The Challenges and Opportunities of Pharmacoepidemiology in Bone Diseases.
Pharmacoepidemiology is used extensively in osteoporosis research and involves the study of the use and effects of drugs in large numbers of people. Randomized controlled trials are considered the gold standard in assessing treatment efficacy and safety. However, their results can have limited external validity when applied to day-to-day patients. Pharmacoepidemiological studies aim to assess the effect/s of treatments in actual practice conditions, but they are limited by the quality, completeness, and inherent bias due to confounding. Sources of information include prospectively collected (primary) as well as readily available routinely collected (secondary) (eg, electronic medical records, administrative/claims databases) data. Although the former enable the collection of ad hoc measurements, the latter provide a unique opportunity for the study of large representative populations and for the assessment of rare events at relatively low cost. Observational cohort and case-control studies, the most commonly implemented study designs in pharmacoepidemiology, each have their strengths and limitations. However, the choice of the study design depends on the research question that needs to be answered. Despite the many advantages of observational studies, they also have limitations. First, missing data is a common issue in routine data, frequently dealt with using multiple imputation. Second, confounding by indication arises because of the lack of randomization; multivariable regression and more specific techniques such as propensity scores (adjustment, matching, stratification, trimming, or weighting) are used to minimize such biases. In addition, immortal time bias (time period during which a subject is artefactually event-free by study design) and time-varying confounding (patient characteristics changing over time) are other types of biases usually accounted for using time-dependent modeling. Finally, residual "uncontrolled" confounding is difficult to assess, and hence to account for it, sensitivity analyses and specific methods (eg, instrumental variables) should be considered.


Production of reactive oxygen species is turned on and rapidly shut down in epithelial cells infected with Chlamydia trachomatis.
Reactive oxygen species (ROS) are many-faceted compounds involved in cell defense against pathogens, as well as in cell signaling. Their involvement in the response to infection in epithelial cells remains poorly documented. Here, we investigated the production of ROS during infection with Chlamydia trachomatis, a strict intracellular pathogen, in HeLa cells. C. trachomatis induced a transient increase in the ROS level within a few hours, followed by a return to basal level 9 hours after infection. At this time point, the host enzyme dedicated to ROS production, NADPH oxidase, could no longer be activated by external stimuli, such as interleukin-1beta. In addition, Rac, a regulatory subunit of the NADPH oxidase complex, was relocated to the membrane of the compartment in which the bacteria develop, the inclusion, while other subunits were not. Altogether, these results indicate that C. trachomatis infection elicits the production of ROS and that the bacteria rapidly target the activity of NADPH oxidase to shut it down. Prevention of ROS production at the onset of the bacterial developmental cycle might delay the host response to infection.


Interventions to enhance informed choices among invitees of screening programmes-a systematic review.
Informed decision making about participation has become an explicit purpose in invitations for screening programmes in western countries. An informed choice is commonly defined as based on: (i) adequate levels of knowledge of the screening and (ii) agreement between the invitee's values towards own screening participation and actual (intention to) participation. We systematically reviewed published studies that empirically evaluated the effects of interventions aiming at enhancing informed decision making in screening programmes targeted at the general population. We focused on prenatal screening and neonatal screening for diseases of the foetus/new-born and screening for breast cancer, cervical cancer and colorectal cancer. The Medline, EMBASE and Cochrane databases were searched for studies published till April 2012, using the terms 'informed choice', 'decision making' and 'mass screening' separately and in combination and terms referring to the specific screening programmes. Of the 2238 titles identified, 15 studies were included, which evaluated decision aids (DAs), information leaflets, film, video, counselling and a specific screening visit for informed decision making in prenatal screening, breast and colorectal cancer screening. Most of the included studies evaluated DAs and showed improved knowledge and informed decision making. Due to the limited number of studies the results could not be synthesized. The empirical evidence regarding interventions to improve informed decision making in screening is limited. It is unknown which strategies to enhance informed decision making are most effective, although DAs are promising. Systematic development of interventions to enhance informed choices in screening deserves priority, especially in disadvantaged groups.


Enhanced light focusing in self-assembled optoplasmonic clusters with subwavelength dimensions.
Compact metallo-dielectric hybrid clusters with subwavelength dimensions are fabricated by template guided self-assembly. Elastic and inelastic scattering spectroscopy and electromagnetic simulations reveal that hybrid clusters comprising TiO(2) nanoparticles on top of a cluster of strongly coupled gold nanoparticles harness synergistic electromagnetic interactions between the building blocks. This results in a boost of the peak electric field intensity and a redistribution of the field in the ambient medium. The complex phase landscape in the clusters features optical vortices that enhance the magnetic field.


Tracing anti-cancer and cancer-promoting actions of all-trans retinoic acid in breast cancer to a RARα epigenetic mechanism of mammary epithelial cell fate.
A hallmark of cancer cells is the ability to evade the growth inhibitory/pro-apoptotic action of physiological all-trans retinoic acid (RA) signal, the bioactive derivative of Vitamin A. However, as we and others reported, RA can also promote cancer cell growth and invasion. Here we show that anticancer and cancer-promoting RA actions in breast cancer have roots in a mechanism of mammary epithelial cell morphogenesis that involves both transcriptional (epigenetic) and non-transcriptional RARα (RARA) functions. We found that the mammary epithelial cell-context specific degree of functionality of the RARA transcriptional (epigenetic) component of this mechanism, by tuning the effects of the non-transcriptional RARA component, determines different cell fate decisions during mammary morphogenesis. Indeed, factors that hamper the RARA epigenetic function make physiological RA drive aberrant morphogenesis via non-transcriptional RARA, thus leading to cell transformation. Remarkably, also the cell context-specific degree of functionality of the RARA epigenetic component retained by breast cancer cells is critical to determine cell fate decisions in response to physiological as well as supraphysiological RA variation. Overall this study supports the proof of principle that the epigenetic functional plasticity of the mammary epithelial cell RARA mechanism, which is essential for normal morphogenetic processes, is necessary to deter breast cancer onset/progression consequent to the insidious action of physiological RA.


Ictal-Interictal Continuum.
The term "ictal-interictal" continuum has seen wide adoption in the critical care EEG domain, referring to the presence of abnormal periodic activity on the scalp EEG variably associated with seizures. The historical origin of the ictal-interictal continuum concept is discussed with a review of known and surmised physiological mechanisms for their origin and relationship to seizures. Therapeutic approaches to patients exhibiting ictal-interictal continuum EEG patterns are reviewed, and some open scientific questions highlighted. Further understanding of the ictal-interictal continuum is likely to significantly improve the care of the critically ill neurological patient.


The effect of antibody against TNF alpha on cytokine response in Jarisch-Herxheimer reactions of louse-borne relapsing fever.
Severe Jarisch Herxheimer reaction (J-HR) precipitated by antibiotic treatment of louse-borne relapsing fever (LBRF) is associated with a transient, marked rise in circulating tumour necrosis factor alpha (TNF alpha), interleukin 6 (IL-6) and interleukin 8 (IL-8). Ovine polyclonal anti-TNF alpha antibody fragments (Fab) were used in a randomized double blind placebo controlled trial in an attempt to prevent this reaction. Within 4 h after penicillin, in controls (n = 29), a several-fold rise in cytokines occurred, concomitant with a fall in spirochaetes and maximal clinical manifestations of the J-HR. An intravenous infusion of anti-TNF alpha Fab, 30 min before penicillin in 20 patients reduced peak plasma levels of IL-6 and IL-8 (but not IL-1 beta) compared with controls (p = 0.01 and < 0.001, respectively) and the incidence of the J-HR, indicating some neutralization of TNF alpha. An apparent fall in TNF alpha reflected interference of anti-TNF alpha in the immunoassay.


Laparoscopic-assisted endoscopic sigmoidopexy: a new surgical option for sigmoid volvulus.
Sigmoid volvulus accounts for 5% of cases of large-bowel obstruction. Here, we present the development of a new minimally invasive treatment method that aims to reduce the complication rate of standard percutaneous endoscopic colostomy tube insertion and negate the need for prolonged anesthesia or colectomy. Six patients underwent laparoscopic-assisted endoscopic sigmoidopexy for recurrent sigmoid volvulus at a mean age of 80.5 years (range, 76-83). The volvulus was decompressed endoscopically before laparoscopic adhesiolysis and detorsion of the sigmoid. Finally, the sigmoid was approximated with the anterior abdominal wall, and 2 endoscopically placed percutaneous endoscopic colostomy tubes were inserted. Later, the external component of the percutaneous endoscopic colostomy tubes was removed, and the internal parts were passed by way of the rectum. Each operation was completed successfully in a mean time of 69 minutes and with no intraoperative complication. The mean postoperative stay was 20 days (range, 4-54). At median follow-up of 10.8 months, all patients were tube free with no incidence of recurrent volvulus, inadvertent tube traction, or leak. This preliminary report shows laparoscopic-assisted endoscopic sigmoidopexy to be a safe treatment modality for recurrent sigmoid volvulus. As such, it is particularly useful for elderly patients, for whom colectomy is a high-risk procedure.


TM9SF4 is required for Drosophila cellular immunity via cell adhesion and phagocytosis.
Nonaspanins are characterised by a large N-terminal extracellular domain and nine putative transmembrane domains. This evolutionarily conserved family comprises three members in Dictyostelium discoideum (Phg1A, Phg1B and Phg1C) and Drosophila melanogaster, and four in mammals (TM9SF1-TM9SF4), the function of which is essentially unknown. Genetic studies in Dictyostelium demonstrated that Phg1A is required for cell adhesion and phagocytosis. We created Phg1A/TM9SF4-null mutant flies and showed that they were sensitive to pathogenic Gram-negative, but not Gram-positive, bacteria. This increased sensitivity was not due to impaired Toll or Imd signalling, but rather to a defective cellular immune response. TM9SF4-null larval macrophages phagocytosed Gram-negative E. coli inefficiently, although Gram-positive S. aureus were phagocytosed normally. Mutant larvae also had a decreased wasp egg encapsulation rate, a process requiring haemocyte-dependent adhesion to parasitoids. Defective cellular immunity was coupled to morphological and adhesion defects in mutant larval haemocytes, which had an abnormal actin cytoskeleton. TM9SF4, and its closest paralogue TM9SF2, were both required for bacterial internalisation in S2 cells, where they displayed partial redundancy. Our study highlights the contribution of phagocytes to host defence in an organism possessing a complex innate immune response and suggests an evolutionarily conserved function of TM9SF4 in eukaryotic phagocytes.


