Single Cell Omics for Musculoskeletal Research.
The ability to analyze the molecular events occurring within individual cells as opposed to populations of cells is revolutionizing our understanding of musculoskeletal tissue development and disease. Single cell studies have the great potential of identifying cellular subpopulations that work in a synchronized fashion to regenerate and repair damaged tissues during normal homeostasis. In addition, such studies can elucidate how these processes break down in disease as well as identify cellular subpopulations that drive the disease. This review highlights three emerging technologies: single cell RNA sequencing (scRNA-seq), Assay for Transposase-Accessible Chromatin using sequencing (ATAC-seq), and Cytometry by Time-Of-Flight (CyTOF) mass cytometry. Technological and bioinformatic tools to analyze the transcriptome, epigenome, and proteome at the individual cell level have advanced rapidly making data collection relatively easy; however, understanding how to access and interpret the data remains a challenge for many scientists. It is, therefore, of paramount significance to educate the musculoskeletal community on how single cell technologies can be used to answer research questions and advance translation. This article summarizes talks given during a workshop on "Single Cell Omics" at the 2020 annual meeting of the Orthopedic Research Society. Studies that applied scRNA-seq, ATAC-seq, and CyTOF mass cytometry to cartilage development and osteoarthritis are reviewed. This body of work shows how these cutting-edge tools can advance our understanding of the cellular heterogeneity and trajectories of lineage specification during development and disease.


Apparent stability of herpes simplex virus genomes isolated from primary and recurrent infections and dorsal root ganglia in the mouse.
Following natural or experimental primary infection, herpes simplex virus (HSV) becomes latent in sensory ganglia. Reactivation of latent virus may lead to recurrent disease. If HSV DNA remains stable during primary, recurrent and latent infections, that stability would enable us to trace the transmission of HSV from one individual to another. We inoculated mice in the ear pinna with HSV and collected virus at various intervals during primary infection. In mice surviving primary infections, recurrent disease was induced from which virus was isolated. Virus was also recovered from explanted dorsal root ganglia. Virus isolates were characterised by restriction endonuclease digestion and compared with the original inoculate(s). The data indicate that in all cases except two, the isolates from primary and recurrent infections remained identical to the original inoculates.


Multilocus analyses of admixture and introgression among hybridizing Heliconius butterflies.
Introgressive hybridization is an important evolutionary process and new analytical methods provide substantial power to detect and quantify it. In this study we use variation in the frequency of 657 AFLP fragments and DNA sequence variation from 15 genes to measure the extent of admixture and the direction of interspecific gene flow among three Heliconius butterfly species that diverged recently as a result of natural selection for Miillerian mimicry, and which continue to hybridize. Bayesian clustering based on AFLP genotypes correctly delineated the three species and identified four H. cydno, three H. pachinus, and three H. melpomene individuals that were of mixed ancestry. Gene genealogies revealed substantial shared DNA sequence variation among all three species and coalescent simulations based on the Isolation with Migration (IM) model pointed to interspecific gene flow as its cause. The IM simulations further indicated that interspecific gene flow was significantly asymmetrical, with greater gene flow from H. pachinus into H. cydno (2Nm = 4.326) than the reverse (2Nm = 0.502), and unidirectional gene flow from H. cydno and H. pachinus into H. melpomene (2Nm = 0.294 and 0.252, respectively). These asymmetries are in the directions expected based on the genetics of wing patterning and the probability that hybrids of various phenotypes will survive and reproduce in different mimetic environments. This empirical demonstration of extensive interspecific gene flow is in contrast to a previous study which found little evidence of gene flow between another pair of hybridizing Heliconius species, H. himera and H. erato, and it highlights the critical role of natural selection in maintaining species diversity. Furthermore, these results lend support to the hypotheses that phenotypic diversification in the genus Heliconius has been fueled by introgressive hybridization and that reinforcement has driven the evolution of assortative mate preferences.


MdbHLH93, an apple activator regulating leaf senescence, is regulated by ABA and MdBT2 in antagonistic ways.
The molecular mechanism of leaf senescence in apple (Malus domestica) is still not fully understood. We used gene expression analysis and protein-protein interactions to decipher the relationships of abscisic acid (ABA) and two proteins, MdbHLH93 and MdBT2, in the senescence process. We found that MdbHLH93 promoted leaf senescence and the expression of senescence-related genes, which exhibited similar effects to ABA on leaf senescence. MdbHLH93 activated directly the transcription of MdSAG18. We also found that an ABA-responsive protein, MdBT2, interacted directly with MdbHLH93, and induced the ubiquitination and degradation of the MdbHLH93 protein, and thus delayed leaf senescence. Our findings provide new insights into the regulatory network of leaf senescence through the functional interactions among ABA, MdbHLH93 and MdBT2.


Development of an Aboriginal Resilience and Recovery Questionnaire - a collaboration between practitioners and help-seeking clients of a Victorian Aboriginal community controlled health service.
Indigenous experiences and perspectives of resilience, healing and recovery from trauma is gaining increasing attention, with a growing qualitative literature that spans multiple indigenous cultural groups. However, few quantitative measures are available. In this article, development of a preliminary version of the Aboriginal Resilience and Recovery Questionnaire is described. The first aim of this study was to describe findings from two focus groups that provided theoretical knowledge and development of items for a draft version of an Aboriginal Resilience Recovery Questionnaire. The second aim of the study was to conduct a preliminary psychometric analysis of the properties of the measure. Multi-method research design grounded in indigenous research methodologies. Aboriginal Resilience and Recovery Questionnaire, Australian Aboriginal Version of the Harvard Trauma Questionnaire Trauma symptom subscale, Growth and Empowerment Measure. (1) Two focus groups with six counselling staff from an Aboriginal health service were run that explored Victorian Aboriginal understandings of resilience, healing, and recovery from trauma. Sixty different protective factors viewed as potentially important to resilience, healing and recovery from trauma were identified by participants. (2) Following a review of the resilience literature, 75 items were reviewed and revised, with additional items developed by the focus group. (3) The final outcome was 60 items selected for a preliminary version of the Aboriginal Resilience Recovery Questionnaire, 50 of which made up 19 different subscales in addition to 10 single items. (4) Structured interviews were conducted with 81 help seeking Aboriginal clients recruited from the same health service. Preliminary psychometric assessment of the Aboriginal Resilience Recovery Questionnaire was undertaken using Principal Components Analysis. Two component subscales were extracted with adequate internal consistency and good convergent and discriminant validity. For both subscales there were moderate to strong positive associations with empowerment, and moderate to strong negative associations with trauma symptom severity. The preliminary results are promising for a strength-based resilience measure developed from the knowledge of Aboriginal practitioners and staff of a counselling service. Further research to address some psychometric limitations in the measure is required. A larger sample size will allow for a common factor analysis to be conducted. The Aboriginal Resilience Recovery Questionnaire has potential to assist Aboriginal Community Controlled Health Organisations and other organisations to evaluate whether services and programs can effectively support community members to strengthen individual, relational, community and cultural resilience resources.


Expression of endogenous retroviruses is negatively regulated by the pluripotency marker Rex1/Zfp42.
Rex1/Zfp42 is a Yy1-related zinc-finger protein whose expression is frequently used to identify pluripotent stem cells. We show that depletion of Rex1 levels notably affected self-renewal of mouse embryonic stem (ES) cells in clonal assays, in the absence of evident differences in expression of marker genes for pluripotency or differentiation. By contrast, marked differences in expression of several endogenous retroviral elements (ERVs) were evident upon Rex1 depletion. We demonstrate association of REX1 to specific elements in chromatin-immunoprecipitation assays, most strongly to muERV-L and to a lower extent to IAP and musD elements. Rex1 regulates muERV-L expression in vivo, as we show altered levels upon transient gain-and-loss of Rex1 function in pre-implantation embryos. We also find REX1 can associate with the lysine-demethylase LSD1/KDM1A, suggesting they act in concert. Similar to REX1 binding to retrotransposable elements (REs) in ES cells, we also detected binding of the REX1 related proteins YY1 and YY2 to REs, although the binding preferences of the two proteins were slightly different. Altogether, we show that Rex1 regulates ERV expression in mouse ES cells and during pre-implantation development and suggest that Rex1 and its relatives have evolved as regulators of endogenous retroviral transcription.


Deregulation of cell growth and apoptosis in UV-induced melanomagenesis.
We have previously characterized the role of p16/Rb in coordinating the early events in UVB-irradiated skin. As an extension to this work, normal melanocytes and mutant p16-inducible melanoma cell models were employed to elucidate further the coordinated molecular mechanisms occurring during early UVB exposure. Our results showed that melanocytes expressed p16 only at a high UVB dose, with undetectable p53. The Bax/Bcl2 ratio increased at higher dose, indicating that the cells had selected apoptosis program. In the wt-p16 melanoma cells, while low UVB dose upregulated p16, the high dose suppressed it, and further abrogated Cdk6 but not Cdk4. Interestingly, while induction of mutant-p16 increased Cdk4, cdk6 and pRb proteins, UVB exposure did not affect this increase. More interestingly, p16 mutant cells increased their resistance to apoptosis at high UVB-dose, associated with decreased Bax and increased Bcl2 expression. Thus, mutant-p16 appears to dictate a deregulation of cell cycle and increased resistance to apoptosis in melanoma cells. Together, the data indicate a deregulation of p16INK4/Rb pathway as an early event in UVB-induced melanomagenesis.


Demographic and clinical profile of vernal keratoconjunctivitis at a tertiary eye care center in India.
To study the demographic and clinical profile of patients with vernal keratoconjunctivitis (VKC) at a tertiary eye care center in India. Retrospective chart analysis of 468 patients of VKC seen from January 2006 to December 2006. Mean age at presentation was 12 years. Majority of the patients had mixed pattern disease (72%). Chronic perennial disease was seen in 36% patients. Personal or family history of allergies was noted in 5% patients. Severe disease based on clinical grading was present in 37% patients. Moderate to severe vision loss was seen in 12% of total population. Persistent disease beyond 20 years of age was found in 12% patients. VKC-related complications such as corneal scarring (11%), shield ulcer (3%), keratoconus (6%), and limbal stem cell deficiency (1.2%) were seen. Treatment-related complications like corticosteroid-induced cataract and glaucoma were seen in 6% and 4% of patients, respectively. Clinical pattern of VKC seen in the tropical climate of India is essentially similar to that seen in other tropical countries. Few distinct features that we noted represent chronic perennial disease, low association with atopy, and higher propensity for disease and treatment-related complications.


