Investigation of efficacy and safety of low-dose sodium glucose transporter 2 inhibitors and differences between two agents, canagliflozin and ipragliflozin, in patients with type 2 diabetes mellitus.
Sodium glucose transporter 2 inhibitors (SGLT2is), new antidiabetic agents, were reported to improve not only glycemic parameters but also metabolic and circulatory parameters. Whereas, several adverse events caused by SGLT2is were also reported. We aimed to investigate the changes of glycemic, metabolic, and circulatory parameters as well as safety with low-dose administration of two SGLT2is, canagliflozin and ipragliflozin, and also the difference between the two agents. 25 individuals with type-2 diabetes mellitus (T2DM) were recruited and administered with low-dose SGLT2is, canagliflozin (n = 10, 50 mg/day) and ipragliflozin (n = 15, 25 mg/day). We examined glycemic, metabolic, and circulatory parameters at baseline and 24 weeks after administration. All patients completed the study without complications. Compared with baseline, levels of glycated hemoglobin, fasting plasma glucose, and homeostasis model assessment of β-cell function improved significantly at 24 weeks after administration (p < 0.05). Levels of body weight, low-density lipoproteincholesterol, aspartate transaminase, γ-glutamyl transferase, and urinary excretion of albumin also improved significantly (p < 0.05). Moreover, systolic/diastolic blood pressure and levels of brain natriuretic peptide improved significantly (p < 0.05). The comparison of improvement ratio (values of improvement/values of basement) of each agent revealed that there was a significant difference between low-dose canagliflozin and low-dose ipragliflozin for brain natriuretic peptide (0.4404 vs. 0.0970, p = 0.0275). Hence, low-dose SGLT2is could be useful for patients of T2DM not only for hyperglycemia but also for metabolic and circulatory disorders without eliciting adverse events. In addition, with regard to the efficacy upon cardiovascular function, canagliflozin could be more suitable than ipragliflozin.


Transgenic mouse models for tumors of melanocytes and retinal pigment epithelium.
Cutaneous and ocular melanomas are due to malignant transformation of neural crest-derived melanocytes. The rising incidence of this tumor in humans has stimulated experiments to devise suitable mouse models. In the past years, transgenic mouse lines have been generated using different oncogenes - Ha-ras, SV40 T antigen (Tag), ret - which develop benign lesions of melanocytes, melanoma, and/or eye tumors. Pigment cell tumors in humans, although rather rare, can also develop from the retinal pigment epithelium (RPE), a cell layer of neuroectodermal origin. We, therefore, established transgenic models for this ocular tumor. Regulated by the promoter of tyrosinase-related protein-1 (TRP-1), two oncogenes, ret and SV40 Tag, were targeted to the developing RPE in transgenic mice. The TRP-1/ret transgenic mice displayed microphthalmia and benign tumors of the RPE. Expression of SV40 T antigen (TRP-1/Tag) led to malignant tumors, which were invasive and metastasized to inguinal lymph node and spleen.


Starvation and recovery in the deep-sea methanotroph Methyloprofundus sedimenti.
In the deep ocean, the conversion of methane into derived carbon and energy drives the establishment of diverse faunal communities. Yet specific biological mechanisms underlying the introduction of methane-derived carbon into the food web remain poorly described, due to a lack of cultured representative deep-sea methanotrophic prokaryotes. Here, the response of the deep-sea aerobic methanotroph Methyloprofundus sedimenti to methane starvation and recovery was characterized. By combining lipid analysis, RNA analysis, and electron cryotomography, it was shown that M. sedimenti undergoes discrete cellular shifts in response to methane starvation, including changes in headgroup-specific fatty acid saturation levels, and reductions in cytoplasmic storage granules. Methane starvation is associated with a significant increase in the abundance of gene transcripts pertinent to methane oxidation. Methane reintroduction to starved cells stimulates a rapid, transient extracellular accumulation of methanol, revealing a way in which methane-derived carbon may be routed to community members. This study provides new understanding of methanotrophic responses to methane starvation and recovery, and lays the initial groundwork to develop Methyloprofundus as a model chemosynthesizing bacterium from the deep sea.


A comparison of left and right atrial fibroblasts reveals different collagen production activity and stress-induced mitogen-activated protein kinase signalling in rats.
Atrial fibrosis plays a pivotal role in the pathophysiology of heart failure (HF). The left atrium (LA) experiences greater fibrosis than the right atrium (RA) during HF. It is not clear whether LA cardiac fibroblasts contain distinctive activities that predispose LA to fibrosis. LA and RA fibrosis were evaluated in healthy and isoproterenol-induced HF Sprague Dawley rats. Rat LA and RA primary isolated fibroblasts were subjected to proliferation assay, oxidative stress assay, cell migration analysis, collagen measurement, cytokine array and Western blot. Healthy rat LA and RA had a similar extent of collagen deposition. HF significantly increased fibrosis to a greater severity in LA than in RA. Compared to isolated RA fibroblasts, the in vitro experiments showed that isolated LA fibroblasts had higher oxidative stress and exhibited higher collagen, transforming growth factor-β1, connective tissue growth factor production and less vascular endothelial growth factor (VEGF) production, but had similar migration, myofibroblast differentiation and proliferation activities. VEGF significantly increased the collagen production ability of LA fibroblasts, but not RA fibroblasts. LA fibroblasts had more phosphorylated ERK1/2 and P38 expression. ERK inhibitor (PD98059, 50 μmol L-1 ) significantly attenuated collagen production and increased VEGF production in RA fibroblasts but not in LA fibroblasts. P38 inhibitor (SB203580, 30 μmol L-1 ) significantly attenuated collagen production in LA fibroblasts but not in RA fibroblasts. P38 inhibitor also significantly increased VEGF production in RA and LA fibroblasts. Differences in profibrotic activity between LA and RA fibroblasts may be caused by different responses to mitogen-activated protein kinase signalling.


Transition states. Trapping a transition state in a computationally designed protein bottle.
The fleeting lifetimes of the transition states (TSs) of chemical reactions make determination of their three-dimensional structures by diffraction methods a challenge. Here, we used packing interactions within the core of a protein to stabilize the planar TS conformation for rotation around the central carbon-carbon bond of biphenyl so that it could be directly observed by x-ray crystallography. The computational protein design software Rosetta was used to design a pocket within threonyl-transfer RNA synthetase from the thermophile Pyrococcus abyssi that forms complementary van der Waals interactions with a planar biphenyl. This latter moiety was introduced biosynthetically as the side chain of the noncanonical amino acid p-biphenylalanine. Through iterative rounds of computational design and structural analysis, we identified a protein in which the side chain of p-biphenylalanine is trapped in the energetically disfavored, coplanar conformation of the TS of the bond rotation reaction.


The Effects of Repetitive Transcranial Magnetic Stimulation in Patients with Chronic Schizophrenia: Insights from EEG Microstates.
The objective of this study was to investigate the effects of 10 Hz repetitive transcranial magnetic stimulation (rTMS) in patients with schizophrenia using EEG microstates. Thirty-eight patients with chronic schizophrenia were included in a double-blind, randomized and sham-controlled trial (19 participants in the active group and 19 participants in the sham group) and received 10 Hz active or sham rTMS stimulation to the left dorsolateral prefrontal cortex (left DLPFC) 5 days per week over for 4 weeks. Four classical microstate classes (i.e., classes A, B, C and D) were identified by clustering, and the parameters (i.e., duration, occurrence and contribution) of each class were computed. Our results showed that (1) after stimulation, the positive and negative syndrome scale (PANSS) positive scores decreased significantly in the active group; (2) the duration of the microstate of class C derived from EEG data decreased significantly in the active group; and (3) the change of the duration of class D in the active group was significantly higher than that in the sham group. Our findings demonstrated that 10 Hz active rTMS stimulation was beneficial to improving the positive symptoms of patients with chronic schizophrenia, and the EEG microstate could be an effective indicator of symptom improvements.


Evaluating the generalizability of a large streamlined cardiovascular trial: comparing hospitals and patients in the dual antiplatelet therapy study versus the National Cardiovascular Data Registry.
The Dual Antiplatelet Therapy Study is large streamlined clinical trial designed to evaluate antiplatelet treatment strategies in a broadly inclusive population of subjects treated with coronary stents. Whether large streamlined trials can successfully include a representative group of study sites and patients has not been formally assessed. Within the National Cardiovascular Data Registry CathPCI Registry, we compared characteristics and outcomes of hospitals participating versus not participating in the Dual Antiplatelet Therapy Study. We also compared clinical and procedural characteristics of trial subjects undergoing percutaneous coronary intervention (PCI) with drug-eluting stents to contemporaneous patients within the National Cardiovascular Data Registry CathPCI Registry. Standardized differences between groups were estimated. Between September 2009 and July 2011, 1.1 million PCIs were performed among 1276 hospitals, of which 309 (24.2%) participated in the Dual Antiplatelet Therapy Study. Participating hospitals were larger (468 versus 311 beds), more frequently located in urban settings (61.2% versus 42.6%), and had higher annual PCI volumes (858 versus 378) compared with nonparticipating hospitals, although hospital case mix and procedural outcomes were similar. Compared with CathPCI patients, trial patients undergoing PCI with drug-eluting stents were similar with respect to race, sex, and rates of diabetes mellitus, hypertension, and smoking, although they had lower rates of prior cardiovascular disease. Within the Dual Antiplatelet Therapy Study, clinical trial sites had similar patient case mix and clinical outcomes as nonparticipating sites. Although trial participants were representative of PCI patients with respect to race, sex and most comorbidities, they had a lower prevalence of chronic cardiovascular disease compared with registry patients. Although a streamlined cardiovascular clinical trial may successfully involve a large number of hospitals and rapidly enroll a diverse population of patients, differences between eligible patients and those actually enrolled remained. http://www.clinicaltrials.gov. Unique identifier: NCT00977938.