Malaria transmission modelling: a network perspective.
Malaria transmission can be affected by multiple or even hidden factors, making it difficult to timely and accurately predict the impact of elimination and eradication programs that have been undertaken and the potential resurgence and spread that may continue to emerge. One approach at the moment is to develop and deploy surveillance systems in an attempt to identify them as timely as possible and thus to enable policy makers to modify and implement strategies for further preventing the transmission. Most of the surveillance data will be of temporal and spatial nature. From an interdisciplinary point of view, it would be interesting to ask the following important as well as challenging question: Based on the available surveillance data in temporal and spatial forms, how can we build a more effective surveillance mechanism for monitoring and early detecting the relative prevalence and transmission patterns of malaria? What we can note from the existing clustering-based surveillance software systems is that they do not infer the underlying transmission networks of malaria. However, such networks can be quite informative and insightful as they characterize how malaria transmits from one place to another. They can also in turn allow public health policy makers and researchers to uncover the hidden and interacting factors such as environment, genetics and ecology and to discover/predict malaria transmission patterns/trends. The network perspective further extends the present approaches to modelling malaria transmission based on a set of chosen factors. In this article, we survey the related work on transmission network inference, discuss how such an approach can be utilized in developing an effective computational means for inferring malaria transmission networks based on partial surveillance data, and what methodological steps and issues may be involved in its formulation and validation.


Blindsight modulation of motion perception.
Monkey data suggest that of all perceptual abilities, motion perception is the most likely to survive striate damage. The results of studies on motion blindsight in humans, though, are mixed. We used an indirect strategy to examine how responses to visible stimuli were modulated by blind-field stimuli. In a 26-year-old man with focal striate lesions, discrimination of visible optic flow was enhanced about 7% by blind-field flow, even though discrimination of optic flow in the blind field alone (the direct strategy) was at chance. Pursuit of an imagined target using peripheral cues showed reduced variance but not increased gain with blind-field cues. Preceding blind-field prompts shortened reaction times to visible targets by about 10 msec, but there was no attentional crowding of visible stimuli by blind-field distractors. A similar efficacy of indirect blind-field optic flow modulation was found in a second patient with residual vision after focal striate damage, but not in a third with more extensive medial occipito-temporal damage. We conclude that indirect modulatory strategies are more effective than direct forced-choice methods at revealing residual motion perception after focal striate lesions.


Prospective study on ocular motility limitation due to orbital muscle entrapment or impingement associated with orbital wall fracture.
The recommended urgent surgical management of ocular motility restriction due to orbital muscle entrapment or impingement associated with orbital wall fracture needs to be elucidated. To evaluate the importance of the time from injury to surgery for the outcome in ocular motility and diplopia, the time lapse of ocular motility, diplopia and hypesthesia recovery. Patients with entrapment or impingement of orbital contents due to orbital wall fracture were followed up prospectively over 1year regarding ocular motility, diplopia, hypesthesia and cosmetic deformity. 21 patients (10 entrapments and 11 impingements) were included and treated surgically. The median time from injury to surgery was 36 (8-413)h for the entrapment group and 168 (48-326)h for the impingement group. The median time from study inclusion to surgery was 0 (0-1) days for the entrapment group and 1.0 (0.2-4.8) days for the impingement group. All the patients had ocular motility limitation and diplopia at the inclusion. Ocular motility improved gradually and was normal at final visit. Diplopia resolved gradually in all patients except in two with non-disturbing diplopia, at the final visit. Forced duction test was positive in 90% of the patients in the entrapment group and 70% in impingement group. At final visit, hypesthesia was found in none of the patients in the entrapment group but in 4 patients in the impingement group. In this, the first prospective long term follow up of orbital wall fractures with ocular motility restriction, we did not find any significant correlation between the time from injury to surgery and the outcomes in ocular motility and diplopia. An entrapment requires surgery as soon as possible; however, the surgical reduction is at least as important as surgical timing. Surgery should be delayed until it can be performed by an experienced surgeon. Ocular motility restriction causing diplopia due to impingement is not an ophthalmologic emergency and surgery is recommended if the diplopia and ocular motility has not improved over time. Clinical examination of ocular motility and not CT scan findings is crucial to determine whether a limitation of ocular motility exists or not.


Associations Among Child Maltreatment, Mental Health, and Police Contact in Adulthood: Findings From a National Canadian Sample.
The purpose of this study was to examine associations between three subtypes of childhood maltreatment (physical abuse, sexual abuse, and exposure to intimate partner violence) and two forms of adult police contact (criminality, victimization) using nationally representative Canadian data. Presence of a mental health disorder was also explored as a potential mediating variable in these associations. The weighted sample included 23,846 adult participants from the 2012 Canadian Community Health Survey-Mental Health. Logistic and multinomial regression analyses examined associations among individual and multiple exposures to subtypes of childhood maltreatment with police contact. The Sobel test was used to assess the mediating effect of mental health disorders. Adjusting for sociodemographic variables, results indicated that all maltreatment subtypes were significantly associated with increased odds of both forms of police contact (adjusted odds ratios ranged from 2.06 to 2.95). Presence of a mental health disorder was a partial mediator in the associations between child maltreatment and both forms of adult police contact (adjusted odds ratios ranged from 1.52 to 2.32). In addition, a dose-response relationship was observed for victimization; as the number of subtypes of maltreatment increased, there was an incremental increase in risk of victimization. Future efforts are needed to prioritize child maltreatment prevention, trauma-informed approaches, mental health awareness, and training in law enforcement.


Polymorphism of HLA-A, -B, -DRB1, -DQA1 and -DQB1 haplotypes in a Croatian population.
We describe for the first time extended haplotypes in a Croatian population. The present study gives the HLA-A, -B, -DRB1, -DQA1 and -DQB1 allele and haplotype frequencies in 105 families with at least two offspring. All individuals were studied by conventional serology for HLA class I antigens (A and B), while class II alleles (DRB1, DQA1, DQB1) were typed using the PCR-SSOP method. HLA genotyping was performed by segregation in all 105 families. For extended haplotype analysis, 420 independent parental haplotypes were included. Fourteen HLA-A, 18 HLA-B, 28 DRB1, 9 DQA1 and 11 DQB1 alleles were found in the studied population. Most of the DRB1 alleles in our population had an exclusive association with one specific DQA1-DQB1 combination. This strong linkage disequilibrium within the HLA class II region is often extended to the HLA-B locus. A total of 10 HLA-A, -B, -DRB1, -DQA1, -DQB1 haplotypes were observed with a frequency </= 1.0%. The three most frequent haplotypes were HLA-A1, B8, DRB1*0301, DQA1*0501, DQB1*0201; HLA-A3, B7, DRB1*1501, DQA1*0102, DQB1*0602 and HLA-A24, B44, DRB1*0701, DQA1*0201, DQB1*02. These results should provide a useful reference for further anthropological studies, transplantation studies, and studies of associations between HLA and diseases.


Genome Sequence Analysis of the Naphthenic Acid Degrading and Metal Resistant Bacterium Cupriavidus gilardii CR3.
Cupriavidus sp. are generally heavy metal tolerant bacteria with the ability to degrade a variety of aromatic hydrocarbon compounds, although the degradation pathways and substrate versatilities remain largely unknown. Here we studied the bacterium Cupriavidus gilardii strain CR3, which was isolated from a natural asphalt deposit, and which was shown to utilize naphthenic acids as a sole carbon source. Genome sequencing of C. gilardii CR3 was carried out to elucidate possible mechanisms for the naphthenic acid biodegradation. The genome of C. gilardii CR3 was composed of two circular chromosomes chr1 and chr2 of respectively 3,539,530 bp and 2,039,213 bp in size. The genome for strain CR3 encoded 4,502 putative protein-coding genes, 59 tRNA genes, and many other non-coding genes. Many genes were associated with xenobiotic biodegradation and metal resistance functions. Pathway prediction for degradation of cyclohexanecarboxylic acid, a representative naphthenic acid, suggested that naphthenic acid undergoes initial ring-cleavage, after which the ring fission products can be degraded via several plausible degradation pathways including a mechanism similar to that used for fatty acid oxidation. The final metabolic products of these pathways are unstable or volatile compounds that were not toxic to CR3. Strain CR3 was also shown to have tolerance to at least 10 heavy metals, which was mainly achieved by self-detoxification through ion efflux, metal-complexation and metal-reduction, and a powerful DNA self-repair mechanism. Our genomic analysis suggests that CR3 is well adapted to survive the harsh environment in natural asphalts containing naphthenic acids and high concentrations of heavy metals.


New insights into gestational glucose metabolism: lessons learned from 21st century approaches.
Pregnancy presents a unique physiological challenge that requires changes coordinated by placentally and non-placentally derived hormones to prepare the mother for the metabolic stress presented by fetal development and to ensure appropriate nutrient allocation between mother and fetus. Of particular importance is the maintenance of normal glucose metabolism during pregnancy. Here, we describe physiological changes in glucose metabolism during pregnancy and highlight new insights into these adaptations that have emerged over the past decade using novel methodologies, specifically genome-wide association studies (GWAS) and metabolomics. While GWAS have identified some novel associations with metabolic traits during pregnancy, the majority of the findings overlap with those observed in nonpregnant populations and individuals with type 2 diabetes (T2D). Metabolomics studies have provided new insight into key metabolites involved in gestational diabetes mellitus (GDM). Both of these approaches have suggested that a strong link exists between GDM and T2D. Most recently, a role of the gut microbiome in pregnancy has been observed, with changes in the microbiome during the third trimester having metabolic consequences for the mother. In this Perspectives in Diabetes article, we highlight how these new data have broadened our understanding of gestational metabolism, and emphasize the importance of future studies to elucidate differences between GDM and T2D.


