Challenges and Obstacles of Long-Term Home Parenteral Nutrition.
Patients receiving long-term home parenteral nutrition (HPN) and clinicians managing their care face complex challenges not fully addressed by existing clinical practice guidelines. This review aims to increase awareness of some of the challenges encountered when managing patients receiving HPN and provide strategies for management. The ability to optimally manage these patients starts with involvement of a qualified team of clinicians, which is sometimes difficult to find. There are unique challenges related to the parenteral nutrition (PN) prescribing and compounding process that are not typically encountered with inpatient use. Clear communication is required between the HPN prescriber/team and the home infusion pharmacist to prevent errors related to misinterpretation of the order and PN product shortages. Dependency on HPN and living with chronic disease create a number of psychosocial, financial, and other lifestyle restrictions that can negatively impact a patient's quality of life. HPN nonadherence is a challenge that complicates the clinician's ability to accurately assess and make appropriate adjustments to therapy. HPN adherence may be improved by incorporating a patient-centered approach to care that allows patients to prioritize those issues most meaningful and valuable to them. Patient-centered care also encourages self-care and relies on a high level of HPN education. Clinicians are encouraged to use an interactive interview style when engaging patients to prioritize goals of care and make self-motivated decisions for change. In spite of challenges, HPN therapy has allowed patients the ability to maintain adequate nutrition and thrive in the home setting when the oral/enteral route fails.


Effectiveness of diclofenac, ketorolac and etoricoxib in the treatment of acute pain from ankle fracture.
Tissue degeneration, infection, inflammation, cancer, trauma, surgery and limb fractures all produce pain. Each of these physiological abnormalities requires a therapeutic approach different from the last. In acute pain, caused by fracture, several classes of analgesics have been utilized. These basic remedies for analgesia, however, are still confined to a small number of medications, including nonsteroidal anti-inflammatory drugs (NSAIDs), local anesthetics and opioids. In addition, most of these drugs have side effects, limiting their use in clinical practice. The purpose of this study was to compare the efficacy of three NSAIDs to relief acute pain caused by ankle fracture. Sixty subjects with ankle fracture were randomized to receive ketorolac, diclofenac, or etoricoxib, every 12 hours in a prospective, double-blind study. Forty-nine patients completed the study. The subjects' assessments of ankle pain on the visual analog scale and a Likert scale showed a significant reduction from baseline over 24 hr, regardless the treatment group. All treatments showed a similar profile in pain reduction. Etoricoxib, diclofenac and ketorolac twice daily are a rapid and effective treatment for acute pain. All the regimens were well tolerated in this study.


NUP98/NSD1 characterizes a novel poor prognostic group in acute myeloid leukemia with a distinct HOX gene expression pattern.
Translocations involving nucleoporin 98kD (NUP98) on chromosome 11p15 occur at relatively low frequency in acute myeloid leukemia (AML) but can be missed with routine karyotyping. In this study, high-resolution genome-wide copy number analyses revealed cryptic NUP98/NSD1 translocations in 3 of 92 cytogenetically normal (CN)-AML cases. To determine their exact frequency, we screened > 1000 well-characterized pediatric and adult AML cases using a NUP98/NSD1-specific RT-PCR. Twenty-three cases harbored the NUP98/NSD1 fusion, representing 16.1% of pediatric and 2.3% of adult CN-AML patients. NUP98/NSD1-positive AML cases had significantly higher white blood cell counts (median, 147 × 10⁹/L), more frequent FAB-M4/M5 morphology (in 63%), and more CN-AML (in 78%), FLT3/internal tandem duplication (in 91%) and WT1 mutations (in 45%) than NUP98/NSD1-negative cases. NUP98/NSD1 was mutually exclusive with all recurrent type-II aberrations. Importantly, NUP98/NSD1 was an independent predictor for poor prognosis; 4-year event-free survival was < 10% for both pediatric and adult NUP98/NSD1-positive AML patients. NUP98/NSD1-positive AML showed a characteristic HOX-gene expression pattern, distinct from, for example, MLL-rearranged AML, and the fusion protein was aberrantly localized in nuclear aggregates, providing insight into the leukemogenic pathways of these AMLs. Taken together, NUP98/NSD1 identifies a previously unrecognized group of young AML patients, with distinct characteristics and dismal prognosis, for whom new treatment strategies are urgently needed.


A veni-venomotor response to local congestion
Using the occluded venous segment technique a venoconstrictor response to local congestion was demonstrated in human limbs. Though the response appears to be dependent upon nerves, it may or may not involve the central nervous system. Results are discussed in terms of the mechanism and functional significance of the local vascular ℌreflex.ℍ


Involvement of dynamin-related protein 1 in free fatty acid-induced INS-1-derived cell apoptosis.
Elevated extracellular free fatty acids (FFAs) can induce pancreatic beta cell apoptosis, thereby contributing to the pathogenesis of type 2 diabetes mellitus (T2D). Mitochondrial dysfunction has been implicated in FFA-induced beta cell apoptosis. However, molecular mechanisms linking mitochondrial dysfunction and FFA-induced beta cell apoptosis are not clear. Dynamin-related protein 1 (DRP-1) is a mitochondrial fission modulator. In this study, we investigated its role in FFA-induced INS-1 beta cell apoptosis. DRP-1 protein was promptly induced in INS-1 cells and rat islets after stimulation by FFAs, and this DRP-1 upregulation was accompanied by increased INS-1 cell apoptosis. Induction of DRP-1 expression significantly promoted FFA-induced apoptosis in DRP-1 WT (DRP-1 wild type) inducible INS-1-derived cell line, but not in DRP-1K38A (a dominant negative mutant of DRP-1) inducible INS-1-derived cell line. To validate these in vitro results, we transplanted DRP-1 WT or DRP-1 K38A cells into renal capsules of streptozotocin (STZ)-treated diabetic mice to study the apoptosis in xenografts. Consistent with the in vitro results, the over-expression of DRP-1 led to aggravated INS-1-derived cell apoptosis triggered by FFAs. In contrast, dominant-negative suppression of DRP-1 function as represented by DRP-1 K38A significantly prevented FFA-induced apoptosis in xenografts. It was further demonstrated that mitochondrial membrane potential decreased, while cytochrome c release, caspase-3 activation, and generation of reactive oxygen species (ROS) were enhanced by the induction of DRP-1WT, but prevented by DRP-1 K38A in INS-1-derived cells under FFA stimulation. These results indicated that DRP-1 mediates FFA-induced INS-1-derived cell apoptosis, suggesting that suppression of DRP-1 is a potentially useful therapeutic strategy for protecting against beta cell loss that leads to type 2 diabetes.


Total body and hepatic clearance in rats of recombinant tissue-type plasminogen activator expressed in mouse C127 and Chinese hamster ovary cells.
The total body and hepatic clearance of two recombinant human tissue-type plasminogen activators (t-PA) produced in mouse C127 cells [t-PA(C127)] and in Chinese hamster ovary cells [t-PA(CHO)] were determined in rats. Recombinant t-PA was administered as a constant iv or intraportal venous infusion for 60 min. The plasma t-PA level over 120 min was measured by an ELISA. Whole-body pharmacokinetics were characterized in terms of the total body clearance (CLt), total volume of distribution, and mean residence time in the body. Hepatic disposition was described by the hepatic clearance (CL(hep)) and the mean hepatoportal transit time. Whereas a marked hepatic extraction was observed with both t-PAs, the estimated CL(hep) accounted for only one-half the Clt. No elimination took place in the lung, because no difference was observed between iv and intraarterial administration. Extrahepatic elimination, such as that in circulating blood, appeared to play a significant role in the disposition of t-PA in vivo. t-PA(C127) is known to possess the Gal alpha 1-3Gal epitope in its complex-type carbohydrate chains, whereas this structure is not involved in t-PA(CHO). In the presence of Bandeiraea simplicifolia lectin I isolectin B4, which is specific to the Gal alpha 1-3Gal, the Clt of t-PA(C127) was significantly decreased, whereas that of t-PA(CHO) was unchanged. These results imply that the formation of a high molecular weight complex by this carbohydrate epitope effectively reduces the rate of catabolism of t-PA in vivo.


Heterogeneous spine loss in layer 5 cortical neurons after spinal cord injury.
A large thoracic spinal cord injury disconnects the hindlimb (HL) sensory-motor cortex from its target, the lumbar spinal cord. The fate of the synaptic structures of the axotomized cortical neurons is not well studied. We evaluated the density of spines on axotomized corticospinal neurons at 3, 7, and 21 days after the injury in adult mice expressing yellow fluorescence protein in a subset of layer 5 neurons. Spine density of the dendritic segment proximal to the soma (in layer 5) declined as early as 3 days after injury, far preceding the onset of somatic atrophy. In the distal segment (in layer 2/3), spine loss was slower and less severe than in the proximal segment. Axotomy of corticospinal axons in the brainstem (pyramidotomy) induced a comparable reduction of spine density, demonstrating that the loss is not restricted to the neurons axotomized in the thoracic spinal cord. Surprisingly, in both forms of injury, the spine density of putative non-axotomized layer 5 neurons was reduced as well. The spine loss may reflect fast rearrangements of cortical circuits after axotomy, for example, by a disconnection of HL cortical neurons from synaptic inputs that no longer provide useful information.


Monotreme glucagon-like peptide-1 in venom and gut: one gene - two very different functions.
The importance of Glucagon like peptide 1 (GLP-1) for metabolic control and insulin release sparked the evolution of genes mimicking GLP-1 action in venomous species (e.g. Exendin-4 in Heloderma suspectum (gila monster)). We discovered that platypus and echidna express a single GLP-1 peptide in both intestine and venom. Specific changes in GLP-1 of monotreme mammals result in resistance to DPP-4 cleavage which is also observed in the GLP-1 like Exendin-4 expressed in Heloderma venom. Remarkably we discovered that monotremes evolved an alternative mechanism to degrade GLP-1. We also show that monotreme GLP-1 stimulates insulin release in cultured rodent islets, but surprisingly shows low receptor affinity and bias toward Erk signaling. We propose that these changes in monotreme GLP-1 are the result of conflicting function of this peptide in metabolic control and venom. This evolutionary path is fundamentally different from the generally accepted idea that conflicting functions in a single gene favour duplication and diversification, as is the case for Exendin-4 in gila monster. This provides novel insight into the remarkably different metabolic control mechanism and venom function in monotremes and an unique example of how different selective pressures act upon a single gene in the absence of gene duplication.


