Evolution via recombination: Cell-to-cell contact facilitates larger recombination events in Streptococcus pneumoniae.
Homologous recombination in the genetic transformation model organism Streptococcus pneumoniae is thought to be important in the adaptation and evolution of this pathogen. While competent pneumococci are able to scavenge DNA added to laboratory cultures, large-scale transfers of multiple kb are rare under these conditions. We used whole genome sequencing (WGS) to map transfers in recombinants arising from contact of competent cells with non-competent 'target' cells, using strains with known genomes, distinguished by a total of ~16,000 SNPs. Experiments designed to explore the effect of environment on large scale recombination events used saturating purified donor DNA, short-term cell assemblages on Millipore filters, and mature biofilm mixed cultures. WGS of 22 recombinants for each environment mapped all SNPs that were identical between the recombinant and the donor but not the recipient. The mean recombination event size was found to be significantly larger in cell-to-cell contact cultures (4051 bp in filter assemblage and 3938 bp in biofilm co-culture versus 1815 bp with saturating DNA). Up to 5.8% of the genome was transferred, through 20 recombination events, to a single recipient, with the largest single event incorporating 29,971 bp. We also found that some recombination events are clustered, that these clusters are more likely to occur in cell-to-cell contact environments, and that they cause significantly increased linkage of genes as far apart as 60,000 bp. We conclude that pneumococcal evolution through homologous recombination is more likely to occur on a larger scale in environments that permit cell-to-cell contact.


Interactions Among Vitamin D, Atrial Fibrillation, and the Renin-Angiotensin-Aldosterone System.
Blockade of the renin-angiotensin-aldosterone system (RAAS) with angiotensin-converting enzyme inhibitors (ACEIs) and angiotensin receptor blockers (ARB) has been shown to decrease incident atrial fibrillation (AF). Vitamin D deficiency has been implicated but associations between vitamin D and AF are inconsistent. However, vitamin D deficiency results in renin and angiotensin upregulation. This study seeks to determine the relation between vitamin D deficiency and incident AF and characterize this relation in the setting of RAAS inhibition. A total of 47,062 patients with 25-hydroxyvitamin D [25(OH)D] laboratory testing were retrospectively identified. Clinical information was extracted from the medical record. The primary outcome was incidence of AF. Given significant patient variation, propensity score matching was used to select comparable groups of those who initiated ACEI/ARB ("AI") versus those who did not ("NAI"). Odds ratios (ORs) for incident AF associated with ACEI/ARB initiation and 25(OH)D level were estimated using generalized linear mixed models. AI patients had less incident AF than NAI patients (OR 0.36, 95% confidence interval [CI] 0.32 to 0.39) consistent with previous studies, but 25(OH)D deficiency was not associated with incident AF (OR 1.08, 95% CI 0.95 to 1.22). When stratified by 25(OH)D, there was a statistically significant decreased rate of AF in AI patients. Interestingly, this benefit was attenuated in 25(OH)D deficiency (deficient: OR 0.48, 95% CI 0.38 to 0.60 vs normal: OR 0.34, 95% CI 0.30 to 0.37). In conclusion, 25(OH)D deficiency was not associated with incident AF, and the benefit of ACE/ARB use was attenuated in 25(OH)D deficiency; this suggests that 25(OH)D may act as a cofactor in the mitigation of AF by RAAS inhibition.


Synthesis of 2,6-trans- and 3,3,6-trisubstituted tetrahydropyran-4-ones from Maitland-Japp derived 2H-dihydropyran-4-ones: a total synthesis of diospongin B.
6-Substituted-2H-dihydropyran-4-one products of the Maitland-Japp reaction have been converted into tetrahydropyrans containing uncommon substitution patterns. Treatment of 6-substituted-2H-dihydropyran-4-ones with carbon nucleophiles led to the formation of tetrahydropyran rings with the 2,6-trans-stereochemical arrangement. Reaction of the same 6-substituted-2H-dihydropyran-4-ones with l-Selectride led to the formation of 3,6-disubstituted tetrahydropyran rings, while trapping of the intermediate enolate with carbon electrophiles in turn led to the formation 3,3,6-trisubstituted tetrahydropyran rings. The relative stereochemical configuration of the new substituents was controlled by the stereoelectronic preference for pseudo-axial addition of the nucleophile and trapping of the enolate from the opposite face. Application of these methods led to a synthesis of the potent anti-osteoporotic diarylheptanoid natural product diospongin B.


Frontal cortex and the discovery of abstract action rules.
Although we often encounter circumstances with which we have no prior experience, we rapidly learn how to behave in these novel situations. Such adaptive behavior relies on abstract behavioral rules that are generalizable, rather than concrete rules mapping specific cues to specific responses. Although the frontal cortex is known to support concrete rule learning, less well understood are the neural mechanisms supporting the acquisition of abstract rules. Here, we use a reinforcement learning paradigm to demonstrate that more anterior regions along the rostro-caudal axis of frontal cortex support rule learning at higher levels of abstraction. Moreover, these results indicate that when humans confront new rule learning problems, this rostro-caudal division of labor supports the search for relationships between context and action at multiple levels of abstraction simultaneously.