Design of freeze-drying processes for pharmaceuticals: practical advice.
Design of freeze-drying processes is often approached with a "trial and error" experimental plan or, worse yet, the protocol used in the first laboratory run is adopted without further attempts at optimization. Consequently, commercial freeze-drying processes are often neither robust nor efficient. It is our thesis that design of an "optimized" freeze-drying process is not particularly difficult for most products, as long as some simple rules based on well-accepted scientific principles are followed. It is the purpose of this review to discuss the scientific foundations of the freeze-drying process design and then to consolidate these principles into a set of guidelines for rational process design and optimization. General advice is given concerning common stability issues with proteins, but unusual and difficult stability issues are beyond the scope of this review. Control of ice nucleation and crystallization during the freezing step is discussed, and the impact of freezing on the rest of the process and final product quality is reviewed. Representative freezing protocols are presented. The significance of the collapse temperature and the thermal transition, denoted Tg', are discussed, and procedures for the selection of the "target product temperature" for primary drying are presented. Furthermore, guidelines are given for selection of the optimal shelf temperature and chamber pressure settings required to achieve the target product temperature without thermal and/or mass transfer overload of the freeze dryer. Finally, guidelines and "rules" for optimization of secondary drying and representative secondary drying protocols are presented.


Measuring spore settling velocity for an improved assessment of dispersal rates in mosses.
The settling velocity of diaspores is a key parameter for the measurement of dispersal ability in wind-dispersed plants and one of the most relevant parameters in explicit dispersal models, but remains largely undocumented in bryophytes. The settling velocities of moss spores were measured and it was determined whether settling velocities can be derived from spore diameter using Stokes' Law or if specific traits of spore ornamentation cause departures from theoretical expectations. A fall tower design combined with a high-speed camera was used to document spore settling velocities in nine moss species selected to cover the range of spore diameters within the group. Linear mixed effect models were employed to determine whether settling velocity can be predicted from spore diameter, taking specific variation in shape and surface roughness into account. Average settling velocity of moss spores ranged from 0·49 to 8·52 cm s(-1) There was a significant positive relationship between spore settling velocity and size, but the inclusion of variables of shape and texture of spores in the best-fit models provides evidence for their role in shaping spore settling velocities. Settling velocities in mosses can significantly depart from expectations derived from Stokes' Law. We suggest that variation in spore shape and ornamentation affects the balance between density and drag, and results in different dispersal capacities, which may be correlated with different life-history traits or ecological requirements. Further studies on spore ultrastructure would be necessary to determine the role of complex spore ornamentation patterns in the drag-to-mass ratio and ultimately identify what is the still poorly understood function of the striking and highly variable ornamentation patterns of the perine layer on moss spores.


Weak memory for future-oriented feedback: investigating the roles of attention and improvement focus.
Recent research showed that people recall past-oriented, evaluative feedback more fully and accurately than future-oriented, directive feedback. Here we investigated whether these memory biases arise from preferential attention toward evaluative feedback during encoding. We also attempted to counter the biases via manipulations intended to focus participants on improvement. Participants received bogus evaluative and directive feedback on their writing. Before reading the feedback, some participants set goals for improvement (experiments 1 and 2), or they wrote about their past or future use of the writing skills, and/or were incentivised to improve (experiment 3); we objectively measured participants' attention during feedback reading using eyetracking. Finally, all participants completed a recall test. We successfully replicated the preferential remembering of evaluative feedback, but found little support for an attentional explanation. Goal-setting reduced participants' tendency to reproduce feedback in an evaluative style, but not their preferential remembering of evaluative feedback. Neither orienting participants toward their past or future use of the writing skills, nor incentivising them to improve, influenced their attention toward or memory for the feedback. These findings advance the search for a mechanism to explain people's weaker memory for future-oriented feedback, demonstrating that attentional and improvement-oriented accounts cannot adequately explain the effect.


Reduced repressive epigenetic marks, increased DNA damage and Alzheimer's disease hallmarks in the brain of humans and mice exposed to particulate urban air pollution.
Exposure to air pollutants is associated with an increased risk of developing Alzheimer's disease (AD). AD pathological hallmarks and cognitive deficits are documented in children and young adults in polluted cities (e.g. Metropolitan Mexico City, MMC). Iron-rich combustion- and friction-derived nanoparticles (CFDNPs) that are abundantly present in airborne particulate matter pollution have been detected in abundance in the brains of young urbanites. Epigenetic gene regulation has emerged as a candidate mechanism linking exposure to air pollution and brain diseases. A global decrease of the repressive histone post-translational modifications (HPTMs) H3K9me2 and H3K9me3 (H3K9me2/me3) has been described both in AD patients and animal models. Here, we evaluated nuclear levels of H3K9me2/me3 and the DNA double-strand-break marker γ-H2AX by immunostaining in post-mortem prefrontal white matter samples from 23 young adults (age 29 ± 6 years) who resided in MMC (n = 13) versus low-pollution areas (n = 10). Lower H3K9me2/me3 and higher γ-H2A.X staining were present in MMC urbanites, who also displayed the presence of hyperphosphorylated tau and amyloid-β (Aβ) plaques. Transmission electron microscopy revealed abundant CFDNPs in neuronal, glial and endothelial nuclei in MMC residents' frontal samples. In addition, mice exposed to particulate air pollution (for 7 months) in urban Santiago (Chile) displayed similar brain impacts; reduced H3K9me2/me3 and increased γ-H2A.X staining, together with increased levels of AD-related tau phosphorylation. Together, these findings suggest that particulate air pollution, including metal-rich CFDNPs, impairs brain chromatin silencing and reduces DNA integrity, increasing the risk of developing AD in young individuals exposed to high levels of particulate air pollution.


Evidence for exacerbation of irritable bowel syndrome during menses.
Many women report that bowel symptoms are associated with menstruation, but neither the prevalence of these complaints nor their physiological basis is known. This study aimed to estimate prevalence, to determine whether patients with irritable bowel syndrome are more likely to make such complaints, and to determine whether bowel complaints during menstruation are attributable to psychological traits such as increased somatization. To estimate prevalence, 369 clients of Planned Parenthood of Maryland were asked whether gas, diarrhea, or constipation occurred during menstruation. These subjects were compared with women referred to a gastroenterology clinic and found to have irritable bowel syndrome or functional bowel disorder (abdominal pain plus altered bowel habits but not satisfying restrictive criteria for irritable bowel syndrome). Thirty-four percent of 233 Planned Parenthood clients who denied symptoms of irritable bowel syndrome or functional bowel disorder reported that menstruation was associated with one or more bowel symptoms. Gastroenterology clinic patients with irritable bowel syndrome were significantly more likely to experience exacerbations of each of these bowel symptoms, but especially increased bowel gas. Self-reports of bowel symptoms during menstruation were not associated with psychological traits or with menses-related changes in affect.


Activation of a phosphotyrosine phosphatase by tyrosine phosphorylation.
Regulation of cell proliferation, differentiation, and metabolic homeostasis is associated with the phosphorylation and dephosphorylation of specific tyrosine residues of key regulatory proteins. The phosphotyrosine phosphatase 1D (PTP 1D) contains two amino terminally located Src homology 2 (SH2) domains and is similar to the Drosophila corkscrew gene product, which positively regulates the torso tyrosine kinase signal transduction pathway. PTP activity was found to be regulated by physical interaction with a protein tyrosine kinase. PTP 1D did not dephosphorylate receptor tyrosine kinases, despite the fact that it associated with the epidermal growth factor receptor and chimeric receptors containing the extracellular domain of the epidermal growth factor receptor and the cytoplasmic domain of either the HER2-neu, kit-SCF, or platelet-derived growth factor beta (beta PDGF) receptors. PTP 1D was phosphorylated on tyrosine in cells overexpressing the beta PDGF receptor kinase and this tyrosine phosphorylation correlated with an enhancement of its catalytic activity. Thus, protein tyrosine kinases and phosphatases do not simply oppose each other's action; rather, they may work in concert to maintain a fine balance of effector activation needed for the regulation of cell growth and differentiation.


Arthroscopic Resection of a Large Bony Exostosis Arising from the Anterior Inferior Iliac Spine Causing Extra-articular Hip Impingement: A Case Series.
Abnormal morphology of the anterior inferior iliac spine (AIIS) is a rarely recognized but important source of extra-articular hip impingement. Chronic progressive symptoms of stiffness and limitation of hip motion with persistent groin pain may place significant restriction on activity. Concomitant femoroacetabular impingement is often present but recognition and effective treatment of the uncommon extra-articular component is important for successful outcome. Three cases of symptomatic extra-articular hip impingement secondary to AIIS deformity and in conjunction with mild underlying femoroacetabular impingement (FAI) are presented. They include two athletic Caucasian males aged 27 and 35-years old with a history of prior rectus tendon injury and secondary bony exostosis formation and a 53-year-old Caucasian male with a nontraumatic, developmental AIIS deformity. In all cases, an excellent clinical outcome with a full return to pain free activity was achieved postoperation. Their clinical presentation, diagnosis and post-operative outcome at 1.5-2 years (mean 1.7 years) following arthroscopic AIIS resection are discussed. Arthroscopic management of AIIS extra-articular hip impingement has been rarely reported and the longer-term outcome is unknown. We report the successful clinical outcome in a case series of three patients up to 2 years following arthroscopic AIIS resection. This case series demonstrates the sustainable benefits of arthroscopic correction of AIIS bony exostosis as a cause of extra-articular FAI.


Pathogen-Specific Binding Soluble Down Syndrome Cell Adhesion Molecule (Dscam) Regulates Phagocytosis via Membrane-Bound Dscam in Crab.
The Down syndrome cell adhesion molecule (Dscam) gene is an extraordinary example of diversity that can produce thousands of isoforms and has so far been found only in insects and crustaceans. Cumulative evidence indicates that Dscam may contribute to the mechanistic foundations of specific immune responses in insects. However, the mechanism and functions of Dscam in relation to pathogens and immunity remain largely unknown. In this study, we identified the genome organization and alternative Dscam exons from Chinese mitten crab, Eriocheir sinensis. These variants, designated EsDscam, potentially produce 30,600 isoforms due to three alternatively spliced immunoglobulin (Ig) domains and a transmembrane domain. EsDscam was significantly upregulated after bacterial challenge at both mRNA and protein levels. Moreover, bacterial specific EsDscam isoforms were found to bind specifically with the original bacteria to facilitate efficient clearance. Furthermore, bacteria-specific binding of soluble EsDscam via the complete Ig1-Ig4 domain significantly enhanced elimination of the original bacteria via phagocytosis by hemocytes; this function was abolished by partial Ig1-Ig4 domain truncation. Further studies showed that knockdown of membrane-bound EsDscam inhibited the ability of EsDscam with the same extracellular region to promote bacterial phagocytosis. Immunocytochemistry indicated colocalization of the soluble and membrane-bound forms of EsDscam at the hemocyte surface. Far-Western and coimmunoprecipitation assays demonstrated homotypic interactions between EsDscam isoforms. This study provides insights into a mechanism by which soluble Dscam regulates hemocyte phagocytosis via bacteria-specific binding and specific interactions with membrane-bound Dscam as a phagocytic receptor.