Continuous assessment of hemodynamic control by complex demodulation of cardiovascular variability.
Usefulness of complex demodulation (CDM) in assessing the frequency components of cardiovascular variability was assessed and, subsequently, this technique was utilized to determine the time-dependent responses of the low-frequency (LF) and high-frequency (HF) amplitudes of heart rate and blood pressure variabilities during postural tilt. CDM provides the time-dependent changes in amplitude of a particular frequency component on a continuous basis. Analysis of simulated data showed that CDM has sufficient frequency resolution to separately measure LF and HF amplitudes with a time resolution < 15 s and that CDM is robust to alterations in the frequency of the components. Analysis of actual data during postural tilt test in 23 young healthy subjects demonstrated that the HF amplitude of heart rate, an index of cardiac parasympathetic tone, rapidly decayed with head-up tilt (P < 0.01) and increased quickly showing an overshoot with tilt back to the supine position (P < 0.01). The LF amplitude of blood pressure, an index of vasomotor sympathetic activity, showed marked rhythmic fluctuation at an interval of 48-100 s during head-up tilt (P < 0.01), synchronizing with similar fluctuation in the LF amplitude of heart rate (P < 0.01). These results suggest that CDM can be used to provide a continuous assessment of cardiovascular variability components and that the dynamic responses of autonomic circulatory control to upright posture result in a phasic modulation of LF amplitude.


Interpretation of gypsy moth frontal advance using meteorology in a conditional algorithm.
The gypsy moth, Lymantria dispar, is a non-native species that continues to invade areas in North America. It spreads generally through stratified dispersal where local growth and diffusive spread are coupled with long-distance jumps ahead of the leading edge. Long-distance jumps due to anthropogenic movement of life stages is a well-documented spread mechanism. Another mechanism is the atmospheric transport of early instars and adult males, believed to occur over short distances. However, empirical gypsy moth population data continue to support the possibility of alternative methods of long-range dispersal. Such dispersal events seemed to have occurred in the mid- to late-1990s with spread across Lake Michigan to Wisconsin. Such dispersal would be against the prevailing wind flow for the area and would have crossed a significant physical barrier (Lake Michigan). The climatology of the region shows that vigorous cyclones can result in strong easterly winds in the area at the time when early instars are present. It is hypothesized that these storms would enable individuals to be blown across the Lake and explain the appearance of new population centers observed at several locations on the western shore of Lake Michigan nearly simultaneously. A synoptic climatology model coupled with population dynamics data from the area was parameterized to show an association between transport events and population spread from 1996 to 2007. This work highlights the importance of atmospheric transport events relative to the invasion dynamics of the gypsy moth, and serves as a model for understanding this mechanism of spread in other related biological invasions.


Time-dependent regional myocardial denervation as a nonspecific response to transmyocardial laser revascularization.
It is known that denervation occurs in the regions of myocardium treated by laser transmyocardial revascularization (TMR). The purpose of this study was to determine when regional denervation occurs in the early postoperative period and whether or not it is specific to laser TMR when compared with TMR using ultrasonically activated energy. Dogs with normal myocardium underwent either holmium:yttrium-aluminum-garnet laser TMR, TMR using an ultrasonic activated surgical blade, or a thoracotomy as sham operation. The responses of mean arterial pressure to topical application of bradykinin were examined at 3 time points: before, 1 hour after, and 2 weeks after surgery. The hearts were excised for Western blot and immunohistochemical analysis. The response of mean arterial pressure to bradykinin was similarly attenuated in both TMR groups 1 hour after treatment and decreased to almost none after 2 weeks compared with pretreatment values. By comparison, the sham group showed persistent responses at both time points. Tissue tyrosine hydroxylase content of the treated area decreased significantly compared with the non-treated area in both TMR groups. Immunohistochemistry using anti-Protein Gene Product 9.5 and anti-synaptophysin antibodies showed a significant decrease in the number of positive nerve fibers in both TMR treatment groups compared with the sham group. Transmyocardial revascularization caused partial alteration in myocardial innervation immediately after TMR. Tissue responses may continue to occur for the first 2 weeks after treatment. Tissue responses may also contribute to the development of denervation regardless of the energy source in non-ischemic canine myocardium.


High Yield of Pathogenic Germline Mutations Causative or Likely Causative of the Cancer Phenotype in Selected Children with Cancer.
Purpose: In many children with cancer and characteristics suggestive of a genetic predisposition syndrome, the genetic cause is still unknown. We studied the yield of pathogenic mutations by applying whole-exome sequencing on a selected cohort of children with cancer.Experimental Design: To identify mutations in known and novel cancer-predisposing genes, we performed trio-based whole-exome sequencing on germline DNA of 40 selected children and their parents. These children were diagnosed with cancer and had at least one of the following features: (1) intellectual disability and/or congenital anomalies, (2) multiple malignancies, (3) family history of cancer, or (4) an adult type of cancer. We first analyzed the sequence data for germline mutations in 146 known cancer-predisposing genes. If no causative mutation was found, the analysis was extended to the whole exome.Results: Four patients carried causative mutations in a known cancer-predisposing gene: TP53 and DICER1 (n = 3). In another 4 patients, exome sequencing revealed mutations causing syndromes that might have contributed to the malignancy (EP300-based Rubinstein-Taybi syndrome, ARID1A-based Coffin-Siris syndrome, ACTB-based Baraitser-Winter syndrome, and EZH2-based Weaver syndrome). In addition, we identified two genes, KDM3B and TYK2, which are possibly involved in genetic cancer predisposition.Conclusions: In our selected cohort of patients, pathogenic germline mutations causative or likely causative of the cancer phenotype were found in 8 patients, and two possible novel cancer-predisposing genes were identified. Therewith, our study shows the added value of sequencing beyond a cancer gene panel in selected patients, to recognize childhood cancer predisposition. Clin Cancer Res; 24(7); 1594-603. ©2018 AACR.


Flint arrowhead embedded in a human humerus from the Bronze Age site in the Tollense valley, Germany - A high-resolution micro-CT study to distinguish antemortem from perimortem projectile trauma to bone.
The Bronze Age site in the Tollense valley, Germany, has yielded thousands of human and animal bones and a number of archaeological artifacts. Several of the human bones exhibit blunt and sharp force lesions, and the assemblage has been interpreted as representing victims of a large scale conflict. One of the earliest finds is a human humerus with an embedded flint arrowhead. Alleged signs of healing initially reported for this humerus based on clinical CT imaging were interpreted as evidence of an antemortem lesion. The present study, using micro-CT imaging, revealed that the arrowhead lesion in the humerus, contrary to the previous interpretation, shows no signs of healing. The structure previously assumed to represent a sclerotic margin around the wound canal was shown to actually represent compacted trabecular debris. Thus, our re-analysis of the specimen led to a re-classification of the arrow wound as a perimortem lesion. The findings of the present study demonstrate the value of micro-CT imaging as a non-destructive method for obtaining information on the nature of bone lesions and healing reactions critical for the reconstruction of interpersonal conflict scenarios in the past.


Comprehensive model of transport and metabolism of adenosine and S-adenosylhomocysteine in the guinea pig heart.
Regulation of blood flow and mitochondrial respiration in the heart would be clarified by improved knowledge of interstitial concentrations and cellular production rates of adenosine; however, these variables cannot be measured directly. To interpret indexes that are available, a comprehensive mathematical model was developed, based on a large body of experimental data. The model describes most of the important pathways of capillary-tissue transport and cellular metabolism of adenosine in the guinea pig heart. It includes capillary flow, solute transport between tissue regions, nonlinear enzyme kinetics for adenosine kinase and adenosine deaminase, and reversible biunireactant kinetics for S-adenosylhomocysteine hydrolase in cardiomyocytes and endothelial cells, intracellular production of adenosine via AMP hydrolysis and transmethylation, and extracellular production of adenosine. A single set of parameter values for the model was obtained in the first stage of the analysis by taking certain values directly from published sources, other values were subject to specific constraints, and other values were determined by parameter optimization. The effects of flow and endothelial metabolism on the relation between interstitial and venous adenosine concentrations were determined. The relation between myocardial adenosine production rate and S-adenosylhomocysteine accumulation in the presence of excess homocysteine was estimated. In the second stage of the analysis, the model was used to investigate the mechanism of myocardial adenosine production, without changing the parameter values. Cellular adenosine production rates were estimated by fitting measurements of venous adenosine release obtained during altered energetic conditions in experiments by different investigators. The original results showed a dissociation between measurements of cytosolic AMP concentrations and venous adenosine release. It is concluded that 1) it is essential to account for the effect of flow on interstitial and venous adenosine concentrations, since decreased flow may produce effects outwardly resembling inhibition of the enzyme 5'-nucleotidase, 2) adenosine concentrations in epicardial transudate are not in equilibrium with interstitial fluid, and 3) the rate of cellular adenosine production increases monotonically with free cytosolic concentrations of AMP during a variety of alterations in energy balance of the guinea pig heart.


Nic96p is required for nuclear pore formation and functionally interacts with a novel nucleoporin, Nup188p.
The amino-terminal domain of Nic96p physically interacts with the Nsp1p complex which is involved in nucleocytoplasmic transport. Here we show that thermosensitive mutations mapping in the central domain of Nic96p inhibit nuclear pore formation at the nonpermissive temperature. Furthermore, the carboxyterminal domain of Nic96p functionally interacts with a novel nucleoporin Nup188p in an allele-specific fashion, and when ProtA-Nup188p was affinity purified, a fraction of Nic96p was found in physical interaction. Although NUP188 is not essential for viability, a null mutant exhibits striking abnormalities in nuclear envelope and nuclear pore morphology. We propose that Nic96p is a multivalent protein of the nuclear pore complex linked to several nuclear pore proteins via its different domains.