A quality risk management model approach for cell therapy manufacturing.
International regulatory authorities view risk management as an essential production need for the development of innovative, somatic cell-based therapies in regenerative medicine. The available risk management guidelines, however, provide little guidance on specific risk analysis approaches and procedures applicable in clinical cell therapy manufacturing. This raises a number of problems. Cell manufacturing is a poorly automated process, prone to operator-introduced variations, and affected by heterogeneity of the processed organs/tissues and lot-dependent variability of reagent (e.g., collagenase) efficiency. In this study, the principal challenges faced in a cell-based product manufacturing context (i.e., high dependence on human intervention and absence of reference standards for acceptable risk levels) are identified and addressed, and a risk management model approach applicable to manufacturing of cells for clinical use is described for the first time. The use of the heuristic and pseudo-quantitative failure mode and effect analysis/failure mode and critical effect analysis risk analysis technique associated with direct estimation of severity, occurrence, and detection is, in this specific context, as effective as, but more efficient than, the analytic hierarchy process. Moreover, a severity/occurrence matrix and Pareto analysis can be successfully adopted to identify priority failure modes on which to act to mitigate risks. The application of this approach to clinical cell therapy manufacturing in regenerative medicine is also discussed.


Ocular symptoms in COVID-19 infection: a survey study.
Coronavirus disease 2019 (COVID-19) systemic symptoms and sequelae have been studied extensively, but less is known about the characterization, duration, and long-term sequelae of ocular symptoms associated with COVID-19 infection. The purpose of this study was to analyze the frequency, spectrum, and duration of ocular symptoms in participants with COVID-19 infection treated in inpatient and outpatient settings. A retrospective electronic survey was distributed to NIH employees and the public who reported testing positive for SARS-CoV-2. The anonymous survey collected information on demographics, past ocular history, systemic COVID-19 symptoms, and ocular symptoms. A total of 229 (21.9% male and 78.1% female, mean age 42.5 ± 13.9) survey responses were included. Ocular symptoms were reported by 165 participants with a mean of 2.31 ± 2.42 symptoms. The most commonly reported ocular symptoms were light sensitivity (31.0%), itchy eyes (24.9%), tearing (24.9%), eye redness (24.5%), and eye pain (24.5%). Participants with ocular symptoms had a higher number of systemic symptoms compared to participants without ocular symptoms (mean 9.17 ± 4.19 vs 6.22 ± 3.63; OR: 1.21; 95% CI: 1.11 - 1.32; p < 0.001). Ocular symptoms were more common in those who reported a past ocular history compared to those who did not (81.8% vs 67.1%; OR: 2.17; 95% CI: 1.08 - 4.37; p = 0.03). Additionally, the onset of ocular symptoms occurred most frequently at the same time as systemic symptoms (47.5%), and 21.8% reported symptoms lasting ≥ 14 days. Ocular surface-related symptoms are the most frequent ocular manifestations, and systemic disease severity is associated with the presence of ocular symptoms. Additionally, our results show that ocular symptoms can persist post-COVID-19 infection. Further work is needed to better understand ocular symptoms in COVID-19 and long-term sequelae.


Translation fidelity coevolves with longevity.
Whether errors in protein synthesis play a role in aging has been a subject of intense debate. It has been suggested that rare mistakes in protein synthesis in young organisms may result in errors in the protein synthesis machinery, eventually leading to an increasing cascade of errors as organisms age. Studies that followed generally failed to identify a dramatic increase in translation errors with aging. However, whether translation fidelity plays a role in aging remained an open question. To address this issue, we examined the relationship between translation fidelity and maximum lifespan across 17 rodent species with diverse lifespans. To measure translation fidelity, we utilized sensitive luciferase-based reporter constructs with mutations in an amino acid residue critical to luciferase activity, wherein misincorporation of amino acids at this mutated codon re-activated the luciferase. The frequency of amino acid misincorporation at the first and second codon positions showed strong negative correlation with maximum lifespan. This correlation remained significant after phylogenetic correction, indicating that translation fidelity coevolves with longevity. These results give new life to the role of protein synthesis errors in aging: Although the error rate may not significantly change with age, the basal rate of translation errors is important in defining lifespan across mammals.


Phlegmasia cerulea dolens with compartment syndrome: a complication of femoral vein catheterization.
Central venous catheterization is commonly performed in the critically ill. The femoral vein is widely accepted as an insertion site with complications thought to be comparable to other central access sites. We used serial ultrasound examinations with Doppler to examine the evolution of a heretofore undescribed complication of femoral vein catheterization, phlegmasia cerulea dolens with compartment syndrome. Serial ultrasounds were performed in patients before the insertion of femoral venous catheters and sequentially every 48 hrs while the catheters were in place. The noncatheterized leg served as a control. A trauma and life support center of a tertiary multidisciplinary critical care unit. A 32-yr-old man with respiratory failure as a consequence of a severe community-acquired pneumonia that required central venous access for antibiotics because no peripheral sites could be obtained. None. The initial ultrasound examination of both legs before femoral catheter insertion revealed no sign of venous thrombosis. Ultrasound of the catheterized leg at 48 hrs revealed a small nonocclusive thrombosis, whereas the opposite leg remained normal. At 72 hrs, the catheterized leg had clinical and ultrasonographic evidence of a massive thrombosis. A compartment syndrome defined by pressure measurements soon ensued and required emergent surgical release. This case report and a review of the available literature suggest that thrombosis associated with femoral vein catheterization should be considered when clinicians decide where to obtain central venous access when multiple sites are available. This report also suggests the utility of serial ultrasound examinations to define clinically nonapparent thrombosis as an early indicator of a potentially catastrophic complication.