[Reform of long-term care in the Netherlands: solidarity maintained?].
The reform of long-term care (LTC) in the Netherlands is a much debated topic. The reform essentially comes down to a shift in healthcare claims and a cutback. As of 1 January 2015, the Long-Term Care Act (WLZ) shall replace the Exceptional Medical Expenses Act (AWBZ). In doing so, parts of the AWBZ will shift to the Health Care Insurance Act (ZVW) and the renewed Social Support Act (WMO 2015), which will be carried out by municipalities. This is a significant change: whereas the AWBZ provides a right to care, the WMO commands delivery of tailor-made support. Care that falls under the WMO is only awarded if the capacity of persons seeking care, among others their financial resources and social network, are insufficient. Higher contributions than in the AWBZ may also be requested. These developments influence the experienced level of solidarity.


Patient empowerment in intensive care--an interview study.
Intensive care patients often experience a lack of control, as well as inner chaos. Experiences from intensive care can continue to affect patients for a long time. Empowerment is a positive and dynamic process that focuses on people's strengths, rights and abilities. It takes on different expressions for different people in different environments and must be described by the people involved. The aim of this study was to describe patient empowerment in an intensive care situation. The study was based on open-ended interviews with 11 patients in two intensive care units. The interviews were analysed according to the empirical phenomenological psychological method. The results showed that patient empowerment in intensive care consists of strengthening and stimulating the patients' own inherent joy of life and will to fight. A positive environment that encouraged feelings of value and motivation and in which the patient felt safe, received additional care and participated as he/she wished had a positive influence.


Time-Resolved Diffuse Optical Spectroscopy and Imaging Using Solid-State Detectors: Characteristics, Present Status, and Research Challenges.
Diffuse optical spectroscopy (DOS) and diffuse optical imaging (DOI) are emerging non-invasive imaging modalities that have wide spread potential applications in many fields, particularly for structural and functional imaging in medicine. In this article, we review time-resolved diffuse optical imaging (TR-DOI) systems using solid-state detectors with a special focus on Single-Photon Avalanche Diodes (SPADs) and Silicon Photomultipliers (SiPMs). These TR-DOI systems can be categorized into two types based on the operation mode of the detector (free-running or time-gated). For the TR-DOI prototypes, the physical concepts, main components, figures-of-merit of detectors, and evaluation parameters are described. The performance of TR-DOI prototypes is evaluated according to the parameters used in common protocols to test DOI systems particularly basic instrumental performance (BIP). In addition, the potential features of SPADs and SiPMs to improve TR-DOI systems and expand their applications in the foreseeable future are discussed. Lastly, research challenges and future developments for TR-DOI are discussed for each component in the prototype separately and also for the entire system.


Role of phosphoinositide 3-kinases in regulating cardiac function.
Phosphoinositide 3-kinases (PI3Ks) are important signaling proteins in the heart. Class IA PI3Ks (p110alpha, beta) are critical regulators of physiological heart growth and cell survival, and are generally considered to be beneficial for heart function. In contrast, activation of class IB PI3K(p110gamma) is detrimental for heart function, reducing cardiac contractility. This may have implications for the treatment of heart disease and failure. In vitro, ex vivo and in vivo studies have contributed to our understanding of PI3K signaling in the heart. This review summarizes class I PI3K signaling in the regulation of cardiac function, with a particular focus on the role of different PI3K isoforms in settings of heart disease.


Peripheral Leukocytosis Is Inversely Correlated with Intratumoral CD8+ T-Cell Infiltration and Associated with Worse Outcome after Chemoradiotherapy in Anal Cancer.
Peripheral blood leukocytosis has been implicated in promoting tumor progression leading to worse survival, but the mechanisms behind this phenomenon remain unexplored. Here, we examined the prognostic role of pretreatment white blood cell (WBC) count and clinicopathologic parameters in the context of CD8+ tumor-infiltrating lymphocytes (TIL) and myeloperoxidase+ tumor-associated neutrophils (TANs) in patients with anal squamous cell carcinoma (ASCC) treated with definitive chemoradiotherapy (CRT). After a median follow-up of 26 months, leukocytosis correlated with advanced T-stage (p < 0.001) and N-stage (p < 0.001), and predicted for worse distant-metastasis-free survival (p = 0.006), disease-free-survival (DFS, p = 0.029), and overall survival (p = 0.013). Importantly, leukocytosis was associated with a lower intraepithelial CD8+ TIL density (p = 0.014), whereas low CD8+ TIL expression in the intraepithelial compartment was associated with worse DFS (p = 0.028). Additionally, high TAN expression in the peritumoral compartment was associated with a significantly lower density of CD8+ TIL (p = 0.039), albeit, TAN expression lacked prognostic value. In conclusion, leukocytosis constitutes an important prognostic marker in ASCC patients treated with CRT. In conjunction with intratumoral TIL and TAN, these data provide for the first time important insight on the correlation of peripheral blood leukocytosis with the intratumoral immune contexture and could be relevant for future patient stratification using immunotherapies in ASCC.


A sequence variant at 4p16.3 confers susceptibility to urinary bladder cancer.
Previously, we reported germline DNA variants associated with risk of urinary bladder cancer (UBC) in Dutch and Icelandic subjects. Here we expanded the Icelandic sample set and tested the top 20 markers from the combined analysis in several European case-control sample sets, with a total of 4,739 cases and 45,549 controls. The T allele of rs798766 on 4p16.3 was found to associate with UBC (odds ratio = 1.24, P = 9.9 x 10(-12)). rs798766 is located in an intron of TACC3, 70 kb from FGFR3, which often harbors activating somatic mutations in low-grade, noninvasive UBC. Notably, rs798766[T] shows stronger association with low-grade and low-stage UBC than with more aggressive forms of the disease and is associated with higher risk of recurrence in low-grade stage Ta tumors. The frequency of rs798766[T] is higher in Ta tumors that carry an activating mutation in FGFR3 than in Ta tumors with wild-type FGFR3. Our results show a link between germline variants, somatic mutations of FGFR3 and risk of UBC.


EEVEE: the Empathy-Enhancing Virtual Evolving Environment.
Empathy is a multifaceted emotional and mental faculty that is often found to be affected in a great number of psychopathologies, such as schizophrenia, yet it remains very difficult to measure in an ecological context. The challenge stems partly from the complexity and fluidity of this social process, but also from its covert nature. One powerful tool to enhance experimental control over such dynamic social interactions has been the use of avatars in virtual reality (VR); information about an individual in such an interaction can be collected through the analysis of his or her neurophysiological and behavioral responses. We have developed a unique platform, the Empathy-Enhancing Virtual Evolving Environment (EEVEE), which is built around three main components: (1) different avatars capable of expressing feelings and emotions at various levels based on the Facial Action Coding System (FACS); (2) systems for measuring the physiological responses of the observer (heart and respiration rate, skin conductance, gaze and eye movements, facial expression); and (3) a multimodal interface linking the avatar's behavior to the observer's neurophysiological response. In this article, we provide a detailed description of the components of this innovative platform and validation data from the first phases of development. Our data show that healthy adults can discriminate different negative emotions, including pain, expressed by avatars at varying intensities. We also provide evidence that masking part of an avatar's face (top or bottom half) does not prevent the detection of different levels of pain. This innovative and flexible platform provides a unique tool to study and even modulate empathy in a comprehensive and ecological manner in various populations, notably individuals suffering from neurological or psychiatric disorders.


The impact of fissure depth and enamel conditioning protocols on glass-ionomer and resin-based fissure sealant penetration.
To evaluate the effect that fissure depth and enamel conditioning protocols have on penetration abilities of resin-based and glass-ionomer fissure sealants. Fifty extracted human molars were randomized into 10 experimental groups. Conditioning was performed as follows: groups I and VI, no pretreatment; groups II and VII, 10% polyacrylic acid and 37% phosphoric acid, respectively; groups III and VIII, self-etching adhesive; groups IV and IX, Er:YAG laser; groups V and X, Er:YAG laser/10% polyacrylic acid and laser/37% phosphoric acid, respectively. In groups I to V, teeth were sealed with a glass-ionomer material, and in groups VI to X a resin-based sealant was used. The sealed teeth were sectioned and the penetration ability was evaluated using scanning electron microscopy (SEM). Pretreatment with a suitable acid resulted in superior penetration of glass-ionomer and resin-based sealant when compared with other conditioning protocols (p < 0.0001), except for the teeth treated with both Er:YAG laser and acid. Both conditioning protocol and fissure depth significantly affected the sealant penetration (p < 0.0001), but fissure depth had a greater impact on fissure penetration than did conditioning protocol. Resin-based and glass-ionomer sealing materials show similar potential for pit-and-fissure penetration. Penetration of a fissure sealant is largely influenced by the fissure depth. Regarding the conditioning protocol, a pretreatment with a suitable acid is essential to obtain an adequate penetration of a sealing material.


Evolutionary aspects of emerging Lyme disease in Canada.
In North America, Lyme disease (LD) is a tick-borne zoonosis caused by the spirochete bacterium Borrelia burgdorferi sensu stricto, which is maintained by wildlife. Tick vectors and bacteria are currently spreading into Canada and causing increasing numbers of cases of LD in humans and raising a pressing need for public health responses. There is no vaccine, and LD prevention depends on knowing who is at risk and informing them how to protect themselves from infection. Recently, it was found in the United States that some strains of B. burgdorferi sensu stricto cause severe disease, whereas others cause mild, self-limiting disease. While many strains occurring in the United States also occur in Canada, strains in some parts of Canada are different from those in the United States. We therefore recognize a need to identify which strains specific to Canada can cause severe disease and to characterize their geographic distribution to determine which Canadians are particularly at risk. In this review, we summarize the history of emergence of LD in North America, our current knowledge of B. burgdorferi sensu stricto diversity, its intriguing origins in the ecology and evolution of the bacterium, and its importance for the epidemiology and clinical and laboratory diagnosis of LD. We propose methods for investigating associations between B. burgdorferi sensu stricto diversity, ecology, and pathogenicity and for developing predictive tools to guide public health interventions. We also highlight the emergence of B. burgdorferi sensu stricto in Canada as a unique opportunity for exploring the evolutionary aspects of tick-borne pathogen emergence.