Biotransformation of primary nicotine metabolites. I. In vivo metabolism of R-(+)-[14C-NCH3]N-methylnicotinium ion in the guinea pig.
The in vivo biotransformation and tissue distribution of the methylated nicotine metabolite R-(+)-[14C-NCH3]N-methylnicotinium acetate was studied in the guinea pig. The detection and quantification of 24-hr urinary metabolites after ip injection was determined by cation-exchange HPLC interfaced to a radiochemical flowthrough detector. The urinary metabolite profile consisted of five peaks. One eluted close to the void, and three coeluted with authentic standards of N-methylcotininium ion, N-methylnornicotinium ion, and N-methylnicotinium ion. A fifth, and as yet unidentified, metabolite was also detected. Tissue distribution of 14C label after 24 hr was highest in the adrenal gland and epididymis followed by the gallbladder, bladder, kidney, spleen, and heart. No significant amounts of 14C were found in the brain. The results indicate that N-methylcotininium ion and N-methylnornicotinium ion are both formed subsequent to the formation of N-methylnicotinium ion in the metabolism of R-(+)-nicotine in the guinea pig.


The Wnt1 ligand/Frizzled 3 receptor system plays a regulatory role in the achievement of the 'in vitro' capacitation and subsequent 'in vitro' acrosome exocytosis of porcine spermatozoa.
The aim of this work was to determine the existence of a functional Wnt/β-catenin signaling pathway in boar spermatozoa, which would be linked with the already well-known GSK-3 signaling pathway. This was first confirmed by detecting the presence of the specific Frizzled 3 receptor in these cells. Furthermore, this signaling pathway was activated in boar spermatozoa subjected to 'in vitro' capacitation (IVC) and subsequent progesterone-induced 'in vitro' acrosome exocytosis (IVAE) by incubating cells with separate concentrations of Wnt1 ligand, the Wnt/β-catenin signaling pathway-specific effector. Incubation with the Wnt1 ligand increased the rhythm of the time-dependent reduction in sperm viability during the achievement of both IVC and IVAE. This finding was concomitant with an increase in the percentage of spermatozoa with altered membrane fluidity and permeability determined through both merocyanine-540 and YO-PRO-1 stains. While the Wnt1 ligand did not affect total sperm motility during the achievement of the IVC, it induced a fast and transient increase in the overall motility patterns in spermatozoa subjected to IVAE. This IVAE-linked action was related to a decrease in the percentage of cells with high mitochondrial membrane potential and an increase in the percentage of cells with high intracellular Fluo-3-marked Ca(2+) content. In conclusion, our results suggest that the Wnt1 ligand-modulated Wnt/β-catenin signaling pathway plays a relevant role in the modulation of both IVC and subsequent, progesterone-induced IVAE. Furthermore, our data indicate that the transduction pathways by which the Wnt1 ligand acts on IVC and IVAE are different, and that the Wnt/β-catenin pathway is independent from GSK-3 activity in the achievement of IVC.


Thymic enlargement in childhood.
Thymic masses constitute one of the least common mediastinal masses in childhood. While producing symptoms of airway compromise, they also raise the suspicion of malignancy when detected. Radiological, operative and pathological findings of patients that have been operated for thymic masses in our institution is presented in this paper. Nine patients were operated in our institution during a 12-year-period between 1985-1997 for thymic masses. Ages of the patients ranged from four months to 13 years. With the exception of one, who was diagnosed with a routine chest x-ray, all the patients had respiratory complaints. All the patients had been evaluated with computed tomography preoperatively. In total, seven sternotomies and four thoracotomies were performed to reach the anterior mediastinum. The distribution of masses was as follows two malignant thymomas, three thymic hyperplasia, one lymphocyte-rich thymoma, one epithelial thymoma, one cystic thymoma and one lymphoblastic lymphoma. Although rare, thymic enlargement may be a cause of intractable respiratory complaints in childhood. Because of the high incidence of primary malignancy of the mediastinal neoplasms in childhood, thymic enlargement requires accurate pathological diagnosis and treatment. Median sternotomy with intensive anesthetical care allows proper tumoral exposure.


Training and transfer of clustering and cumulative rehearsal strategies in retarded individuals.
Forty-four retarded adolescents varying in MA, IQ, and etiology were taught one of three active rehearsal strategies or assigned to a control condition. Immediate, short-term (1 day), and long-term (2 weeks) transfer of the strategy was assessed by presenting new 9-item serial lists on each day. Exposure durations, which indicated the rate of subject-pacing, showed that the two highest MA-IQ (mean IQ = 55) groups, one diagnosed as organically impaired and the other as cultural-familially retarded, successfully transferred their strategies over three sessions, while no transfer occurred in the lower MA-IQ group (mean IQ = 40). Greater consistency in the expected pattern of exposure durations of high MA-IQ subjects was found in the cumulative-rehearsal group than in the cumulative-clustering group across sessions. During immediate transfer, the best recall was found for the cumulative group, while on short-term transfer both the cumulative and the cumulative-clustering groups learned more rapidly than the clustering or control groups. No significant recall differences during long-term transfer were apparent for high MA-IQ subjects, probably due to a ceiling effect. For retarded individuals with IQs above 50, the cumulative-rehearsal strategy was the most efficient technique for inducing strategy transfer to a categorized serial list.


Magnetic anisotropy and mechanism of magnetic relaxation in Er(III) single-ion magnets.
Magnetic anisotropy is a key component in the design of single-molecule magnets (SMMs) possessing a large barrier height for magnetization reversal. Lanthanide-based SMMs are the most promising candidates in this arena as they offer a large magnetic anisotropy due to the presence of strong spin-orbit coupling. Among lanthanides, Er(III) complexes are gaining attention in the area of SMMs, because of their intriguing magnetic properties and attractive blocking temperatures. Here, we have undertaken detailed ab initio calculations on four structurally diverse Er(III) SMMs to shed light on how the magnetic anisotropy is influenced by the role of symmetry and structural distortions. The employed CASSCF+RASSI calculations have offered rationale for the observed differences in the estimated Ueff values for the studied complexes and also offered hints to the mechanism of magnetic relaxation. The differences in the mechanism of magnetic relaxations are further analyzed based on the Er-ligand interactions, which is obtained by analyzing the charges, densities, luminescent behavior and the frontier molecular orbitals. Our calculations, for the first time, have highlighted the importance of high symmetry environment and ligand donor strength in obtaining large Ueff values for the Er(III) complexes. We have examined these possibilities by modeling several structures with variable coordination numbers and point group symmetry. These results signify the need of a detailed understanding on the shape of the anisotropy and the point group symmetry in order to achieve large Ueff values in Er(III) single-ion magnets.


Effects of Esmolol on the Esophagogastric Junction: A Double-Blind, Randomized, Crossover Study on 14 Healthy Volunteers.
Passive regurgitation may occur throughout the perioperative period, increasing the risk for pulmonary aspiration and postoperative pulmonary complications. Hypnotics and opioids, especially remifentanil, that are used during anesthesia have been shown to decrease the pressure in the esophagogastric junction (EGJ), that otherwise acts as a barrier against passive regurgitation of gastric contents. Esmolol, usually used to counteract tachycardia and hypertension, has been shown to possess properties useful during general anesthesia. Like remifentanil, the β-1-adrenoreceptor antagonist may be used to attenuate the stress reaction to tracheal intubation and to modify perioperative anesthetic requirements. It may also reduce the need for opioids in the postoperative period. Its action on the EGJ is however unknown.The aim of this trial was to compare the effects of esmolol and remifentanil on EGJ pressures in healthy volunteers, when administrated as single drugs. Measurements of EGJ pressures were made in 14 healthy volunteers using high-resolution solid-state manometry. Interventions were administered in a randomized sequence and consisted of esmolol that was given IV as a bolus dose of 1 mg/kg followed by an infusion of 10 μg·kg·minute over 15 minutes, and remifentanil with target-controlled infusion of 4 ng/mL over 15 minutes. Interventions were separated by a 20-minute washout period. Analyses of EGJ pressures were performed at baseline, and during drug administration at 2 (T2) and 15 minutes (T15). The primary outcome was the inspiratory EGJ augmentation, while the inspiratory and expiratory EGJ pressures were secondary outcomes. There was no effect on inspiratory EGJ augmentation when comparing remifentanil and esmolol (mean difference -4.0 mm Hg [-9.7 to 1.7]; P= .15). In contrast, remifentanil significantly decreased both inspiratory and expiratory pressures compared to esmolol (-12.2 [-18.6 to -5.7]; P= .003 and -8.0 [-13.3 to -2.8]; P= .006). Esmolol, compared with remifentanil, does not affect EGJ function. This may be an advantage regarding passive regurgitation and esmolol may thus have a role to play in anesthesia where maintenance of EGJ barrier function is of outmost importance.


Two alleles of the 175-kilodalton Plasmodium falciparum erythrocyte binding antigen.
EBA-175, erythrocyte binding antigen 175, is a 175-kDa antigen of Plasmodium falciparum which has been shown to be involved in the recognition of erythrocytes by merozoites and may be involved in the process of erythrocyte invasion. Invasion of erythrocytes by Camp strain merozoites is inhibited by pre-treatment of red blood cells by EBA-175 from the heterologous strain, FCR-3. The sequence of the Camp strain has been published and we report here the sequence of the FCR-3 strain. The sequences are nearly identical except for a 423-bp segment in the FCR-3 strain, F-segment, that is not found in the Camp strain and a 342-bp segment, C-segment, present in the Camp strain but not in the FCR-3 strain. The locations of these two segments are different in Camp and FCR-3 EBA-175 genes and there is little DNA or amino acid sequence homology between them. The essentially dimorphic alleles, F-segment and C-segment, are conserved in all isolates examined to date. Evidence of genetic cross-over between the FCR-3 and the Camp EBA-175 genes was not observed in the analysis of a limited number of wild isolates. The continued study of the biological relevance of these sequence divergences in EBA-175 may further elucidate the sequence of events resulting in merozoite invasion of erythrocytes.