Dysregulation of lipolysis and lipid metabolism in visceral and subcutaneous adipocytes by high-fat diet: role of ATGL, HSL, and AMPK.
This study investigated the molecular mechanisms by which a high-fat diet (HFD) dysregulates lipolysis and lipid metabolism in mouse epididymal (visceral, VC) and inguinal (subcutaneous, SC) adipocytes. Eight-weeks of HFD feeding increased adipose triglyceride lipase (ATGL) content and comparative gene identification-58 (CGI-58) expression, whereas hormone-sensitive lipase (HSL) phosphorylation and perilipin content were severely reduced. Adipocytes from HFD mice elicited increased basal but blunted epinephrine-stimulated lipolysis and increased diacylglycerol content in both fat depots. Consistent with impaired adrenergic receptor signaling, HFD also increased adipose-specific phospholipase A(2) expression in both fat depots. Inhibition of E-prostanoid 3 receptor increased basal lipolysis in control adipocytes but failed to acutely alter the effects of HFD on lipolysis in both fat depots. In HFD visceral adipocytes, activation of adenylyl cyclases by forskolin increased HSL phosphorylation and surpassed the lipolytic response of control cells. However, in HFD subcutaneous adipocytes, forskolin induced lipolysis without detectable HSL phosphorylation, suggesting activation of an alternative lipase in response to HFD-induced suppression of HSL in VC and SC adipocytes. HFD also powerfully inhibited basal, epinephrine-, and forskolin-induced AMP kinase (AMPK) activation as well peroxisome proliferator-activated receptor gamma coactivator-1alpha expression, citrate synthase activity, and palmitate oxidation in both fat depots. In summary, novel evidence is provided that defective adrenergic receptor signaling combined with upregulation of ATGL and suppression of HSL and AMPK signaling mediate HFD-induced alterations in lipolysis and lipid utilization in VC and SC adipocytes, which may play an important role in defective lipid mobilization and metabolism seen in diet-induced obesity.


[Indicators of the persistent pro-inflammatory activation of the immune system in depression].
The aetiology of depression remains tentative. Current hypotheses on the aetiology of the depressive disorder tend to integrate monoaminoergic, neuroendocrine and immunological concepts of depression. A number of research papers emphasise the altered hormonal and immune status of patients with depression with pronounced cytokine level variations. Those studies tend to link the variable course of depression in relation to the altered proinflammatory activity of the immune system. The results of the studies on the activity of the selected elements of the immune system are ambiguous indicating both increased and decreased activities of its selected elements. However, a number of basic and psychopharmacological studies support the hypothesis of the increased proinflammatory activity of the immune system in the course of depression which is the foundation for the immunological hypothesis of depression. The aim of this paper is to review the functional abnormalities that are observed in depression focusing on the monoaminoergic deficiency and increased immune activation as well as endocrine dysregulation. This paper puts together and discusses current studies related to this subject with a detailed insight into interactions involving nervous, endocrine and immune systems.


Pre-operative echocardiographic features associated with persistent mitral regurgitation after left ventricular assist device implantation.
Previous studies have shown remarkable decrease in size of the left ventricle after left ventricular assist device (LVAD) implantation due to mechanical unloading. However, a certain number of patients continue to have significant mitral regurgitation (MR) under LVAD support. We investigated pre-operative echocardiographic features associated with persistent MR after LVAD implantation. We retrospectively reviewed 82 consecutive patients undergoing continuous-flow LVAD implantation between 2007 and 2010. We obtained echocardiograms performed within 2 weeks before and 1 week after surgery. We investigated the pre-operative echocardiographic findings associated with significant MR post-LVAD and compared 1-year mortality after LVAD surgery between patients with and without significant MR post-LVAD. MR was significant in 43 patients (52.4%) before LVAD surgery. Among those, 5 underwent concomitant mitral valve repair (MVr) at the time of LVAD implantation. Of the remaining 38 patients, 25 (65.8%) showed improvement of MR, whereas 13 patients (34.2%) continued to have significant MR post-LVAD. Multivariate analysis revealed that posterior displacement of the coaptation point of mitral leaflets was significantly associated with significant MR post-LVAD (hazard ratio, 1.335; 95% confidence interval, 1.035-1.721; p = 0.026) even after adjusting for the amount of pre-operative MR flow. Post-LVAD 1-year survival of patients with and without significant MR post-LVAD was not significantly different (92.3% vs 89.1%, p = 0.826). Pre-LVAD posterior displacement of mitral leaflets may be indicative of post-operative significant MR, which would help identify echocardiographic features of functional MR refractory to simple volume reduction of the ventricle.


Occurrence and Distribution of Triticum mosaic virus in the Central Great Plains.
Wheat curl mite (WCM)-transmitted viruses-namely, Wheat streak mosaic virus (WSMV), Triticum mosaic virus (TriMV), and the High Plains virus (HPV)-are three of the wheat-infecting viruses in the central Great Plains of the United States. TriMV is newly discovered and its prevalence and incidence are largely unknown. Field surveys were carried out in Colorado, Kansas, Nebraska, and South Dakota in spring and fall 2010 and 2011 to determine TriMV prevalence and incidence and the frequency of TriMV co-infection with WSMV or HPV in winter wheat. WSMV was the most prevalent and was detected in 83% of 185 season-counties (= s-counties), 73% of 420 season-fields (= s-fields), and 35% of 12,973 samples. TriMV was detected in 32, 6, and 6% of s-counties, s-fields, and samples, respectively. HPV was detected in 34, 15, and 4% of s-counties, s-fields, and samples, respectively. TriMV was detected in all four states. In all, 91% of TriMV-positive samples were co-infected with WSMV, whereas WSMV and HPV were mainly detected as single infections. The results from this study indicate that TriMV occurs in winter wheat predominantly as a double infection with WSMV, which will complicate breeding for resistance to WCM-transmitted viruses.