Effect of golimumab on patient-reported outcomes in rheumatoid arthritis: results from the GO-FORWARD study.
To evaluate the effect of golimumab on physical function, general health, and fatigue in patients with active rheumatoid arthritis (RA) despite methotrexate (MTX) therapy. In the multicenter, randomized, placebo-controlled GO-FORWARD study, 444 adults with active RA despite MTX received subcutaneous placebo + MTX (crossover to golimumab 50 mg at Week 24), golimumab 100 mg + placebo, golimumab 50 mg + MTX, or golimumab 100 mg + MTX every 4 weeks. Physical function and general health were assessed using the Health Assessment Questionnaire-Disability Index (HAQ-DI) and Physical and Mental Component Summary (PCS, MCS) scores of the Medical Outcomes Study Short Form-36 questionnaire (SF-36), respectively, through Week 52. Fatigue was measured through Week 24 using the Functional Assessment of Chronic Illness Therapy-Fatigue (FACIT-Fatigue) questionnaire. Mean improvements from baseline in HAQ-DI, SF-36 PCS, and FACIT-Fatigue scores (Weeks 14 and 24) were significantly greater for golimumab 50 mg + MTX and 100 mg + MTX versus placebo + MTX. Significantly greater proportions of patients treated with golimumab + MTX achieved clinically meaningful improvements from baseline to Weeks 14 and 24 in HAQ-DI, PCS, and FACIT-Fatigue scores. Mean improvements in SF-36 PCS (Week 14), MCS (Week 24), and FACIT-Fatigue (Weeks 14 and 24) scores were significantly greater for golimumab 100 mg + placebo versus placebo + MTX. Mean improvements from baseline in HAQ-DI, SF-36 PCS, and MCS scores through Week 24 were sustained through Week 52. Patients with active RA despite MTX had significant improvement in physical function, general health, and fatigue following golimumab + MTX therapy; improvements in physical function and general health were maintained through Week 52. (Clinical Trials Registration NCT00264550).


Development and validation of a cross-cultural knowledge, attitudes, and practices survey instrument for chronic kidney disease in a Swahili-speaking population.
Non-communicable diseases are a growing global burden, and structured surveys can identify critical gaps to address this epidemic. In sub-Saharan Africa, there are very few well-tested survey instruments measuring population attributes related to non-communicable diseases. To meet this need, we have developed and validated the first instrument evaluating knowledge, attitudes and practices pertaining to chronic kidney disease in a Swahili-speaking population. Between December 2013 and June 2014, we conducted a four-stage, mixed-methods study among adults from the general population of northern Tanzania. In stage 1, the survey instrument was constructed in English by a group of cross-cultural experts from multiple disciplines and through content analysis of focus group discussions to ensure local significance. Following translation, in stage 2, we piloted the survey through cognitive and structured interviews, and in stage 3, in order to obtain initial evidence of reliability and construct validity, we recruited and then administered the instrument to a random sample of 606 adults. In stage 4, we conducted analyses to establish test-retest reliability and known-groups validity which was informed by thematic analysis of the qualitative data in stages 1 and 2. The final version consisted of 25 items divided into three conceptual domains: knowledge, attitudes and practices. Each item demonstrated excellent test-retest reliability with established content and construct validity. We have developed a reliable and valid cross-cultural survey instrument designed to measure knowledge, attitudes and practices of chronic kidney disease in a Swahili-speaking population of Northern Tanzania. This instrument may be valuable for addressing gaps in non-communicable diseases care by understanding preferences regarding healthcare, formulating educational initiatives, and directing development of chronic disease management programs that incorporate chronic kidney disease across sub-Saharan Africa.


Colonoscopy screening among US adults aged 40 or older with a family history of colorectal cancer.
Colonoscopy screening reduces colorectal cancer (CRC) incidence and mortality. CRC screening is recommended at age 50 for average-risk people. Screening of first-degree relatives of CRC patients is recommended to begin at age 40 or 10 years before the age at diagnosis of the youngest relative diagnosed with CRC. CRC incidence has increased recently among younger Americans while it has declined among older Americans. The objective of this study was to determine whether first-degree relatives of CRC patients are being screened according to recommended guidelines. We studied colonoscopy screening rates among the US population reporting a CRC family history using 2005 and 2010 National Health Interview Survey data. Of 26,064 study-eligible respondents, 2,470 reported a CRC family history; of those with a family history, 45.6% had a colonoscopy (25.2% in 2005 and 65.8% 2010). The colonoscopy rate among first-degree relatives aged 40 to 49 in 2010 (38.3%) was about half that of first-degree relatives aged 50 or older (69.7%). First-degree relatives were nearly twice as likely as nonfirst-degree relatives to have a colonoscopy (adjusted odds ratio [AOR], 1.7; 95% confidence interval, 1.5-1.9), but those aged 40 to 49 were less likely to have a colonoscopy than those in older age groups (AOR, 2.6 for age 50-64; AOR, 3.6 for age ≥65). Interactions with age, insurance, and race/ethnicity were not significant. Having health insurance tripled the likelihood of screening. Despite a 5-fold increase in colonoscopy screening rates since 2005, rates among first-degree relatives younger than the conventional screening age have lagged. Screening promotion targeted to this group may halt the recent rising trend of CRC among younger Americans.


Suppressing the Chills: Effects of Musical Manipulation on the Chills Response.
Research on musical chills has linked the response to multiple musical features; however, there exists no study that has attempted to manipulate musical stimuli to enable causal inferences, meaning current understanding is based mainly on correlational evidence. In the current study, participants who regularly experience chills (N = 24) listened to an original and manipulated version of three pieces reported to elicit chills in a previous survey. Predefined chills sections were removed to create manipulated conditions. The effects of these manipulations on the chills response were assessed through continuous self-reports, and skin conductance measurements. Results show that chills were significantly less frequent following stimulus manipulation across all three pieces. Continuous measurements of chills intensity were significantly higher in the chills sections compared with control sections in the pieces; similar patterns were found for phasic skin conductance, although some differences emerged. Continuous measurements also correlated with psychoacoustic features such as loudness, brightness and roughness in two of the three pieces. Findings are discussed in terms of understanding structural and acoustic features and chills experiences within their local music contexts, the necessity of experimental approaches to musical chills, and the possibility of different features activating different underlying mechanisms.


"Oncogenic shock": explaining oncogene addiction through differential signal attenuation.
"Oncogene addiction" describes the curious acquired dependence of tumor cells on an activated oncogene for their survival and/or proliferation, a phenomenon that has important implications for the success of targeted cancer therapies. However, the mechanisms explaining oncogene addiction remain elusive. We propose that "addiction" may be an illusion generated as a consequence of differential attenuation rates of prosurvival and proapoptotic signals emanating from an oncoprotein acutely following its inactivation. According to this model, which we call "oncogenic shock," prosurvival signals dissipate quickly on oncoprotein inactivation whereas proapoptotic signals linger sufficiently long to commit the cell to an apoptotic death. This mechanism may contribute to the rapid and dramatic clinical responses observed in some cancer patients treated with selective tyrosine kinase inhibitors and could yield additional drug targets that determine the balance of signaling outputs from activated oncoproteins.


Characteristics and circumstances of volatile solvent misuse-related death in Australia, 2000-2021.
Volatile solvent misuse-related death is associated with neuropsychiatric, cardiovascular, respiratory and renal pathology, as well as sudden death. The study aimed to determine: (1) the circumstances of death and case characteristics of volatile solvent misuse-related death in Australia, 2000-2021; (2) the toxicological profile of cases; and (3) the major autopsy findings. Retrospective study of volatile solvent misuse-related death in Australia, 2000-2021 retrieved from the National Coronial Information System. One hundred and sixty-four cases were identified, 79.9% male, mean age 26.5 years (8.5% aged 40 years or older). Circumstances of death were unintentional toxicity (61.0%), unintentional asphyxia (20.1%), intentional self-harm (12.2%) and traumatic accident (6.7%). The most commonly reported acute presentation prior to death was sudden collapse (22 of 47 witnessed events). The most frequently used solvents at the fatal incident were gas fuels (35.4%), gasoline (petrol) (19.5%) adhesives/paints (19.5%), aerosol propellants (12.8%), and volatile anaesthetics (12.8%). The most commonly detected volatile substances were butane (40.7%), toluene (29.6%), and propane (25.9%). Cannabis was present in 27.6% and alcohol in 24.6%. The prevalence of acute pneumonia amongst autopsied cases was low (5.8%) which, together with reports of sudden collapse, suggests that in many cases, death was extremely rapid. There were low levels of major organ pathology. While the average age of volatile solvent misuse-related death was in the mid-twenties, a substantial proportion occurred amongst people aged 40 years or older. Reflecting availability, gas fuels predominated. In many cases, death appeared to have been rapid.


Unravelling the enigma of Perthes disease.
Perthes disease is an idiopathic avascular necrosis of a juvenile hip. Although 2010 marked a century since it was first described, the aetiology remains unknown. It is suggested that adverse socioeconomic circumstances may be a key precipitant. This work describes recent studies that explore the disease epidemiology. Descriptive studies include a case register from Merseyside, hospital discharge data from Scotland, analysis of the world's largest community disease register (General Practice Research Database [GPRD]) and a systematic review of incidence. Analytical studies include a nested case-controlled study in the GPRD and a hospital case-controlled study. The studies demonstrated a striking north-south divide in the UK incidence of Perthes disease, similar to that seen in many adult diseases. There was a sustained fall in disease frequency in all studies, with a narrowing of the north-south divide. There was a strong association with area deprivation, independent of living in an urban environment. Internationally, equatorial regions were unaffected by disease and northern Europe had the highest incidence, which was primarily a function of race although latitude was an independent predictor. Individual characteristics associated with the disease were congenital anomalies of the genitourinary tract and a structural abnormality of arterial calibre. Despite a falling incidence, Perthes disease remains an important cause of child morbidity and exemplifies socioeconomic inequalities. A deprivation-related exposure, acting early in development, appears critical. The aetiological factor in Perthes disease remains elusive but it is likely that unravelling this enigma may unlock additional secrets pertaining to the developmental origins of this and other diseases.