Time Spent Gaming, Device Type, Addiction Scores, and Well-being of Adolescent English Gamers in the 2021 OxWell Survey: Latent Profile Analysis.
The shift in the last decades to screen-based and increasingly web-based gaming activity has raised concerns about its impact on the development of children and adolescents. Despite decades of research into gaming and related psychosocial effects, the question remains how best to identify what degree or context of gaming may be a cause for concern. This study aimed to classify adolescents into gamer profiles based on both gaming behaviors and well-being. Once we distinguished the different gamer profiles, we aimed to explore whether membership to a specific profile could be predicted based on a range of personal characteristics and experiences that could then help identify those at risk. We explored gaming and well-being in an adolescent school population (aged 12-18 years) in England as part of the 2021 OxWell student survey. Self-report measures of time spent playing games on computers or consoles, time spent playing games on mobile phones, the Game Addiction Scale, and the Warwick-Edinburgh Mental Well-being Scale were used to classify adolescent heavy gamers (playing games for at least 3.5 hours a day) using latent profile analysis. We used multinomial logistic regression analysis to predict the profile membership based on a range of personal characteristics and experiences. In total, 12,725 participants answered the OxWell gaming questions. Almost one-third (3970/12,725, 31.2%) indicated that they play games for at least 3.5 hours a day. The correlation between time spent playing video games overall and well-being was not significant (P=.41). The latent profile analysis distinguished 6 profiles of adolescent heavy gamers: adaptive computer gamers (1747/3970, 44%); casual computer gamers (873/3970, 22%); casual phone gamers (595/3970, 15%); unknown device gamers (476/3970, 12%); maladaptive computer gamers (238/3970, 6%); and maladaptive phone gamers (79/3970, 2%). In comparison with adaptive computer gamers, maladaptive phone gamers were mostly female (odds ratio [OR] 0.08, 95% CI 0.03-0.21) and were more likely to have experienced abuse or neglect (OR 3.18, 95% CI 1.34-7.55). Maladaptive computer gamers, who reported gaming both on their mobile phones and on the computer, were mostly male and more likely to report anxiety (OR 2.25, 95% CI 1.23-4.12), aggressive behavior (OR 2.83, 95% CI 1.65-4.88), and web-based gambling (OR 2.18, 95% CI 1.24-3.81). A substantial number of adolescents are spending ≥3.5 hours gaming each day, with almost 1 in 10 (317/3970, 8%) reporting co-occurring gaming and well-being issues. Long hours gaming using mobile phones, particularly common in female gamers, may signal poorer functioning and indicate a need for additional support. Although increased time gaming might be changing how adolescents spend their free time and might thus have public health implications, it does not seem to relate to co-occurring well-being issues or mental ill-health for the majority of adolescent gamers.


Deep Learning for Prediction and Optimization of Fast-Flow Peptide Synthesis.
The chemical synthesis of polypeptides involves stepwise formation of amide bonds on an immobilized solid support. The high yields required for efficient incorporation of each individual amino acid in the growing chain are often impacted by sequence-dependent events such as aggregation. Here, we apply deep learning over ultraviolet-visible (UV-vis) analytical data collected from 35 427 individual fluorenylmethyloxycarbonyl (Fmoc) deprotection reactions performed with an automated fast-flow peptide synthesizer. The integral, height, and width of these time-resolved UV-vis deprotection traces indirectly allow for analysis of the iterative amide coupling cycles on resin. The computational model maps structural representations of amino acids and peptide sequences to experimental synthesis parameters and predicts the outcome of deprotection reactions with less than 6% error. Our deep-learning approach enables experimentally aware computational design for prediction of Fmoc deprotection efficiency and minimization of aggregation events, building the foundation for real-time optimization of peptide synthesis in flow.


Limb-specific autonomic dysfunction in complex regional pain syndrome modulated by wearing prism glasses.
In unilateral upper-limb complex regional pain syndrome (CRPS), the temperature of the hands is modulated by where the arms are located relative to the body midline. We hypothesized that this effect depends on the perceived location of the hands, not on their actual location, nor on their anatomical alignment. In 2 separate cross-sectional randomized experiments, 10 (6 female) unilateral CRPS patients wore prism glasses that laterally shifted the visual field by 20°. Skin temperature was measured before and after 9-minute periods in which the position of one hand was changed. Placing the affected hand on the healthy side of the body midline increased its temperature (Δ°C=+0.47 ± 0.14°C), but not if prism glasses made the hand appear to be on the body midline (Δ°C=+0.07 ± 0.06°C). Similarly, when prism glasses made the affected hand appear to be on the healthy side of the body midline, even though it was not, the affected hand warmed up (Δ°C=+0.28 ± 0.14°C). When prism glasses made the healthy hand appear to be on the affected side of the body midline, even though it was not, the healthy hand cooled down (Δ°C=-0.30 ± 0.15°C). Friedman's analysis of variance and Wilcoxon pairs tests upheld the results (P<0.01 for all). We conclude that, in CRPS, cortical mechanisms responsible for encoding the perceived location of the limbs in space modulate the temperature of the hands.


Heavily and fully modified RNAs guide efficient SpyCas9-mediated genome editing.
RNA-based drugs depend on chemical modifications to increase potency and to decrease immunogenicity in vivo. Chemical modification will likely improve the guide RNAs involved in CRISPR-Cas9-based therapeutics as well. Cas9 orthologs are RNA-guided microbial effectors that cleave DNA. Here, we explore chemical modifications at all positions of the crRNA guide and tracrRNA cofactor. We identify several heavily modified versions of crRNA and tracrRNA that are more potent than their unmodified counterparts. In addition, we describe fully chemically modified crRNAs and tracrRNAs (containing no 2'-OH groups) that are functional in human cells. These designs will contribute to Cas9-based therapeutics since heavily modified RNAs tend to be more stable in vivo (thus increasing potency). We anticipate that our designs will improve the use of Cas9 via RNP and mRNA delivery for in vivo and ex vivo purposes.


Barriers to racemization in C3-symmetric complexes containing the hydrotris(2-mercapto-1-ethylimidazolyl)borate (Tm(Et)) ligand.
The tripodal ligands hydrotris(N-ethyl-2-mercaptoimidazol-1-yl)borate (NaTm(Et)) (1) and hydrotris(N-benzyl-2-mercaptoimidazol-1-yl)borate (NaTm(Bn)) (2), analogues of the hydrotris(N-methyl-2-mercaptoimidazol-1-yl)borate ligand (Tm) containing alternative nitrogen substituents, have been employed to examine the racemization of their C3-symmetric complexes with both four- and six-coordinate metals. The ligands react at room temperature with metal halides to provide C3-symmetric metal complexes. The syntheses of the four-coordinate complexes [Tm(Et)ZnCl] (3), [Tm(Et)CdBr] (4), [Tm(Et)HgCl] (5), [Tm(Et)CuPPh3] (6), [Tm(Et)AgPPh3] (7), and [Tm(Bn)ZnCl] (8) are reported. The six-coordinate complexes [Tm(Et)Ru(p-cymene)]Cl (9), [Tm(Et)Ru(p-cymene)]PF(6) (10), and [Tm(Et)Mn(CO)3] (11) were also synthesized. The X-ray crystal structures of 3, 4, 6, and 9 are reported. The diastereotopic nature of the ethyl and benzyl hydrogen atoms in the ligands allows the enantiomeric forms of these complexes to be distinguished by 1H NMR spectroscopy. Variable-temperature (VT) 1H NMR spectra have thus been used to investigate the energies of the racemization processes occurring in these chiral complexes. In solvents the activation energies to racemization for the four-coordinate complexes lay in the range of 53-77 kJ mol(-1). In non-donor solvents the energies are reduced and a dissociative mechanism is therefore implicated. No interconversion could be observed by VT NMR for the six-coordinate complexes in any solvent. To further explore the racemization mechanisms ab initio density functional theory calculations have been conducted on the ground- and transition-state structures of representative six-coordinate [Mn(I)] and four-coordinate [Zn(II)] complexes following a proposed nondissociative mechanism of racemization. The calculated energy barriers to racemization are 163 and 121 kJ mol(-1), respectively. It is concluded that the low-energy racemization of substitution-labile four-coordinate complexes occurs via a dissociative mechanism, while substitution-inert six-coordinate complexes experience a significantly higher barrier to racemization. Whether this is due to the operation of a dissociative mechanism with a higher activation barrier or to a nondissociative mechanism remains unknown.


p16(INK4a) immunohistochemistry is a promising biomarker to predict the outcome of low grade cervical intraepithelial neoplasia: comparison study with HPV genotyping.
In cervical intraepithelial neoplasia (CIN), p16(INK4a) immunohistochemistry has been reported to be a useful diagnostic biomarker. However, limited information is available about the association between the p16(INK4a) immunohistochemistry and the outcomes of CIN. Here, we report p16(INK4a) immunohistochemistry as an effective biomarker to predict the outcomes of CIN. p16(INK4a) immunohistochemistry was performed in patients with CIN from January 2000 to August 2009. Among these patients, we have performed a retrospective analysis of the medical records to evaluate the outcome of CIN 1-2 and performed statistical analysis to determine the correlation between p16(INK4a) expression and the outcomes. We also performed HPV genotyping and analyzed the relation between the infecting human papillomavirus (HPV) genotype and the outcomes. A total of 244 patients, including 82 with CIN 1, 60 with CIN 2, and 102 with CIN 3, were examined. The rate of p16(INK4a) overexpression increased with increasing CIN grade, 20.7% for CIN 1, 80.0% for CIN 2, and 89.2% for CIN 3, with significant differences between CIN 1 and CIN 2-3 group. In the 131 CIN 1-2 patients, the progression rate was significantly higher for the patients showing p16(INK4a) overexpression than for those not showing p16(INK4a) overexpression (p=0.005); the regression rate was also found to be significantly lower for the patients showing p16(INK4a) overexpression (p=0.003). High-risk HPV genotypes were detected in 73 patients (73.7%). Both progression and regression rates were not significantly different between the high-risk HPV-positive and HPV-negative groups (p=0.401 and p=0.381, respectively). p16(INK4a) overexpression was correlated with the outcome of CIN 1-2, and p16(INK4a) is considered to be a superior biomarker for predicting the outcome of CIN 1-2 compared with HPV genotyping.


A proposed grading system for arteriovenous malformations.
An important factor in making a recommendation for treatment of a patient with arteriovenous malformation (AVM) is to estimate the risk of surgery for that patient. A simple, broadly applicable grading system that is designed to predict the risk of morbidity and mortality attending the operative treatment of specific AVM's is proposed. The lesion is graded on the basis of size, pattern of venous drainage, and neurological eloquence of adjacent brain. All AVM's fall into one of six grades. Grade I malformations are small, superficial, and located in non-eloquent cortex; Grade V lesions are large, deep, and situated in neurologically critical areas; and Grade VI lesions are essentially inoperable AVM's. Retrospective application of this grading scheme to a series of surgically excised AVM's has demonstrated its correlation with the incidence of postoperative neurological complications. The application of a standardized grading scheme will enable a comparison of results between various clinical series and between different treatment techniques, and will assist in the process of management decision-making.