Activation of c-Jun N-terminal protein kinase is a common mechanism underlying paraquat- and rotenone-induced dopaminergic cell apoptosis.
Parkinson's disease (PD) is characterized by selective loss of dopaminergic neurons in the substantia nigra of the brain. Although the underlying causes are not well characterized, epidemiological studies suggest an elevated risk of PD with occupational pesticide exposure. Here, we utilized pheochromocytoma (PC) 12 and SH-SY5Y cells as well as rat primary cultured dopaminergic neurons to investigate mechanisms for dopaminergic cell death induced by paraquat and rotenone, pesticides that are used to model PD in rodents. Both paraquat and rotenone induce selective loss of dopaminergic neurons in primary cultures. We discovered that paraquat induces apoptosis in PC12 cells but not in SH-SY5Y cells, while rotenone exposure causes apoptosis in SH-SY5Y cells but not in PC12 cells. The selective ability of paraquat and rotenone to induce apoptosis in different cell lines correlates with their ability to activate c-Jun N-terminal protein kinase (JNK) and p38 mitogen-activated protein kinases. Furthermore, JNK and p38 are required for rotenone-induced apoptosis in SH-SY5Y cells (K. Newhouse et al., 2004, Toxicol. Sci. 79, 137-146) as well as primary neurons, and for paraquat-induced apoptosis in PC12 cells. However, JNK but not p38 plays a role in paraquat-induced loss of primary cultured dopaminergic neurons. Our data identify JNK activation as a common mechanism underlying dopaminergic cell death induced by both paraquat and rotenone in model cell lines and primary cultures.


Temporal Changes in Fibrinolysis following Injury.
Trauma patients present to the emergency department with a spectrum of fibrinolytic activity. This wide variance in fibrinolysis activity is a complex multifactorial process impacted by the degree of hemorrhagic shock and the amount of tissue injury the individual sustains. The fibrinolytic activity of the trauma patient at presentation to the hospital has prognostic and therapeutic implications. Those patients with high fibrinolytic activity (hyperfibrinolysis) are at risk of mortality from hemorrhage, whereas those patients with low fibrinolytic activity (shutdown or hypofibrinolysis) are at an increased risk of delayed mortality from traumatic brain injury or organ failure. These phenotypes of fibrinolysis acutely following injury change with resuscitation, and the majority of trauma patients will transition to a fibrinolytic resistant state several hours after injury. The mechanism for this near-global transition to this acquired fibrinolysis appears to be related to the generation of plasminogen activator inhibitor-1 in the liver. Those patients who do not recover from this fibrinolytic state 24 hours after injury have a poor prognosis. The purpose of this article is to review the different states of fibrinolytic activity following injury and how they change over time following resuscitation and in the intensive care unit.


The Tumor Microenvironment as a Regulator of Endocrine Resistance in Breast Cancer.
Estrogen receptor positive breast neoplasias represent over 70% of diagnosed breast cancers. Depending on the stage at which the tumor is detected, HER2 status and genomic risk, endocrine therapy is combined with either radio, chemo and/or targeted therapy. A growing amount of evidence supports the notion that components of the tumor microenvironment play specific roles in response to treatment and that strategies targeting these key interactions with tumor cells could pave the way to a new generation of therapies. In this review, we analyze the evidence suggesting different components of the tumor microenvironment play a role in hormone receptor positive breast cancer progression. In particular we focus on the immune system, carcinoma associated fibroblasts and the extracellular matrix. Further insight into the cross talk between these constituents of the microenvironment and the tumor cells may lead to therapies that eliminate disseminated metastatic cells early on, and thus reduce distant disease relapse which is the leading cause of death for patients who are diagnosed with this illness.


Long-read sequence assembly: a technical evaluation in barley.
Sequence assembly of large and repeat-rich plant genomes has been challenging, requiring substantial computational resources and often several complementary sequence assembly and genome mapping approaches. The recent development of fast and accurate long-read sequencing by circular consensus sequencing (CCS) on the PacBio platform may greatly increase the scope of plant pan-genome projects. Here, we compare current long-read sequencing platforms regarding their ability to rapidly generate contiguous sequence assemblies in pan-genome studies of barley (Hordeum vulgare). Most long-read assemblies are clearly superior to the current barley reference sequence based on short-reads. Assemblies derived from accurate long reads excel in most metrics, but the CCS approach was the most cost-effective strategy for assembling tens of barley genomes. A downsampling analysis indicated that 20-fold CCS coverage can yield very good sequence assemblies, while even five-fold CCS data may capture the complete sequence of most genes. We present an updated reference genome assembly for barley with near-complete representation of the repeat-rich intergenic space. Long-read assembly can underpin the construction of accurate and complete sequences of multiple genomes of a species to build pan-genome infrastructures in Triticeae crops and their wild relatives.


Antibiotic resistance in uncomplicated urinary tract infection: problems with interpreting cumulative resistance rates from local community laboratories.
To determine the resistance rates and patterns in bacteria causing uncomplicated urinary tract infections (UTIs) presenting to general practitioners (GPs) in Christchurch. 82 randomly selected GPs in Christchurch participated in the study. Midstream urine (MSU) samples were prospectively collected for standard microbiological analysis on all women between the ages of 16 and 50 years presenting with symptoms of dysuria and frequency and who had positive dipstick testing to either (or both) nitrites or leucocytes. MSUs were submitted for bacterial colony counts and resistance testing of isolates present in adequate numbers. 374 specimens were collected. 299 filled the inclusion criteria, of which 94 fulfilled criteria for significant infection. Trimethoprim resistance was found in 8, (8.5%) (95%CI 2.8,14.2) overall with a resistance rate for Escberichia coli (E. coli) to trimethoprim of 11.5%. This compared with cumulative resistance rates from local community laboratories for E. coli to trimethoprim of 19%. For a woman in this age group presenting with symptoms of UTI we estimated that her probability of having a trimethoprim resistant organism was 2.7%. Trimethoprim remains a reasonable first line treatment for uncomplicated UTI in Christchurch. Actual resistance rates are significantly less than those derived from routine pooled laboratory specimens, and when used in an intention to treat calculation to inform empiric prescribing, become even less significant. While collection of these routine data is essential to provide early warning of emergent resistance, a truly representative rate should be determined to inform prescribing decisions if resistance appears to be increasing.


Systematic Review of Neighborhood Factors Impacting HIV Care Continuum Participation in the United States.
Social determinants have been increasingly implicated in accelerating HIV vulnerability, particularly for disenfranchised communities. Among these determinants, neighborhood factors play an important role in undermining HIV prevention. However, there has been little research comprehensively examining the impact of neighborhood factors on HIV care continuum participation in the US. To address this, we conducted a systematic review (PROSPERO registration number CRD42022359787) to determine neighborhood factors most frequently associated with diminished HIV care continuum participation. Peer-reviewed studies were included if published between 2013 - 2022, centralized in the US, and analyzed a neighborhood factor with at least one aspect of the HIV care continuum. The review was guided by the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) protocol. Study quality was guided by LEGEND (Let Evidence Guide Every New Decision) evaluation guidelines. Systematic review analysis was conducted using Covidence software. There were 3,192 studies identified for initial screening. Forty-four were included for review after eliminating duplicates, title/abstract screening, and eligibility assessment. Social and economic disenfranchisement of neighborhoods negatively impacts HIV care continuum participation among persons living with HIV. In particular, five key neighborhood factors (socioeconomic status, segregation, social disorder, stigma, and care access) were associated with challenged HIV care continuum participation. Race moderated relationships between neighborhood quality and HIV care continuum participation. Structural interventions addressing neighborhood social and economic challenges may have favorable downstream effects for improving HIV care continuum participation.


Nutrition education and infant growth in rural Indian infants: narrowing the gender gap?
Growth faltering and malnutrition are highly prevalent in most South Asian countries. Among the serious consequences of malnutrition are increased risks of morbidity and mortality in children as well as deficits in physical stature and lowered cognitive measures. Childhood malnutrition in poor households has been well documented in India, with the highest rates observed in those aged 12-23 months. Countrywide National Family Health Survey II (NFHS II) data show mean underweight prevalence increases from 11.9% among infants under six months of age to 58.4% at 12-23 months of age. This paper discusses the results of an analysis of a longitudinal community-based study of infant growth feeding and care practices in rural south India, for which data have been collected since 1997 and completed in May 2001. In 1998 an intervention began in which a monthly visit for nutrition counselling was made by a trained worker. Mothers were counselled about the need for exclusive breastfeeding for four to six months, choice of appropriate complementary foods, and feeding frequency. Infants enrolled in the study prior to 1998, did not receive this intervention in the first year of life. Using bivariate analysis and regression modelling, weight velocity and feeding practices for intervention and non-intervention infants, controlling for a range of socio-economic and demographic factors for infants aged 6 to 10 months, are compared. Both descriptive and multivariate analyses show a statistically significant positive association between intervention and weight velocity for female infants. The intervention group as a whole had improved feeding practices such as avoidance of feeding bottle and increased variety of foods. The implication of this association, in a culture where discrimination against females exists, is promising for the development of low-cost educational interventions to improve nutritional status without the need to provide supplementary food.


Proglumide: selective antagonism of excitatory effects of cholecystokinin in central nervous system.
Extracellular single-unit recording techniques were used to test the ability of proglumide to block cholecystokinin-induced excitation of rat midbrain dopaminergic neurons and dopamine-sensitive prefrontal cortex cells. Intravenous and iontophoretic proglumide administration consistently blocked cholecystokinin-induced excitations while having no effect on glutamic acid-induced increases in activity. This selective blockade of central cholecystokinin effects by proglumide suggests that this drug may be valuable for studying the possible role of cholecystokinin as a neurotransmitter or neuromodulator in the central nervous system.


Notch ligand delta-like1: X-ray crystal structure and binding affinity.
The Notch pathway is a fundamental signalling system in most multicellular animals. We have determined the X-ray crystal structure of the extracellular domain of the Notch ligand delta-like ligand-1 (Dll-1). The structure incorporates the N-terminal C2 domain, receptor-binding DSL domain and the first six (of eight) EGF (epidermal growth factor)-like repeats, which form a highly extended conformation, confirmed by analytical ultracentrifugation. Comparison of our structure with a fragment of Jagged1 ligand allows us to dissect the similarities and differences between the ligand families. Differences in the C2 domains of Dll-1 and Jagged1 suggest their lipid-binding properties are likely to differ. A conserved hydrophobic patch on the surface of both Dll-1 and Jagged1 provides a likely receptor-interaction site that is common to both ligands. We also explore the binding affinity of Dll-1 for a fragment of Notch1 using different techniques. Apparent binding affinities vary when different techniques are used, explaining discrepancies in the literature. Using analytical ultracentrifugation, we perform for the first time binding analyses where both receptor and ligand are in solution, which confirms a Kd of 10 μM for this interaction.