Knee stability following anterior cruciate ligament rupture and surgery. The contribution of irreducible tibial subluxation.
Knee stability after anterior cruciate ligament reconstruction is generally determined by measuring total anteroposterior tibial motion. In spite of a decrease in excessive anteroposterior tibial motion after anterior cruciate ligament reconstruction, problems can still develop. In the present study, we sought to define the tibiofemoral relationship more accurately with use of stress radiographs of human knees after anterior cruciate ligament rupture and after anterior cruciate ligament reconstruction. A previously described radiographic technique was used to evaluate the position of the tibia relative to the femur with the application of an anteriorly directed tibial force and subsequently with the application of a posteriorly directed tibial force. Tibial position and total tibial translation were calculated from these radiographs. In addition, KT-1000 measurements were obtained. Three groups of patients were studied: Group 1 included twenty-eight patients with an untreated anterior cruciate ligament rupture, Group 2 included nineteen patients who had undergone a clinically successful anterior cruciate ligament reconstruction, and Group 3 included twenty-five control subjects with normal knees. KT-1000 testing showed that the average side-to-side differences in Group 1 (5.8 mm) and Group 2 (2.7 mm) were significantly different from that in Group 3 (0.8 mm) (p < 0.01 and p < 0.05, respectively). Stress radiographs showed that the average total tibial translation in Group 1 (9.8 mm) was significantly different from those in Group 2 (5.6 mm) and Group 3 (4.3 mm) (p < 0.05 and p < 0.001, respectively). Within Group 1, knees with radiographic signs of osteoarthritis were more stable, with an average total tibial excursion of 6.8 mm. The improved stability of the reconstructed knees in Group 2 and the osteoarthritic knees in Group 1 was not entirely the result of decreased anterior tibial translation; it was, in part, due to an irreducible anterior subluxation of the tibia. A posteriorly directed stress in these knees did not reduce the tibia to the anatomic position relative to the femur; the osteoarthritic knees in Group 1 were 9.9 mm short of full reduction and the knees in Group 2 were 3.1 mm short of full reduction (p < 0.01) Irreducible tibial subluxation can be present in the knee following surgical reconstruction of the anterior cruciate ligament. Osteoarthritic changes following an untreated anterior cruciate ligament rupture are also associated with uncorrectable tibial subluxation along with a decrease in instability. The irreducible tibial subluxation could explain why osteoarthritic changes still may develop in stable, reconstructed knees in spite of the improved stability. Currently used arthrometric measurements, such as KT-1000 scores, do not measure this phenomenon.


Effects of cypermethrin on some haematological, biochemical and histopathological parameters of common carp (Cyprinus carpio L.).
The study aimed at assessing the effects of a pyrethroid cypermethrin on common carp (Cyprinus carpio L.) on the basis of the results of acute toxicity tests and haematological, biochemical and histological examinations. Experimental carp were exposed to the pesticide preparation Alimetrin 10 EM in the concentration of 29.1 microg/l(-1) (96hLC(50)) corresponding to 29.1 microg/l(-1) of cypermethrin in a 96 h acute toxicity test. The results of the test and of the examinations of exposed carp were compared with the results of carp from the control group. In haematological profile, the experimental group of carp showed a significant (P<0.01) increase in the number of erythrocytes, segmented neutrophile granulocytes, developmental forms of myeloid sequence and eosinophiles, and a significant (P<0.01) decrease in mean erythrocyte volume, mean erythrocyte haemoglobin and lymphocyte count. As far as biochemical indices are concerned, the exposure caused a significant (P<0.01) increase in glucose and lactate concentration and creatine kinase activity, and a significant (P<0.01) decrease in total proteins, albumins, total globulins, ammonia, lactate dehydrogenase and alkaline phosphatase in blood plasma of exposed carp compared to the control group. Histopathological tissue examinations proved random hyperaemia and perivascular lymphocyte infiltration in skin, respiratory epithelium hyperplasia and chloride cell activation in gills, and pancreas exocrine cell vacuolisation in common carp after the exposure. The exposure of common carp to 29.1 microg/l(-1) of Alimetrin 10 EM caused significant shifts in haematological and biochemical profile and histopathological changes in organ tissues. The cypermethrin-based pesticide Alimetrin 10 EM was classified among substances strongly toxic for fish.


Analysis of the native conformation of the LIR/AIM motif in the Atg8/LC3/GABARAP-binding proteins.
The Atg8/LC3/GABARAP family of proteins, a group that has structural homology with ubiquitin, connects with a large set of binding partners to function in macroautophagy (hereafter autophagy). This interaction occurs primarily via a conserved motif termed the LC3-interacting region (LIR), or the Atg8-interacting motif (AIM). The consensus sequence for this motif, [W/F/Y]xx[L/I/V], can be found in many proteins, but only some of them are physiological partners containing a functional LIR/AIM. Because the structure of many full-length partners has not been, or cannot be, solved, the structural context of the LIR/AIM within the native protein conformation is not obvious. Here we suggest that the functional LIR/AIM is a short linear motif (SLiM) protein-binding module, arising from an intrinsically disordered region. This finding enables the rapid elimination of some false Atg8/LC3/GABARAP-binding proteins, and connects the exponentially growing knowledge on disordered SLiMs with autophagy.


Non-Toxic Dimeric Peptides Derived from the Bothropstoxin-I Are Potent SARS-CoV-2 and Papain-like Protease Inhibitors.
The COVID-19 outbreak has rapidly spread on a global scale, affecting the economy and public health systems throughout the world. In recent years, peptide-based therapeutics have been widely studied and developed to treat infectious diseases, including viral infections. Herein, the antiviral effects of the lysine linked dimer des-Cys11, Lys12,Lys13-(pBthTX-I)2K ((pBthTX-I)2K)) and derivatives against SARS-CoV-2 are reported. The lead peptide (pBthTX-I)2K and derivatives showed attractive inhibitory activities against SARS-CoV-2 (EC50 = 28-65 µM) and mostly low cytotoxic effect (CC50 > 100 µM). To shed light on the mechanism of action underlying the peptides' antiviral activity, the Main Protease (Mpro) and Papain-Like protease (PLpro) inhibitory activities of the peptides were assessed. The synthetic peptides showed PLpro inhibition potencies (IC50s = 1.0-3.5 µM) and binding affinities (Kd = 0.9-7 µM) at the low micromolar range but poor inhibitory activity against Mpro (IC50 > 10 µM). The modeled binding mode of a representative peptide of the series indicated that the compound blocked the entry of the PLpro substrate toward the protease catalytic cleft. Our findings indicated that non-toxic dimeric peptides derived from the Bothropstoxin-I have attractive cellular and enzymatic inhibitory activities, thereby suggesting that they are promising prototypes for the discovery and development of new drugs against SARS-CoV-2 infection.


Mindfulness: a systematic review of instruments to measure an emergent patient-reported outcome (PRO).
Mindfulness has emerged as an important health concept based on evidence that mindfulness interventions reduce symptoms and improve health-related quality of life. The objectives of this study were to systematically assess and compare the properties of instruments to measure self-reported mindfulness. Ovid Medline(®), CINAHL(®), and PsycINFO(®) were searched through May 2012, and articles were selected if their primary purpose was development or evaluation of the measurement properties (validity, reliability, responsiveness) of a self-report mindfulness scale. Two reviewers independently evaluated the methodological quality of the selected studies using the COnsensus-based Standards for the selection of health status Measurement INstruments checklist. Discrepancies were discussed with a third reviewer and scored by consensus. Finally, a level of evidence approach was used to synthesize the results and study quality. Our search strategy identified a total of 2,588 articles. Forty-six articles, reporting 79 unique studies, met inclusion criteria. Ten instruments quantifying mindfulness as a unidimensional scale (n = 5) or as a set of 2-5 subscales (n = 5) were reviewed. The Mindful Attention Awareness Scale was evaluated by the most studies (n = 27) and had positive overall quality ratings for most of the psychometric properties reviewed. The Five Facet Mindfulness Questionnaire received the highest possible rating ("consistent findings in multiple studies of good methodological quality") for two properties, internal consistency and construct validation by hypothesis testing. However, none of the instruments had sufficient evidence of content validity. Comprehensiveness of construct coverage had not been assessed; qualitative methods to confirm understanding and relevance were absent. In addition, estimates of test-retest reliability, responsiveness, or measurement error to guide users in protocol development or interpretation of scores were lacking. Current mindfulness scales have important conceptual differences, and none can be strongly recommended based solely on superior psychometric properties. Important limitations in the field are the absence of qualitative evaluations and accepted external referents to support construct validity. Investigators need to proceed cautiously before optimizing any mindfulness intervention based on the existing scales.


Expression, purification and characterization of human proton-coupled oligopeptide transporter 1 hPEPT1.
The human peptide transporter hPEPT1 (SLC15A1) is responsible for uptake of dietary di- and tripeptides and a number of drugs from the small intestine by utilizing the proton electrochemical gradient, and hence an important target for peptide-like drug design and drug delivery. hPEPT1 belongs to the ubiquitous major facilitator superfamily that all contain a 12TM core structure, with global conformational changes occurring during the transport cycle. Several bacterial homologues of these transporters have been characterized, providing valuable insight into the transport mechanism of this family. Here we report the overexpression and purification of recombinant hPEPT1 in a detergent-solubilized state. Thermostability profiling of hPEPT1 at different pH values revealed that hPEPT1 is more stable at pH 6 as compared to pH 7 and 8. Micro-scale thermophoresis (MST) confirmed that the purified hPEPT1 was able to bind di- and tripeptides respectively. To assess the in-solution oligomeric state of hPEPT1, negative stain electron microscopy was performed, demonstrating a predominantly monomeric state.