Trends in nitrogen, phosphorus, and sediment concentrations and loads in streams draining to Lake Tahoe, California, Nevada, USA.
Lake Tahoe, a large freshwater lake of the eastern Sierra Nevada in California and Nevada, has 63 tributaries that are sources of nutrients and sediment to the lake. The Tahoe watershed is relatively small, and the surface area of the lake occupies about 38% of the watershed area (1313 km2). Only about 6% of the watershed is urbanized or residential land, and as part of a plan to maintain water clarity, wastewater is exported out of the basin. The lake's clarity has been diminishing due to algae and fine sediment, prompting development of management plans. Much of the annual discharge and nutrient load to the lake results from snowmelt in the spring and summer months. To understand the relative importance of land use, climate, forest management, and other factors affecting trends in nutrient stream concentrations and loads, a Weighted Regression on Time Discharge and Season (WRTDS) model simulated these trends over a time frame of >25 years (mid-1970s to 2017). All studied locations generally show nitrate concentration and load trending down. Ammonium concentration and load initially trended down then increased continuously after 2005. Some locations show initially decreasing orthophosphate trends, followed by small significant increases in concentration and loads starting around 2000 to 2005. Total Kjeldahl nitrogen, total phosphorus and suspended sediment mostly trended downward. Overall, the trends in various forms of nitrogen were observed at most sites irrespective of the degree of development and indicate a change in ecological conditions is affecting the nitrogen cycle throughout the watershed, most likely attributable to forest aggradation and fire suppression. Ratios of bioavailable nitrogen in the form of nitrate and ammonium to orthophosphate have also trended downward during the period of record suggesting a shift of these streams from phosphorus limited to nitrogen limited.


Fluoroquinolone Impact on Nasal Methicillin-Resistant and Methicillin-Sensitive Staphylococcus aureus Colonization Durations in Neurologic Long-Term-Care Facilities.
Staphylococcus aureus nasal carriage is a risk factor for subsequent infection. Estimates of colonization duration vary widely among studies, and factors influencing the time to loss of colonization, especially the impact of antibiotics, remain unclear. We conducted a prospective study on patients naive for S. aureus colonization in 4 French long-term-care facilities. Data on nasal colonization status and potential factors for loss of colonization were collected weekly. We estimated methicillin-resistant S. aureus (MRSA) and methicillin-sensitive S. aureus (MSSA) colonization durations using the Kaplan-Meier method and investigated factors for loss of colonization using shared-frailty Cox proportional hazards models. A total of 285 S. aureus colonization episodes were identified in 149 patients. The median time to loss of MRSA or MSSA colonization was 3 weeks (95% confidence interval, 2 to 8 weeks) or 2 weeks (95% confidence interval, 2 to 3 weeks), respectively. In multivariable analyses, the methicillin resistance phenotype was not associated with S. aureus colonization duration (P = 0.21); the use of fluoroquinolones (hazard ratio, 3.37; 95% confidence interval, 1.31 to 8.71) and having a wound positive for a nonnasal strain (hazard ratio, 2.17; 95% confidence interval, 1.15 to 4.07) were associated with earlier loss of MSSA colonization, while no factor was associated with loss of MRSA colonization. These results suggest that the methicillin resistance phenotype does not influence the S. aureus colonization duration and that fluoroquinolones are associated with loss of MSSA colonization but not with loss of MRSA colonization.


Computational strategies for understanding the nature of interaction in dioxin imprinted nanoporous trappers.
A new computational model capable of understanding the nature of interactions in signature complexes formed between the template (2,3,7,8-tetrachlorodibenzo-p dioxin (TCDD)) and the functional monomers (methacrylic acid (MAA)) using density functional theory (DFT) has been designed. The polymer precursors were optimized for geometries in polymerization media, computing the interaction energies between template molecules and functional monomers of transient pre-polymerized complexes (PPC), and structural and vibrational properties reference to theoretical infrared spectra were computed using DFT of B3LYP/6 311+G(d,p) hybrid functional method. Atom in molecule theory was used to analyze the hydrogen-bonding characteristics of PPC of MAA-TCDD. Considering the theoretical titrations conducted in a virtual solvent box, it was found that the 1:4 molar ratio was required to form the most stable PPC in a given solvent system. The electron density plots indicate strong hydrogen bonding as shown by the 2pz dominant highest occupied molecular orbital (HOMO) character that could be the preferable sites of binding for target molecule, TCDD. Considering HOMO approach, the active adsorption sites in molecularly imprinted polymer was modeled to get insight on molecular recognition property for targeted molecule, TCDD. The proposed computational protocol is simple, accurate, and novel to design the polymer and is useful to predict the properties of polymer systems than the conventional theoretical analysis of template-monomer interactions.


Effect of Timing of Fungicide Applications on Development of Rusts on Daylily, Geranium, and Sunflower.
Integrated disease management should provide the most effective means of controlling rusts on ornamental crops over time, and fungicides are an important component of an integrated rust management program. Proper timing of fungicide applications is critical for effective disease management; however, information about application timing is lacking for rusts on ornamental crops. The objective of this study was to determine how fungicides affected rust development on daylily, geranium, and sunflower plants when applied several days before or after inoculation. Five fungicides registered for use against rusts on ornamental crops were evaluated: the strobilurin azoxystrobin; three sterol biosynthesis inhibiting fungicides-myclobutanil, propiconazole, and triadimefon; and the broad spectrum protectant chlorothalonil. All five fungicides significantly reduced lesion development by rust pathogens on daylily, geranium, and sunflower plants when these compounds were applied preventatively up to 15 days before inoculation and infection with a few exceptions (e.g., propiconazole on geranium and triadimefon on sunflower). Curative activity, which resulted from fungicide application after inoculation, was observed for the three rusts with some products (azoxystrobin on all three plants and myclobutanil, propiconazole, and triadimefon on geranium) when applied up to 7 days postinoculation. In general, fungicide efficacy with several of the products decreased as the time from application to inoculation (preventative activity) or inoculation to application (curative activity) increased.


Impact of kinesiophobia on initiation of cardiac rehabilitation: a prospective cohort path analysis.
To identify factors associated with kinesiophobia (fear of movement) after cardiac hospitalisation and to assess the impact of kinesiophobia on cardiac rehabilitation (CR) initiation. Prospective cohort study. Academic Medical Centre, Department of Cardiology. We performed a prospective cohort study in cardiac patients recruited at hospital discharge. In total, 149 patients (78.5% male) with a median age of 65 years were included, of which 82 (59%) were referred for CR. We assessed kinesiophobia with the Tampa Scale for Kinesiophobia (TSK). For this study, the total score was used (range 13-52). We assessed baseline factors (demographics, cardiac disease history, questionnaire data on anxiety, biopsychosocial complexity and self-efficacy) associated with kinesiophobia using linear regression with backward elimination. For linear regression, the standardised beta (β) was reported. Prospectively, the impact of kinesiophobia on probability of CR initiation, in the first 3 months after hospital discharge (subsample referred for CR), was assessed with logistic regression. For logistic regression, the OR was reported. Moderate and severe levels of kinesiophobia were found in 22.8%. In the total sample, kinesiophobia was associated with cardiac anxiety (β=0.33, 95% CI: 0.19 to 0.48), social complexity (β=0.23, 95% CI: 0.06 to 0.39) and higher education (β=-0.18, 95% CI: -0.34 to -0.02). In those referred for CR, kinesiophobia was negatively associated with self-efficacy (β=-0.29, 95% CI: -0.47 to -0.12) and positively with cardiac anxiety (β=0.43, 95% CI: 0.24 to 0.62). Kinesiophobia decreased the probability of CR initiation (OR Range13-52 points =0.92, 95% CI: 0.85 to 0.99). In patients hospitalised for cardiovascular disease, kinesiophobia is associated with cardiac anxiety, social complexity, educational level and self-efficacy. Kinesiophobia decreased the likelihood of CR initiation with 8% per point on the TSK.


Structural analysis of protein folding by the long-chain archaeal chaperone FKBP26.
In the cell, protein folding is mediated by folding catalysts and chaperones. The two functions are often linked, especially when the catalytic module forms part of a multidomain protein, as in Methanococcus jannaschii peptidyl-prolyl cis/trans isomerase FKBP26. Here, we show that FKBP26 chaperone activity requires both a 50-residue insertion in the catalytic FKBP domain, also called 'Insert-in-Flap' or IF domain, and an 80-residue C-terminal domain. We determined FKBP26 structures from four crystal forms and analyzed chaperone domains in light of their ability to mediate protein-protein interactions. FKBP26 is a crescent-shaped homodimer. We reason that folding proteins are bound inside the large crescent cleft, thus enabling their access to inward-facing peptidyl-prolyl cis/trans isomerase catalytic sites and ipsilateral chaperone domain surfaces. As these chaperone surfaces participate extensively in crystal lattice contacts, we speculate that the observed lattice contacts reflect a proclivity for protein associations and represent substrate interactions by FKBP26 chaperone domains. Finally, we find that FKBP26 is an exceptionally flexible molecule, suggesting a mechanism for nonspecific substrate recognition.


A prospective study of gynecological cancer risk in relation to adiposity factors: cumulative incidence and association with plasma adipokine levels.
Associations of obesity and obesity-related metabolic factors (adiposity factors) with uterine corpus cancer (UCC) and ovarian cancer (OVC) risk have been described. Still, a cause-effect relationship and the underlying mediators remain unclear, particularly for low-incidence populations. We aimed to prospectively determine whether adiposity factors could predict the development of UCC and OVC in Taiwanese women. To explore the biological mediators linking adiposity factors to cancer risk, we examined the association of two adipokines, leptin and adiponectin, with the gynecological cancers. Totally, 11,258 women, aged 30-65, were recruited into the Community-Based Cancer Screening Program (CBCSP) study during 1991-1993, and were followed for UCC and OVC cases until December 31, 2011. Cox proportional hazard models were used to estimate hazard ratios (HRs). Adiposity factors and risk covariates were assessed at recruitment. Newly-developed cancer cases were determined from data in the government's National Cancer Registry and Death Certification System. For adipokienes study, a nested case-control study was conducted within the cohort. Baseline plasma samples of 40 incident gynecological cancer cases and 240 age-menopause-matched controls were assayed for adipokines levels. There were 38 and 30 incident cases of UCC and OVC, respectively, diagnosed during a median 19.9 years of follow-up. Multivariate analysis showed that alcohol intake (HR = 16.00, 95% = 4.83-53.00), high triglyceride levels (HR = 2.58, 95% = 1.28-5.17), and years of endogenous estrogen exposure per 5-year increment (HR = 1.91, 95% = 1.08-3.38) were associated with increased UCC risk. High body mass index (BMI ≥ 27 kg/m(2), HR = 2.90, 95% = 1.30-6.46) was associated with increased OVC risk. Analysis further showed an independent effect of adipokines on UCC and OVC risk after adjustment of the risk covariates. We provided evidence that alcohol intake, high triglyceride levels and long endogenous estrogen exposure increase UCC risk, whereas obesity positively predicts OVC risk. Circulating adipokines may mediate the link of adiposity factors to gynecological cancer risk.