The inhibitory effect of 7,7″-dimethoxyagastisflavone on the metastasis of melanoma cells via the suppression of F-actin polymerization.
7,7″-Dimethoxyagastisflavone (DMGF), a biflavonoid isolated from Taxus × media cv. Hicksii, induces apoptotic and autophagic cell death. However, whether DMGF suppresses tumor metastasis is unclear. The aim of this study was to investigate the anti-metastatic activities of DMGF on the metastatic processes of melanoma cells in vivo and in vitro. A transwell assay showed that DMGF could effectively attenuate the motility of B16F10 cells, and the results of real-time PCR revealed that DMGF also suppressed the expressions of matrix metalloproteinase-2 (MMP-2). Moreover, DMGF did not influence tube formation but inhibited the migration of endothelial cells. Furthermore, animal models were used to monitor the effects of DMGF on tumor metastasis, and all models showed that DMGF significantly suppressed the metastatic behaviors of B16F10 cells, including intravasation, colonization, and invasion of the lymphatic duct. In addition, DMGF could also reduce the densities of the blood vessels in the tumor area in vivo. Further investigation of the molecular mechanisms of anti-metastatic activity revealed that DMGF can down-regulate the levels of key modulators of the Cdc42/Rac1 pathway to interfere in F-actin polymerization and suppress the formation of lamellipodia by reducing the phosphorylation of CREB. These data suggested that DMGF presents anti-metastatic activities in B16F10 melanoma cells. Here, we demonstrated that DMGF can inhibit the metastasis of highly invasive melanoma cancer cells through the down-regulation of F-actin polymerization. Considering these findings, DMGF may be further developed to serve as a chemoprevention drug for patients with metastatic melanoma.


Variability and reproducibility of flow-mediated dilatation in a multicentre clinical trial.
The aim of this study was to assess the reproducibility of flow-mediated dilatation (FMD) in a multicentre setting. This study was performed as part of the dal-VESSEL trial in which FMD was measured in 19 vascular imaging centres in six European countries. A subgroup of patients who were allocated in the placebo group and scanned twice at each trial time point (substudy) was analysed. Intra-sonographer variability was calculated from FMD measurements 48 h apart. Centre variability and short-, medium-, and long-term reproducibility of FMD were calculated at 48 h and at 3 and 9 months intervals, respectively. Intra- and inter-reader variability was assessed by re-analysing the FMD images by three certified readers at two time intervals, 7 days apart. Sixty-seven patients were included. Variability between centres was comparable at 48 h and 3 months interval but almost doubled at 9 months. The mean absolute difference in %FMD was 1.04, 0.99, and 1.45% at the three time intervals, respectively. Curves were generated to indicate the number of patients required for adequate power in crossover and parallel study designs. This study demonstrates for the first time that in a multicentre setting reproducible FMD measurements can be achieved for short- and medium-term evaluation, which are comparable with those reported from specialized laboratories. These findings justify the use of FMD as an outcome measure for short- and medium-term assessment of pharmacological interventions.


[Functional assessment, using micturition cystourethrography, of the unstable bladder in childhood].
The accuracy and reliability of a modified method of micturating urethrocystography for the diagnosis of uncontrolled detrusor contraction in unstable bladders (sphincter-detrusor dyssynergy during bladder filling) was studied in 100 patients aged between four and 15 years. A comparison with a urodynamic method showed an accuracy of 92%, sensitivity of 93% and specificity of 90.7%. The technique and indications are described and the clinical relevance of the results is discussed.


Increased expression of intracisternal A-particle RNA in regenerated myeloid cells after X irradiation in C3H/He inbred mice.
Myeloid Cells after X Irradiation in C3H/He Inbred Mice. Myeloid leukemia cells were derived from regenerated hematopoietic cells damaged by sublethal doses of X radiation in C3H/He inbred mice. We previously found that within the genome of the myeloid leukemia cells, a retrotransposon, the intracisternal A-particle (IAP) element, is integrated. Levels of IAP RNA, the source of cDNA for the integration, were analyzed quantitatively in C3H mice. Higher levels of IAP transcripts were observed in normal cells, particularly in hematopoietic cells, from C3H/He mice, than in those from C57BL/6J and STS/A mice. In the C3H/He mice, an approximately twofold increase in IAP RNA was found in the regenerated spleen and bone marrow cells at 5 days and from 12 to 90 days after whole-body X irradiation. In addition, an increased level of IAP RNA was observed in all the myeloid leukemia cells derived from C3H/He mice. This suggests that the elevated levels of IAP RNA in the regenerated hematopoietic cells after irradiation contribute to the increase in retrotransposition of IAP found in myeloid leukemia cells from C3H/He mice.


The impact of sustained and intermittent docetaxel chemotherapy regimens on cognition and neural morphology in healthy mice.
A subset of cancer survivors demonstrates impairments in cognition long after chemotherapy completion. At present, it is unclear whether these changes are due to direct neurotoxic effects of chemotherapy. This study examined the impact of variable docetaxel (DTX) chemotherapy dosing on brain DTX exposure via analyses of neural morphology and changes in cognition. Male CD-1 mice were treated with DTX either intermittently (8 mg/kg i.p. weekly) or via a sustained delivery system (DTX-PoLigel), which continuously releases DTX. Both groups received total DTX doses of 32 mg/kg. Mice were assessed on the novel object recognition (NOR) task and the Morris water maze (MWM) shortly after treatment. Post-treatment behavioral testing demonstrated impaired NOR in mice treated with either dosing schedule relative to controls. No differences were observed between groups in MWM training and initial testing, though control mice performed better than chance while DTX-treated mice did not. Appreciable amounts of DTX were found in the brain after both treatment regimens. DTX treatment did not significantly increase levels of apoptosis within the CNS. However, some elevation in neural autophagy was observed following DTX treatment. Analysis of astrocytic activation demonstrated that intermittent DTX treatment resulted in an elevation of GFAP-positive astrocytes for 48 h after administration. Sustained chemotherapy demonstrated prolonged but lower levels of astrocyte activation over 9 days following implantation. DTX treatment induced cognitive impairment shortly after treatment. Further, these findings suggest an association between DTX dosing, neurotoxicity, and cognitive effects.


Maintaining PGC-1α expression following pressure overload-induced cardiac hypertrophy preserves angiogenesis but not contractile or mitochondrial function.
During pathological hypertrophy, peroxisome proliferator-activated receptor coactivator 1α (PGC-1α) is repressed in concert with reduced mitochondrial oxidative capacity and fatty acid oxidation (FAO). We therefore sought to determine if maintaining or increasing PGC-1α levels in the context of pressure overload hypertrophy (POH) would preserve mitochondrial function and prevent contractile dysfunction. Pathological cardiac hypertrophy was induced using 4 wk of transverse aortic constriction (TAC) in mice overexpressing the human PGC-1α genomic locus via a bacterial artificial chromosome (TG) and nontransgenic controls (Cont). PGC-1α levels were increased by 40% in TG mice and were sustained following TAC. Although TAC-induced repression of FAO genes and oxidative phosphorylation (oxphos) genes was prevented in TG mice, mitochondrial function and ATP synthesis were equivalently impaired in Cont and TG mice after TAC. Contractile function was also equally impaired in Cont and TG mice following TAC, as demonstrated by decreased +dP/dt and ejection fraction and increased left ventricular developed pressure and end diastolic pressure. Conversely, capillary density was preserved, in concert with increased VEGF expression, while apoptosis and fibrosis were reduced in TG relative to Cont mice after TAC. Hence, sustaining physiological levels of PGC-1α expression following POH, while preserving myocardial vascularity, does not prevent mitochondrial and contractile dysfunction.


18F-FDG PET or PET-CT to evaluate prognosis for head and neck cancer: a meta-analysis.
The purpose of this meta-analysis was to evaluate the prognostic value of standard uptake value (SUV) from serial Fluorine-18-fluorodeoxyglucose positron emission tomography/computed tomography ((18)F-FDG PET/CT) in patients with head and neck cancer. We searched for articles limited to head and neck cancer, dealt with the impact of SUV on survival and published in English. The endpoints were disease-free survival (DFS), overall survival (OS), and local control (LC). Two reviewers extracted data independently. Thirty-five studies were identified; of which, 26 studies involving 1,415 patients met the inclusion criteria. Pooled survival data suggested better DFS, OS, and LC in patients with low SUV of pre-treatment, and the odds ratio (OR) was 0.23, 0.24, and 0.27, respectively. Patients having tumors with low SUV of post-treatment also had significantly better DFS (OR = 0.17) and OS (OR = 0.28) than those with high SUV. The present meta-analysis showed that (18)F-FDG uptake, as measured by the SUV before treatment and metabolic response after treatment, are valuable for predicting long-term survival in head and neck cancer. High (18)F-FDG uptake may be useful for identifying patients requiring more aggressive treatment.


Sarcoidosis Presenting as Cranial Multinevritis.
A 29-year-old woman presented to the emergency department with red eyes and hypoacusia since the previous weeks associated with a sudden right facial palsy. Ophthalmologic examination revealed bilateral uveitis and lumbar puncture was compatible with aseptic meningitis. Thoracic computed tomography scan showed pulmonary nodules and mediastinal and hilar lymphadenopathy. Bronchoalveolar lavage revealed a CD4/CD8 ratio of 8, and lymph node biopsy confirmed non-caseating granulomas. The diagnosis of sarcoidosis with ocular, pulmonary, and neurologic involvement was made. Due to sarcoidosis severity at presentation, the patient was started on systemic steroids and methotrexate with improvement. Sarcoidosis involvement of the central nervous system is a rare condition (5-10%) but can be the first manifestation of the disease. Its presentation can be heterogeneous; therefore, symptoms are frequently devalued by patients and physicians, leading to a late diagnosis and permanent damage. Clinicians' awareness is crucial to early diagnosis and treatment.