RNA:(guanine-N2) methyltransferases RsmC/RsmD and their homologs revisited--bioinformatic analysis and prediction of the active site based on the uncharacterized Mj0882 protein structure.
Escherichia coli guanine-N2 (m2G) methyltransferases (MTases) RsmC and RsmD modify nucleosides G1207 and G966 of 16S rRNA. They possess a common MTase domain in the C-terminus and a variable region in the N-terminus. Their C-terminal domain is related to the YbiN family of hypothetical MTases, but nothing is known about the structure or function of the N-terminal domain. Using a combination of sequence database searches and fold recognition methods it has been demonstrated that the N-termini of RsmC and RsmD are related to each other and that they represent a "degenerated" version of the C-terminal MTase domain. Novel members of the YbiN family from Archaea and Eukaryota were also indentified. It is inferred that YbiN and both domains of RsmC and RsmD are closely related to a family of putative MTases from Gram-positive bacteria and Archaea, typified by the Mj0882 protein from M. jannaschii (1dus in PDB). Based on the results of sequence analysis and structure prediction, the residues involved in cofactor binding, target recognition and catalysis were identified, and the mechanism of the guanine-N2 methyltransfer reaction was proposed. Using the known Mj0882 structure, a comprehensive analysis of sequence-structure-function relationships in the family of genuine and putative m2G MTases was performed. The results provide novel insight into the mechanism of m2G methylation and will serve as a platform for experimental analysis of numerous uncharacterized N-MTases.


Phenotype, cancer risk, and surveillance in Beckwith-Wiedemann syndrome depending on molecular genetic subgroups.
Patients with Beckwith-Wiedemann syndrome (BWS) have an increased risk to develop cancer in childhood, especially Wilms tumor and hepatoblastoma. The risk varies depending on the cause of BWS. We obtained clinical and molecular data in our cohort of children with BWS, including tumor occurrences, and correlated phenotype and genotype. We obtained similar data from larger cohorts reported in the literature. Phenotype, genotype and tumor occurrence were available in 229 of our own patients. Minor differences in phenotype existed depending on genotype/epigenotype, similar to earlier studies. By adding patients from the literature, we obtained data on genotype and tumor occurrence of in total 1,971 BWS patients. Tumor risks were highest in the IC1 (H19/IGF2:IG-DMR) hypermethylation subgroup (28%) and pUPD subgroup (16%) and were lower in the KCNQ1OT1:TSS-DMR (IC2) subgroup (2.6%), CDKN1C (6.9%) subgroup, and the group in whom no molecular defect was detectable (6.7%). Wilms tumors (median age 24 months) were frequent in the IC1 (24%) and pUPD (7.9%) subgroups. Hepatoblastoma occurred mostly in the pUPD (3.5%) and IC2 (0.7%) subgroups, never in the IC1 and CDKN1C subgroups, and always before 30 months of age. In the CDKN1C subgroup 2.8% of patients developed neuroblastoma. We conclude tumor risks in BWS differ markedly depending on molecular background. We propose a differentiated surveillance protocol, based on tumor risks in the various molecular subgroups causing BWS. © 2016 Wiley Periodicals, Inc.


CRISPR-Cas Defense System and Potential Prophages in Cyanobacteria Associated with the Coral Black Band Disease.
Understanding how pathogens maintain their virulence is critical to developing tools to mitigate disease in animal populations. We sequenced and assembled the first draft genome of Roseofilum reptotaenium AO1, the dominant cyanobacterium underlying pathogenicity of the virulent coral black band disease (BBD), and analyzed parts of the BBD-associated Geitlerinema sp. BBD_1991 genome in silico. Both cyanobacteria are equipped with an adaptive, heritable clustered regularly interspaced short palindromic repeats (CRISPR)-Cas defense system type I-D and have potential virulence genes located within several prophage regions. The defense system helps to prevent infection by viruses and mobile genetic elements via identification of short fingerprints of the intruding DNA, which are stored as templates in the bacterial genome, in so-called "CRISPRs." Analysis of CRISPR target sequences (protospacers) revealed an unusually high number of self-targeting spacers in R. reptotaenium AO1 and extraordinary long CRIPSR arrays of up to 260 spacers in Geitlerinema sp. BBD_1991. The self-targeting spacers are unlikely to be a form of autoimmunity; instead these target an incomplete lysogenic bacteriophage. Lysogenic virus induction experiments with mitomycin C and UV light did not reveal an actively replicating virus population in R. reptotaenium AO1 cultures, suggesting that phage functionality is compromised or excision could be blocked by the CRISPR-Cas system. Potential prophages were identified in three regions of R. reptotaenium AO1 and five regions of Geitlerinema sp. BBD_1991, containing putative BBD relevant virulence genes, such as an NAD-dependent epimerase/dehydratase (a homolog in terms of functionality to the third and fourth most expressed gene in BBD), lysozyme/metalloendopeptidases and other lipopolysaccharide modification genes. To date, viruses have not been considered to be a component of the BBD consortium or a contributor to the virulence of R. reptotaenium AO1 and Geitlerinema sp. BBD_1991. We suggest that the presence of virulence genes in potential prophage regions, and the CRISPR-Cas defense systems are evidence of an arms race between the respective cyanobacteria and their bacteriophage predators. The presence of such a defense system likely reduces the number of successful bacteriophage infections and mortality in the cyanobacteria, facilitating the progress of BBD.