Selective inhibition of mutant IDH1 by DS-1001b ameliorates aberrant histone modifications and impairs tumor activity in chondrosarcoma.
Chondrosarcoma is the second most common malignant bone tumor. It is characterized by low vascularity and an abundant extracellular matrix, which confer these tumors resistance to chemotherapy and radiotherapy. There are currently no effective treatment options for relapsed or dedifferentiated chondrosarcoma, and new targeted therapies need to be identified. Isocitrate dehydrogenase (IDH) mutations, which are detected in ~50% of chondrosarcoma patients, contribute to malignant transformation by catalyzing the production of 2-hydroxyglutarate (2-HG), a competitive inhibitor of α-ketoglutarate-dependent dioxygenases. Mutant IDH inhibitors are therefore potential novel anticancer drugs in IDH mutant tumors. Here, we examined the efficacy of the inhibition of mutant IDH1 as an antitumor approach in chondrosarcoma cells in vitro and in vivo, and investigated the association between the IDH mutation and chondrosarcoma cells. DS-1001b, a novel, orally bioavailable, selective mutant IDH1 inhibitor, impaired the proliferation of chondrosarcoma cells with IDH1 mutations in vitro and in vivo, and decreased 2-HG levels. RNA-seq analysis showed that inhibition of mutant IDH1 promoted chondrocyte differentiation in the conventional chondrosarcoma L835 cell line and caused cell cycle arrest in the dedifferentiated JJ012 cell line. Mutant IDH1-mediated modulation of SOX9 and CDKN1C expression regulated chondrosarcoma tumor progression, and DS-1001b upregulated the expression of these genes via a common mechanism involving the demethylation of H3K9me3. DS-1001b treatment reversed the epigenetic changes caused by aberrant histone modifications. The present data strongly suggest that inhibition of mutant IDH1 is a promising therapeutic approach in chondrosarcoma, particularly for the treatment of relapsed or dedifferentiated chondrosarcoma.


Relationship of fruit and vegetable intake with adiposity: a systematic review.
Fruit and vegetable (FV) intake has been proposed to protect against obesity. The purpose of this paper was to assess the FV consumption to adiposity relationship. Twenty-three publications were included. longitudinal or experimental designs; FV intake tested in relation to adiposity; child, adolescent or adult participants; published in English-language peer-reviewed journals. dietary pattern and cross-sectional designs; participants with health concerns. Experimental studies found increased FV consumption (in conjunction with other behaviours) contributed to reduced adiposity among overweight or obese adults, but no association was shown among children. Longitudinal studies among overweight adults found greater F and/or V consumption was associated with slower weight gain, but only half of child longitudinal studies found a significant inverse association. Limitations in methods prevented a thorough examination of the role of increased FV intake alone or mechanisms of effect. An inverse relationship between FV intake and adiposity among overweight adults appears weak; this relationship among children is unclear. Research needs to clarify the nature of, and mechanisms for, the effects of FV consumption on adiposity. Whether increases in FV intake in isolation from lower caloric intake or increased physical activity will result in declines or slower growth in adiposity remains unclear.


Paclitaxel-carboplatin alone or with bevacizumab for non-small-cell lung cancer.
Bevacizumab, a monoclonal antibody against vascular endothelial growth factor, has been shown to benefit patients with a variety of cancers. Between July 2001 and April 2004, the Eastern Cooperative Oncology Group (ECOG) conducted a randomized study in which 878 patients with recurrent or advanced non-small-cell lung cancer (stage IIIB or IV) were assigned to chemotherapy with paclitaxel and carboplatin alone (444) or paclitaxel and carboplatin plus bevacizumab (434). Chemotherapy was administered every 3 weeks for six cycles, and bevacizumab was administered every 3 weeks until disease progression was evident or toxic effects were intolerable. Patients with squamous-cell tumors, brain metastases, clinically significant hemoptysis, or inadequate organ function or performance status (ECOG performance status, >1) were excluded. The primary end point was overall survival. The median survival was 12.3 months in the group assigned to chemotherapy plus bevacizumab, as compared with 10.3 months in the chemotherapy-alone group (hazard ratio for death, 0.79; P=0.003). The median progression-free survival in the two groups was 6.2 and 4.5 months, respectively (hazard ratio for disease progression, 0.66; P<0.001), with corresponding response rates of 35% and 15% (P<0.001). Rates of clinically significant bleeding were 4.4% and 0.7%, respectively (P<0.001). There were 15 treatment-related deaths in the chemotherapy-plus-bevacizumab group, including 5 from pulmonary hemorrhage. The addition of bevacizumab to paclitaxel plus carboplatin in the treatment of selected patients with non-small-cell lung cancer has a significant survival benefit with the risk of increased treatment-related deaths. (ClinicalTrials.gov number, NCT00021060.)


pH-induced variations in the TK1 gene model.
A physiological decrease in extracellular pH (pHe) alters the efficiency of DNA repair and increases formation of DNA double-strand breaks (DSBs). Whether this could translate into genetic instability and variations, was investigated using the TK6 cell model, in which positive selection of the TK1 gene loss-of-function mutations can be achieved from resistance to trifluorothymidine. Cell exposure to suboptimal pH (down to 6.9) for 3 weeks resulted in the 100 % frequency of a stronger frameshift mutation that has spread to both TK1 alleles, whereas weaker frameshift mutations within the 3'exon were eliminated during the selection. Suboptimal pHe values were also found to alter the proportion of the TK1 splicing variant expressed as percent spliced in index values and promote selection of truncated exons as well as intron retention. Although recovery at pH 7.4 did not reverse the selected frameshift mutation, reversal of splice variants and exon truncation towards control values were observed. Hence, suboptimal pHe can induce a combination of mutational events and splicing alterations within the same gene in the resistant clones. This model of positive selection for loss-of-function clearly demonstrates that suboptimal pHe may confer a similar growth advantage when such instability occurs within tumor suppressor genes.


Axial ligand complexes of the Rhodnius nitrophorins: reduction potentials, binding constants, EPR spectra, and structures of the 4-iodopyrazole and imidazole complexes of NP4.
Previously, we utilized 4-iodopyrazole (4IPzH) as a heavy atom derivative for the initial solution of the crystal structure of the nitrophorin from Rhodnius prolixus, NP1, where it was found to bind to the heme with the iodo group disordered in two positions. We have now determined the structure of the 4IPzH complex of NP4 at pH 7.5 and find that the geometry and bond lengths at the iron center are extremely similar to those of the imidazole (ImH) complex of the same protein (structure determined at pH 5.6), except that the G-H loop is not in the closed conformation. 4IPzH binds to the heme of NP4 in an ordered manner, with the iodo substituent pointed toward the opening of the heme pocket, near the surface of the protein. In order to understand the solution chemistry in terms of the relative binding abilities of 4IPzH, ImH, and histamine (Hm, a physiological ligand for the nitrophorins), we have also investigated the equilibrium binding constants and reduction potentials of these three ligand complexes of the four Rhodnius nitrophorins as a function of pH. We have found that, unlike the other Lewis bases, 4IPzH forms less stable complexes with the Fe(III) than the Fe(II) oxidation states of NP1 and NP4, and similar stability for the two oxidation states of NP2 and NP3, suggesting that this ligand is a softer base than ImH or Hm, for both of which the Fe(III) complexes are more stable than those of Fe(II) for all four nitrophorins. Surprisingly, in spite of this and the much lower basicity of 4IPzH than imidazole and histamine, the EPR g-values of all three ligand complexes are very similar.


Bevacizumab treatment induces metabolic adaptation toward anaerobic metabolism in glioblastomas.
Anti-angiogenic therapy in glioblastoma (GBM) has unfortunately not led to the anticipated improvement in patient prognosis. We here describe how human GBM adapts to bevacizumab treatment at the metabolic level. By performing (13)C6-glucose metabolic flux analysis, we show for the first time that the tumors undergo metabolic re-programming toward anaerobic metabolism, thereby uncoupling glycolysis from oxidative phosphorylation. Following treatment, an increased influx of (13)C6-glucose was observed into the tumors, concomitant to increased lactate levels and a reduction of metabolites associated with the tricarboxylic acid cycle. This was confirmed by increased expression of glycolytic enzymes including pyruvate dehydrogenase kinase in the treated tumors. Interestingly, L-glutamine levels were also reduced. These results were further confirmed by the assessment of in vivo metabolic data obtained by magnetic resonance spectroscopy and positron emission tomography. Moreover, bevacizumab led to a depletion in glutathione levels indicating that the treatment caused oxidative stress in the tumors. Confirming the metabolic flux results, immunohistochemical analysis showed an up-regulation of lactate dehydrogenase in the bevacizumab-treated tumor core as well as in single tumor cells infiltrating the brain, which may explain the increased invasion observed after bevacizumab treatment. These observations were further validated in a panel of eight human GBM patients in which paired biopsy samples were obtained before and after bevacizumab treatment. Importantly, we show that the GBM adaptation to bevacizumab therapy is not mediated by clonal selection mechanisms, but represents an adaptive response to therapy.


Nuclear magnetic resonance studies on the interaction of avoparcin with model receptors of bacterial cell walls.
On the basis of nuclear Overhauser enhancement and 1H chemical shift data obtained in aqueous solution, a model is proposed for the interaction of beta-avoparcin and epi-beta-avoparcin with acetyl-D-alanyl-D-alanine (Ac-D-Ala-D-Ala) and diacetyl-L-lysyl-D-alanyl-D-alanine (Ac2-L-Lys-D-Ala-D-Ala). For the beta-avoparcin: Ac2-L-Lys-D-Ala-Ala complex, the COOH-terminal end of the tripeptide is located near the NH2 terminus of the antibiotic with the tripeptide extending across the peptide backbone of beta-avoparcin toward its COOH-terminal end. In our proposed structure, the three amino acid residues of the peptide span the entire length of the antibiotic, and the aliphatic side chain of the lysine residue extends over the D-ring of beta-avoparcin. The structure of the epi-beta-avoparcin:Ac2-L-Lys-D-Ala-D-Ala complex was found to be similar to the beta-avoparcin complex at the binding site for the lysine residue at the COOH-terminal end of the antibiotic, but differed in the interactions at the NH2 terminus. These results are consistent with the similarities in the COOH-terminal conformations and the differences in conformations at the NH2 terminus found for beta-avoparcin and epi-beta-avoparcin which were described in the preceding paper [Fesik, S. W., I. M. Armitage, G. A. Ellestad, and W. J. McGahren, Mol. Pharmacol. 25:275-280 (1984)]. The association constants (measured by UV methods) for both beta-avoparcin:peptide complexes were greater than those measured for epi-beta-avoparcin and correlated with their differences in antibacterial activity. Epi-beta-avoparcin exhibited no measurable binding to the dipeptide: however, a significant affinity was measured for the tripeptide, indicating that the interactions with the NH2 terminus of the antibiotics provide binding energy for the antibiotic peptide complex but that the COOH-terminal end of the antibiotics also plays an important role in the binding interaction. These results are interesting in light of the similarities in the structural and conformational features at the COOH terminus for all of the glycopeptide antibiotics.