Severe iron deficiency anemia in transgenic mice expressing liver hepcidin.
We recently reported the hemochromatosis-like phenotype observed in our Usf2 knockout mice. In these mice, as in murine models of hemochromatosis and patients with hereditary hemochromatosis, iron accumulates in parenchymal cells (in particular, liver and pancreas), whereas the reticuloendothelial system is spared from this iron loading. We suggested that this phenotypic trait could be attributed to the absence, in the Usf2 knockout mice, of a secreted liver-specific peptide, hepcidin. We conjectured that the reverse situation, namely overexpression of hepcidin, might result in phenotypic traits of iron deficiency. This question was addressed by generating transgenic mice expressing hepcidin under the control of the liver-specific transthyretin promoter. We found that the majority of the transgenic mice were born with a pale skin and died within a few hours after birth. These transgenic animals had decreased body iron levels and presented severe microcytic hypochromic anemia. So far, three mosaic transgenic animals have survived. They were unequivocally identified by physical features, including reduced body size, pallor, hairless and crumpled skin. These pleiotropic effects were found to be associated with erythrocyte abnormalities, with marked anisocytosis, poikylocytosis and hypochromia, which are features characteristic of iron-deficiency anemia. These results strongly support the proposed role of hepcidin as a putative iron-regulatory hormone. The animal models devoid of hepcidin (the Usf2 knockout mice) or overexpressing the peptide (the transgenic mice presented in this paper) represent valuable tools for investigating iron homeostasis in vivo and for deciphering the molecular mechanisms of hepcidin action.


Fucoidan Exerts Anticancer Effects Against Head and Neck Squamous Cell Carcinoma In Vitro.
Fucoidans have been reported to exert anticancer effects with simultaneous low toxicity against healthy tissue. That correlation was observed in several cancer models, however, it has never been investigated in head and neck cancer before. To magnify the efficacy of conventional therapy, the administration of agents like fucoidan could be beneficial. The aim of this study was to evaluate the anticancer effect of Fucus vesiculosus (FV) extract alone and with co-administration of cisplatin in head and neck squamous cell carcinoma (HNSCC) in vitro. MTT assay results revealed an FV-induced inhibition of proliferation in all tested cell lines (H103, FaDu, KB). Flow cytometric cell cycle analysis showed an FV-induced, dose-dependent arrest in either S/G2 phase (H103, FaDu) or G1 arrest (KB). Furthermore, a dose-dependent gain in apoptotic fraction was observed. Western blot analysis confirmed the induction of apoptosis. A significant dose-dependent increase in reactive oxygen species (ROS) production was revealed in the H103 cell line, while FaDu cells remained unresponsive. On the contrary, an HPV-positive cell line, KB, demonstrated a dose-dependent decrease in ROS synthesis. Moreover, fucoidan enhanced the response to cisplatin (synergistic effect) in all cell lines with the HPV-positive one (KB) being the most sensitive. These results have been confirmed by flow-cytometric apoptosis analysis. In conclusion, we confirmed that fucoidan exhibits anticancer properties against HNSCC, which are manifested by the induction of apoptosis, regulation of ROS production, cell cycle arrest, and inhibition of proliferation.


Twenty Thousand Needles Under the Sea. Trigger Point Dry Needling Aboard an Israeli Navy Submarine: A Case Report.
Nonspecific lower back pain affects a major part of the population at a certain point of their life. The intensity of pain can be debilitating and it causes a significant burden on society. Trigger point dry needling is a method of alleviating such pain by the introduction of needles into trigger points in muscles. A growing body of evidence supports its use in myofascial pain and specifically lower back pain. Submarine Medicine is a unique field due to the special characteristics and the environment of the submarine. It poses challenges that are not always seen by primary care physicians. Here, we present a case of a 40-yr-old senior submarine officer who complained of pain in his lower back and pelvis before departing on a mission. The pain persisted in spite of treatment with nonsteroidal anti-inflammatory drugs and he was then treated by the submarine's physician with trigger point dry needling. The officer showed rapid improvement following this treatment, both regarding pain and the range of motion.


Interactions of actin, myosin, and a new actin-binding protein of rabbit pulmonary macrophages. II. Role in cytoplasmic movement and phagocytosis.
Actin and myosin of rabbit pulmonary macrophages are influenced by two other proteins. A protein cofactor is required for the actin activation of macrophage myosin Mg2 ATPase activity, and a high molecular weight actin-binding protein aggregates actin filaments (Stossel T.P., and J.H. Hartwig. 1975. J. Biol. Chem. 250:5706-5711)9 When warmed in 0.34 M sucrose solution containing Mg2-ATP and dithiothreitol, these four proteins interact cooperatively. Acin-binding protein in the presence of actin causes the actin to form a gel, which liquifies when cooled. The myosin contracts the gel into an aggregate, and the rate of aggregation is accelerated by the cofactor. Therefore, we believe that these four proteins also effec the temperature-dependent gelation and aggregation of crude sucrose extracts pulmonary macrophages containing Mg2-ATP and dithiothreitol. The gelled extracts are composed of tangled filaments. Relative to homogenates of resting macrophages, the distribution of actin-binding protein in homogenates of phagocytizing macrophages is altered such that 2-6 times more actin-binding protein is soluble. Sucrose extracts of phagocytizing macrophages gel more rapidly than extracts of resting macrophages. Phagocytosis by pulmonary macrophages involves the formation of peripheral pseudopods containing filaments. The findings suggest that the actin-binding protein initiates a cooperative interaction of contractile proteins to generate cytoplasmic gelation, and that phagocytosis influences the behavior of the actin-binding protein.


Developing a set of patient-centered outcomes for routine use in endometriosis: An international Delphi study.
There is large variation in individual patient care for endometriosis. A uniform approach to measure outcomes could be incorporated into routine clinical practice to personalize and monitor treatments and potentially improve the quality of care. The aim of this study is to identify a group of patient-centered outcomes for use in routine endometriosis care which are relevant to all patient profiles. By means of a modified two-round Delphi study with international representation including healthcare professionals, researchers and patient representatives (51 participants, 16 countries) we developed a set of patient-centered measurements. The participants evaluated 47 Patient Reported Outcome Measures (PROMs) and 30 Clinician Reported Outcome Measures (CROMs) regarding their feasibility and relevance for their use in routine endometriosis care. After the two rounds of quotation, meetings of the experts were convened to participate in a final discussion to finalize the consensus of the final set of included measures. The final set of patient-centered outcomes includes six PROMs (measuring symptomatic impact, pain, work productivity and quality of life) and 10 CROMs (measuring clinical, imaging and surgical indicators). A supplementary list of outcomes was added to include important dimensions that were considered essential by the expert panel but are not relevant to all patients. In addition the need for development of specific tools (PROMs) measuring the psychological impact and the impact in sexual activity of endometriosis was highlighted. We have developed a set of patient-centered outcomes measures in endometriosis care. The selected outcomes comprise the common features for all patients suffering from endometriosis. adapted for use in routine practice. The list of outcomes has been adapted for use in routine practice from which clinicians can chose, depending on their needs.


Acidity promotes degradation of multi-species environmental DNA in lotic mesocosms.
Accurate quantification of biodiversity is fundamental to understanding ecosystem function and for environmental assessment. Molecular methods using environmental DNA (eDNA) offer a non-invasive, rapid, and cost-effective alternative to traditional biodiversity assessments, which require high levels of expertise. While eDNA analyses are increasingly being utilized, there remains considerable uncertainty regarding the dynamics of multispecies eDNA, especially in variable systems such as rivers. Here, we utilize four sets of upland stream mesocosms, across an acid-base gradient, to assess the temporal and environmental degradation of multispecies eDNA. Sampling included water column and biofilm sampling over time with eDNA quantified using qPCR. Our findings show that the persistence of lotic multispecies eDNA, sampled from water and biofilm, decays to non-detectable levels within 2 days and that acidic environments accelerate the degradation process. Collectively, the results provide the basis for a predictive framework for the relationship between lotic eDNA degradation dynamics in spatio-temporally dynamic river ecosystems.


Recombinant human erythropoietin for treating treatment-resistant depression: a double-blind, randomized, placebo-controlled phase 2 trial.
Pharmacological treatments for depression have insufficient efficacy in 30-40% of patients and fail to reverse cognitive deficits. Erythropoietin (EPO) has neurotrophic actions and aids neurocognitive function. The aim of this exploratory study was to determine whether recombinant human EPO improves mood and memory in treatment-resistant depression. Forty treatment-resistant depressed unipolar patients with Hamilton Depression Rating Scale-17 (HDRS-17) score ≥ 17 were randomized to eight weekly EPO (Eprex; 40,000 IU) or saline infusions in a double-blind, placebo-controlled, parallel-group design. Patients were assessed at baseline and at weeks 5, 9, and 14. Primary outcome was reduction in HDRS-17 score. Global assessment of function (GAF) was reported in addition. Secondary outcome was remission rate, and tertiary outcomes were changes in Rey Auditory Verbal Learning Test (RAVLT), Beck Depression Inventory-21 (BDI-21), and World Health Organization Quality of life-BREF (WHOQOL-BREF). Exploratory outcomes were depression and cognition composite scores. HDRS-17, GAF, and remission rates showed no effects of EPO over saline at week 9 (P-value ≥ 0.09). However, EPO improved BDI (P=0.02) and WHOQOL-BREF (P=0.01), and this was maintained at follow-up week 14 (P-values ≤ 0.04). EPO enhanced verbal recall (P=0.02) and recognition (P=0.03), which was sustained at follow-up (P-values ≤ 0.04). Exploratory analysis in patients fulfilling depression severity criteria at trial start revealed ameliorated HDRS-17 in EPO (N=14) vs saline groups (N=17), which was sustained at week 14 (P-values ≤ 0.05). Exploratory analysis in the complete cohort showed that EPO reduced depression composite at weeks 9 and 14 (P-values=0.02). The findings of this exploratory study highlight EPO as an interesting compound for treatment-resistant depression, which deserves further investigation.


Population structure drives cultural diversity in finite populations: A hypothesis for localized community patterns on Rapa Nui (Easter Island, Chile).
Understanding how and why cultural diversity changes in human populations remains a central topic of debate in cultural evolutionary studies. Due to the effects of drift, small and isolated populations face evolutionary challenges in the retention of richness and diversity of cultural information. Such variation, however, can have significant fitness consequences, particularly when environmental conditions change unpredictably, such that knowledge about past environments may be key to long-term persistence. Factors that can shape the outcomes of drift within a population include the semantics of the traits as well as spatially structured social networks. Here, we use cultural transmission simulations to explore how social network structure and interaction affect the rate of trait retention and extinction. Using Rapa Nui (Easter Island, Chile) as an example, we develop a model-based hypothesis for how the structural constraints of communities living in small, isolated populations had dramatic effects and likely led to preventing the loss of cultural information in both community patterning and technology.