Twist1-Induced Epithelial Dissemination Requires Prkd1 Signaling.
Dissemination is an essential early step in metastasis but its molecular basis remains incompletely understood. To define the essential targetable effectors of this process, we developed a 3D mammary epithelial culture model, in which dissemination is induced by overexpression of the transcription factor Twist1. Transcriptomic analysis and ChIP-PCR together demonstrated that protein kinase D1 (Prkd1) is a direct transcriptional target of Twist1 and is not expressed in the normal mammary epithelium. Pharmacologic and genetic inhibition of Prkd1 in the Twist1-induced dissemination model demonstrated that Prkd1 was required for cells to initiate extracellular matrix (ECM)-directed protrusions, release from the epithelium, and migrate through the ECM. Antibody-based protein profiling revealed that Prkd1 induced broad phosphorylation changes, including an inactivating phosphorylation of β-catenin and two microtubule depolymerizing phosphorylations of Tau, potentially explaining the release of cell-cell contacts and persistent activation of Prkd1. In patients with breast cancer, TWIST1 and PRKD1 expression correlated with metastatic recurrence, particularly in basal breast cancer. Prkd1 knockdown was sufficient to block dissemination of both murine and human mammary tumor organoids. Finally, Prkd1 knockdown in vivo blocked primary tumor invasion and distant metastasis in a mouse model of basal breast cancer. Collectively, these data identify Prkd1 as a novel and targetable signaling node downstream of Twist1 that is required for epithelial invasion and dissemination. SIGNIFICANCE: Twist1 is a known regulator of metastatic cell behaviors but not directly targetable. This study provides a molecular explanation for how Twist1-induced dissemination works and demonstrates that it can be targeted. GRAPHICAL ABSTRACT: http://cancerres.aacrjournals.org/content/canres/80/2/204/F1.large.jpg.


Poverty, AIDS and child health: identifying highest-risk children in South Africa.
Identifying children at the highest risk of negative health effects is a prerequisite to effective public health policies in Southern Africa. A central ongoing debate is whether poverty, orphanhood or parental AIDS most reliably indicates child health risks. Attempts to address this key question have been constrained by a lack of data allowing distinction of AIDS-specific parental death or morbidity from other causes of orphanhood and chronic illness. To examine whether household poverty, orphanhood and parental illness (by AIDS or other causes) independently or interactively predict child health, developmental and HIV-infection risks. We interviewed 6 002 children aged 10 - 17 years in 2009 - 2011, using stratified random sampling in six urban and rural sites across three South African provinces. Outcomes were child mental health risks, educational risks and HIV-infection risks. Regression models that controlled for socio-demographic co-factors tested potential impacts and interactions of poverty, AIDS-specific and other orphanhood and parental illness status. Household poverty independently predicted child mental health and educational risks, AIDS orphanhood independently predicted mental health risks and parental AIDS illness independently predicted mental health, educational and HIV-infection risks. Interaction effects of poverty with AIDS orphanhood and parental AIDS illness were found across all outcomes. No effects, or interactions with poverty, were shown by AIDS-unrelated orphanhood or parental illness. The identification of children at highest risk requires recognition and measurement of both poverty and parental AIDS. This study shows negative impacts of poverty and AIDS-specific vulnerabilities distinct from orphanhood and adult illness more generally. Additionally, effects of interaction between family AIDS and poverty suggest that, where these co-exist, children are at highest risk of all.


Automated left atrial time-resolved segmentation in MRI long-axis cine images using active contours.
Segmentation of the left atrium (LA) is required to evaluate atrial size and function, which are important imaging biomarkers for a wide range of cardiovascular conditions, such as atrial fibrillation, stroke, and diastolic dysfunction. LA segmentations are currently being performed manually, which is time-consuming and observer-dependent. This study presents an automated image processing algorithm for time-resolved LA segmentation in cardiac magnetic resonance imaging (MRI) long-axis cine images of the 2-chamber (2ch) and 4-chamber (4ch) views using active contours. The proposed algorithm combines mitral valve tracking, automated threshold calculation, edge detection on a radially resampled image, edge tracking based on Dijkstra's algorithm, and post-processing involving smoothing and interpolation. The algorithm was evaluated in 37 patients diagnosed mainly with paroxysmal atrial fibrillation. Segmentation accuracy was assessed using the Dice similarity coefficient (DSC) and Hausdorff distance (HD), with manual segmentations in all time frames as the reference standard. For inter-observer variability analysis, a second observer performed manual segmentations at end-diastole and end-systole on all subjects. The proposed automated method achieved high performance in segmenting the LA in long-axis cine sequences, with a DSC of 0.96 for 2ch and 0.95 for 4ch, and an HD of 5.5 mm for 2ch and 6.4 mm for 4ch. The manual inter-observer variability analysis had an average DSC of 0.95 and an average HD of 4.9 mm. The proposed automated method achieved performance on par with human experts analyzing MRI images for evaluation of atrial size and function. Video Abstract.


Requirement of ErbB2 for signalling by interleukin-6 in prostate carcinoma cells.
Interleukin-6 (IL-6) is a cytokine that was initially recognized as a regulator of immune and inflammatory responses, but it also regulates the growth of many tumour cells, including prostrate carcinoma. Overexpression of the growth-factor receptors ErbB2/neu and ErbB3 has been implicated in the neoplastic transformation of prostate carcinoma. Here we show that treatment of the prostate cancer cell line LNCaP with IL-6 induces tyrosine phosphorylation of ErbB2 and ErbB3, but not ErbB1/EGFR. We also show that ErbB2 forms a complex with the gp130 subunit of the IL-6 receptor in an IL-6-dependent manner. This association is important because the inhibition of ErbB2 activity results in abrogation of IL-6-induced MAPK activation. Thus ErbB2 is a critical component of IL-6 signalling through the MAP kinase pathway. These data show how a cytokine receptor can diversify its signalling pathways by engaging with a growth-factor receptor kinase.


Type 2 Diabetes Mellitus Trajectories and Associated Risks.
Disease progression models, statistical models that assess a patient's risk of diabetes progression, are popular tools in clinical practice for prevention and management of chronic conditions. Most, if not all, models currently in use are based on gold standard clinical trial data. The relatively small sample size available from clinical trial limits these models only considering the patient's state at the time of the assessment and ignoring the trajectory, the sequence of events, that led up to the state. Recent advances in the adoption of electronic health record (EHR) systems and the large sample size they contain have paved the way to build disease progression models that can take trajectories into account, leading to increasingly accurate and personalized assessment. To address these problems, we present a novel method to observe trajectories directly. We demonstrate the effectiveness of the proposed method by studying type 2 diabetes mellitus (T2DM) trajectories. Specifically, using EHR data for a large population-based cohort, we identified a typical trajectory that most people follow, which is a sequence of diseases from hyperlipidemia (HLD) to hypertension (HTN), impaired fasting glucose (IFG), and T2DM. In addition, we also show that patients who follow different trajectories can face significantly increased or decreased risk.


Race as a physiosocial phenomenon.
This paper offers both a criticism of and a novel alternative perspective on current ontologies that take race to be something that is either static and wholly evident at one's birth or preformed prior to it. In it I survey and critically assess six of the most popular conceptions of race, concluding with an outline of my own suggestion for an alternative account. I suggest that race can be best understood in terms of one's experience of his or her body, one's interactions with other individuals, and one's experiences within particular cultures and societies. This embeddedness of human experience has been left out of most discussions of race which tie race to a set of characteristics (either biologically or sociologically defined). To rectify this omission, I articulate what I call the "physiosocial" view of race. This emphasizes the situatedness of human experience, the reciprocal and dynamic nature of the racial identities of individuals and groups. Approaching racial identity in this way entails a union of two historically uncomfortable partners: biological and sociological conceptions of race. If successful, this philosophical stance may illuminate the process of racial self-ascription as well as provide an explanation for the potential changeability of an individual's racial identity at different times and at different places.


AMDA: an R package for the automated microarray data analysis.
Microarrays are routinely used to assess mRNA transcript levels on a genome-wide scale. Large amount of microarray datasets are now available in several databases, and new experiments are constantly being performed. In spite of this fact, few and limited tools exist for quickly and easily analyzing the results. Microarray analysis can be challenging for researchers without the necessary training and it can be time-consuming for service providers with many users. To address these problems we have developed an automated microarray data analysis (AMDA) software, which provides scientists with an easy and integrated system for the analysis of Affymetrix microarray experiments. AMDA is free and it is available as an R package. It is based on the Bioconductor project that provides a number of powerful bioinformatics and microarray analysis tools. This automated pipeline integrates different functions available in the R and Bioconductor projects with newly developed functions. AMDA covers all of the steps, performing a full data analysis, including image analysis, quality controls, normalization, selection of differentially expressed genes, clustering, correspondence analysis and functional evaluation. Finally a LaTEX document is dynamically generated depending on the performed analysis steps. The generated report contains comments and analysis results as well as the references to several files for a deeper investigation. AMDA is freely available as an R package under the GPL license. The package as well as an example analysis report can be downloaded in the Services/Bioinformatics section of the Genopolis http://www.genopolis.it/.


A suburban focus of endemic typhus in Los Angeles County: association with seropositive domestic cats and opossums.
Thirty-three cases of locally acquired murine typhus were reported in Los Angeles County residents from May 1984 through February 1988. Only eight cases were reported over the previous 20-year period. Thirty (91%) cases resided within a suburban area encompassing approximately 50 km2 in northcentral Los Angeles or had contact with an animal from this area. Serologic testing (complement fixation and indirect fluorescent antibody) of selected animals in close association with human cases revealed a high prevalence of seropositivity among domestic cats and opossums. Nine (90%) of 10 resident cats tested had demonstrable antibody titers compared with none (0%) of 20 cats from a control area (P < 0.001). Suburban typhus cases were more likely than neighborhood controls to own a cat or dog (odds ratio = 6.9, 95% confidence interval = 1.8, 25.9, P = 0.002). Sixteen (42%) of 38 opossums trapped in close proximity to the residences of cases were seropositive versus none (0%) of 36 opossums from control areas (P < 0.001) A low frequency (2.8%) of seropositivity was found in commensal rodents, and the classic vector of murine typhus, Xenopsylla cheopis, was not found. Ectoparasite indices form seropositive opossums revealed heavy infestations with the cat flea, Ctenocephalides felis (mean flea count = 104.7), a species that readily bites humans. These data provide evidence that a suburban focus of murine typhus exists in Los Angeles that differs substantially from the classic transmission cycle, and that cats, opossums and C. felis may play an important role in the occurrence of human cases.