Spectral-overlap approach to multiframe superresolution image reconstruction.
Various techniques and algorithms have been developed to improve the resolution of sensor-aliased imagery captured with multiple subpixel-displaced frames on an undersampled pixelated image plane. These dealiasing algorithms are typically known as multiframe superresolution (SR), or geometric SR to emphasize the role of the focal-plane array. Multiple low-resolution (LR) aliased frames of the same scene are captured and allocated to a common high-resolution (HR) reconstruction grid, leading to the possibility of an alias-free reconstruction, as long as the HR sampling rate is above the Nyquist rate. Allocating LR-frame irradiances to HR frames requires the use of appropriate weights. Here we present a novel approach in the spectral domain to calculating exactly weights based on spatial overlap areas, which we call the spectral-overlap (SO) method. We emphasize that the SO method is not a spectral approach but rather an approach to calculating spatial weights that uses spectral decompositions to exploit the array properties of the HR and LR pixels. The method is capable of dealing with arbitrary aliasing factors and interframe motions consisting of in-plane translations and rotations. We calculate example reconstructed HR images (the inverse problem) from synthetic aliased images for integer and for fractional aliasing factors. We show the utility of the SO-generated overlap-area weights in both noniterative and iterative reconstructions with known or unknown aliasing factor. We show how the overlap weights can be used to generate the Green's function (pixel response function) for noniterative dealiasing. In addition, we show how the overlap-area weights can be used to generate synthetic aliased images (the forward problem). We compare the SO approach to the spatial-domain geometric approach of O'Rourke and find virtually identical high accuracy but with significant enhancements in speed for SO. We also compare the SO weights to interpolated weights and find that the overlap-area weights lead to significantly smaller errors in iterative reconstructions. We consider how the SO method might be extended to account for the influence of the optical transfer function, more complex or space-variant motions, the registration process, and noise.


Microgroove and Collagen-poly(ε-caprolactone) Nanofiber Mesh Coating Improves the Mechanical Stability and Osseointegration of Titanium Implants.
The effect of depositing a collagen (CG)-poly-ε-caprolactone (PCL) nanofiber mesh (NFM) at the microgrooves of titanium (Ti) on the mechanical stability and osseointegration of the implant with bone was investigated using a rabbit model. Three groups of Ti samples were produced: control Ti samples where there were no microgrooves or CG-PCL NFM, groove Ti samples where microgrooves were machined on the circumference of Ti, and groove-NFM Ti samples where CG-PCL NFM was deposited on the machined microgrooves. Each group of Ti samples was implanted in the rabbit femurs for eight weeks. The mechanical stability of the Ti/bone samples were quantified by shear strength from a pullout tension test. Implant osseointegration was evaluated by a histomorphometric analysis of the percentage of bone and connective tissue contact with the implant surface. The bone density around the Ti was measured by micro-computed tomography (μCT) analysis. This study found that the shear strength of groove-NFM Ti/bone samples was significantly higher compared to control and groove Ti/bone samples (p < 0.05) and NFM coating influenced the bone density around Ti samples. In vivo histomorphometric analyses show that bone growth into the Ti surface increased by filling the microgrooves with CG-PCL NFM. The study concludes that a microgroove assisted CG-PCL NFM coating may benefit orthopedic implants.


Parent and child perspectives on family interactions related to melanoma risk and prevention after CDKN2A/p16 testing of minor children.
Predispositional genetic testing of children for adult-onset health risks is typically only used when prevention and screening measures have utility during childhood. Little is known about how children and their parents may use predispositional risk information, including whether it changes their interactions around risk-reducing prevention and screening behaviors. The current study examined perspectives on family interactions around skin cancer prevention and control practices through 1 year after test reporting and counseling among children who received melanoma predispositional genetic testing and their parents. Eighteen children (50% carriers, 56% male, mean age = 12.4 years) and 11 parents from 11 families participated in semi-structured interviews 1 month and 1 year after receiving the child's test result. Both parents (73%) and children (50%) reported making changes to family skin cancer prevention and control practices after receiving the test result. Parent- and child-reported discussions about melanoma prevention increased over time (36% parents and 61% children at 1 month, 73% parents and 67% at 1 year). One-quarter (27%) of parents and no children reported having conflicts about sun protection or screening 1 year after test reporting. A majority of parents (63%) reported treating their child differently at the 1-year follow-up, especially among carriers. Predispositional genetic testing for melanoma was associated with reported changes to plans for and discussions about sun protection, and high levels of parent-child collaboration to implement child sun protection. Future work could seek to identify child and parent factors and interactions that predict improved prevention and screening behaviors following pediatric predispositional genetic testing.


Intravenous vitamin C in the treatment of allergies: an interim subgroup analysis of a long-term observational study.
Objective Oxidative stress appears to be a key factor in the pathogenesis of allergic diseases and a potential therapeutic target in allergy treatment. Allergic diseases are reportedly associated with reduced plasma levels of ascorbate, which is a key physiological antioxidant. Ascorbate prevents excessive inflammation without reducing the defensive capacity of the immune system. Methods An interim analysis of a multicenter, prospective, observational study was conducted to investigate the change in disease-specific and nonspecific symptoms (fatigue, sleep disorders, depression, and lack of mental concentration) during adjuvant treatment with intravenous vitamin C (Pascorbin®; Pascoe, Giessen, Germany) in 71 patients with allergy-related respiratory or cutaneous indications. Results Between the start and end of treatment, the mean sum score of three disease-specific symptoms decreased significantly by 4.71 points and that of four nonspecific symptoms decreased significantly by 4.84 points. More than 50% of patients took no other allergy-related medication besides vitamin C. Conclusions Our observations suggest that treatment with intravenous high-dose vitamin C reduces allergy-related symptoms. Our observations form a basis for planning a randomized controlled clinical trial to obtain more definitive evidence of the clinical relevance of our findings. We also obtained evidence of ascorbate deficiency in allergy-related diseases. Clinical Trials NCT02422901.


Meal Delivery Programs Reduce The Use Of Costly Health Care In Dually Eligible Medicare And Medicaid Beneficiaries.
Delivering food to nutritionally vulnerable patients is important for addressing these patients' social determinants of health. However, it is not known whether food delivery programs can reduce the use of costly health services and decrease medical spending among these patients. We sought to determine whether home delivery of either medically tailored meals or nontailored food reduces the use of selected health care services and medical spending in a sample of adults dually eligible for Medicare and Medicaid. Compared with matched nonparticipants, participants had fewer emergency department visits in both the medically tailored meal program and the nontailored food program. Participants in the medically tailored meal program also had fewer inpatient admissions and lower medical spending. Participation in the nontailored food program was not associated with fewer inpatient admissions but was associated with lower medical spending. These findings suggest the potential for meal delivery programs to reduce the use of costly health care and decrease spending for vulnerable patients.


Efficient eradication of subcutaneous but not of autochthonous gastric tumors by adoptive T cell transfer in an SV40 T antigen mouse model.
In stomach cancer, there is a need for new therapeutic strategies, in particular for the treatment of unresectable tumors and micrometastases. We investigated the efficacy of immunotherapy in an autochthonous model of gastric cancer, the CEA424-SV40 T Ag (TAg) transgenic mice. Treatment efficacy against both the autochthonous tumors and s.c. tumors induced by the derived cell line mGC3 were assessed. In wild-type mice, a dendritic cell vaccine loaded with irradiated tumor cells combined with CpG oligonucleotides induced efficient cytotoxic T cell and memory responses against mGC3 s.c. tumors. In contrast, neither s.c. nor autochthonous tumors responded to vaccination in CEA424-SV40 TAg mice, indicating tolerance to the SV40 TAg. To examine whether tumors in these mice were principally accessible to immunotherapy, splenocytes from immune wild-type mice were adoptively transferred into CEA424-SV40 TAg transgenic mice. Treated mice showed complete regression of the s.c. tumors associated with intratumoral infiltrates of CD8 and CD4 T cells. In contrast, the autochthonous gastric tumors in the same mice were poorly infiltrated and did not regress. Thus, even in the presence of an active anti-tumoral T cell response, autochthonous gastric tumors do not respond to immunotherapy. This is the first comparison of the efficacy of adoptive T cell transfer between transplanted s.c. tumors and autochthonous tumors in the same animals. Our results suggest that in gastric cancer patients, even a strong anti-tumor T cell response will not efficiently penetrate the tumor in the absence of additional therapeutic strategies targeting the tumor microenvironment.


Nitric oxide is the mediator of both endothelium-dependent relaxation and hyperpolarization of the rabbit carotid artery.
It is controversial whether the endothelial cell release of nitric oxide (NO) or a different factor(s) accounts for endothelium-dependent hyperpolarization, because in many arteries endothelium-dependent relaxation and hyperpolarization resists inhibitors of NO synthase. The contribution of NO to acetylcholine-induced endothelium-dependent hyperpolarization and relaxation of the rabbit carotid artery was determined by measuring NO with electrochemical and chemiluminescence techniques. In the presence of phenylephrine to depolarize and contract the smooth muscle cells, acetylcholine caused concentration-dependent hyperpolarization and relaxation which were closely correlated to the release of NO. N(omega)-nitro-L-arginine methyl ester (30 microM) partially reduced the release of NO and caused a similar reduction in smooth muscle cell relaxation and hyperpolarization. To determine if the residual responses were mediated by another endothelium-derived mediator or NO released despite treatment with N(omega)-nitro-L-arginine methyl ester, N(omega)-nitro-L-arginine (300 microM) was added. The combined inhibitors further reduced, but did not eliminate, NO release, smooth muscle relaxation, and hyperpolarization. Hyperpolarization and relaxation to acetylcholine remained closely correlated with the release of NO in the presence of the inhibitors. In addition, the NO donor, SIN-1, caused hyperpolarization and relaxation which correlated with the concentrations of NO that it released. These studies indicate that (i) the release of NO by acetylcholine is only partially inhibited by these inhibitors of NO synthase when used even at high concentrations, and (ii) NO rather than another factor accounts fully for endothelium-dependent responses of the rabbit carotid artery.