Chlorthalidone for poorly controlled hypertension in chronic kidney disease: an interventional pilot study.
To test the hypothesis that thiazide-type diuretics effectively lower blood pressure (BP) in moderate to advanced chronic kidney disease (CKD; estimated GFR 20-45 ml/min/ 1.73 m(2)), after confirming poorly controlled hypertension with 24-hour ambulatory BP monitoring, chlorthalidone was added to existing medications in a dose of 25 mg/day, and the dose doubled every 4 weeks if the BP remained elevated. The average age of the 14 subjects was 67.5 years, a median of 4 antihypertensive drugs were used and estimated GFR was 26.8 ± 8.8 ml/min/1.73 m(2). Twelve subjects completed the 12-week treatment phase, and the 24-hour BP, which was 143.1/75.1 mm Hg at baseline, was reduced by 10.5/ 3.1 mm Hg (p = 0.01/p = 0.17). Home BP prior to initiating chlorthalidone was 152.4/82.6 mm Hg and fell at 4, 8, and 12 weeks by 10.2/4.8, 13.4/6.0, and 9.4/3.7 mm Hg (all p < 0.05). Maximal reduction in body weight and total body volume (measured by air displacement plethysmography) was seen at 8 weeks, concurrent with the maximal elevation in serum creatinine concentration and plasma renin activity. Albuminuria was significantly reduced by 40-45%. Adverse events were seen following chlorthalidone therapy in 7 subjects who experienced 18 events as follows: hypokalemia (n = 4), hyperuricemia (4), hyponatremia (3), transient creatinine changes (3), dizziness (2), hyperglycemia (1), and constipation (1). One subject had ischemic stroke during the study. In conclusion, among people with moderate to advanced CKD with poorly controlled hypertension, chlorthalidone may significantly reduce BP via volume contraction; a randomized trial is needed to define the risks and benefits. Adverse effects may occur within a few weeks and should be carefully monitored.


Lung cancer: chemoprevention and intermediate effect markers.
Even after smoking cessation, genetic damage in the airways epithelium may lead to focal progression of lung carcinogenesis. Some centres now report as many new lung cancer cases among former smokers as among current smokers. Chemoprevention is a potential approach to diminish the progression of pre-clinical genetic damage. The most intensively studied lung cancer chemoprevention agents are the retinoids, including vitamin A and its synthetic analogues and precursors. While effective in suppressing lung carcinogenesis in animal models, retinoids have failed to inhibit carcinogenesis in human chemoprevention trials with premalignant end-points (sputum atypia, bronchial metaplasia). In trials with lung cancer end-points, administration of retinoids either was ineffective or, in the case of beta-carotene, led to greater lung cancer incidence and mortality. In view of these findings, markers of specific retinoid effect (i.e., levels of RAR-beta) become less relevant. Other markers of genetic instability and proliferation may be useful for both early detection and potentially as intermediate-effect markers for new chemoprevention trials. Cytological atypia, bronchial metaplasia, protein (hnRNP A2/B1) overexpression, ras oncogene activation and tumour-suppressor gene deletion, genomic instability (loss of heterozygosity, microsatellite alterations), abnormal methylation, helical CT detection of atypical adenomatous hyperplasia and fluorescent bronchoscopic detection of angiogenic squamous dysplasia offer great promise for molecular diagnosis of lung cancer far in advance of clinical presentation. These end-points can now be evaluated as monitors of response to chemoprevention as potential intermediate-effect markers.


Atherosclerosis--epidemiological studies on the health effects of a Mediterranean diet.
Mediterranean diets are characterized by olive oil, as the dominant fat source and a high to moderate consumption of fruit and vegetables, cereal products, fish, legumes, in combination with little meat and wine with meals. The "reference" Mediterranean diet seems to differ according to country, but is associated with good health and a long life expectancy. From the Seven Countries Studies, it has been shown that especially the traditional Cretan diet was associated with very low 25-year mortality rates for coronary heart disease, cancer and all-causes. In terms of nutrients and bioactive compounds the "reference" Mediterranean diet is low in saturated fat and high in monounsaturated fatty acids, high in antioxidants especially vitamin C and E, and high in fiber and folic acid. Several epidemiological studies have investigated these dietary components either separately or in combination in other than Mediterranean populations. In general, also in other populations beneficial effects on the coronary risk profile have been observed, which gives further support to the positive health effects of the Mediterranean diet. Intervention studies in East Finland and Southern Italy have convincingly shown that the coronary risk profile (lower LDL cholesterol and blood pressure levels) is improved by a Mediterranean diet. Moreover, the Cretan diet was tested in cardiac patients and showed a 70 % lower cardiac and all-causes mortality compared to the control diet. In conclusion, epidemiological studies and intervention trials suggest that the Cretan Mediterranean diet lowers the risk of coronary heart disease.


Description of a new species of Gymnotocinclus from the rio Tocantins basin with phylogenetic analysis of the subfamily Hypoptopomatinae (Siluriformes: Loricariidae).
Gymnotocinclus canoeiro n. sp. of the Hypoptopomatinae is described from small tributaries of the upper rio Tocantins basin. It is distinguished from G. anosteos by having five characters: (1) the presence of body dermal plates, (2) the pectoral girdle not exposed in ventral view, (3) the pelvic spine longer than pectoral spine in males, (4) the pectoral fin with seven to eight branched rays, and (5) the presence of an adipose fin. Furthermore, maximum likelihood (ML) analysis was used to estimate a molecular phylogeny from previously published data of one nuclear (F-Reticulon 4) and three mitochondrial (16S RNA, COI and CytB) genes. The phylogenetic results revealed the new species as a sister taxon of Gymnotocinclus anosteos within the Otothyrini. We also included samples of Nannoplecostomus eleonorae that appeared sister group to all other Neoplecostomini species, and Plesioptopoma curvidens that appeared within the Neoplecostomini forming a sister clade to all species of Neoplecostomus, except N. ribeirensis and the species of Pareiorhina, except P. rudolphi.


