oxide synthase expressing neurogliaform cells.
GABAergic interneurons critically regulate cortical computation through exquisite spatiotemporal control over excitatory networks. Precision of this inhibitory control requires a remarkable diversity within interneuron populations that is largely specified during embryogenesis. Although interneurons expressing the neuronal isoform of nitric oxide synthase (nNOS) constitute the largest hippocampal interneuron cohort their origin and specification remain unknown. Thus, as neurogliaform cells (NGC) and Ivy cells (IvC) represent the main nNOS(+) interneurons, we investigated their developmental origins. Although considered distinct interneuron subtypes, NGCs and IvCs exhibited similar neurochemical and electrophysiological signatures, including NPY expression and late spiking. Moreover, lineage analyses, including loss-of-function experiments and inducible fate-mapping, indicated that nNOS(+) IvCs and NGCs are both derived from medial ganglionic eminence (MGE) progenitors under control of the transcription factor Nkx2-1. Surprisingly, a subset of NGCs lacking nNOS arises from caudal ganglionic eminence (CGE) progenitors. Thus, while nNOS(+) NGCs and IvCs arise from MGE progenitors, a CGE origin distinguishes a discrete population of nNOS(-) NGCs.


Dissociating effect of upper limb non-use and overuse on space and body representations.
Accurate and updated representations of the space where the body acts, i.e. the peripersonal space (PPS), and the location and dimension of body parts (body representation, BR) are essential to perform actions. Because both PPS and BR are involved in motor execution and display the same plastic proprieties after the use of a tool to reach far objects, it has been suggested that they overlap in a unique representation of the body in a space devoted to action. Here we determined whether manipulating actions in space, without modifying body metrics, i.e. through immobilization, induces a dissociation of the plastic properties of PPS and BR. In 39 healthy subjects we evaluated PPS and BR for the non-used right limb and the overused left limb before and after 10 h of right arm immobilization. We observed that non-use reduces PPS representation around the immobilized arm, without affecting the metric representation (i.e. perceived length) of that limb. In contrast, overuse modulates the metric representation of the free arm, leaving PPS unchanged around that limb. These results suggest that the plasticity in PPS and BR depends on different mechanisms; while PPS representation is shaped as a function of the dimension of the acting space, metric characteristics of BR are forged on a complex interplay between visual and sensorimotor information related to the body. This behavioral dissociation between PPS and BR defines a new scenario for the role of action in shaping space and body representations.


Determinants of patient satisfaction with outcome after rotator cuff surgery.
The outcome after rotator cuff surgery has been reported frequently with use of condition-specific measures, but patient satisfaction has not been well studied. The purpose of this study was to identify the determinants of patient satisfaction with the outcome after rotator cuff surgery. A cohort of 311 patients undergoing rotator cuff surgery was studied prospectively. Two hundred and fifty-four patients had subjective follow-up for a minimum of one year, and fifty-seven patients had subjective and objective follow-up for a minimum of one year. Diagnoses included tendinitis or impingement (eighty patients), a partial-thickness rotator cuff tear (fifty-five patients), and a full-thickness rotator cuff tear (176 patients). The dependent variable was patient satisfaction with the outcome, graded ordinally on a scale of 1 to 10. Independent variables included demographic, surgical, objective follow-up, and subjective follow-up parameters. Univariate and multivariate analyses were performed to identify the determinants of satisfaction. Univariate analysis of demographic variables demonstrated no significant differences (p > 0.05) with respect to patient satisfaction. Analysis of the surgical variables showed significantly decreased satisfaction (p < 0.05) for patients who had debridement for a massive, irreparable cuff tear; those who had subscapularis tears; and those with larger supraspinatus and infraspinatus tears. Analysis of the objective variables at follow-up demonstrated significantly decreased satisfaction (p < 0.05) for patients with diminished and weakened forward elevation, impingement signs, and acromioclavicular joint pain and tenderness. Univariate analysis of subjective variables at follow-up showed significantly decreased satisfaction (p < 0.001) for patients with pain, functional difficulty, and work disability. Multivariate analysis demonstrated significantly decreased satisfaction associated with pain (p < 0.001) and dysfunction (p < 0.05). A significant relationship (p < 0.05) was detected between satisfaction and a willingness to recommend the surgery to another. A significant relationship (p < 0.05) was also found between satisfaction and the American Shoulder and Elbow Surgeons score. Subjective variables of symptoms and function have the most robust associations with patient satisfaction following rotator cuff surgery. Thus, in assessing the outcome of rotator cuff surgery from the perspective of patient satisfaction with outcome, we emphasize the importance of patient-derived subjective assessment of symptoms and function.