Reorganization of genera in the families Rickettsiaceae and Anaplasmataceae in the order Rickettsiales: unification of some species of Ehrlichia with Anaplasma, Cowdria with Ehrlichia and Ehrlichia with Neorickettsia, descriptions of six new species combinations and designation of Ehrlichia equi and 'HGE agent' as subjective synonyms of Ehrlichia phagocytophila.
The genera Anaplasma, Ehrlichia, Cowdria, Neorickettsia and Wolbachia encompass a group of obligate intracellular bacteria that reside in vacuoles of eukaryotic cells and were previously placed in taxa based upon morphological, ecological, epidemiological and clinical characteristics. Recent genetic analyses of 16S rRNA genes, groESL and surface protein genes have indicated that the existing taxa designations are flawed. All 16S rRNA gene and groESL sequences deposited in GenBank prior to 2000 and selected sequences deposited thereafter were aligned and phylogenetic trees and bootstrap values were calculated using the neighbour-joining method and compared with trees generated with maximum-probability, maximum-likelihood, majority-rule consensus and parsimony methods. Supported by bootstrap probabilities of at least 54%, 16S rRNA gene comparisons consistently clustered to yield four distinct clades characterized roughly as Anaplasma (including the Ehrlichia phagocytophila group, Ehrlichia platys and Ehrlichia bovis) with a minimum of 96.1% similarity, Ehrlichia (including Cowdria ruminantium) with a minimum of 97.7% similarity, Wolbachia with a minimum of 95.6% similarity and Neorickettsia (including Ehrlichia sennetsu and Ehrlichia risticii) with a minimum of 94.9% similarity. Maximum similarity between clades ranged from 87.1 to 94.9%. Insufficient differences existed among E. phagocytophila, Ehrlichia equi and the human granulocytic ehrlichiosis (HGE) agent to support separate species designations, and this group was at least 98.2% similar to any Anaplasma species. These 16S rRNA gene analyses are strongly supported by similar groESL clades, as well as biological and antigenic characteristics. It is proposed that all members of the tribes Ehrlichieae and Wolbachieae be transferred to the family Anaplasmataceae and that the tribe structure of the family Rickettsiaceae be eliminated. The genus Anaplasma should be emended to include Anaplasma (Ehrlichia) phagocytophila comb. nov. (which also encompasses the former E. equi and the HGE agent), Anaplasma (Ehrlichia) bovis comb. nov. and Anaplasma (Ehrlichia) platys comb. nov., the genus Ehrlichia should be emended to include Ehrlichia (Cowdria) ruminantium comb. nov. and the genus Neorickettsia should be emended to include Neorickettsia (Ehrlichia) risticii comb. nov. and Neorickettsia (Ehrlichia) sennetsu comb. nov.


Implications of stx loss for clinical diagnostics of Shiga toxin-producing Escherichia coli.
The dynamics related to the loss of stx genes from Shiga toxin-producing Escherichia coli remain unclear. Current diagnostic procedures have shortcomings in the detection and identification of STEC. This is partly owing to the fact that stx genes may be lost during an infection or in the laboratory. The aim of the present study was to provide new insight into in vivo and in vitro stx loss in order to improve diagnostic procedures. Results from the study support the theory that loss of stx is a strain-related phenomenon and not induced by patient factors. It was observed that one strain could lose stx both in vivo and in vitro. Whole genome comparison of stx-positive and stx-negative isolates from the same patient revealed that different genomic rearrangements, such as complete or partial loss of the parent prophage, may be factors in the loss of stx. Of diagnostic interest, it was shown that patients can be co-infected with different E. coli pathotypes. Therefore, identification of eae-positive, but stx-negative isolates should not be interpreted as "Shiga toxin-lost" E. coli without further testing. Growth and recovery of STEC were supported by different selective agar media for different strains, arguing for inclusion of several media in STEC diagnostics.


Transcriptome profiling analysis of the response to walnut polyphenol extract in Helicobacter pylori-infected cells.
Dietary intervention to prevent Helicobacter pylori (H. pylori)-associated gastric diseases seems to be ideal with no risk of bacterial resistance, safe long-term intervention, and correcting pathogenic mechanisms including rejuvenation of precancerous atrophic gastritis and anti-mutagenesis. A transcriptome as set of all RNAs transcribed by certain tissues or cells demonstrates gene functions and reveals the molecular mechanism of specific biological processes against diseases. Here, we have performed RNAseq and bioinformatic analysis to explain proof of concept that walnut intake can rescue from H. pylori infection and explore unidentified mode of actions of walnut polyphenol extract (WPE). As results, BIRC3, SLC25A4, f3 transcription, VEGFA, AZU1, HMOX1, RAB3A, RELBTNIP1, ETFB, INPP5J, PPME1, RHOB, TPI1, FOSL1, JUND.RELB, KLF2, MUC1, NDRG1, ALDOA, ENO1, PFKP, GPI, GDF15, and NRTN genes were newly discovered to be enriched with WPE, whereas CCR4, BLNK, CCR7, CXCR4, CDO1, KLSG1, SELE, RASGRP2, PIK3R3, TSPAN32, HOXC-AS3, HCG8, BTNL8, and CXCL3 genes as inhibitory targets by WPE in H. pylori infection. We identified additional genes what WPE afforded actions of avoiding H. pylori-driven onco-inflammation and rejuvenating precancerous atrophic gastritis. Conclusively, after applying RNAseq analysis in order to document walnut intake for precision medicine against H. pylori infection, significant transcriptomic profiling applicable for validation were drawn.


Determination of the molecular weight of neuronectin, a conditioned medium-derived, substrate-binding neurite-extension factor: comparison with laminin using radiation-inactivation analysis.
Neurite outgrowth from a wide variety of peripheral neurons is stimulated and may be directed in culture by a substrate-binding factor(s) derived from medium conditioned over numerous types of cells. This factor, or family of factors, which we shall call neuronectin by reason of its ability to serve as an attachment molecule for neurons, has been studied by target-size analysis using radiation inactivation. The radiation-inactivation method has the unique advantage of providing a means for determining the actual functional size of a biologically active molecule irrespective of its state of purification. By this method, the functional size of the major neurite outgrowth-promoting activity (neuronectin) from mouse heart cell conditioned medium has been found to be 350,000 Da. While neuronectin has not yet been purified, determination of the actual functional size provides a framework within which possible models must fit. Thus, although neurite outgrowth-promoting activity in this system is found to be associated with a complex containing laminin, fibronectin, heparan sulfate proteoglycan, and other extracellular matrix molecules, the total size of the functional molecule or molecular complex serving as the major source of activity is limited to 350,000 Da. Consequently, our results suggest that neuronectin from mouse heart cell-conditioned medium is different from laminin (Mr approximately 900,000), a molecule that also exhibits neurite-promoting activity. In addition to the difference in molecular size, neuronectin and laminin differ in that laminin, unlike neuronectin, gives rise to toxic or inhibitory products when exposed to high-energy radiation.


Long-term memory search across the visual brain.
Signal transmission from the human retina to visual cortex and connectivity of visual brain areas are relatively well understood. How specific visual perceptions transform into corresponding long-term memories remains unknown. Here, I will review recent Blood Oxygenation Level-Dependent functional Magnetic Resonance Imaging (BOLD fMRI) in humans together with molecular biology studies (animal models) aiming to understand how the retinal image gets transformed into so-called visual (retinotropic) maps. The broken object paradigm has been chosen in order to illustrate the complexity of multisensory perception of simple objects subject to visual--rather than semantic--type of memory encoding. The author explores how amygdala projections to the visual cortex affect the memory formation and proposes the choice of experimental techniques needed to explain our massive visual memory capacity. Maintenance of the visual long-term memories is suggested to require recycling of GluR2-containing α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors (AMPAR) and β(2)-adrenoreceptors at the postsynaptic membrane, which critically depends on the catalytic activity of the N-ethylmaleimide-sensitive factor (NSF) and protein kinase PKMζ.


Regional brain abnormalities associated with long-term heavy cannabis use.
Cannabis is the most widely used illicit drug in the developed world. Despite this, there is a paucity of research examining its long-term effect on the human brain. To determine whether long-term heavy cannabis use is associated with gross anatomical abnormalities in 2 cannabinoid receptor-rich regions of the brain, the hippocampus and the amygdala. Cross-sectional design using high-resolution (3-T) structural magnetic resonance imaging. Participants were recruited from the general community and underwent imaging at a hospital research facility. Fifteen carefully selected long-term (>10 years) and heavy (>5 joints daily) cannabis-using men (mean age, 39.8 years; mean duration of regular use, 19.7 years) with no history of polydrug abuse or neurologic/mental disorder and 16 matched nonusing control subjects (mean age, 36.4 years). Volumetric measures of the hippocampus and the amygdala combined with measures of cannabis use. Subthreshold psychotic symptoms and verbal learning ability were also measured. Cannabis users had bilaterally reduced hippocampal and amygdala volumes (P = .001), with a relatively (and significantly [P = .02]) greater magnitude of reduction in the former (12.0% vs 7.1%). Left hemisphere hippocampal volume was inversely associated with cumulative exposure to cannabis during the previous 10 years (P = .01) and subthreshold positive psychotic symptoms (P < .001). Positive symptom scores were also associated with cumulative exposure to cannabis (P = .048). Although cannabis users performed significantly worse than controls on verbal learning (P < .001), this did not correlate with regional brain volumes in either group. These results provide new evidence of exposure-related structural abnormalities in the hippocampus and amygdala in long-term heavy cannabis users and corroborate similar findings in the animal literature. These findings indicate that heavy daily cannabis use across protracted periods exerts harmful effects on brain tissue and mental health.


Chronic alcohol intake differently influences glucose metabolism according to nutritional status.
We investigated the potential different effects of a chronic alcohol intake on glucose metabolism according to nutritional status in growing rats. Eighty weanling 4-week-old male Sprague Dawley rats were fed with low (5%, wt/wt) or control (22%) protein diet for 8 weeks. Each group was subdivided into alcohol (5 g/kg(-1) x day(-1)) or saline gavage rats during the last 4 weeks. At 12 weeks of age, we measured the weights of the body, pancreas, and epididymal fat; glycogen synthase activity of gastrocnemius muscle; and insulin content of the pancreas. We performed an ip glucose tolerance test and a euglycemic hyperinsulinemic clamp test. Weight gain was almost arrested in protein-deficient rats. The relative weight and insulin content of the pancreas and glycogen synthase activity were not different among the four groups, but the relative amount of epididymal fat decreased only in protein-deficient saline rats. Insulin response after glucose challenge and glucose disposal rate during the euglycemic clamp were both markedly decreased in protein-deficient saline rats, but not changed in protein-deficient alcohol rats. Protein-deficiency per se causes deterioration both in insulin secretory function and in sensitivity, but these defects are protected by a chronic alcohol intake. In a protein-sufficient state, alcohol intake gives no additional effects on glucose metabolism. Therefore, according to individual nutritional status, the metabolic effect of alcohol intake appears differently.