Posterior instrumentation using compressive laminar hooks and anterior interbody arthrodesis for the treatment of tuberculosis of the lower lumbar spine.
A retrospective review with clinical and radiologic assessment was conducted. To demonstrate the feasibility of posterior short segment augmentation with lamina hook and anterior lumbar interbody arthrodesis in tuberculosis of the lumbar spine by evaluating clinical results, fusion status, and sagittal angle. There are few articles in the literature describing circumferential treatment of lumbar tuberculosis. We have had significant experience with a uniform operative technique for the treatment of this relatively rare condition and report our results with a minimum 2-year follow-up. Twenty-three patients with tuberculosis of the lower lumbar spine underwent posterior instrumentation with laminar hooks and anterior interbody arthrodesis by a single surgeon. The clinical outcomes were evaluated with preoperative and postoperative questionnaires, and the radiographs were independently analyzed with respect to fusion status and sagittal angle. RESULTS.: The mean follow-up period was 28.7 months (range, 24-39 months). The average preoperative, immediate postoperative, and final follow-up sagittal angles were 2.7 degrees, -14.1 degrees, and -11.5 degrees, respectively. There was a mean reduction of 16.7 degrees (range, 9 degrees-23 degrees) after surgery. Two patients had a correction loss more than 5 degrees during the follow-up period. Bony fusion was obtained in all patients. There was no recurrence of the disease. CONCLUSION.: This technique appears to be effective in stabilizing the vertebrae, restoring lordosis, achieving a solid fusion, and improving clinical outcome without sacrificing additional motion segments.


Discovery of CTSK+ Periosteal Stem Cells Mediating Bone Repair in Orbital Reconstruction.
The purpose of this study was to explore the role of cathepsin K positive (CTSK+) periosteal stem cells (PSCs) in orbital bone repair and to clarify the source of endogenous stem cells for orbital bone self-repair. Periosteum samples obtained by clinical orbital bone repair surgery were analyzed, after which immunofluorescence and immunohistochemical staining were used to detect the content of bone marrow-derived cells and CTSK+ PSCs in periosteum as well as the mobilization of PSCs. CTSK+ PSCs were characterized by flow cytometry. Transcriptome sequencing was used to compare the transcriptomic characteristics of CTSK+ PSCs and bone marrow mesenchymal stem cells (BMSCs). The orbital periosteum contained CTSK+CD200+ cell lineage, including CD200+CD105- PSCs and CD200+CD105+ progenitor cells. CTSK and osteocalcin (OCN) colocalized in the inner layer of the orbital periosteum, suggesting the osteogenic differentiation potential of CTSK+ PSCs. CTSK expression was much higher in periosteum after mobilization. Immunofluorescence showed low amounts of scattered CD31+ and CD45+ cells in the orbital periosteum. The stem cell characteristics of CTSK+ PSCs were verified by multidirectional differentiation. Flow cytometry found CD200+CD105- CTSK+ PSCs and CD200variantCD105+ progenitor cells. Transcriptome sequencing of CTSK+ PSCs and BMSCs found 3613 differential genes with significant differences. Gene Ontology (GO) analysis showed the differences between the two types of stem cells, revealing that PSCs were more suitable for intramembranous osteogenesis. CTSK+ PSCs may be endogenous stem cells for orbital bone repair. They are mobilized after orbital fracture and have unique features suitable for intramembranous osteogenesis, completely different from BMSCs.


A novel role for the NLRC4 inflammasome in mucosal defenses against the fungal pathogen Candida albicans.
Candida sp. are opportunistic fungal pathogens that colonize the skin and oral cavity and, when overgrown under permissive conditions, cause inflammation and disease. Previously, we identified a central role for the NLRP3 inflammasome in regulating IL-1β production and resistance to dissemination from oral infection with Candida albicans. Here we show that mucosal expression of NLRP3 and NLRC4 is induced by Candida infection, and up-regulation of these molecules is impaired in NLRP3 and NLRC4 deficient mice. Additionally, we reveal a role for the NLRC4 inflammasome in anti-fungal defenses. NLRC4 is important for control of mucosal Candida infection and impacts inflammatory cell recruitment to infected tissues, as well as protects against systemic dissemination of infection. Deficiency in either NLRC4 or NLRP3 results in severely attenuated pro-inflammatory and antimicrobial peptide responses in the oral cavity. Using bone marrow chimeric mouse models, we show that, in contrast to NLRP3 which limits the severity of infection when present in either the hematopoietic or stromal compartments, NLRC4 plays an important role in limiting mucosal candidiasis when functioning at the level of the mucosal stroma. Collectively, these studies reveal the tissue specific roles of the NLRP3 and NLRC4 inflammasome in innate immune responses against mucosal Candida infection.


Abnormal baseline brain activity in drug-naïve patients with Tourette syndrome: a resting-state fMRI study.
Tourette syndrome (TS) is a childhood-onset chronic disorder characterized by the presence of multiple motor and vocal tics. This study investigated spontaneous low-frequency fluctuations in TS patients during resting-state functional magnetic resonance imaging (rs-fMRI) scans. We obtained rs-fMRI scans from 17 drug-naïve TS children and 15 demographically matched healthy children. We computed the amplitude of low-frequency fluctuation (ALFF) and fractional ALFF (fALFF) of rs-fMRI data to measure spontaneous brain activity, and assessed the between-group differences in ALFF/fALFF and the relationship between ALFF/fALFF and tic severity scores. Our results showed that the children with TS exhibited significantly decreased ALFF in the posterior cingulate gyrus/precuneus and bilateral parietal gyrus. fALFF was decreased in TS children in the anterior cingulated cortex, bilateral middle and superior frontal cortices and superior parietal lobule, and increased in the left putamen and bilateral thalamus. Moreover, we found significantly positive correlations between fALFF and tic severity scores in the right thalamus. Our study provides empirical evidence for abnormal spontaneous neuronal activity in TS patients, which may implicate the underlying neurophysiological mechanism in TS and demonstrate the possibility of applying ALFF/fALFF for clinical TS studies.


[Appllication of human acellular dermal matrix in surgical treatment of genitourinary disease].
Humanacellular dermal matrix (HADM) is widely used in the field of burn wound repair and tissue engineering plastic surgery. HADM is manufactored by physical and chemical decellular process to remove the antigenic components that might cause immune rejection in dermis.The extracellular matrix of three-dimensional cell scaffold structure with collagen fibers had been used for wound repair and tissue regeneration, while HADM characterized with low absorption rate after implantation and strong ability to induce angiogenesis in host tissue. Studies reported that after the HADM was implanted into the patient, the host cells, such as fibroblasts and myofibroblasts, as well as lymphocytes, macrophages, granulocytes and mast cells, rapidly infiltrated the graft. The connective tissue and neovascularization were then formed within the HADM three-dimensional cell scaffold, the lymphatic system also appears after vascular reconstruction. Traditional urethral reconstruction using autologous skin flaps has some defects, such as complexity of the technology, risk of necrosis of the skin flaps after transplantation, and failure to achieve functional repair of the urethral epithelium. It has been reported that using HADM to reconstruct the urethra in patients with urethral stricture, hypospadias and bladder-vaginal fistula, showed promising results. Others have reported the experience of using HADM to repair and reconstruct congenital classic bladder exstrophy. HADM has also been used for tissue repair in patients with penile skin defect caused by Fonier's gangrene and hidradenitis suppurativa, and implanted under Bucks' fascia to enlarge the penis. The report of HADM implantation for treating premature ejaculation also deserves attention. Researchers found that HADM implantation can form a tissue barrier between the skin and corpus cavernosum, which can effectively reduce penile sensitivity and treat premature ejaculation. The safety and effectiveness of HADM implantation in the treatment of premature ejaculation need to be further standardized by data from multi-center, large-sample clinical studies. In summary, HADM is the extracellular matrix and three-dimensional cell scaffold of human dermis. As a new type of tissue repair material, new blood vessels are formed actively after implantation, which shows good histocompatibility. HADM has shown increasingly broad application prospects in treatment of genitourinary diseases including penis, urethra and bladder diseases. HADM has also been used in the treatment of premature ejaculation in recent clinical studies, and its long-term safety and efficacy need to be further investigated.


Understanding bias in relationships between the food environment and diet quality: the Coronary Artery Risk Development in Young Adults (CARDIA) study.
The relationship between food environment exposures and diet behaviours is unclear, possibly because the majority of studies ignore potential residual confounding. We used 20 years (1985-1986, 1992-1993 2005-2006) of data from the Coronary Artery Risk Development in Young Adults (CARDIA) study across four US cities (Birmingham, Alabama; Chicago, Illinois; Minneapolis, Minnesota; Oakland, California) and instrumental variables (IV) regression to obtain causal estimates of longitudinal associations between the percentage of neighbourhood food outlets (per total food outlets within 1 km network distance of respondent residence) and an a priori diet quality score, with higher scores indicating higher diet quality. To assess the presence and magnitude of bias related to residual confounding, we compared results from causal models (IV regression) to non-causal models, including ordinary least squares regression, which does not account for residual confounding at all and fixed-effects regression, which only controls for time-invariant unmeasured characteristics. The mean diet quality score across follow-up was 63.4 (SD=12.7). A 10% increase in fast food restaurants (relative to full-service restaurants) was associated with a lower diet quality score over time using IV regression (β=-1.01, 95% CI -1.99 to -0.04); estimates were attenuated using non-causal models. The percentage of neighbourhood convenience and grocery stores (relative to supermarkets) was not associated with diet quality in any model, but estimates from non-causal models were similarly attenuated compared with causal models. Ignoring residual confounding may generate biased estimated effects of neighbourhood food outlets on diet outcomes and may have contributed to weak findings in the food environment literature.


Value of single-injection or continuous sciatic nerve block in addition to a continuous femoral nerve block in patients undergoing total knee arthroplasty: a prospective, randomized, controlled trial.
Continuous femoral nerve block in patients undergoing total knee arthroplasty (TKA) improves and shortens postoperative rehabilitation. The primary aim of this study was to investigate whether the addition of sciatic nerve block to continuous femoral nerve block will shorten the time-to-discharge readiness. Ninety patients undergoing TKA were prospectively randomized to 1 of 3 groups: patient-controlled analgesia via femoral nerve catheter alone (F group) or combined with a single-injection (Fs group) or continuous sciatic nerve block (FCS group) until the second postoperative day. Discharge readiness was defined as the ability to walk and climb stairs independently, average pain on a numerical rating scale at rest lower than 4, and no complications. In addition, knee function, pain, supplemental morphine requirement, local anesthetic consumption, and postoperative nausea and vomiting (PONV) were evaluated. Median time-to-discharge readiness was similar: F group, 4 days (range, 2-16 days); Fs group, 4 days (range, 2-7 days); and FCS group, 4 days (range, 2-9 days; P = 0.631). No significant differences were found regarding knee function, local anesthetic consumption, or postoperative nausea and vomiting. During the day of surgery, pain was moderate to severe in the F group, whereas Fs and FCS groups experienced minimal pain (P < 0.01). Patients in the F group required significantly more supplemental morphine on the day of surgery and the first postoperative day. Until the second postoperative day, pain was significantly less in the FCS group (P < 0.01). A single-injection or continuous sciatic nerve block in addition to a femoral nerve block did not influence time-to-discharge readiness. A single-injection sciatic nerve block can reduce severe pain on the day of the surgery, whereas a continuous sciatic nerve block reduces moderate pain during mobilization on the first 2 postoperative days.