Expression of messenger ribonucleic acids of luteinizing hormone and follicle-stimulating hormone receptors in granulosa and theca layers of chicken preovulatory follicles.
Expression of luteinizing hormone receptor (LHR) and follicle-stimulating hormone receptor (FSHR) mRNAs was demonstrated in the granulosa and theca layers of the large preovulatory follicles of the chicken ovary by Northern hybridization and reverse transcription-polymerase chain reaction (RT-PCR). Northern hybridization results showed multiple LHR and FSHR mRNA transcripts and the predominant species were 2.3 and 2.5 kb, respectively. The highest abundance of LHR mRNA was found in the granulosa layer of the largest follicle (F1 follicle), while the abundance remained low in the granulosa layers of the third (F3) and fifth largest (F5) follicles. FSH mRNA abundance was the highest in the granulosa layer of F5 follicle, but decreased in the granulosa layers of F3 and F1 follicles. In the theca layers of all the three follicles examined LHR and FSHR mRNAs were extremely low. These results were confirmed by RT-PCR experiment which involved coamplification of LHR or FSHR mRNA and beta-actin mRNA as the internal control. The LHR PCR product was sequenced and indicated 92.2% homology with the corresponding region of the quail LHR cDNA. This study indicated that the marked increase in expression of LHR mRNA in granulosa layer of the F1 follicle might be important for LHR protein synthesis and succeeding bonus progesterone production in F1 follicle destined to ovulation. However, higher expression of FSHR mRNA in the granulosa layer of the less mature follicles may be involved in the differentiation and maturation of granulosa cells in these follicles.


Multidisciplinary vascular malformations clinic in Hong Kong.
To review clinical characteristics, imaging modalities, and treatment outcomes of patients referred to a multidisciplinary clinic for management of vascular malformations. Retrospective case series. Multidisciplinary vascular malformation out-patient referral clinic in a teaching hospital in Hong Kong. The 141 attendees of the clinic from August 2005 to November 2011. Management and treatments offered, and responses to treatment. Of the 141 patients, 46% were diagnosed to have low-flow vascular malformations, 16% were diagnosed to have high-flow vascular malformations, and 15% were diagnosed to have a haemangioma. Prior to attending the clinic, approximately one third (32%) of the patients had a clinical diagnosis that was consistent with the final diagnosis. Overall, the radiological and clinical diagnoses were consistent in 43% of the patients. Magnetic resonance imaging and ultrasonography were the most commonly used imaging modalities. Of the 73 patients who received active treatment, 70% had a good response, 12% had minimal improvement, 8% had no change, and 7% had a recurrence or a major complication; in 3% of the patients the outcome was unknown. From this retrospective case series, it is evident that confusion still exists over vascular malformations and haemangiomas. Multidisciplinary clinics have a role in providing an accurate diagnosis and facilitating appropriate management and treatment plans. Magnetic resonance imaging and ultrasonography had demonstrable utility in determining the extent of the lesions and flow type.


GLUCOMANNAN AND GLUCOMANNAN PLUS SPIRULINA-ENRICHED SQUID-SURIMI ADDED TO HIGH SATURATED DIET AFFECT GLYCEMIA, PLASMA AND ADIPOSE LEPTIN AND ADIPONECTIN LEVELS IN GROWING FA/FA RATS.
Type 2 diabetes is a very prevalent chronic disease. Among dietary factors for its prevention and treatment, interest has grown in satiating fibre (konjac glucomannan) and spirulina. Our previous studies suggest that glucomannan itself and/or in conjunction to spirulina displayed hypolipemic and antioxidant effects when incorporated to squid surimi as functional ingredients. The present study aims to determine whether glucomannan- enriched or glucomannan plus spirulina-enriched squid-surimi improve plasma glucose and insulin levels in Zucker fa/fa rats fed a high saturated fat diet. Twenty four growing rats, divided into three groups, were given modified AIN-93M diets for seven weeks: 30% squid-surimi control diet (C), 30% glucomannan-enriched squid-surimi diet (G) and 30% glucomannan plus spirulina-enriched squid-surimi diet (GS). All rats became hyperglycemics and hyperinsulinemics, but G and GS diets induced significantly lower glucose levels (20%; p < 0.05) but did not modify insulinemia with respect to C diet. GS animals showed higher HOMA-D (p < 0.05) than C ones suggesting increased insulin availability. Plasma leptin and adiponectin decreased in G and GS vs. C group (p < 0.05). Adipose adiponectin increased significantly in G and GS vs. C rats (16-20 times, p < 0.01). Leptin in adipose tissue was higher in GS vs. G group (p < 0.05). In conclusion, both glucomannan-diets were able to reduce hyperglycemia and increase adipose tissue adiponectin levels in fa/fa rats, suggesting an anti-hypertrophic and insulin-sensitizing adipokine effect in this tissue. Spirulina inclusion increased insulin availability. Although results are promising, the utility of consuming glucomannan surimis as part of usual diets demands future studies.


Defining the antigenic diversity of Plasmodium falciparum apical membrane antigen 1 and the requirements for a multi-allele vaccine against malaria.
Apical Membrane Antigen 1 (AMA1) is a leading malaria vaccine candidate and a target of naturally-acquired human immunity. Plasmodium falciparum AMA1 is polymorphic and in vaccine trials it induces strain-specific protection. This antigenic diversity is a major roadblock to development of AMA1 as a malaria vaccine and understanding how to overcome it is essential. To assess how AMA1 antigenic diversity limits cross-strain growth inhibition, we assembled a panel of 18 different P. falciparum isolates which are broadly representative of global AMA1 sequence diversity. Antibodies raised against four well studied AMA1 alleles (W2Mef, 3D7, HB3 and FVO) were tested for growth inhibition of the 18 different P. falciparum isolates in growth inhibition assays (GIA). All antibodies demonstrated substantial cross-inhibitory activity against different isolates and a mixture of the four different AMA1 antibodies inhibited all 18 isolates tested, suggesting significant antigenic overlap between AMA1 alleles and limited antigenic diversity of AMA1. Cross-strain inhibition by antibodies was only moderately and inconsistently correlated with the level of sequence diversity between AMA1 alleles, suggesting that sequence differences are not a strong predictor of antigenic differences or the cross-inhibitory activity of anti-allele antibodies. The importance of the highly polymorphic C1-L region for inhibitory antibodies and potential vaccine escape was assessed by generating novel transgenic P. falciparum lines for testing in GIA. While the polymorphic C1-L epitope was identified as a significant target of some growth-inhibitory antibodies, these antibodies only constituted a minor proportion of the total inhibitory antibody repertoire, suggesting that the antigenic diversity of inhibitory epitopes is limited. Our findings support the concept that a multi-allele AMA1 vaccine would give broad coverage against the diversity of AMA1 alleles and establish new tools to define polymorphisms important for vaccine escape.


Structure, specificity, and mode of interaction for bacterial albumin-binding modules.
We have determined the solution structure of an albumin binding domain of protein G, a surface protein of group C and G streptococci. We find that it folds into a left handed three-helix bundle similar to the albumin binding domain of protein PAB from Peptostreptococcus magnus. The two domains share 59% sequence identity, are thermally very stable, and bind to the same site on human serum albumin. The albumin binding site, the first determined for this structural motif known as the GA module, comprises residues spanning the first loop to the beginning of the third helix and includes the most conserved region of GA modules. The two GA modules have different affinities for albumin from different species, and their albumin binding patterns correspond directly to the host specificity of C/G streptococci and P. magnus, respectively. These studies of the evolution, structure, and binding properties of the GA module emphasize the power of bacterial adaptation and underline ecological and medical problems connected with the use of antibiotics.


Enzymatic synthesis of a quorum-sensing autoinducer through use of defined substrates.
Many bacteria, including several pathogens of plants and humans, use a pheromone called an autoinducer to regulate gene expression in a cell density-dependent manner. Agrobacterium autoinducer [AAI, N-(3-oxo-octanoyl)-L-homoserine lactone] of A. tumefaciens is synthesized by the Tral protein, which is encoded by the tumor-inducing plasmid. Purified hexahistidinyl-Tral (H6-Tral) used S-adenosylmethionine to make the homoserine lactone moiety of AAI, but did not use related compounds. H6-Tral used 3-oxo-octanoyl-acyl carrier protein to make the 3-oxo-octanoyl moiety of AAI, but did not use 3-oxo-octanoyl-coenzyme A. These results demonstrate the enzymatic synthesis of an autoinducer through the use of purified substrates.


Ischemic lesion volumes in acute stroke by diffusion-weighted magnetic resonance imaging correlate with clinical outcome.
Diffusion-weighted magnetic resonance imaging detects ischemic injury within minutes after onset, and has been used to demonstrate drug efficacy in animal models of stroke. In 50 patients diagnosed with acute ischemic stroke (<24-hour duration) within the middle cerebral artery territory, lesion volume was measured by diffusion-weighted imaging. Thirty-four patients also had volumes measured by T2-weighted imaging chronically (median time, 7.5 weeks; mean, 15.9 weeks). Clinical severity was measured by the National Institutes of Health Stroke Scale Score and the Barthel index. Acute lesion volumes correlated with the acute stroke scale score (r = 0.56), the chronic stroke scale score (r = 0.63), and chronic lesion volumes (r = 0.84). Chronic volumes correlated with the chronic stroke scale score (r = 0.86) and the Barthel index (r = -0.60). When only cortically based lesions were considered, the correlations relating acute lesion volume measured by diffusion-weighted imaging (r = 0.61) and chronic lesion volume measured by T2-weighted imaging (r = 0.90) to the chronic stroke scale score were higher. These results provide evidence that lesion volumes determined by diffusion-weighted imaging acutely may be predictive of clinical severity and outcome, and may support a role for diffusion-weighted imaging in the assessment of acute stroke therapies in clinical trials.