Demographical Aspects of Central Large Lumbar Disc Herniation.
The purpose of this study was to investigate the risk factors for the development of the central large disc herniations and to compare the demographic data between central mass prolapse and broad-based central disc herniation. Between 2002 and 2007, 1630 patients underwent surgery and a large disc herniation was the main problem in 59 patients (3.6%). We performed a retrospective analysis of the demographic data of these patients. Magnetic resonance (MR) images were evaluated according to the disc type and level. Variables were evaluated both at baseline and follow-up, with special emphasis on physical job characteristics, sports activities, and MR - based morphologic findings. Central large disc herniation was diagnosed in 59 patients consisting of 41 males and 18 females. The average age was 34.7 years. 36 patients had a central mass prolapse that occupied more than 50% of the spinal canal. Intraoperative observations confirmed that 29 out of 36 central disc prolapse patients (80.5%) had intact posterior longitudinal ligaments. Interestingly, the condition in these 29 patients was found to have a direct relation with age and occupation or other body training sports activities. The size of the large central disc herniation, physical activity, age and gender are major factors in the development of disc herniation.


Mouse models for hereditary nonpolyposis colorectal cancer.
Hemizygous germ-line defects in mismatch repair (MMR) genes underlie hereditary nonpolyposis colorectal cancer (HNPCC). Loss of the wild-type allele results in a mutator phenotype, accelerating tumorigenesis. Tumorigenesis specifically occurs in the gastrointestinal and genitourinary tracts; the cause of this tissue specificity is elusive. To understand the etiology and tissue distribution of tumors in HNPCC, we have developed mouse models carrying a deficiency in the MMR gene Msh2. Most of the completely Msh2-deficient mice succumbed to lymphomas at an early age; lymphomagenesis was synergistically enhanced by exposure to ethylnitrosourea. Lymphomas were absent in immunocompromised Tap1-/-;Msh2-/- mice; these mice generally succumbed to HNPCC-like tumors. Together, these data suggest that the HNPCC tumor spectrum is determined by exposure of MMR-deficient cells to exogenous mutagens, rather than by tissue-specific loss of the wild-type MMR allele or by immune surveillance. Msh2 hemizygous mice had an elevated tumor incidence that, surprisingly, was rarely correlated with loss of the Msh2+ allele. To develop a model for intestinal tumorigenesis in HNPCC, we introduced the Min allele of the Apc tumor suppressor gene. We observed loss of the wild-type Msh2 allele in a significant fraction of intestinal tumors in Apc+/Min;Msh2+/- mice. In some of the latter tumors, one area of the tumor displayed loss of the Msh2+ allele, but not of the Apc+ allele, whereas another area displayed the inverse genotype. This apparent biclonality might indicate a requirement for collaboration between independent tumor clones during intestinal tumorigenesis.


Detection of K-ras mutations in lung carcinomas: relationship to prognosis.
The K-ras mutation is one of the most common genetic alterations found in human lung cancer. To evaluate the prognostic value of ras gene alterations in lung cancer in a U.S. population, we have screened 173 human lung tumors, which included 127 adenocarcinomas, 37 squamous carcinomas, and 9 adenosquamous carcinomas, for mutations in the K-ras gene using the combination of the PCR and denaturing gradient gel electrophoresis. Forty-three tumors contained K-ras mutations. Of these, 41 were identified among the adenocarcinomas (32%), 1 among the squamous carcinomas (2.7%), and 1 among the adenosquamous carcinomas (11%). Forty of these mutations were found in codon 12 and consisted of 24 G to T transversions, 12 G to A transitions, 2 G to C transversions, and 1 double GG to TT mutation. Two other G to T transversions were found in codon 13, and 1 A to C transversion was found in codon 61. The data showed that gender did not seem to affect the incidence and the types of the K-ras mutations or amino acid changes. Examination of the mutations in adenocarcinomas in relation to overall survival showed no difference in adenocarcinomas with K-ras mutations compared with K-ras-negative adenocarcinomas. However, the substitution of the wild-type GGT (glycine) at codon 12 with a GTT (valine) or a CGT (arginine) showed a strong trend (P = 0.07) toward a poorer prognosis compared with wild-type or other amino acid substitutions. Substitution of the wild-type glycine for aspartate (GAT) showed a strong trend (P = 0.06) for a better outcome than the valine or arginine substitution. Although these trends will require larger patient populations for verification, these data suggest that the prognostic significance of K-ras mutations may depend on the amino acid substitution in the p21(ras) protein.


Target organs for carcinogenicity of chemicals and industrial exposures in humans: a review of results in the IARC monographs on the evaluation of the carcinogenic risk of chemicals to humans.
Epidemiological observations indicate that cancers affecting different organs and systems in humans have different causes. At the descriptive level, cancer incidence and mortality rates exhibit patterns of geographic and temporal variation which are distinct and separate for each cancer site and even, at a given site, for different histological types (for instance, increasing squamous cell carcinoma of the lung and decreasing stomach cancer in most developed countries in recent decades). The existence of these distinct patterns in itself indicates that different causes are at the origin of cancers at different sites. Hence, it is of scientific and practical importance not only to identify agents that are carcinogenic to humans but also to specify as definitely as possible the target organ(s) of their action. This is done in the present review of results in the International Agency for Research on Cancer Monographs on the Evaluation of the Carcinogenic Risk of Chemicals to Humans.


Nosocomial diarrhoea due to Clostridium difficile.
The purpose of this review is to summarize recent developments in the diagnosis, epidemiology, treatment and prevention of nosocomial diarrhoea due to Clostridium difficile. Twenty-five years after its discovery, the diagnosis of C. difficile-associated diarrhoea is still problematic with laboratories trying to reconcile the time and expense of the diagnostic process. Newer molecular techniques may offer hope. With the introduction of new antibiotics into clinical practice, confusion has arisen about the risk they pose for C. difficile-associated diarrhoea. Strains of C. difficile that fail to produce an active toxin A are an emerging problem and good molecular epidemiology is required to determine whether highly infectious clones exist. Little progress has been made in the treatment of recurrent C. difficile-associated diarrhoea; however, the development of a vaccine is imminent. More effort is being made to rid the hospital environment of C. difficile through infection-control procedures or changes in antibiotic-prescribing policies. C. difficile continues to be a major nosocomial infection in many health-care institutions throughout the world. Strategies that reduce exposure to the organism or to antibiotics will have an impact on rates of C. difficile-associated diarrhoea.


Comparison of Polydopamine-Coated Mesoporous Silica Nanorods and Spheres for the Delivery of Hydrophilic and Hydrophobic Anticancer Drugs.
Mesoporous silica nanoparticles (MSNs) have been widely studied as drug delivery systems in nanomedicine. Surface coating of MSNs have enabled them to perform efficiently in terms of bioavailability, biocompatibility, therapeutic efficacy and targeting capability. Recent studies have suggested the use of polydopamine (PDA) as a facilitative coating for MSNs that provides sustained and pH-responsive drug release, owing to the adhesive "molecular-glue" function of PDA. This further endows these hybrid MSN@PDA particles with the ability to carry large amounts of hydrophilic drugs. In this study, we expand the feasibility of this platform in terms of exploring its ability to also deliver hydrophobic drugs, as well as investigate the effect of particle shape on intracellular delivery of both a hydrophilic and hydrophobic anticancer drug. MSN@PDA loaded with doxorubicin (hydrophilic) and fingolimod (hydrophobic) was studied via a systematic in vitro approach (cellular internalization, intracellular drug distribution and cytotoxicity). To promote the cellular uptake of the MSN@PDA particles, they were further coated with a polyethylene imine (PEI)-polyethylene glycol (PEG) copolymer. Drug-loaded, copolymer-coated MSN@PDA showed effective cellular uptake, intracellular release and an amplified cytotoxic effect with both doxorubicin and fingolimod. Additionally, rods exhibited delayed intracellular drug release and superior intracellular uptake compared to spheres. Hence, the study provides an example of how the choice and design of drug delivery systems can be tuned by the need for performance, and confirms the PDA coating of MSNs as a useful drug delivery platform beyond hydrophilic drugs.


A novel POSS-coated quantum dot for biological application.
Quantum dots (QDs) are fluorescent semiconductor nanocrystals that have the potential for major advancements in the field of nanomedicine through their unique photophysical properties. They can potentially be used as fluorescent probes for various biomedical imaging applications, including cancer localization, detection of micrometastasis, image guided surgery, and targeted drug delivery. Their main limitation is toxicity, which requires a biologically compatible surface coating to shield the toxic core from the surrounding environment. However, this leads to an increase in QD size that may lead to problems of excretion and systemic sequestration. We describe a one pot synthesis, characterization, and in vitro cytotoxicity of a novel polyhedral oligomeric silsesquioxane (POSS)-coated CdTe-cored QD using mercaptosuccinic acid (MSA) and D-cysteine as stabilizing agents. Characterization was performed using transmission electron microscopy Fourier transform infrared spectroscopy, and photoluminescence studies. POSS-coated QDs demonstrated high colloidal stability and enhanced photostability on high degrees of ultraviolet (UV) excitation compared to QDs coated with MSA and D-cysteine alone (P value < 0.05). In vitro toxicity studies showed that both POSS and MSA-QDs were significantly less toxic than ionized salts of Cd(+2) and Te(-2). Confocal microscopy confirmed high brightness of POSS-QDs in cells at both 1 and 24 hours, indicating that these QDs are rapidly taken up by cells and remain photostable in a biological environment. We therefore conclude that a POSS coating confers biological compatibility, photostability, and colloidal stability while retaining the small size and unique photophysical properties of the QDs. The amphiphilic nature of the coating allows solubility in aqueous solutions and rapid transfer across cell membranes, enabling the use of lower concentrations of the QDs for an overall reduced toxicity particularly for prolonged live cell and in vivo imaging applications.