Correlation of peak Doppler frequency with lumen narrowing in carotid stenosis.
The peak Doppler-shifted frequency and degree of lumen narrowing were compared in 75 cervical carotid stenoses. Peak frequency was not found to precisely indicate severity of stenosis, but it was possible to divide stenoses into four ranges of severity on the basis of peak frequency. Peak frequencies of less than 5 KHz, in most instances, indicated stenoses of less than 50% decrease in lumen area. Frequencies from 5 to 8 KHz were generally associated with stenoses of 50-75% decrease in lumen area, and frequencies of 8-12 KHz were most often associated with stenoses of 75-90% decrease in area. Frequencies greater than 12 KHz almost invariably occurred in very severe lesions of greater than 90% decrease in lumen area (greater than 70% decrease in diameter). The use of peak frequency as an indicator of severity of stenosis, while not specific, is felt to be of considerable clinical benefit as it provides a more quantitative evaluation of stenosis than auditory assessment of Doppler frequencies.


Clinical and radiographic outcome of revision surgery for failed triple arthrodesis.
Between 1987 and 1994, we treated 33 patients with surgical revision for failed triple arthrodesis, 28 (29 feet) of whom returned for final examination (mean, 4.4 years; range, 2-7 years). The average age of these 16 women and 12 men was 46 years (range, 14-69 years). Before the revision procedure, patients had undergone nonoperative therapies for an average of 3.7 years (range, 0.5-12 years) and an average of three foot operations (range, 1-6 operations) after the primary triple arthrodesis. All patients were managed with rigid internal fixation via cannulated screws and power staples. Calcaneal osteotomy and/or revision of the transverse tarsal arthrodesis via appropriate saw cuts and bone wedges were used. Iliac crest bone graft was added, when a bone block arthrodesis was required, for those patients with nonunion or ankle impingement. Arthrodesis was achieved in all 29 feet, although 4 patients (4 feet) (14%) required additional procedures for malunion (2 patients), deformity recurrence (1 patient), deep infection (1 patient), and skin graft (1 patient). Comparison of the average pre- (retrospective) and postoperative American Orthopaedic Foot and Ankle Society 94-point hindfoot and ankle scores showed a significant improvement: 31 points (range, 13-61 points) versus 59 points (range, 24-91 points), respectively (P < 0.05). On a scale of 0 to 10 points, average patient satisfaction was 7.8 points (range, 2-10 points). This study demonstrated a satisfactory improvement in patient outcome after surgical correction of failed triple arthrodesis. We conclude that such a revision, although complex, may be attempted to establish a plantigrade foot free of infection and able to wear shoes without and orthosis or brace.


GeNuIne (gene-nutrient interactions) Collaboration: towards implementing multi-ethnic population-based nutrigenetic studies of vitamin B12 and D deficiencies and metabolic diseases.
Gene-nutrient interactions (GeNuIne) collaboration, a large-scale collaborative project, has been initiated to investigate the impact of gene-nutrient interactions on cardiometabolic diseases using population-based studies from ethnically diverse populations. In this project, the relationship between deficiencies of vitamins B12 and D, and metabolic diseases was explored using a nutrigenetic approach. A genetic risk score (GRS) analysis was used to examine the combined effect of several genetic variations that have been shown to be associated with metabolic diseases and vitamin B12 and D deficiencies, respectively. In Sri Lankan, Indonesian and Brazilian populations, those carrying a high B12-GRS had an increased risk of metabolic diseases under the influence of dietary protein, fibre and carbohydrate intakes, respectively; however, in Asian Indians, genetically instrumented metabolic disease risk showed a significant association with low vitamin B12 status. With regards to nutrigenetic studies on vitamin D status, although high metabolic-GRS showed an interaction with dietary carbohydrate intake on vitamin D status, the study in Indonesian women demonstrated a vitamin D GRS-carbohydrate interaction on body fat percentage. In summary, these nutrigenetic studies from multiple ethnic groups have provided evidence for the influence of the dietary factors on the relationship between vitamin B12/D deficiency and metabolic outcomes. Furthermore, these studies highlight the existence of genetic heterogeneity in gene-diet interactions across ethnically diverse populations, which further implicates the significance of personalised dietary approaches for the prevention of these micronutrient deficiencies and metabolic diseases.


Herpes simplex virus non-structural proteins. IV. Purification of the virus-induced deoxyribonuclease and characterization of the enzyme using monoclonal antibodies.
The alkaline nucleases induced by herpes simplex virus type 1 (HSV-1) and type 2 (HSV-2) have been purified from high salt extracts of virus-infected cells. The purification used three types of column chromatography and resulted in apparently homogeneous DNase preparations with good recovery. The enzyme from HSV-2-infected cells has been characterized. It had both exonuclease and endonuclease activity, each with an unusually high pH optimum. The enzyme had an absolute requirement for magnesium which could not be replaced by other divalent cations. Analysis of the sedimentation characteristics and electrophoretic properties of the purified enzyme indicated that it was composed of a single subunit of mol. wt. 85 000. The purified HSV-2 enzyme was used as an immunogen to prime BALB/c mice which were used to prepare monoclonal antibodies. Three monoclonal antibodies were shown by several criteria to react with the enzyme. Thus, we were able to confirm that the 85K polypeptide did indeed have nuclease activity. This polypeptide was designated ICSP 22 in earlier studies and is a major polypeptide of virus-infected cells.


Applying mechanisms of chemical toxicity to predict drug safety.
Toxicology can no longer be used only as a science that reacts to problems but must be more proactive in predicting potential human safety issues with new drug candidates. Success in this area must be based on an understanding of the mechanisms of toxicity. This review summarizes and extends some of the concepts of an American Chemical Society ProSpectives meeting on the title subject held in June 2006. One important area is the discernment of the exact nature of the most common problems in drug toxicity. Knowledge of chemical structure alerts and relevant biological pathways are important. Biological activation to reactive products and off-target pharmacology are considered to be major contexts of drug toxicity, although defining exactly what the contributions are is not trivial. Some newer approaches to screening for both have been developed. A goal in predictive toxicology is the use of in vitro methods and database development to make predictions concerning potential modes of toxicity and to stratify drug candidates for further development. Such predictions are desirable for several economic and other reasons but are certainly not routine yet. However, progress has been made using several approaches. Some examples of the application of studies of wide-scale biological responses are now available, with incorporation into development paradigms.


Pharmacological treatment of Alzheimer's disease: effect of race and demographic variables.
Existing research shows differences in medication use for Alzheimer's disease (AD) based on demographics such as race, ethnicity, and geographical location. To determine individual and community characteristics associated with differences in acetylcholinesterase inhibitor (AChEI) and memantine use in AD, 3,049 AD subjects were drawn from 30 centers and evaluated using the Uniform data set (UDS). Cases were evaluated at the individual level within the context of 31 communities (one center encompassed two separate geographical regions). Multivariate analysis was used to determine the significance of individual variables on medication use. Compared to non-Hispanic Whites, Blacks were less likely to use AChEI and memantine with odds ratios (OR) of 0.59 (95% CI 0.46-0.76) and 0.43 (95% CI 0.32-0.57), respectively. Compared to non-Hispanic Whites, non-Black Hispanics were less likely to use memantine (OR=0.69 (95% CI 0.49-0.98)). No association was found between the proportion of Blacks or non-Black Hispanics versus non-Hispanic Whites at an Alzheimer Disease Center and individual use of AChEI or memantine. Other significant variables include gender, age, marital status, dementia severity, source of referral, AChEI use, and education. Education and age somewhat mitigated disparity. Significant racial and ethnic differences in AChEI and memantine use exist at the individual level regardless of the racial and ethnic composition of the individual's community. Research and initiatives at the societal level may be an important consideration toward addressing these differences.


Activation of MRTF-A-dependent gene expression with a small molecule promotes myofibroblast differentiation and wound healing.
Myocardin-related transcription factors (MRTFs) regulate cellular contractility and motility by associating with serum response factor (SRF) and activating genes involved in cytoskeletal dynamics. We reported previously that MRTF-A contributes to pathological cardiac remodeling by promoting differentiation of fibroblasts to myofibroblasts following myocardial infarction. Here, we show that forced expression of MRTF-A in dermal fibroblasts stimulates contraction of a collagen matrix, whereas contractility of MRTF-A null fibroblasts is impaired under basal conditions and in response to TGF-β1 stimulation. We also identify an isoxazole ring-containing small molecule, previously shown to induce smooth muscle α-actin gene expression in cardiac progenitor cells, as an agonist of myofibroblast differentiation. Isoxazole stimulates myofibroblast differentiation via induction of MRTF-A-dependent gene expression. The MRTF-SRF signaling axis is activated in response to skin injury, and treatment of dermal wounds with isoxazole accelerates wound closure and suppresses the inflammatory response. These results reveal an important role for MRTF-SRF signaling in dermal myofibroblast differentiation and wound healing and suggest that targeting MRTFs pharmacologically may prove useful in treating diseases associated with inappropriate myofibroblast activity.


Annular structures as intermediates in fibril formation of Alzheimer Abeta17-42.
We report all-atom molecular dynamics simulations of annular beta-amyloid (17-42) structures, single- and double-layered, in solution. We assess the structural stability and association force of Abeta annular oligomers associated through different interfaces, with a mutated sequence (M35A), and with the oxidation state (M35O). Simulation results show that single-layered annular models display inherent structural instability: one is broken down into linear-like oligomers, and the other collapses. On the other hand, a double-layered annular structure where the two layers interact through their C-termini to form an NC-CN interface (where N and C are the N and C termini, respectively) exhibits high structural stability over the simulation time due to strong hydrophobic interactions and geometrical constraints induced by the closed circular shape. The observed dimensions and molecular weight of the oligomers from atomic force microscopy (AFM) experiments are found to correspond well to our stable double-layered model with the NC-CN interface. Comparison with K3 annular structures derived from the beta 2-microglobulin suggests that the driving force for amyloid formation is sequence specific, strongly dependent on side-chain packing arrangements, structural morphologies, sequence composition, and residue positions. Combined with our previous simulations of linear-like Abeta, K3 peptide, and sup35-derived GNNQQNY peptide, the annular structures provide useful insight into oligomeric structures and driving forces that are critical in amyloid fibril formation.