Ethanol reward and aversion in mice bred for sensitivity to ethanol withdrawal.
The present study examined mice selectively bred for sensitivity to ethanol withdrawal for differences in the conditioned place preference (CPP) and conditioned taste aversion (CTA) paradigms. Withdrawal Seizure-Prone (WSP) and Withdrawal Seizure-Resistant (WSR) mice and High Alcohol Withdrawal (HAW) and Low Alcohol Withdrawal (LAW) mice were selectively bred for differences in chronic and acute ethanol withdrawal, respectively. For the CPP experiment, male HAW and LAW (generation 5) mice received four pairings of ethanol (2 g/kg), with a distinctive floor stimulus. On intervening days, mice received saline paired with an alternate floor type. During the preference test, all mice received an injection of saline before 60-min access to both floor types. HAW mice showed conditioned preference for the ethanol-paired floor, whereas LAW mice did not. For the CTA experiments, male HAW, LAW, WSP, and WSR mice were adapted to a 2-hr/day water restriction regimen and subsequently received ethanol injections (0, 2, 2.5, or 4 g/kg, i.p.) immediately after 1-hr access to a NaCl-flavored solution. Dose-dependent reductions in NaCl intake reflected the development of CTA in both HAW/LAW and WSP/WSR lines. However, a smaller magnitude of CTA was observed in WSP mice relative to WSR mice after the first ethanol-NaCl pairing. WSP/WSR mice showed similar reductions of NaCl intake on subsequent conditioning trials. Overall, these data suggest that HAW mice selectively bred for high sensitivity to acute ethanol withdrawal are more sensitive to the rewarding effects of ethanol in the CPP paradigm. This outcome is consistent with a previous study showing greater CPP in WSP mice relative to WSR mice. In the CTA paradigm, sensitivity to ethanol withdrawal in the HAW/ LAW selected lines does not appear to be genetically correlated with sensitivity to the aversive properties of ethanol. However, the difference in acquisition of CTA in WSP/WSR lines suggest that some genes determining ethanol withdrawal severity may also influence initial sensitivity to ethanol's aversive effects.


Toward a quantitative estimate of future heat wave mortality under global climate change.
Climate change is anticipated to affect human health by changing the distribution of known risk factors. Heat waves have had debilitating effects on human mortality, and global climate models predict an increase in the frequency and severity of heat waves. The extent to which climate change will harm human health through changes in the distribution of heat waves and the sources of uncertainty in estimating these effects have not been studied extensively. We estimated the future excess mortality attributable to heat waves under global climate change for a major U.S. city. We used a database comprising daily data from 1987 through 2005 on mortality from all nonaccidental causes, ambient levels of particulate matter and ozone, temperature, and dew point temperature for the city of Chicago, Illinois. We estimated the associations between heat waves and mortality in Chicago using Poisson regression models. Under three different climate change scenarios for 2081-2100 and in the absence of adaptation, the city of Chicago could experience between 166 and 2,217 excess deaths per year attributable to heat waves, based on estimates from seven global climate models. We noted considerable variability in the projections of annual heat wave mortality; the largest source of variation was the choice of climate model. The impact of future heat waves on human health will likely be profound, and significant gains can be expected by lowering future carbon dioxide emissions.


An ancestral core haplotype defines the critical region harbouring the North Carolina macular dystrophy gene (MCDR1).
Autosomal dominant North Carolina macular dystrophy (NCMD) or central areolar pigment epithelial dystrophy (CAPED) is an allelic disorder that maps to an approximately 7.2 cM interval between DNA markers at D6S424 and D6S1671 on 6q14-q16.2. The further refinement of the disease locus has been hindered by the lack of additional recombination events involving the critical region. In this study, we have identified three multigeneration families of German descent who express the NCMD phenotype. Genotyping was carried out with a series of markers spanning approximately 53 cM around the NCMD locus, MCDR1. Genetic linkage between the markers and the disease phenotype in each of the families could be shown. Disease associated haplotypes were constructed and provide evidence for an ancestral founder for the German NCMD families. This haplotype analysis suggests that a 4.0 cM interval flanked by markers at D6S249 and D6S475 harbours the gene causing NCMD, facilitating further positional cloning approaches.


Control of vertebrate Hox clusters by remote and global cis-acting regulatory sequences.
Despite apparently shared structural organisation and functional roles, vertebrate Hox genes are controlled by regulatory mechanisms rather distinct from those of the prototypic Drosophila Antennapedia (ANT-C) and Bithorax (BX-C) Complexes. If individual regulatory modules have been shown to recapitulate specific Hox expression patterns, other experimental studies underscore that vertebrate Hox clusters are controlled in many of their functions in a global manner, through distinct mechanisms. We will discuss the different models that have been proposed to account for these global regulatory modes. In this context, the studies of the regulation of the HoxD complex during limb development highlighted the role of global regulatory elements and the different mechanisms associated to transform a structural organisation into distinct temporal and spatial expression domains. We will further discuss how these mechanisms may have benefited from the structure of the vertebrate homeotic clusters and reciprocally contribute to shape their evolution towards an increased level of organisation and compaction.


On-site test for cannabinoids in oral fluid.
Oral fluid (OF) testing offers noninvasive sample collection for on-site drug testing; however, to date, test performance for Δ(9)-tetrahydrocannabinol (THC) detection has had unacceptable diagnostic sensitivity. On-site tests must accurately identify cannabis exposure because this drug accounts for the highest prevalence in workplace drug testing and driving under the influence of drugs (DUID) programs. Ten cannabis smokers (9 males, 1 female) provided written informed consent to participate in this institutional review board-approved study and smoked 1 6.8%-THC cigarette ad libitum. OF was collected with the Draeger DrugTest(®) 5000 test cassette and Quantisal™ device 0.5 h before and up to 22 h after smoking. Test cassettes were analyzed within 15 min (n = 66), and Quantisal GC-MS THC results obtained within 24 h. Final THC detection times and test performances were assessed at different cannabinoid cutoffs. Diagnostic sensitivity, diagnostic specificity, and efficiency at DrugTest 5000's 5 μg/L screening cutoff and various THC confirmation cutoffs were 86.2-90.7, 75.0-77.8, and 84.8-87.9%, respectively. Last detection times were >22 h, longer than previously suggested. Confirmation of 11-nor-9-carboxy-THC, absent in THC smoke, minimized the potential for passive OF contamination and still provided 22-h windows of detection, appropriate for workplace drug testing, whereas confirmation of cannabidiol, and/or cannabinol yielded shorter 6-h windows of detection, appropriate for DUID OF testing. The DrugTest 5000 on-site device provided high diagnostic sensitivity for detection of cannabinoid exposure, and the selection of OF confirmation analytes and cutoffs provided appropriate windows of detection to meet the goals of different drug testing programs.


Biological Maturation and Physical Activity in South Korean Adolescent Girls.
To examine whether body fatness (i.e., body mass index and percent body fat) and psychosocial correlates of physical activity (PA) mediate the relationships between pubertal status and PA and sedentary behavior (SB) among South Korean adolescent girls. Pubertal status, body fatness, sport competence, perceived barriers to PA, self-efficacy, parental support for PA, self-reported and step-determined PA, and self-reported SB were assessed among 236 South Korean adolescent girls (mean age = 13.56 ± 1.2 yr). A direct association was identified between pubertal status and PA in all models; more advanced pubertal status predicted lower PA among South Korean adolescents (b = -0.51 to -0.57; P < 0.001). Indirect effects on the pubertal status-PA relationship were also found: body fatness, perceived barriers to PA, and self-efficacy mediated the relationship between pubertal status and PA. Parental support for PA did not moderate the relationship between maturation and PA. SB was not associated with pubertal status. This study found mediated effects of body fatness, and psychosocial correlates (i.e., perceived barriers to PA, self-efficacy) on the relationship between maturity and PA among South Korean adolescent girls. Efforts to promote PA among South Korean adolescent girls should focus on reducing perceived barriers to PA, and increasing self-efficacy by providing more opportunities to have positive, and enjoyable experiences in PA among early-maturing girls.


Conformational properties of the G.G mismatch in d(CGCGAATTGGCG)2 determined by NMR.
The conformational properties of the DNA duplex d(CGCGAATTGGCG)2, which contains two noncomplementary G.G base pairs, have been examined in aqueous solution by 1H and 31P NMR as a function of temperature. The G.G mismatch is highly destabilizing, with a Tm value 35 K below that observed for the native EcoRI dodecamer. The dodecamer appears symmetric in the NMR spectra and exists largely as an average B-type DNA conformation. However, the 1H and 31P NMR spectra give evidence of considerable conformational heterogeneity at the mismatched nucleotides and their nearest neighbors, which increases with increasing temperature. There is no evidence for a significant population of the syn purine conformation. The imino protons of the mispaired bases G4 and G9 are degenerate, resonate at high field, and exchange readily with solvent. These results indicate that the mispaired bases are only weakly hydrogen-bonded and are only partially stacked into the helix. On raising the temperature, the duplex shows increasing exchange between two or more conformations originating from the mismatch sites. However, these additional conformations maintain their Watson-Crick hydrogen bonding. The increase in chemical exchange is consistent with a quasimelting process for which the G.G sites provide local nuclei. Extensive modeling studies by dynamic annealing have confirmed that the G(anti).G(anti) conformation is favored and that the mispairs are poorly stacked within the helix. The results explain both the poor thermal stability and low hypochromicity of this duplex.


Recommendations From the International Colorectal Cancer Screening Network on the Evaluation of the Cost of Screening Programs.
Worldwide, colorectal cancer is the fourth leading cause of death from cancer and the incidence is projected to increase. Many countries are exploring the introduction of organized screening programs, but there is limited information on the resources required and guidance for cost-effective implementation. To facilitate the generating of the economics evidence base for program implementation, we collected and analyzed detailed program cost data from 5 European members of the International Colorectal Cancer Screening Network. The cost per person screened estimates, often used to compare across programs as an overall measure, varied significantly across the programs. In addition, there were substantial differences in the programmatic and clinical cost incurred, even when the same type of screening test was used. Based on these findings, several recommendations are provided to enhance the underlying methodology and validity of the comparative economic assessments. The recommendations include the need for detailed activity-based cost information, the use of a comprehensive set of effectiveness measures to adequately capture differences between programs, and the incorporation of data from multiple programs in cost-effectiveness models to increase generalizability. Economic evaluation of real-world colorectal cancer-screening programs is essential to derive valuable insights to improve program operations and ensure optimal use of available resources.