Celiac disease: from pathogenesis to novel therapies.
Celiac disease has become one of the best-understood HLA-linked disorders. Although it shares many immunologic features with inflammatory bowel disease, celiac disease is uniquely characterized by (1) a defined trigger (gluten proteins from wheat and related cereals), (2) the necessary presence of HLA-DQ2 or HLA-DQ8, and (3) the generation of circulating autoantibodies to the enzyme tissue transglutaminase (TG2). TG2 deamidates certain gluten peptides, increasing their affinity to HLA-DQ2 or HLA-DQ8. This generates a more vigorous CD4(+) T-helper 1 T-cell activation, which can result in intestinal mucosal inflammation, malabsorption, and numerous secondary symptoms and autoimmune diseases. Moreover, gluten elicits innate immune responses that act in concert with the adaptive immunity. Exclusion of gluten from the diet reverses many disease manifestations but is usually not or less efficient in patients with refractory celiac disease or associated autoimmune diseases. Based on the advanced understanding of the pathogenesis of celiac disease, targeted nondietary therapies have been devised, and some of these are already in phase 1 or 2 clinical trials. Examples are modified flours that have been depleted of immunogenic gluten epitopes, degradation of immunodominant gliadin peptides that resist intestinal proteases by exogenous endopeptidases, decrease of intestinal permeability by blockage of the epithelial ZOT receptor, inhibition of intestinal TG2 activity by transglutaminase inhibitors, inhibition of gluten peptide presentation by HLA-DQ2 antagonists, modulation or inhibition of proinflammatory cytokines, and induction of oral tolerance to gluten. These and other experimental therapies will be discussed critically.


Ocular vestibular evoked myogenic potentials as a test for myasthenia gravis.
To explore whether ocular vestibular evoked myogenic potentials (oVEMP) can be used to detect a decrement in the extraocular muscle activity of patients with myasthenia gravis (MG). Twenty-seven patients with MG, including 13 with isolated ocular and 14 with generalized MG, and 28 healthy controls participated. We applied repetitive vibration stimuli to the forehead and recorded the activity of the inferior oblique muscle with 2 surface electrodes placed beneath the eyes. To identify the oVEMP parameters with the highest sensitivity and specificity, we evaluated the decrement over 10 stimulus repetitions at 3 different repetition rates (3 Hz, 10 Hz, and 20 Hz). Repetitive stimulation at 20 Hz yielded the best differentiation between patients with MG and controls with a sensitivity of 89% and a specificity of 64% when using a unilateral decrement of ≥15.2% as cutoff. When using a bilateral decrement of ≥20.4% instead, oVEMP allowed differentiation of MG from healthy controls with 100% specificity, but slightly reduced sensitivity of 63%. For both cutoffs, sensitivity was similar in isolated ocular and generalized MG. Our study demonstrates that the presence of an oVEMP decrement is a sensitive and specific marker for MG. This test allows direct and noninvasive examination of extraocular muscle activity, with similarly good diagnostic accuracy in ocular and generalized MG. Thus, oVEMP represents a promising diagnostic tool for MG. This study provides Class III evidence that oVEMP testing accurately identifies patients with MG with ocular symptoms (sensitivity 89%, specificity 64%).


Poisoned patients as potential organ donors: postal survey of transplant centres and intensive care units.
The number of patients awaiting allograft transplantation in the UK exceeds the number of organs offered for transplantation each year. Most organ donors tend to be young, fit and healthy individuals who die because of trauma or sudden cardiac arrest. Patients who die from drug and poison intoxication tend to have similar characteristics but are less frequently offered as potential organ donors. A postal questionnaire survey of all transplantation centres and an equal number of intensive care units in the UK was undertaken. The use of kidney, heart, lung, liver and pancreas transplants from poisoned patients following deliberate methanol ingestion, cardiac arrest presumed secondary to cocaine overdose, accidental domestic carbon monoxide inhalation and industrial cyanide exposure were used as case scenarios. Response rates were 70% for transplantation centres and 50% for intensive care unit directors. Over 80% of organs would be offered or discussed with transplant coordinators by intensive care unit directors. Transplantation physicians/surgeons would consider transplanting organs in up to 100% of case scenarios, depending on the organ and poisoning or intoxication involved. The postal survey presented here shows that most transplantation physicians and surgeons and intensive care unit directors would consider those who die following acute drug intoxication and poisoning as potential organ donors. The previously reported literature shows in general that transplanted organs from poisoned patients have good long-term survival, although the number of reports is small. Poisoned patients are another pool of organ donors who at present are probably underused by transplantation services.


"It's All about Breaking down Those Barriers…": Exploring Survivors' Perspectives on Services and Treatment Needs following Commercial Sexual Exploitation during Childhood.
Purpose: In recent years, there has been an increase in funds and services available to address the needs of commercially sexually exploited children (CSEC). While previous studies have explored the mental, behavioral, and physical needs of CSEC victims and survivors, few studies have focused on the service needs CSEC victims and survivors themselves deem most important. The current study seeks to bridge this gap by garnering American CSEC victims' and survivors' perspectives regarding CSEC service strengths, weaknesses, and gaps.Method: Semi-structured, in-depth interviews were conducted with 13 adult survivors of CSEC to examine their perceptions of current CSEC services in the United States. All interviews were recorded, transcribed verbatim, and imported into a qualitative data analysis program. Using an inductive approach, two major themes emerged: short-term needs and long-term needs.Results: Short-term needs included victim identification, housing, and emergency medical care. Long-term needs included life-skills, community building, legal assistance, and medical care. The results point to the complex needs of CSEC victims/survivors.Discussion: While CSEC services continue to develop, there remain many gaps in care in the services available. Study findings provide valuable insight to practitioners and researchers alike and identify the most critical needs of CSEC victims and survivors. Implications for practice and research are discussed.


Health Informatics Tools to Improve Utilization of Laboratory Tests.
Herein, we discuss improper test utilization practices and their implications on delivery of health care, as well as providing a brief explanation of the means to reduce such practices by improvement of personnel factors, particularly involving physicians. The article also elaborates on ways to mitigate improperly utilized test practices using appropriate health informatics technologies to their maximum possible capacities.


Contrast-enhanced portal magnetic resonance angiography in dogs with suspected congenital portal vascular anomalies.
Contrast-enhanced multiphase magnetic resonance angiography (CE-MRA) was used in 17 dogs with a suspected congenital portal vascular anomaly. Portal vascular anomalies were identified in 16 of the 17 dogs. Eleven had a single intrahepatic portocaval shunt (two central divisional, three right divisional, and six left divisional), one dog had a double intrahepatic portocaval shunt, one dog had a hepatic arteriovenous malformation, one dog had a complex intrahepatic porto-caval shunt. Two dogs had an extrahepatic portosystemic shunt and no shunt was identified in one dog. Total imaging time was <10 min and image quality was good to excellent in all dogs. Portal CE-MRA is a feasible, fast and non invasive technique to diagnose portal vascular anomalies in dogs, with a large field-of-view and good anatomic depiction of the abnormal vessels. Based on these results, CE-MRA is an efficient imaging technique for the diagnosis of portal vascular anomalies in dogs.


Renal dysfunction in liver transplant recipients: evaluation of the critical issues.
Major progress has been made in the field of liver transplantation since the first procedure was performed nearly 50 years ago. Despite these improvements, renal dysfunction before and after liver transplantation remains a major complicating factor associated with increased health care costs, morbidity, and mortality. Creatinine-based estimates of renal function are inaccurate in the setting of end-stage liver disease and often lead to underdiagnosis and late intervention. This issue is critical in that it is important to understand both the etiology and chronicity of renal dysfunction before liver transplantation because the treatment clearly varies, especially with respect to simultaneous liver-kidney (SLK) transplantation. Because of the scarcity of available grafts, identifying appropriate candidates for SLK transplantation is crucial. Hepatorenal syndrome is common in liver transplant candidates; however, other etiologies of renal dysfunction need to be considered. Renal dysfunction after liver transplantation is common and may have an acute or chronic presentation. Although calcineurin inhibitors (CNIs) have been associated with post-liver transplant nephrotoxicity, their role may be overestimated, and other contributing etiologies should remain in a clinician's differential diagnosis. Alternatives to CNIs have been evaluated; however, a safe immunosuppressive regimen that achieves the preservation of renal function in liver transplant recipients remains to be established. In this review of the literature, renal dysfunction in the setting of liver transplantation is evaluated, and the critical issues that are barriers to improved outcomes are highlighted.


Dabigatran versus warfarin in patients with atrial fibrillation: an analysis of patients undergoing cardioversion.
The Randomized Evaluation of Long-Term Anticoagulation Therapy (RE-LY) trial compared dabigatran 110 mg BID (D110) and 150 mg BID (D150) with warfarin for stroke prevention in 18 113 patients with nonvalvular atrial fibrillation. Cardioversion on randomized treatment was permitted. Precardioversion transesophageal echocardiography was encouraged, particularly in dabigatran-assigned patients. Data from before, during, and 30 days after cardioversion were analyzed. A total of 1983 cardioversions were performed in 1270 patients: 647, 672, and 664 in the D110, D150, and warfarin groups, respectively. For D110, D150, and warfarin, transesophageal echocardiography was performed before 25.5%, 24.1%, and 13.3% of cardioversions, of which 1.8%, 1.2%, and 1.1% were positive for left atrial thrombi. Continuous treatment with study drug for ≥3 weeks before cardioversion was lower in D110 (76.4%) and D150 (79.2%) compared with warfarin (85.5%; P<0.01 for both). Stroke and systemic embolism rates at 30 days were 0.8%, 0.3%, and 0.6% (D110 versus warfarin, P=0.71; D150 versus warfarin, P=0.40) and similar in patients with and without transesophageal echocardiography. Major bleeding rates were 1.7%, 0.6%, and 0.6% (D110 versus warfarin, P=0.06; D150 versus warfarin, P=0.99). This study is the largest cardioversion experience to date and the first to evaluate a novel anticoagulant in this setting. The frequencies of stroke and major bleeding within 30 days of cardioversion on the 2 doses of dabigatran were low and comparable to those on warfarin with or without transesophageal echocardiography guidance. Dabigatran is a reasonable alternative to warfarin in patients requiring cardioversion.