Oxygen-sensing neurons reciprocally regulate peripheral lipid metabolism via neuropeptide signaling in Caenorhabditis elegans.
The mechanisms by which the sensory environment influences metabolic homeostasis remains poorly understood. In this report, we show that oxygen, a potent environmental signal, is an important regulator of whole body lipid metabolism. C. elegans oxygen-sensing neurons reciprocally regulate peripheral lipid metabolism under normoxia in the following way: under high oxygen and food absence, URX sensory neurons are activated, and stimulate fat loss in the intestine, the major metabolic organ for C. elegans. Under lower oxygen conditions or when food is present, the BAG sensory neurons respond by repressing the resting properties of the URX neurons. A genetic screen to identify modulators of this effect led to the identification of a BAG-neuron-specific neuropeptide called FLP-17, whose cognate receptor EGL-6 functions in URX neurons. Thus, BAG sensory neurons counterbalance the metabolic effect of tonically active URX neurons via neuropeptide communication. The combined regulatory actions of these neurons serve to precisely tune the rate and extent of fat loss to the availability of food and oxygen, and provides an interesting example of the myriad mechanisms underlying homeostatic control.


A general Suzuki cross-coupling reaction of heteroaromatics catalyzed by nanopalladium on amino-functionalized siliceous mesocellular foam.
Suzuki-Miyaura cross-coupling reactions of heteroaromatics catalyzed by palladium supported in the cavities of amino-functionalized siliceous mesocellular foam are presented. The nanopalladium catalyst effectively couples not only heteroaryl halides with boronic acids but also heteroaryl halides with boronate esters, potassium trifluoroborates, MIDA boronates, and triolborates, producing a wide range of heterobiaryls in good to excellent yields. Furthermore, the heterogeneous palladium nanocatalyst can easily be removed from the reaction mixture by filtration and recycled several times with minimal loss in activity. This catalyst provides an alternative, environmentally friendly, low-leaching process for the preparation of heterobiaryls.


Thermoresponsive anionic copolymer brush-grafted surfaces for cell separation.
Cell separation methods that do not require surface modification of cells are needed in tissue engineering and regenerative medicine. We developed thermoresponsive anionic polymer brushes for cell separation without modification of the cell surfaces. Copolymer brush poly(N-isopropylacrylamide-co-N-tert-butylacrylamide-co-tert-butyl acrylate, P(NIPAAm-co-tBAAm-co-tBA), was prepared on a cover glass plate through activator regenerated by electron transfer atom transfer radical polymerization (ARGET-ATRP). The tert-butyl group of the copolymer brush was then deprotected and a P(NIPAAm-co-tBAAm-co-acrylic acid (AAc)) brush-modified glass surface was obtained. ARGET-ATRP synthesis achieved polymers with low polydispersity. The negative surface charge of the polymer brush-modified substrates was evaluated using zeta potential measurements and the phase transition temperature of the polymer was modulated between 37-20 °C to perform cell adhesion and detachment, respectively. The adhesion and detachment behavior of cells used in cardiovascular tissue engineering on the thermoresponsive anionic polymer brushes was investigated. Normal human umbilical vein endothelial cells (HUVEC) exhibited prompt detachment from the thermoresponsive anionic polymer brush surfaces. In addition, normal human aortic smooth muscle cells (SMC) exhibited relatively high adhesion on thermoresponsive anionic polymer brush-modified surfaces compared with those modified with thermoresponsive polymer brushes without anionic groups. By utilizing the difference in the cell adhesion and detachment properties of the cell types, a mixture of HUVEC and SMC was separated simply by altering the applied temperature. This result indicated that the prepared thermoresponsive anionic polymer brush-modified glass surface could be used as a tool for the separation of cells in cardiovascular tissue engineering.


Qualitative examination of cognitive change during PTSD treatment for active duty service members.
The current study investigated changes in service members' cognitions over the course of Cognitive Processing Therapy (CPT) for posttraumatic stress disorder (PTSD). Sixty-three active duty service members with PTSD were drawn from 2 randomized controlled trials of CPT-Cognitive Only (CPT-C). Participants wrote an impact statement about the meaning of their index trauma at the beginning and again at the end of therapy. Clauses from each impact statement were qualitatively coded into three categories for analysis: assimilation, accommodation, and overaccommodation. The PTSD Checklist, Posttraumatic Symptom Scale-Interview Version, and the Beck Depression Inventory-II were administered at baseline and posttreatment. Repeated measures analyses documented a significant decrease in the percentage of assimilated or overaccommodated statements and an increase in the percentage of accommodated statements from the beginning to the end of treatment. Changes in accommodated statements over the course of treatment were negatively associated with PTSD and depression symptom severity, while statements indicative of overaccommodation were positively associated with both PTSD and depression symptom severity. Treatment responders had fewer overaccommodated and more accommodated statements. Findings suggest that CPT-C changes cognitions over the course of treatment. Methodological limitations and the lack of association between assimilation and PTSD symptom severity are further discussed.


Genetic regulation of gene expression and splicing during a 10-year period of human aging.
Molecular and cellular changes are intrinsic to aging and age-related diseases. Prior cross-sectional studies have investigated the combined effects of age and genetics on gene expression and alternative splicing; however, there has been no long-term, longitudinal characterization of these molecular changes, especially in older age. We perform RNA sequencing in whole blood from the same individuals at ages 70 and 80 to quantify how gene expression, alternative splicing, and their genetic regulation are altered during this 10-year period of advanced aging at a population and individual level. We observe that individuals are more similar to their own expression profiles later in life than profiles of other individuals their own age. We identify 1291 and 294 genes differentially expressed and alternatively spliced with age, as well as 529 genes with outlying individual trajectories. Further, we observe a strong correlation of genetic effects on expression and splicing between the two ages, with a small subset of tested genes showing a reduction in genetic associations with expression and splicing in older age. These findings demonstrate that, although the transcriptome and its genetic regulation is mostly stable late in life, a small subset of genes is dynamic and is characterized by a reduction in genetic regulation, most likely due to increasing environmental variance with age.