Automating human intuition for protein design.
In the design of new enzymes and binding proteins, human intuition is often used to modify computationally designed amino acid sequences prior to experimental characterization. The manual sequence changes involve both reversions of amino acid mutations back to the identity present in the parent scaffold and the introduction of residues making additional interactions with the binding partner or backing up first shell interactions. Automation of this manual sequence refinement process would allow more systematic evaluation and considerably reduce the amount of human designer effort involved. Here we introduce a benchmark for evaluating the ability of automated methods to recapitulate the sequence changes made to computer-generated models by human designers, and use it to assess alternative computational methods. We find the best performance for a greedy one-position-at-a-time optimization protocol that utilizes metrics (such as shape complementarity) and local refinement methods too computationally expensive for global Monte Carlo (MC) sequence optimization. This protocol should be broadly useful for improving the stability and function of designed binding proteins.


Stigmergy, collective actions, and animal social spacing.
Collective animal behavior studies have led the way in developing models that account for a large number of individuals, but mostly have considered situations in which alignment and attraction play a key role, such as in schooling and flocking. By quantifying how animals react to one another's presence, when interaction is via conspecific avoidance rather than alignment or attraction, we present a mechanistic insight that enables us to link individual behavior and space use patterns. As animals respond to both current and past positions of their neighbors, the assumption that the relative location of individuals is statistically and history independent is not tenable, underscoring the limitations of traditional space use studies. We move beyond that assumption by constructing a framework to analyze spatial segregation of mobile animals when neighbor proximity may elicit a retreat, and by linking conspecific encounter rate to history-dependent avoidance behavior. Our approach rests on the knowledge that animals communicate by modifying the environment in which they live, providing a method to analyze social cohesion as stigmergy, a form of mediated animal-animal interaction. By considering a population of animals that mark the terrain as they move, we predict how the spatiotemporal patterns that emerge depend on the degree of stigmergy of the interaction processes. We find in particular that nonlocal decision rules may generate a nonmonotonic dependence of the animal encounter rate as a function of the tendency to retreat from locations recently visited by other conspecifics, which has fundamental implications for epidemic disease spread and animal sociality.


O-fucosylation of muscle agrin determines its ability to cluster acetylcholine receptors.
Protein O-fucosyltransferase 1 (Pofut1) transfers fucose to serine or threonine on proteins, including Notch receptors, that contain EGF repeats with a particular consensus sequence. Here we demonstrate that agrin is O-fucosylated in a Pofut1-dependent manner, and that this glycosylation can regulate agrin function. Fucosylation of recombinant C45 agrin, both active (neural, z8) and inactive (muscle, z0) splice forms, was eliminated when agrin was overexpressed in Pofut1-deficient cells or by mutation of a consensus site for Pofut1 fucosylation (serine 1726 in the EGF4 domain). Loss of O-fucosylation caused a gain of function for muscle agrin such that it stimulated AChR clustering and MuSK phosphorylation in cultured myotubes at levels normally only found with the neural splice form. Deletion of Pofut1 in cultured primary myotubes and in adult skeletal muscle increased AChR aggregation. In addition, Pofut1 gene and protein expression and Pofut1 activity of the EGF4 domain of agrin were modulated during neuromuscular development. These data are consistent with a role for Pofut1 in AChR aggregation during synaptogenesis via the regulation of the synaptogenic activity of muscle agrin.


FIG4 regulates lysosome membrane homeostasis independent of phosphatase function.
FIG4 is a phosphoinositide phosphatase that is mutated in several diseases including Charcot-Marie-Tooth Disease 4J (CMT4J) and Yunis-Varon syndrome (YVS). To investigate the mechanism of disease pathogenesis, we generated Drosophila models of FIG4-related diseases. Fig4 null mutant animals are viable but exhibit marked enlargement of the lysosomal compartment in muscle cells and neurons, accompanied by an age-related decline in flight ability. Transgenic animals expressing Drosophila Fig4 missense mutations corresponding to human pathogenic mutations can partially rescue lysosomal expansion phenotypes, consistent with these mutations causing decreased FIG4 function. Interestingly, Fig4 mutations predicted to inactivate FIG4 phosphatase activity rescue lysosome expansion phenotypes, and mutations in the phosphoinositide (3) phosphate kinase Fab1 that performs the reverse enzymatic reaction also causes a lysosome expansion phenotype. Since FIG4 and FAB1 are present together in the same biochemical complex, these data are consistent with a model in which FIG4 serves a phosphatase-independent biosynthetic function that is essential for lysosomal membrane homeostasis. Lysosomal phenotypes are suppressed by genetic inhibition of Rab7 or the HOPS complex, demonstrating that FIG4 functions after endosome-to-lysosome fusion. Furthermore, disruption of the retromer complex, implicated in recycling from the lysosome to Golgi, does not lead to similar phenotypes as Fig4, suggesting that the lysosomal defects are not due to compromised retromer-mediated recycling of endolysosomal membranes. These data show that FIG4 plays a critical noncatalytic function in maintaining lysosomal membrane homeostasis, and that this function is disrupted by mutations that cause CMT4J and YVS.