Surgical outcomes of patients with diffuse-type tenosynovial giant-cell tumours: an international, retrospective, cohort study.
Diffuse-type tenosynovial giant-cell tumour is a rare, locally aggressive, and difficult-to-treat soft tissue tumour. Clinical and surgical outcomes depend on multiple factors, including preoperative diagnostic assessment, the localisation and extent of disease, and possibly the choice of treatment modalities by orthopaedic surgeons. We did a retrospective cohort study to characterise global surgical treatment protocols, and assess surgical outcomes, complications, and functional results in patients with diffuse-type tenosynovial giant-cell tumours. In this international, multicentre, retrospective cohort study, we included consecutive patients treated in 31 sarcoma reference centres between Jan 1, 1990, and Dec 31, 2017. Eligible patients were of any age and had histologically proven diffuse-type tenosynovial giant-cell tumour of large joints. Patient data were retrieved from the local databases of participating centres. Patients with localised-type tenosynovial giant-cell tumour were excluded. In the analysis, we only included patients with complete core criteria data regarding admission status, date of treatment, type of treatment at participating centre, and first local recurrence after treatment. We used a non-parametric method to estimate recurrence-free survival at 3, 5, and 10 years after initial surgical resection in a tertiary centre. We used a multivariate Cox regression model to estimate the effect of risk factors. We also present subgroup analyses of disease status at presentation (primary vs recurrent disease) and recurrence-free survival by surgery type (open surgery vs arthroscopic synovectomy), and prespecified risk factors were tested in a univariate and multivariable analyses, with an endpoint of first local recurrence after treatment in a tertiary centre. Data collection for these analyses occurred between January, 2016, and May, 2018. We received the records of 1192 patients, of which 966 (81%) were surgically treated and had complete information on core criteria. 445 patients were admitted with therapy-naive disease of the knee and were primarily treated in a tertiary centre. Since patients with wait and see treatment do not have a starting date of treatment, these patients were excluded in the calculation of median follow-up time for all patients. For this calculation we used time of surgery as a starting date. 758 (64%) of 1192 patients had knee involvement and 628 (54%) of 1163 patients with complete data on type of surgery had one-staged open synovectomy. At a median follow-up of 54 months (IQR 27-97), recurrent disease developed in 425 (44%) of all 966 surgically treated cases, and recurrence-free survival was 62% (95% CI 59-65) at 3 years, 55% (51-58) at 5 years, and 40% (35-45) at 10 years. Surgical complications were reported in 105 (12%) of 906 patients who had complete data on surgical complications. Pain improved after surgical treatment in 255 (59%) of 434 patients and swelling improved in 328 (72%) of 453 patients who had complete data. This study of patients with diffuse-type tenosynovial giant-cell tumour provides a comprehensive and up-to-date disease overview, assessing the clinical profile and management of the disease in multiple specialised referral centres. Surgical treatment of diffuse-type tenosynovial giant cell tumours is not a definitive treatment for every patient because it involves a high risk for local recurrent disease and a relatively high risk for postoperative complications. After surgical treatment in treatment-naive patients, risk factors for recurrent disease in individual patients were not identified in what we believe is the largest cohort to date. Daiichi Sankyo.


Gene transfer in Cryptococcus neoformans by use of biolistic delivery of DNA.
A transformation scheme for Cryptococcus neoformans to yield high-frequency, integrative events was developed. Adenine auxotrophs from a clinical isolate of C. neoformans serotype A were complemented by the cryptococcal phosphoribosylaminoimidazole carboxylase gene (ade2) with a biolistic DNA delivery system. Comparison of two DNA delivery systems (electroporation versus a biolistic system) showed notable differences. The biolistic system did not require linear vectors and transformed each auxotrophic strain at similar frequencies. Examination of randomly selected transformants by biolistics showed that 15 to 40% were stable, depending on the recipient auxotroph, with integrative events identified in all stable transformants by DNA analysis. Although the ade2 cDNA copy transformed at a low frequency, DNA analysis found homologous recombination in each of these transformants. DNA analysis of stable transformants receiving genomic ade2 revealed ectopic integration in a majority of cases, but approximately a quarter of the transformants showed homologous recombination with vector integration or gene replacement. This system has the potential for targeted gene disruption, and its efficiency will also allow for screening of DNA libraries within C. neoformans. Further molecular strategies to study the pathobiology of this pathogenic yeast are now possible with this transformation system.


Oil palm genome sequence reveals divergence of interfertile species in Old and New worlds.
Oil palm is the most productive oil-bearing crop. Although it is planted on only 5% of the total world vegetable oil acreage, palm oil accounts for 33% of vegetable oil and 45% of edible oil worldwide, but increased cultivation competes with dwindling rainforest reserves. We report the 1.8-gigabase (Gb) genome sequence of the African oil palm Elaeis guineensis, the predominant source of worldwide oil production. A total of 1.535 Gb of assembled sequence and transcriptome data from 30 tissue types were used to predict at least 34,802 genes, including oil biosynthesis genes and homologues of WRINKLED1 (WRI1), and other transcriptional regulators, which are highly expressed in the kernel. We also report the draft sequence of the South American oil palm Elaeis oleifera, which has the same number of chromosomes (2n = 32) and produces fertile interspecific hybrids with E. guineensis but seems to have diverged in the New World. Segmental duplications of chromosome arms define the palaeotetraploid origin of palm trees. The oil palm sequence enables the discovery of genes for important traits as well as somaclonal epigenetic alterations that restrict the use of clones in commercial plantings, and should therefore help to achieve sustainability for biofuels and edible oils, reducing the rainforest footprint of this tropical plantation crop.


What outcomes are important to patients with mild cognitive impairment or Alzheimer's disease, their caregivers, and health-care professionals? A systematic review.
Clinical trials involving patients with Alzheimer's disease (AD) continue to try to identify disease-modifying treatments. Although trials are designed to meet regulatory and registration requirements, many do not measure outcomes of the disease most relevant to key stakeholders. A systematic review sought research that elicited information from people with AD, their caregivers, and health-care professionals on which outcomes of the disease were important. Studies published in any language between 2008 and 2017 were included. Participants in 34 studies described 32 outcomes of AD. These included clinical (memory, mental health), practical (ability to undertake activities of daily living, access to health information), and personal (desire for patient autonomy, maintenance of identity) outcomes of the disease. Evidence elicited directly from the people most affected by AD reveals a range of disease outcomes that are relevant to them but are not commonly captured in clinical trials of new treatments.


Outdoor smoking ban at a cancer center: attitudes and smoking behavior among employees and patients.
Policies restricting indoor worksite tobacco use began being implemented more than a decade ago. More recently, the scope of these policies has been expanding to outdoors, with hospitals leading the trend in restricting smoking throughout their grounds. However, research on the effects such bans have on employees is scarce. The purpose of the current study was to examine the impact of a campus-wide smoking ban on employees and patients at a cancer center. Employees completed anonymous questionnaires during the months before (n = 607; 12% smokers) and 3 months after the ban implementation (n = 511; 10% smokers). Patients (n = 278; 23% smokers) completed an anonymous questionnaire preban. Results showed that 86% of nonsmokers, 20% of employees who smoke, and 57% of patients who smoke supported the ban. More than 70% of smokers were planning or thinking about quitting at both time points and nearly one-third were interested in cessation services following the ban. Before the ban, 32% expected the ban to have a negative effect on job performance and 41% thought their smoking before and after work would increase. Postban, 22% reported a negative impact on job performance, 35% increased smoking before and after work, and 7% quit. Overall, these data revealed an overwhelming support for an outdoor smoking ban by nonsmoker employees and patients. Although a majority of employee smokers opposed the ban, a significant proportion was interested in cessation. Compared with preban expectations, a lower proportion experienced negative effects postban. Findings suggest a need for worksite cessation programs to capitalize on the window of opportunity created by tobacco bans, while also addressing concerns about effects on work performance.


Not all are free-living: high-throughput DNA metabarcoding reveals a diverse community of protists parasitizing soil metazoa.
Protists, the most diverse eukaryotes, are largely considered to be free-living bacterivores, but vast numbers of taxa are known to parasitize plants or animals. High-throughput sequencing (HTS) approaches now commonly replace cultivation-based approaches in studying soil protists, but insights into common biases associated with this method are limited to aquatic taxa and samples. We created a mock community of common free-living soil protists (amoebae, flagellates, ciliates), extracted DNA and amplified it in the presence of metazoan DNA using 454 HTS. We aimed at evaluating whether HTS quantitatively reveals true relative abundances of soil protists and at investigating whether the expected protist community structure is altered by the co-amplification of metazoan-associated protist taxa. Indeed, HTS revealed fundamentally different protist communities from those expected. Ciliate sequences were highly over-represented, while those of most amoebae and flagellates were under-represented or totally absent. These results underpin the biases introduced by HTS that prevent reliable quantitative estimations of free-living protist communities. Furthermore, we detected a wide range of nonadded protist taxa probably introduced along with metazoan DNA, which altered the protist community structure. Among those, 20 taxa most closely resembled parasitic, often pathogenic taxa. Therewith, we provide the first HTS data in support of classical observational studies that showed that potential protist parasites are hosted by soil metazoa. Taken together, profound differences in amplification success between protist taxa and an inevitable co-extraction of protist taxa parasitizing soil metazoa obscure the true diversity of free-living soil protist communities.


Pharmacotherapy and family-focused treatment for adolescents with bipolar I and II disorders: a 2-year randomized trial.
Previous studies have found that family-focused treatment is an effective adjunct to pharmacotherapy in stabilizing symptoms in adult bipolar disorder. The authors examined whether pharmacotherapy and family-focused treatment for adolescents with bipolar disorder was more effective than pharmacotherapy and brief psychoeducation (enhanced care) in decreasing time to recovery from a mood episode, increasing time to recurrence, and reducing symptom severity over 2 years. A total of 145 adolescents (mean age, 15.6 years) with bipolar I or II disorder and a DSM-IV-TR manic, hypomanic, depressive, or mixed episode in the previous 3 months were randomly assigned, with family members, either to pharmacotherapy and family-focused treatment, consisting of psychoeducation (i.e., recognition and early intervention with prodromal symptoms), communication enhancement training, and problem-solving skills training, delivered in 21 sessions over 9 months; or to pharmacotherapy and three weekly sessions of enhanced care (family psychoeducation). Independent evaluators assessed participants at baseline, every 3 months during year 1, and every 6 months during year 2, using weekly ratings of mood. Twenty-two participants (15.2%) withdrew shortly after randomization. Time to recovery or recurrence and proportion of weeks ill did not differ between the two treatment groups. Secondary analyses revealed that participants in family-focused treatment had less severe manic symptoms during year 2 than did those in enhanced care. After an illness episode, intensive psychotherapy combined with best-practice pharmacotherapy does not appear to confer advantages over brief psychotherapy and pharmacotherapy in hastening recovery or delaying recurrence among adolescents with bipolar disorder.