[Potential endovascular prophylaxis for pulmonary thromboembolism in the combined treatment of cancer patients].
Our study was concerned with the role of ultrasound examination of the lower extremity veins to detect deep venous thrombosis and to evaluate complex prophylaxis of thromboembolism of pulmonary arteries (TEPA). The procedure involved vena cava filter placement in patients with neoplasia and high risk of TEPA as compared with controls receiving conservative therapy alone. A positive correlation was established between surgery and/or polychemotherapy, on the one hand, and higher stage and high risk of TEPA, on the other. Patients with deep venous thrombosis and neoplasia were referred to groups of extremely high risk of that pathology. Angiological history, physical examination and ultrasound check-ups of the lower vein must be carried out in cancer patients. Vena cava filter placement proved to be an effective and safe measure of TEPA prophylaxis. It lowered the risk of lethal outcome in 24 during surgery and polychemotherapy. Lethality rate among controls was 43.5 +/- 0.51%.


Design of the Trial to Assess Chelation Therapy (TACT).
TACT is an National Institutes of Health-sponsored, randomized, double-blind, placebo-controlled, 2 × 2 factorial clinical trial testing the benefits and risks of 40 infusions of a multicomponent disodium EDTA chelation solution compared with placebo and of an oral, high-dose multivitamin and mineral supplement. TACT has randomized and will follow up 1,708 patients for an average of approximately 4 years. The primary end point is a composite of all-cause mortality, myocardial infarction, stroke, coronary revascularization, and hospitalization for angina. A 900-patient substudy will examine quality-of-life outcomes. The trial is designed to have >85% power to detect a 25% relative reduction in the primary end point for each treatment factor. Enrollment began in September 2003 and was completed in October 2010.


The roles of 6K protein on Getah virus replication and pathogenicity.
Alphavirus is a type of arbovirus that can infect both humans and animals. The amino acid sequence of the 6K protein, being one of the structural proteins of the alphavirus, is not conserved. Deletion of this protein will result in varying effects on different alphaviruses. Our study focuses on the function of the Getah virus (GETV) 6K protein in infected cells and mice. We successfully constructed infectious clone plasmids and created resulting viruses (rGETV and rGETV-Δ6K). Our comprehensive microscopic analysis revealed that the 6 K protein mainly stays in the endoplasmic reticulum. In addition, rGETV-Δ6K has lower thermal stability and sensitivity to temperature than GETV. Although the deletion of the 6K protein does not reduce virion production in ST cells, it affects the release of virions from host cells by inhibiting the process of E2 protein transportation to the plasma membrane. Subsequent in vivo testing demonstrated that neonatal mice infected with rGETV-Δ6K had a lower virus content, less significant pathological changes in tissue slices, and milder disease than those infected with the wild-type virus. Our results indicate that the 6K protein effectively reduces the viral titer by influencing the release of viral particles. Furthermore, the 6K protein play a role in the clinical manifestation of GETV disease.


An endoplasmic reticulum-selective ratiometric fluorescent probe for imaging a copper pool.
Hydrazide-linked naphthalimides undergo copper ion-selective hydrolysis with concomitant ratiometric fluorescence change. In addition, probe 1 is biocompatible and mainly localized to the endoplasmic reticulum (ER) in live HeLa cells. It thus can be used for imaging copper accumulation in the ER of live cells, as implied in copper-overloaded situations.


Inhibition of PKMzeta in nucleus accumbens core abolishes long-term drug reward memory.
During abstinence, memories of drug-associated cues persist for many months, and exposure to these cues often provokes relapse to drug use. The mechanisms underlying the maintenance of these memories are unknown. A constitutively active atypical protein kinase C (PKC) isozyme, protein kinase M ζ (PKMζ), is required for maintenance of spatial memory, conditioned taste aversion, and other memory forms. We used conditioned place preference (CPP) and conditioned place aversion (CPA) procedures to study the role of nucleus accumbens PKMζ in the maintenance of drug reward and aversion memories in rats. Morphine CPP training (10 mg/kg, 4 pairings) increased PKMζ levels in accumbens core but not shell. Injections of the PKMζ inhibitor ζ inhibitory peptide (ZIP) into accumbens core but not shell after CPP training blocked morphine CPP expression for up to 14 d after injections. This effect was mimicked by the PKC inhibitor chelerythrine, which inhibits PKMζ, but not by the conventional and novel PKC inhibitor staurosporine, which does not effectively inhibit PKMζ. ZIP injections into accumbens core after training also blocked the expression of cocaine (10 mg/kg) and high-fat food CPP but had no effect on CPA induced by naloxone-precipitated morphine withdrawal. Accumbens core injections of Tat-GluR2(3Y), which inhibits GluR2-dependent AMPA receptor endocytosis, prevented the impairment in morphine CPP induced by local ZIP injections, indicating that the persistent effect of PKMζ is on GluR2-containing AMPA receptors. Results indicate that PKMζ activity in accumbens core is a critical cellular substrate for the maintenance of memories of relapse-provoking reward cues during prolonged abstinence periods.


Hemangiomas - current therapeutic strategies.
Hemangiomas are benign neoplasms of the vasculature frequently encountered in children. Several studies have shown that these tumors are characterized by excessive angiogenesis. Although benign, the lesions can present with complications, and may thus require treatment. There are multiple therapeutic options available for patients with problematic or life threatening hemangiomas, some of which have serious side effects. Randomized clinical trials and evidence-based studies on the efficacy of these treatments is still lacking. The recognition that excessive angiogenesis underlies hemangiogenesis offers an opportunity for the development of safer therapeutic strategies that are based on the inhibition of angiogenesis. We review medical therapies currently employed in the management of hemangiomas and the role of angiogenesis inhibition in hemangioma therapy.


Single voltage-dependent potassium channels in cultured rat hippocampal neurons.
Single-channel recordings using the gigohm seal patch-clamp technique were carried out on the somatic membranes of dissociated embryonic rat hippocampal neurons grown in cell culture. The recording medium contained tetrodotoxin to block the voltage-dependent Na+ conductance and Cd2+ to block Ca2+ and Ca2+-activated conductances. In the cell-attached configuration, depolarizing voltage steps activated outward directed single-channel currents with conductance 15-20 pS. The channel openings exhibited a moderate degree of flickering. The mean burst lifetimes ranged from 5 to 13 ms with a tendency to increase slightly at more depolarized potentials (T = 21-25 degrees C). Reversal potential measurements using excised membrane patches indicated that the channels behaved as expected of a K+-selective membrane pore. Channel opening occurred in Ca2+-free EGTA-containing solutions but was never observed in the presence of tetraethylammonium (TEA; 20 mM). The frequency of channel opening increased as the membrane was depolarized by up to 50 mV from resting potential; the fraction of time spent in the open state during the first 300 ms following a step depolarization increased e-fold for a 8-25 mV change in potential. First-latency histograms and simulations of the macroscopic current based on channel data obtained during repeated depolarizing voltage steps indicated that the probability of the channel being in the open state increases gradually with time after a step depolarization. During repeated depolarizing steps the channels appeared to randomly enter and exit a long-lived inactive state. It is concluded that these channels may underly the slowly activating, very slowly inactivating, TEA-sensitive voltage-dependent K+ current (IK) in cultured hippocampal neurons.


Factors associated with irritable bowel syndrome symptoms in hemodialysis patients.
To investigate clinical characteristics associated with the presence of irritable bowel syndrome (IBS) symptoms in hemodialysis (HD) patients. This was a cross-sectional study. A questionnaire based on the Bowel Disease Questionnaire that records gastrointestinal symptoms was given to 294 patients in 4 dialysis centers. A total of 196 (67%) subjects returned the survey. A multivariable logistic regression model was used to identify factors significantly associated with IBS symptoms. Symptoms compatible with IBS were present in 27 (13.8%) subjects and independently associated with low post-dialysis serum potassium [OR = 0.258, 95% CI (0.075-0.891), P = 0.032], paracetamol use [OR = 3.159, 95% CI (1.214-8.220), P = 0.018], and Kidney Disease Quality of Life (KDQOL) cognitive function score [OR = 0.977, 95% CI (0.956-0.999), P = 0.042]. Univariate regressions were also performed and the reported significance is for multivariate analysis. No association was detected for age, gender, depressed mood, smoking (present or past), body mass index, albumin level, Kt/V, sodium pre- or post-dialysis level, change in potassium level during HD, proton pump inhibitor or H2 blocker use, aspirin use, residual diuresis, hepatitis B or C infection, diabetes mellitus, marital status and education level. This study examined potential risk factors for symptoms compatible with IBS in HD patients and identified an association with paracetamol use, post-dialysis potassium level and KDQOL-cognitive function score.


Use of schizophrenia and bipolar disorder polygenic risk scores to identify psychotic disorders.
There is increasing evidence for shared genetic susceptibility between schizophrenia and bipolar disorder. Although genetic variants only convey subtle increases in risk individually, their combination into a polygenic risk score constitutes a strong disease predictor.AimsTo investigate whether schizophrenia and bipolar disorder polygenic risk scores can distinguish people with broadly defined psychosis and their unaffected relatives from controls. Using the latest Psychiatric Genomics Consortium data, we calculated schizophrenia and bipolar disorder polygenic risk scores for 1168 people with psychosis, 552 unaffected relatives and 1472 controls. Patients with broadly defined psychosis had dramatic increases in schizophrenia and bipolar polygenic risk scores, as did their relatives, albeit to a lesser degree. However, the accuracy of predictive models was modest. Although polygenic risk scores are not ready for clinical use, it is hoped that as they are refined they could help towards risk reduction advice and early interventions for psychosis.Declaration of interestR.M.M. has received honoraria for lectures from Janssen, Lundbeck, Lilly, Otsuka and Sunovian.