Cell-matrix interaction in bone: type I collagen modulates signal transduction in osteoblast-like cells.
Cell interaction with extracellular matrix (ECM) modulates cell growth and differentiation. By using in vitro culture systems, we tested the effect of type I collagen (Coll-I) on signal transduction mechanisms in the osteosarcoma cell line UMR-106 and in primary cultures from neonatal rat calvariae. Cells were cultured for 72 h on Coll-I gel matrix and compared with control cells plated on plastic surfaces. Agonist-dependent and voltage-dependent rises in cytosolic Ca2+ concentration ([Ca2+]i; measured by fura 2 fluorometry) were significantly blunted in cells cultured on Coll-I compared with cells grown on plastic. In UMR-106 cells, the collagen matrix effect was mimicked by 24-h incubation with soluble Coll-I or short peptides containing the arginine-glycine-aspartate motif. Accumulation of cellular adenosine 3',5'-cyclic monophosphate (cAMP) stimulated by parathyroid hormone, cholera toxin, and forskolin was augmented (50-150%) in cells plated on Coll-I vs. control. The collagen effect on both [Ca2+]i- and adenylate cyclase-signaling pathways in UMR-106 cells was abrogated in the presence of protein kinase C (PKC) depletion or inhibition. Also, Coll-I induced a twofold increase in membrane-bound PKC without changing cytosolic PKC activity. Thus, by altering PKC activity, Coll-I modulates the [Ca2+]i- and cAMP-signaling pathways in osteoblasts. This, in turn, may influence bone remodeling processes.


Induction, maintenance, and reinduction of tumoricidal activity in bone marrow-derived mononuclear phagocytes by Corynebacterium parvum. Evidence for the involvement of a T cell- and interferon-gamma-independent pathway of macrophage activation.
Rat bone marrow-derived mononuclear phagocytes, virtually homogeneous with respect to the cell lineage, do not exhibit spontaneous tumoricidal activity in the resting state. When incubated with macrophage-activating lymphokines, rat recombinant interferon-gamma (IFN), or heat-killed Corynebacterium parvum, bone marrow-derived mononuclear phagocytes readily evolve tumoricidal activity. Whereas tumoricidal activity induced by lymphokines and/or rat recombinant IFN-gamma is short-lived, that elicited by C. parvum is maintained for at least 2 wk, provided that the C. parvum organisms are continuously present in the culture. After washing off extracellular organisms, C. parvum-induced tumoricidal activity decays rapidly, suggesting that sustained extracellular stimulation is required for its maintenance. Induction of tumoricidal activity by macrophage-activating lymphokines and/or rat recombinant IFN-gamma is fully prevented by polyclonal and monoclonal anti-IFN-gamma antibodies; in contrast, induction by C. parvum is not affected by anti-IFN-gamma. Since induction of tumoricidal activity by C. parvum takes place irrespective of the presence of anti-Thy-1 antisera or cyclosporin A, T cells and/or their products appear not to be involved in this type of macrophage activation. Accordingly, present findings provide evidence for the existence of lymphokine-independent pathways of macrophage activation.


Repurposing 18F-FMISO as a PET tracer for translational imaging of nitroreductase-based gene directed enzyme prodrug therapy.
Nitroreductases (NTR) are a family of bacterial enzymes used in gene directed enzyme prodrug therapy (GDEPT) that selectively activate prodrugs containing aromatic nitro groups to exert cytotoxic effects following gene transduction in tumours. The clinical development of NTR-based GDEPT has, in part, been hampered by the lack of translational imaging modalities to assess gene transduction and drug cytotoxicity, non-invasively. This study presents translational preclinical PET imaging to validate and report NTR activity using the clinically approved radiotracer, 18F-FMISO, as substrate for the NTR enzyme. Methods: The efficacy with which 18F-FMISO could be used to report NfsB NTR activity in vivo was investigated using the MDA-MB-231 mammary carcinoma xenograft model. For validation, subcutaneous xenografts of cells constitutively expressing NTR were imaged using 18F-FMISO PET/CT and fluorescence imaging with CytoCy5S, a validated fluorescent NTR substrate. Further, examination of the non-invasive functionality of 18F-FMISO PET/CT in reporting NfsB NTR activity in vivo was assessed in metastatic orthotopic NfsB NTR expressing xenografts and metastasis confirmed by bioluminescence imaging. 18F-FMISO biodistribution was acquired ex vivo by an automatic gamma counter measuring radiotracer retention to confirm in vivo results. To assess the functional imaging of NTR-based GDEPT with 18F-FMISO, PET/CT was performed to assess both gene transduction and cytotoxicity effects of prodrug therapy (CB1954) in subcutaneous models. Results:18F-FMISO retention was detected in NTR+ subcutaneous xenografts, displaying significantly higher PET contrast than NTR- xenografts (p < 0.0001). Substantial 18F-FMISO retention was evident in metastases of orthotopic xenografts (p < 0.05). Accordingly, higher 18F-FMISO biodistribution was prevalent ex vivo in NTR+ xenografts. 18F-FMISO NfsB NTR PET/CT imaging proved useful for monitoring in vivo NTR transduction and the cytotoxic effect of prodrug therapy. Conclusions:18F-FMISO NfsB NTR PET/CT imaging offered significant contrast between NTR+ and NTR- tumours and effective resolution of metastatic progression. Furthermore, 18F-FMISO NfsB NTR PET/CT imaging proved efficient in monitoring the two steps of GDEPT, in vivo NfsB NTR transduction and response to CB1954 prodrug therapy. These results support the repurposing of 18F-FMISO as a readily implementable PET imaging probe to be employed as companion diagnostic test for NTR-based GDEPT systems.