Chronic fatigue syndrome is associated with diminished intracellular perforin.
Chronic fatigue syndrome (CFS) is an illness characterized by unexplained and prolonged fatigue that is often accompanied by abnormalities of immune, endocrine and cognitive functions. Diminished natural killer cell cytotoxicity (NKCC) is a frequently reported finding. However, the molecular basis of this defect of in vitro cytotoxicy has not been described. Perforin is a protein found within intracellular granules of NK and cytotoxic T cells and is a key factor in the lytic processes mediated by these cells. Quantitative fluorescence flow cytometry was used to the intracellular perforin content in CFS subjects and healthy controls. A significant reduction in the NK cell associated perforin levels in samples from CFS patients, compared to healthy controls, was observed. There was also an indication of a reduced perforin level within the cytotoxic T cells of CFS subjects, providing the first evidence, to our knowledge, to suggest a T cell associated cytotoxic deficit in CFS. Because perforin is important in immune surveillance and homeostasis of the immune system, its deficiency may prove to be an important factor in the pathogenesis of CFS and its analysis may prove useful as a biomarker in the study of CFS.


Little evidence for an epidemic of myopia in Australian primary school children over the last 30 years.
Recently reported prevalences of myopia in primary school children vary greatly in different regions of the world. This study aimed to estimate the prevalence of refractive errors in an unselected urban population of young primary school children in eastern Sydney, Australia, between 1998 and 2004, for comparison with our previously published data gathered using the same protocols and other Australian studies over the last 30 years. Right eye refractive data from non-cycloplegic retinoscopy was analysed for 1,936 children aged 4 to 12 years who underwent a full eye examination whilst on a vision science excursion to the Vision Education Centre Clinic at the University of New South Wales. Myopia was defined as spherical equivalents equal to or less than -0.50 D, and hyperopia as spherical equivalents greater than +0.50 D. The mean spherical equivalent decreased significantly (p < 0.0001) with age from +0.73 +/- 0.1D (SE) at age 4 to +0.21 +/- 0.11D at age 12 years. The proportion of children across all ages with myopia of -0.50D or more was 8.4%, ranging from 2.3% of 4 year olds to 14.7% of 12 year olds. Hyperopia greater than +0.50D was present in 38.4%. A 3-way ANOVA for cohort, age and gender of both the current and our previous data showed a significant main effect for age (p < 0.0001) but not for cohort (p = 0.134) or gender (p = 0.61). Comparison of our new data with our early 1990s data and that from studies of over 8,000 Australian non-clinical rural and urban children in the 1970's and 1980's provided no evidence for the rapidly increasing prevalence of myopia described elsewhere in the world. In fact, the prevalence of myopia in Australian children continues to be significantly lower than that reported in Asia and North America despite changing demographics. This raises the issue of whether these results are a reflection of Australia's stable educational system and lifestyle over the last 30 years.


Genome-wide examination of myoblast cell cycle withdrawal during differentiation.
Skeletal and cardiac myocytes cease division within weeks of birth. Although skeletal muscle retains limited capacity for regeneration through recruitment of satellite cells, resident populations of adult myocardial stem cells have not been identified. Because cell cycle withdrawal accompanies myocyte differentiation, we hypothesized that C2C12 cells, a mouse myoblast cell line previously used to characterize myocyte differentiation, also would provide a model for studying cell cycle withdrawal during differentiation. C2C12 cells were differentiated in culture medium containing horse serum and harvested at various time points to characterize the expression profiles of known cell cycle and myogenic regulatory factors by immunoblot analysis. BrdU incorporation decreased dramatically in confluent cultures 48 hr after addition of horse serum, as cells started to form myotubes. This finding was preceded by up-regulation of MyoD, followed by myogenin, and activation of Bcl-2. Cyclin D1 was expressed in proliferating cultures and became undetectable in cultures containing 40% fused myotubes, as levels of p21(WAF1/Cip1) increased and alpha-actin became detectable. Because C2C12 myoblasts withdraw from the cell cycle during myocyte differentiation following a course that recapitulates this process in vivo, we performed a genome-wide screen to identify other gene products involved in this process. Using microarrays containing approximately 10,000 minimally redundant mouse sequences that map to the UniGene database of the National Center for Biotechnology Information, we compared gene expression profiles between proliferating, differentiating, and differentiated C2C12 cells and verified candidate genes demonstrating differential expression by RT-PCR. Cluster analysis of differentially expressed genes revealed groups of gene products involved in cell cycle withdrawal, muscle differentiation, and apoptosis. In addition, we identified several genes, including DDAH2 and Ly-6A, whose expression specifically was up-regulated during cell cycle withdrawal coincident with early myoblast differentiation.


Diagnosis of Hepatopulmonary Syndrome in a Large Integrated Health System.
Data on the accuracy of the diagnosis of hepatopulmonary syndrome (HPS) in cirrhosis is limited. We evaluated the clinical characteristics of patients with International Classification of Diseases (ICD) codes for hepatopulmonary syndrome (HPS) in a large integrated health system. A retrospective review of encounters was performed of all patients with ICD-9-CM and/or ICD-10-CM diagnosis of cirrhosis and HPS from 2014-2019 in a multi-state health system. Demographics and cardiopulmonary testing closest to the time of HPS diagnosis were recorded. HPS was defined using standard criteria. A total of 42,749 unique individuals with cirrhosis were identified. An ICD diagnosis of HPS was found in 194 patients (0.45%), of which 182 had clinically confirmed cirrhosis. 143 (78.5%) underwent contrast-enhanced transthoracic echocardiography, and 98 (54%) had delayed shunting. Among them, 61 patients had a documented arterial blood gas, with 53 showing abnormal oxygenation (A-a gradient of >15 mm Hg). 12 were excluded due to significant pulmonary function test abnormalities and abnormal oxygenation from other cardiopulmonary diseases. Ultimately, 41 (22.5%) fulfilled the criteria for HPS. When stratifying those with an ICD code diagnosis of HPS into HPS, no HPS and indeterminate HPS groups, based on standard diagnostic criteria for HPS, we found that the confirmed HPS patients had similar complications except for less portopulmonary hypertension, worse gas exchange, less cardiopulmonary disease and were more often diagnosed in transplant centers. The diagnosis of HPS by ICD code is made in an extremely small subset of a sizeable cirrhotic cohort. When made, only a minority of these patients meet diagnostic criteria. Our findings highlight the need for improved education and more effective screening algorithms.


Prognostic impact of activated B-cell focused classification in diffuse large B-cell lymphoma patients treated with R-CHOP.
Gene expression profiling studies initially enabled diffuse large B-cell lymphoma to be divided into germinal center and activated B-cell-like subtypes, which define high- and low-risk patient groups when treated with chemotherapy. Attempts to reproduce the prognostic classification immunohistochemically have, however, provided inconsistent results. The aim of this study was to determine whether modified immunohistochemical classification of cell of origin focusing on activated B-cell-like markers could be used to predict the outcome of immunochemotherapy-treated diffuse large B-cell lymphoma patients. The expression of CD10, Bcl-6, MUM1/IRF4, Bcl-2, and FOXP1 was determined immunohistochemically from 88 samples of diffuse large B-cell lymphoma patients treated uniformly with R-CHOP. When the modified classification using MUM1/IRF4 and FOXP1 positivities as activated B-cell-like markers was applied to distinguish the patients between the activated B-cell-like and other diffuse large B-cell lymphoma subtypes, a significantly worse outcome was seen for the patients with the activated B-cell-like phenotype (3-year failure-free survival 63 vs 82%, P=0.048, overall survival 69 vs 85%, P=0.110). Similarly, according to the Muris algorithm, the group 2 patients representing Bcl-2-positive post-germinal center patients showed an inferior outcome in comparison to the group 1 patients (failure-free survival 59 vs 81%, P=0.041, overall survival 67 vs 82%, P=0.159). In contrast, when the classification of the same cohort was performed according to the Hans algorithm, no significant difference in survival was observed between the germinal center and non-germinal center patients. In conclusion, the data suggest that both the modified activated B-cell-like and Muris classifications define the non-germinal center phenotype as an adverse risk factor in R-CHOP-treated diffuse large B-cell lymphoma patients.


Remarkable improvement in detection of readthrough downstream-of-gene transcripts by semi-extractable RNA-sequencing.
The mammalian cell nucleus contains dozens of membrane-less nuclear bodies that play significant roles in various aspects of gene expression. Several nuclear bodies are nucleated by specific architectural noncoding RNAs (arcRNAs) acting as structural scaffolds. We have reported that a minor population of cellular RNAs exhibits an unusual semi-extractable feature upon using the conventional procedure of RNA preparation and that needle shearing or heating of cell lysates remarkably improves extraction of dozens of RNAs. Because semi-extractable RNAs, including known arcRNAs, commonly localize in nuclear bodies, this feature may be a hallmark of arcRNAs. Using the semi-extractability of RNA, we performed genome-wide screening of semi-extractable long noncoding RNAs to identify new candidate arcRNAs for arcRNA under hyperosmotic and heat stress conditions. After screening stress-inducible and semi-extractable RNAs, hundreds of readthrough downstream-of-gene (DoG) transcripts over several hundreds of kilobases, many of which were not detected among RNAs prepared by the conventional extraction procedure, were found to be stress-inducible and semi-extractable. We further characterized some of the abundant DoGs and found that stress-inducible transient extension of the 3'-UTR made DoGs semi-extractable. Furthermore, they were localized in distinct nuclear foci that were sensitive to 1,6-hexanediol. These data suggest that semi-extractable DoGs exhibit arcRNA-like features and our semi-extractable RNA-seq is a powerful tool to extensively monitor DoGs that are induced under specific physiological conditions.


Automatic generation of reaction energy databases from highly accurate atomization energy benchmark sets.
In this contribution, we discuss how reaction energy benchmark sets can automatically be created from arbitrary atomization energy databases. As an example, over 11 000 reaction energies derived from the W4-11 database, as well as some relevant subsets are reported. Importantly, there is only very modest computational overhead involved in computing >11 000 reaction energies compared to 140 atomization energies, since the rate-determining step for either benchmark is performing the same 140 quantum chemical calculations. The performance of commonly used electronic structure methods for the new database is analyzed. This allows investigating the relationship between the performances for atomization and reaction energy benchmarks based on an identical set of molecules. The atomization energy is found to be a weak predictor for the overall usefulness of a method. The performance of density functional approximations in light of the number of empirically optimized parameters used in their design is also discussed.