Factors associated with relative rates of antibiotic resistance in Pseudomonas aeruginosa isolates tested in clinical laboratories in the United States from 1999 to 2002.
For the period from 1999 to 2002 in the United States, the in vitro susceptibilities of 52,637 Pseudomonas aeruginosa isolates to 10 antimicrobial agents were evaluated. The isolates were from 29 laboratories, 11 of which participated in The Surveillance Network for four consecutive years. Isolates were collected from adult patients (> or =18 years of age) in intensive care units (ICU), non-ICU inpatients, nursing home patients, and outpatients; data were analyzed to evaluate factors, such as year of isolation, patient age group, isolate specimen source, and patient type (hospitalized patients [ICU, non-ICU, or nursing home] or outpatients). Rates of resistance for the 4-year period were highest for isolates from patients in ICU and 18- to 39-year-old patients and for isolates from the lower respiratory tract. Resistance decreased with age. Resistance was lowest in isolates from outpatients, in isolates from > or =70-year-old patients, and from specimens from the upper respiratory tract. Multidrug resistance (MDR) (resistance to three or more antimicrobial agents) accounted for 24.9% of all isolates. The MDR rate was highest in isolates from patients in nursing homes (29.9%) and ICU (29.5%).


Menstrual Blood-Derived Mesenchymal Stem Cells: Women's Attitudes, Willingness, and Barriers to Donation of Menstrual Blood.
Background: Menstrual blood contains mesenchymal stem cells (MenSC), considered a potential "off-the-shelf" treatment for a range of diseases and medical conditions. Samples of menstrual blood can be collected painlessly, inexpensively, and as frequently as every month for cell therapy. While there has been considerable previous research into the clinical advantages of MenSC, there is currently little understanding of potential donors' attitudes regarding menstrual blood donation and MenSC. Methods: One hundred women 18 years of age or over were surveyed to understand attitudes and potential barriers to menstrual blood donation. The questionnaire assessed participant age and brief medical history (giving birth, donating blood, donating stem cells), menstrual experience (period rating, preferred menstrual hygiene products), and whether participants would donate MenSC or accept MenSC therapy. Results: MenSC was met with a generally positive response, with 78% of menstruating women willing to donate menstrual blood. No significant relationship was recognized between willingness to donate menstrual blood with age, history of childbirth or blood donation, menstruation perception, and preferred menstrual hygiene product. Women rated their period experience better after being made aware of the ability to donate menstrual blood, meaning MenSC therapy can be beneficial for donors as well as patients. Conclusions: Considering women's attitudes to MenSC and donation of menstrual blood, the future of MenSC therapy is positive; women are generally willing to donate menstrual blood, independent of age, perception of periods, and history of childbirth and blood donation.


The value of plasma cell-free DNA levels as biomarker in patients with eating disorders: A preliminary study.
Circulating cell-free DNA (cfDNA) holds promise as a rapid and convenient biomarker for identifying individuals with eating disorders. To investigate this hypothesis, we measured plasma cfDNA in patients with different eating disorders. In this study, 110 participants (98 patients with eating disorders divided into 30 patients with bulimia nervosa, 33 patients with anorexia nervosa (AN) Restricting subtype, 35 patients with AN Binge-eating/purging subtype and 12 controls) were enrolled. We measured both cell-free nuclear DNA (cf-nDNA) and cell-free mitochondrial DNA (cf-mtDNA) from plasma using two specific droplet digital PCR assays each, referring to two amplicon sizes. Levels of plasma cf-nDNA and cf-mtDNA showed no significant differences between control participants and those with eating disorders. However, we observed a higher proportion of long cf-nDNA fragments in patients with eating disorders, suggesting its potential as a biomarker for eating disorders. This is the first study of cfDNA in patients with eating disorders. Our findings highlight the potential for qualitative exploration of cfDNA, although not of quantitative interest. Full characterization of cfDNA may serve as a valuable biomarker for eating disorders and provide some insights into the hidden mechanisms underlying the chronic development of these conditions. Future studies are needed to confirm or refute this hypothesis.


Acute myositis of the medial rectus muscle: diagnosis by computed tomography.
We have reported a case of myositis of the medial rectus muscle diagnosed and followed up by computerized tomography and orbital ultrasound.


The coat protein of tobamovirus acts as elicitor of both L2 and L4 gene-mediated resistance in Capsicum.
In Capsicum, the resistance conferred by the L(2) gene is effective against all of the pepper-infecting tobamoviruses except Pepper mild mottle virus (PMMoV), whereas that conferred by the L(4) gene is effective against them all. These resistances are expressed by a hypersensitive response, manifested through the formation of necrotic local lesions (NLLs) at the primary site of infection. The Capsicum L(2) gene confers resistance to Paprika mild mottle virus (PaMMV), while the L(4) gene is effective against both PaMMV and PMMoV. The PaMMV and PMMoV coat proteins (CPs) were expressed in Capsicum frutescens (L(2)L(2)) and Capsicum chacoense (L(4)L(4)) plants using the heterologous Potato virus X (PVX)-based expression system. In C. frutescens (L(2)L(2)) plants, the chimeric PVX virus containing the PaMMV CP was localized in the inoculated leaves and produced NLLs, whereas the chimeric PVX containing the PMMoV CP infected the plants systemically. Thus, the data indicated that the PaMMV CP is the only tobamovirus factor required for the induction of the host response mediated by the Capsicum L(2) resistance gene. In C. chacoense (L(4)L(4)) plants, both chimeric viruses were localized to the inoculated leaves and produced NLLs, indicating that either PaMMV or PMMoV CPs are required to elicit the L(4) gene-mediated host response. In addition, transient expression of PaMMV CP into C. frutescens (L(2)L(2)) leaves and PMMoV CP into C. chacoense (L(4)L(4)) leaves by biolistic co-bombardment with a beta-glucuronidase reporter gene led to the induction of cell death and the expression of host defence genes in both hosts. Thus, the tobamovirus CP is the elicitor of the Capsicum L(2) and L(4) gene-mediated hypersensitive response.


Isocitrate dehydrogenase (IDH) mutant gliomas: A Society for Neuro-Oncology (SNO) consensus review on diagnosis, management, and future directions.
Isocitrate dehydrogenase (IDH) mutant gliomas are the most common adult, malignant primary brain tumors diagnosed in patients younger than 50, constituting an important cause of morbidity and mortality. In recent years, there has been significant progress in understanding the molecular pathogenesis and biology of these tumors, sparking multiple efforts to improve their diagnosis and treatment. In this consensus review from the Society for Neuro-Oncology (SNO), the current diagnosis and management of IDH-mutant gliomas will be discussed. In addition, novel therapies, such as targeted molecular therapies and immunotherapies, will be reviewed. Current challenges and future directions for research will be discussed.


Mosquito survey during West Nile virus outbreak 2012 in northeast Croatia.
During the August and September 2012, seven human cases of the West Nile neuro-invasive disease were reported in Croatia. Medical entomology research on a potential vectors during the outbreak was supported by the Ministry of Health. A mosquito survey has been done in 64 sites in three eastern Croatian counties (Osijek-Baranja County, Vukovar-Srijem county and in Brod-Posavina county). Dry ice baited CDC traps were used for mosquito sampling in a period from the 10th to 25th September 2012. A total of 1785 mosquitoes were collected and 5 species were determined. The most numerous species were Aedes vexans with 1634 specimens, a Culex pipiens c., the potential vector of WNV, was present with 6.39%, in 114 specimens. That species was present in 43 out of 64 investigated sites. Vector control included both the control of mosquito larvae and the adults. Treatments have been done on 184 small breeding sites and on 2900 ha of an urban area.


Genome-wide monitoring of wine yeast gene expression during alcoholic fermentation.
The transcriptome of a wine yeast was monitored throughout an alcoholic fermentation under conditions mimicking an enological environment. Major changes in gene expression occurred during fermentation, affecting more than 2000 genes, as the yeast adapted to changing nutritional, environmental and physiological conditions. The genes of many pathways are regulated in a highly coordinated manner, and genes involved in the key metabolic pathways of fermentation are strongly expressed. We showed that, during fermentation of a synthetic medium mimicking a natural must in which growth arrest was caused by nitrogen exhaustion, entry into the stationary phase triggered major transcriptional reprogramming. Many TOR target genes involved in nitrogen utilization or other functions are induced at this stage, suggesting that this signalling pathway plays a critical role in changes in gene expression in response to nitrogen depletion. Entry into stationary phase is a key physiological event and is followed by a general stress response. The superimposition of multiple stresses, including starvation and ethanol stress, gives rise to a unique stress response, involving hundreds of genes encoding proteins involved in various cellular processes, many of unknown function.


Motif independent identification of potential RNA G-quadruplexes by G4RNA screener.
G-quadruplex structures in RNA molecules are known to have regulatory impacts in cells but are difficult to locate in the genome. The minimal requirements for G-quadruplex folding in RNA (G≥3N1-7 G≥3N1-7 G≥3N1-7 G≥3) is being challenged by observations made on specific examples in recent years. The definition of potential G-quadruplex sequences has major repercussions on the observation of the structure since it introduces a bias. The canonical motif only describes a sub-population of the reported G-quadruplexes. To address these issues, we propose an RNA G-quadruplex prediction strategy that does not rely on a motif definition. We trained an artificial neural network with sequences of experimentally validated G-quadruplexes from the G4RNA database encoded using an abstract definition of their sequence. This artificial neural network, G4NN, evaluates the similarity of a given sequence to known G-quadruplexes and reports it as a score. G4NN has a predictive power comparable to the reported G richness and G/C skewness evaluations that are the current state-of-the-art for the identification of potential RNA G-quadruplexes. We combined these approaches in the G4RNA screener, a program designed to manage and evaluate the sequences to identify potential G-quadruplexes. G4RNA screener is available for download at http://gitlabscottgroup.med.usherbrooke.ca/J-Michel/g4rna_screener. email@example.com or firstname.lastname@example.com or firstname.lastname@example.com. Supplementary data are available at Bioinformatics online.


Detection of 14-3-3 brain protein in the cerebrospinal fluid of patients with paraneoplastic neurological disorders
The detection of 14-3-3 protein in cerebrospinal fluid by immunoblotting is useful for the diagnosis of Creutzfeldt-Jakob disease (CJD). We found 14-3-3 protein in 10 of 80 (12.5%) patients with paraneoplastic neurological disorders (PNDs), whose presenting symptoms may mimic those of CJD. In 47 of 48 CJD patients, the 14-3-3 protein was detected as a single band, and it was detected as a double band in 1 patient. The double-band pattern was observed in 9 of the 10 14-3-3 proteinpositive patients with PNDs. The 14-3-3 protein assay may be positive in PND patients, but the immunoblotting pattern distinguishes most PND samples from those of CJD.