Client-perpetrated and husband-perpetrated violence among female sex workers in Andhra Pradesh, India: HIV/STI risk across personal and work contexts.
This study examines violence experienced in work and personal contexts and relation to HIV risk factors in these contexts among female sex workers (FSW) in Andhra Pradesh, India. FSW at least 18 years of age (n=2335) were recruited through three rounds of respondent-driven sampling between 2006 and 2010 for a survey on HIV risk. Using crude and adjusted logistic regression models, any sexual/physical violence (last 6 months) perpetrated by clients and husbands were separately assessed in association with accepting more money for sex without a condom (last 30 days), consistent condom use with clients and husbands (last 30 days), and sexually transmitted infection (STI) symptoms (last 6 months). The mean age among participants was 32, 22% reported being currently married, and 22% and 21% reported physical/sexual violence by clients and husbands, respectively. In adjusted logistic regression models, FSW who experienced client violence were more likely to report accepting more money for unprotected sex trades (adjusted OR (AOR)=1.7; 95% CI 1.4 to 2.2), less likely to report consistent condom use with clients (AOR=0.6; 95% CI 0.5 to 0.7) and more likely to report STI symptoms (AOR=3.5; 95% CI 2.6 to 4.6). Women who reported husband violence were more likely to report accepting more money for unprotected sex trades (AOR=2.1; 95% CI 1.2 to 3.7), less likely to report consistent condom use with clients (AOR=0.5; 95% CI 0.3 to 0.8) and more likely to report STI symptoms (AOR=2.6; 95% CI 1.6 to 4.1). Among FSW, experiences of violence in work and personal contexts are associated with sexual HIV risk behaviours with clients as well as STI symptoms.


Phase II study of weekly vinblastine in recurrent or refractory pediatric low-grade glioma.
To evaluate the efficacy of single-agent vinblastine in pediatric patients with recurrent or refractory low-grade glioma. Patients were eligible if they had experienced previous treatment failure (chemotherapy and/or radiation) for incompletely resected or unresectable low-grade glioma (LGG). Vinblastine (6 mg/m(2)) was administered weekly for 1 year unless unacceptable toxicity or progression (confirmed on two consecutive imaging studies) occurred. Fifty-one patients (age range, 1.4 to 18.2 years; median age, 7.2 years) were prospectively enrolled onto this phase II study. Fifty patients had previously received at least one prior regimen of chemotherapy, and 10 patients had previously received radiation treatment. Fifty patients were evaluable for response; 18 patients (36%) had a complete, partial, or minor response, and 31 patients completed 1 year of treatment. At a median follow-up of 67 months, 23 patients had not experienced progression; three patients have died. Five-year overall survival was 93.2% ± 3.8%, and 5-year progression-free survival was 42.3% ± 7.2%. Toxicity was manageable and mostly hematologic, although a few patients needed transfusions. Weekly vinblastine seems to be a reasonable alternative to radiation for pediatric patients with LGG who have experienced treatment failure with first-line chemotherapy. The 5-year progression-free survival observed in this phase II trial is comparable to results observed with first-line chemotherapy in chemotherapy-naive patients. The role of single-agent vinblastine and other vinca alkaloid in the management of pediatric LGGs deserves further investigation.


Effects of capture stress on free-ranging, reproductively active male Weddell seals.
Physiological stress responses to capture may be an indicator of welfare challenges induced by animal handling. Simultaneously, blood chemistry changes induced by stress responses may confound experimental design by interacting with the biological parameters being measured. Cortisol elevation is a common indicator of stress responses in mammals and reproductive condition can profoundly influence endocrine response. We measured changes in blood cortisol and testosterone induced by handling reproductively active male Weddell seals (Leptonychotes weddellii) early and late in the breeding season. Weddell seals have the highest resting cortisol levels of all mammals yet showed a clear, prolonged elevation in cortisol in response to capture. Responses were similar when first caught and when caught a second time, later in the breeding season. Baseline testosterone levels declined over the breeding season but were not altered by capture. Administering a light dose of diazepam significantly ameliorated the cortisol response of handled animals without affecting testosterone levels. This may be an effective way of reducing acute capture stress responses. Male breeding success in years males were handled was no different to the years they were not, despite the acute capture response, suggesting no long-term impact of handling on male reproductive output.


Ventilatory changes of pulmonary capillary blood volume assessed by arterial density.
By use of an improved density measuring system, we found that the gravimetric density of arterial blood of dogs fluctuates at the same frequency as the spontaneous or mechanical ventilation. Similar density fluctuations were observed in the blood leaving isolated, perfused lobes of dogs that were ventilated cyclicly. Employing an analysis that balanced the erythrocyte and plasma flows through distensible capillaries containing blood with a tube hematocrit lower than the hematocrit in large blood vessels, we derived a relationship to estimate from the density fluctuation the change in pulmonary capillary blood volume (Vc). For mechanical ventilation, the maximum change in density over one ventilation cycle increased from 0.084 +/- 0.01 to 0.47 +/- 0.05 (SE) g/l as the frequency decreased from 29 to 6 cycles/min. These density changes were estimated to be the result of an 1-16% change in Vc. A larger tidal volume for the mechanical ventilation led to a larger density fluctuation. The maximum density change of spontaneous respiration of 6 cycles/min was one-sixth of the mechanical case, indicating a much smaller change in Vc during spontaneous respiration. When the airway flow resistance was increased for spontaneous respiration, larger density fluctuations were observed.


A placebo-controlled, randomized trial of mesenchymal stem cells in COPD.
COPD is a devastating disease affecting millions worldwide. As disease pathogenesis includes both chronic pulmonary and systemic inflammation, antiinflammatory effects of systemically administered mesenchymal stem cells (MSCs) may decrease inflammation, resulting in improved lung function and quality of life. The goal of this study was to assess safety and to perform an initial evaluation of the potential efficacy of systemic MSC administration to patients with moderate to severe COPD. Sixty-two patients at six sites were randomized to double-blinded IV infusions of either allogeneic MSCs (Prochymal; Osiris Therapeutics Inc) or vehicle control. Patients received four monthly infusions (100 × 10⁶ cells/infusion) and were subsequently followed for 2 years after the first infusion. End points included comprehensive safety evaluation, pulmonary function testing (PFT), and quality-of-life indicators including questionnaires, 6MWT, and assessments of systemic inflammation. All study patients completed the full infusion protocol, and 74% completed the 2-year follow-up. There were no infusional toxicities and no deaths or serious adverse events deemed related to MSC administration. There were no significant differences in the overall number of adverse events, frequency of COPD exacerbations, or worsening of disease in patients treated with MSCs. There were no significant differences in PFTs or quality-of-life indicators; however, an early, significant decrease in levels of circulating C-reactive protein (CRP) was observed in patients treated with MSCs who had elevated CRP levels at study entry. Systemic MSC administration appears to be safe in patients with moderate to severe COPD and provides a basis for subsequent cell therapy investigations. ClinicalTrials.gov; No.: NCT00683722; URL: www.clinicaltrials.gov.


Toward a Sensible Single-antigen Bead Cutoff Based on Kidney Graft Survival.
There is no consensus in the literature on the interpretation of single-antigen bead positive for a specific HLA antibody. To inform the debate, we studied the relationship between various single-antigen bead positivity algorithms and the impact of resulting donor-specific HLA antibody (DSA) positivity on long-term kidney graft survival in 3237 deceased-donor transplants. First, we showed that the interassay variability can be greatly reduced when working with signal-to-background ratios instead of absolute median fluorescence intensities (MFIs). Next, we determined pretransplant DSA using various MFI cutoffs, signal-to-background ratios, and combinations thereof. The impact of the various cutoffs was studied by comparing the graft survival between the DSA-positive and DSA-negative groups. We did not observe a strong impact of various cutoff levels on 10-year graft survival. A stronger relationship between the cutoff level and 1-year graft survival for DSA-positive transplants was found when using signal-to-background ratios, most pronounced for the bead of the same HLA locus with lowest MFI taken as background. With respect to pretransplant risk stratification, we propose a signal-to-background ratio-6 (using the bead of the same HLA-locus with lowest MFI as background) cutoff of 15 combined with an MFI cutoff of 500, resulting in 8% and 21% lower 1- and 10-year graft survivals, respectively, for 8% DSA-positive transplants.


Monitoring, removal and risk assessment of cytostatic drugs in hospital wastewater.
Cytostatic agents are applied in cancer therapy and subsequently excreted into hospital wastewater. As these substances are known to be carcinogenic, mutagenic and toxic for reproduction, they should be removed from wastewater at their source of origin. In this study the fate and effects of the cancerostatic platinum compounds (CPC) cisplatin, carboplatin, oxaliplatin, 5-fluorouracil (5-FU) and the anthracyclines doxorubicin, daunorubicin and epirubicin were investigated in hospital wastewater. Wastewater from the in-patient treatment ward of a hospital in Vienna was collected and monitored for the occurrence of the selected drugs. A calculation model was established to spot the correlation between administered dosage and measured concentrations. To investigate the fate of the selected substances during wastewater treatment, the oncologic wastewater was treated in a pilot membrane bioreactor system (MBR) and in downstream advanced wastewater treatment processes (adsorption to activated carbon and UV-treatment). Genotoxic effects of the oncologic wastewater were assessed before and after wastewater treatment followed by a risk assessment. Monitoring concentrations of the selected cytostatics in the oncologic wastewater were in line with calculated concentrations. Due to different mechanisms (adsorption, biodegradation) in the MBR-system 5 - FU and the anthracyclines were removed < LOD, whereas CPC were removed by 60%. In parallel, genotoxic effects could be reduced significantly by the MBR-system. The risk for humans, the aquatic and terrestrial environment by hospital wastewater containing cytostatic drugs was classified as small in a preliminary risk assessment.


Effects of virtual reality on upper extremity function and activities of daily living performance in acute stroke: a double-blind randomized clinical trial.
To examine the effects of conventional therapy (CT) combined with intensive virtual reality (VR) program on upper extremity function and activities of daily living (ADL) in individuals in the acute stage of stroke. Twenty-six individuals were randomly assigned to the control or experimental group. The control group received CT, while the experimental group received VR training in addition to CT on same day. The Fugl-Meyer Assessment (FMA) and the Manual Function Test (MFT) were used to measure improvement of functioning in the affected limb and the Korean version of the Modified Barthel Index (K-MBI) was conducted to evaluate ADL. In tests of upper extremity functioning, VR group showed significant improvement on FMA and MFT (p< 0.05). In the CT group, only FMA score showed significant change (p< 0.05). ADL performance improved significantly in both groups (p< 0.05). However, there were no significant differences between the two groups in upper extremity function and ADL performance (p> 0.05). This study observed that VR training has the advantage of improving intended arm function during intensive training for individuals in the acute stage of stroke.


Anti-CTLA-4 therapy requires an Fc domain for efficacy.
Ipilimumab, a monoclonal antibody that recognizes cytotoxic T lymphocyte antigen (CTLA)-4, was the first approved "checkpoint"-blocking anticancer therapy. In mouse tumor models, the response to antibodies against CTLA-4 depends entirely on expression of the Fcγ receptor (FcγR), which may facilitate antibody-dependent cellular phagocytosis, but the contribution of simple CTLA-4 blockade remains unknown. To understand the role of CTLA-4 blockade in the complete absence of Fc-dependent functions, we developed H11, a high-affinity alpaca heavy chain-only antibody fragment (VHH) against CTLA-4. The VHH H11 lacks an Fc portion, binds monovalently to CTLA-4, and inhibits interactions between CTLA-4 and its ligand by occluding the ligand-binding motif on CTLA-4 as shown crystallographically. We used H11 to visualize CTLA-4 expression in vivo using whole-animal immuno-PET, finding that surface-accessible CTLA-4 is largely confined to the tumor microenvironment. Despite this, H11-mediated CTLA-4 blockade has minimal effects on antitumor responses. Installation of the murine IgG2a constant region on H11 dramatically enhances its antitumor response. Coadministration of the monovalent H11 VHH blocks the efficacy of a full-sized therapeutic antibody. We were thus able to demonstrate that CTLA-4-binding antibodies require an Fc domain for antitumor effect.


Importance of accessibility and opening hours to overall patient experience of general practice: analysis of repeated cross-sectional data from a national patient survey.
The UK government aims to improve the accessibility of general practices in England, particularly by extending opening hours in the evenings and at weekends. It is unclear how important these factors are to patients' overall experiences of general practice. To examine associations between overall experience of general practice and patient experience of making appointments and satisfaction with opening hours. Analysis of repeated cross-sectional data from the General Practice Patient Surveys conducted from 2011-2012 until 2013-2014. These covered 8289 general practice surgeries in England. Data from a national survey conducted three times over consecutive years were analysed. The outcome measure was overall experience, rated on a five-level interval scale. Associations were estimated as standardised regression coefficients, adjusted for responder characteristics and clustering within practices using multilevel linear regression. In total, there were 2 912 535 responders from all practices in England (n = 8289). Experience of making appointments (β 0.24, 95% confidence interval [CI] = 0.24 to 0.25) and satisfaction with opening hours (β 0.15, 95% CI = 0.15 to 0.16) were modestly associated with overall experience. Overall experience was most strongly associated with GP interpersonal quality of care (β 0.34, 95% CI = 0.34 to 0.35) and receptionist helpfulness was positively associated with overall experience (β 0.16, 95% CI = 0.16 to 0.17). Other patient experience measures had minimal associations (β≤0.06). Models explained ≥90% of variation in overall experience between practices. Patient experience of making appointments and satisfaction with opening hours were only modestly associated with overall experience. Policymakers in England should not assume that recent policies to improve access will result in large improvements in patients' overall experience of general practice.


AFLP analysis reveals a lack of phylogenetic structure within Solanum section Petota.
The secondary genepool of our modern cultivated potato (Solanum tuberosum L.) consists of a large number of tuber-bearing wild Solanum species under Solanum section Petota. One of the major taxonomic problems in section Petota is that the series classification (as put forward by Hawkes) is problematic and the boundaries of some series are unclear. In addition, the classification has received only partial cladistic support in all molecular studies carried out to date. The aim of the present study is to describe the structure present in section Petota. When possible, at least 5 accessions from each available species and 5 individual plants per accession (totally approx. 5000 plants) were genotyped using over 200 AFLP markers. This resulted in the largest dataset ever constructed for Solanum section Petota. The data obtained are used to evaluate the 21 series hypothesis put forward by Hawkes and the 4 clade hypothesis of Spooner and co-workers. We constructed a NJ tree for 4929 genotypes. For the other analyses, due to practical reasons, a condensed dataset was created consisting of one representative genotype from each available accession. We show a NJ jackknife and a MP jackknife tree. A large part of both trees consists of a polytomy. Some structure is still visible in both trees, supported by jackknife values above 69. We use these branches with >69 jackknife support in the NJ jackknife tree as a basis for informal species groups. The informal species groups recognized are: Mexican diploids, Acaulia, Iopetala, Longipedicellata, polyploid Conicibaccata, diploid Conicibaccata, Circaeifolia, diploid Piurana and tetraploid Piurana. Most of the series that Hawkes and his predecessors designated can not be accepted as natural groups, based on our study. Neither do we find proof for the 4 clades proposed by Spooner and co-workers. A few species groups have high support and their inner structure displays also supported subdivisions, while a large part of the species cannot be structured at all. We believe that the lack of structure is not due to any methodological problem but represents the real biological situation within section Petota.


A direct role for SNX9 in the biogenesis of filopodia.
Filopodia are finger-like actin-rich protrusions that extend from the cell surface and are important for cell-cell communication and pathogen internalization. The small size and transient nature of filopodia combined with shared usage of actin regulators within cells confounds attempts to identify filopodial proteins. Here, we used phage display phenotypic screening to isolate antibodies that alter the actin morphology of filopodia-like structures (FLS) in vitro. We found that all of the antibodies that cause shorter FLS interact with SNX9, an actin regulator that binds phosphoinositides during endocytosis and at invadopodia. In cells, we discover SNX9 at specialized filopodia in Xenopus development and that SNX9 is an endogenous component of filopodia that are hijacked by Chlamydia entry. We show the use of antibody technology to identify proteins used in filopodia-like structures, and a role for SNX9 in filopodia.


Molecular cloning and characterization of a tobacco MAP kinase kinase that interacts with SIPK.
A tobacco MAP kinase termed SIPK (Salicylic acid-Induced Protein Kinase) is activated in response to a variety of stress signals, including pathogen attack and wounding (S. Zhang and D.F. Klessig, Proc. Natl. Acad. Sci. USA 95:7225-7230, 1998; S. Zhang and D.F. Klessig, Proc. Natl. Acad. Sci. USA 95:7433-7438, 1998). Using the yeast two-hybrid system, we have identified a gene encoding a protein that interacts with SIPK but not the wounding induced protein kinase (WIPK), which is another tobacco MAP kinase. Sequence analysis indicated that this SIPK-interacting protein is a member of the MAP kinase kinase family; thus, it was named SIPK kinase (SIPKK). Co-immunoprecipitation experiments demonstrated that SIPKK and SIPK interact in vitro. Consistent with its putative function as a kinase, SIPKK phosphorylated myelin basic protein in vitro. Interestingly, SIPKK was induced at the mRNA level after Tobacco mosaic virus (TMV) infection or wounding, albeit with kinetics that are too slow to account for the activation of SIPK following these stimuli.


Human epithelial hair follicle stem cells and their progeny: current state of knowledge, the widening gap in translational research and future challenges.
Epithelial hair follicle stem cells (eHFSCs) are required to generate, maintain and renew the continuously cycling hair follicle (HF), supply cells that produce the keratinized hair shaft and aid in the reepithelialization of injured skin. Therefore, their study is biologically and clinically important, from alopecia to carcinogenesis and regenerative medicine. However, human eHFSCs remain ill defined compared to their murine counterparts, and it is unclear which murine eHFSC markers really apply to the human HF. We address this by reviewing current concepts on human eHFSC biology, their immediate progeny and their molecular markers, focusing on Keratin 15 and 19, CD200, CD34, PHLDA1, and EpCAM/Ber-EP4. After delineating how human eHFSCs may be selectively targeted experimentally, we close by defining as yet unmet key challenges in human eHFSC research. The ultimate goal is to transfer emerging concepts from murine epithelial stem cell biology to human HF physiology and pathology.


High resolution mapping of oxygen reduction reaction kinetics at polycrystalline platinum electrodes.
The scanning droplet-based technique, scanning electrochemical cell microscopy (SECCM), combined with electron backscatter diffraction (EBSD), is demonstrated as a powerful approach for visualizing surface structure effects on the rate of the oxygen reduction reaction (ORR) at polycrystalline platinum electrodes. Elucidating the effect of electrode structure on the ORR is of major interest in connection to electrocatalysis for energy-related applications. The attributes of the approach herein stem from: (i) the ease with which the polycrystalline substrate electrode can be prepared; (ii) the wide range of surface character open to study; (iii) the possibility of mapping reactivity within a particular facet (or grain), in a pseudo-single-crystal approach, and acquiring a high volume of data as a consequence; (iv) the ready ability to measure the activity at grain boundaries; and (v) an experimental arrangement (SECCM) that mimics the three-phase boundary in low temperature fuel cells. The kinetics of the ORR was analyzed and a finite element method model was developed to explore the effect of the three-phase boundary, in particular to examine pH variations in the droplet and the differential transport rates of the reactants and products. We have found a significant variation of activity across the platinum substrate, inherently linked to the crystallographic orientation, but do not detect any enhanced activity at grain boundaries. Grains with (111) and (100) contributions exhibit considerably higher activity than those with (110) and (100) contributions. These results, which can be explained by reference to previous single-crystal measurements, enhance our understanding of ORR structure-activity relationships on complex high-index platinum surfaces, and further demonstrate the power of high resolution flux imaging techniques to visualize and understand complex electrocatalyst materials.


Projected Burden of Osteoarthritis and Rheumatoid Arthritis in Australia: A Population-Level Analysis.
To forecast the prevalence and direct health care costs of osteoarthritis (OA) and rheumatoid arthritis (RA) in Australia to the year 2030. An epidemiologic model of the Australian population was developed. Data on the national prevalence of OA and RA were obtained from the Australian Bureau of Statistics (ABS) 2014-2015 National Health Survey. Future prevalence was estimated using ABS population projections for 2020, 2025, and 2030. Available government data on direct health care expenditure for OA and RA were modeled to forecast costs (in Australian $) for the years 2020, 2025, and 2030, from the perspective of the Australian public health care system. The number of people with OA is expected to increase nationally from almost 2.2 million in 2015 to almost 3.1 million Australians in 2030. The number of people with RA is projected to increase from 422,309 in 2015 to 579,915 in 2030. Health care costs for OA were estimated to be over $2.1 billion in 2015; by the year 2030, these are forecast to exceed $2.9 billion ($970 for every person with the condition). Health care costs for RA were estimated to be over $550 million in 2015, including $273 million spent on biologic disease-modifying antirheumatic drugs. Health care costs for RA are projected to rise to over $755 million by the year 2030. OA and RA are costly conditions that will impose an increasing health care burden at the population level. These projections provide tangible data that can be used to map future health service provision to expected need.