Comparison of Dry Needling versus Orthopedic Manual Therapy in Patients with Myofascial Chronic Neck Pain: A Single-Blind, Randomized Pilot Study.
Objective. The aim of this study was to compare the efficacy of three interventions for the treatment of myofascial chronic neck pain. Methods. Thirty-six patients were randomly assigned to one of three intervention groups: orthopedic manual therapy (OMT), dry needling and stretching (DN-S), and soft tissue techniques (STT). All groups received two treatment sessions with a 48 h time interval. Outcome measures included neck pain intensity measured using a visual analogue scale, cervical range of motion (ROM), pressure pain threshold for measuring mechanical hyperalgesia, and two self-reported questionnaires (neck disability index and pain catastrophizing scale). Results. The ANOVA revealed significant differences for the group × time interaction for neck disability, neck pain intensity, and pain catastrophizing. The DN-S and OMT groups reduced neck disability. Only the OMT group showed decreases in mechanical hyperalgesia and pain catastrophizing. The cervical ROM increased in OMT (i.e., flexion, side-bending, and rotation) and DN-S (i.e., side-bending and rotation) groups. Conclusions. The three interventions are all effective in reducing pain intensity. Reduction in mechanical hyperalgesia and pain catastrophizing was only observed in the OMT group. Cervical ROM improved in the DN-S and OMT groups and also neck disability being only clinically relevant for OMT group.


Unsupervised natural experience rapidly alters invariant object representation in visual cortex.
Object recognition is challenging because each object produces myriad retinal images. Responses of neurons from the inferior temporal cortex (IT) are selective to different objects, yet tolerant ("invariant") to changes in object position, scale, and pose. How does the brain construct this neuronal tolerance? We report a form of neuronal learning that suggests the underlying solution. Targeted alteration of the natural temporal contiguity of visual experience caused specific changes in IT position tolerance. This unsupervised temporal slowness learning (UTL) was substantial, increased with experience, and was significant in single IT neurons after just 1 hour. Together with previous theoretical work and human object perception experiments, we speculate that UTL may reflect the mechanism by which the visual stream builds and maintains tolerant object representations.


Axillary evaluation and lymphedema in women with ductal carcinoma in situ.
Axillary evaluation in women with ductal carcinoma in situ (DCIS) is increasing; however, this may introduce additional morbidity with unclear benefit. Our objective was to examine the morbidity and mortality associated with axillary evaluation in DCIS. We conducted a retrospective cohort study of 10,504 women aged 65-90 years with DCIS who underwent breast conserving surgery between 2002 and 2012 using SEER-Medicare database. Patients were categorized by receipt of axillary evaluation with either sentinel lymph node biopsy (SLNB) or axillary node dissection (ALND). We determined the incidence of lymphedema treatment as defined by diagnostic and procedural codes, as well as 10-year breast cancer-specific and all-cause mortality. 18.3 % of those treated with BCS and 69.4 % of those treated with mastectomy had an axillary evaluation. One year after treatment, 8.2 % of women who had an axillary evaluation developed lymphedema, compared to 5.9 % of those who did not. In a multivariable Cox proportional hazard model, the incidence of lymphedema was higher among those who underwent axillary evaluation (HR 1.22, 95 % CI 1.04-1.45). Overall 10-year breast cancer-specific survival was similar between both groups (HR 0.83, 95 % CI 0.40-1.74). Only 44 (0.40 %) women died of breast cancer; receipt of axillary evaluation did not alter overall survival. Axillary evaluation is commonly performed in women with DCIS, especially those undergoing mastectomy. However, women who receive an axillary evaluation have higher rates of lymphedema, without breast cancer-specific or overall survival benefit. Efforts should be made to determine the population of women with DCIS who benefit from this procedure.


Comparison of detection methods of EGFR T790M mutations using plasma, serum, and tumor tissue in EGFR-TKI-resistant non-small cell lung cancer.
Osimertinib, a third-generation epidermal growth factor receptor-tyrosine kinase inhibitor, exerts remarkable effects against EGFR T790M resistance mutation-positive non-small cell lung cancer. Identifying T790M mutation by re-biopsy is essential before prescribing osimertinib. Tissue biopsy is the golden standard for this purpose, but several factors limit its success rate. The liquid biopsy with blood, using circulating tumor DNA, has been an alternative method. However, the true biological meaning and equivalence of liquid biopsy and tumor biopsy are still under investigation. Especially, the usefulness of serum samples to detect T790M mutation is not yet been known. We prospectively evaluated the sensitivity, specificity, and parallelism of the detection of EGFR mutations in tissue re-biopsy and liquid biopsy (plasma and serum), simultaneously, from June 2016 to May 2017. EGFR mutations in tumor re-biopsy were evaluated by COBAS ver2 and PNA-LNA PCR clamp method, and those in liquid biopsy were evaluated with COBAS ver2. Fifteen patients were enrolled. In 10 patients whose EGFR mutation was detected in liquid biopsy, the original EGFR mutation (exon 19 del or L858R) was detected in all patients. Detection of EGFR mutation by COBAS ver2 and by PNA-LNA method was almost the same in tissue re-biopsy. The detection rate of T790M was lower than that of the original EGFR mutation in liquid biopsy compared to that in tissue re-biopsy. The detection of T790M in serum exhibited a higher specificity (67%) and positive predictive value (50%) than that in plasma (50% and 40%, respectively). The detection sensitivity was similar in plasma and serum. Plasma, serum, and tissue genotyping can have complementary roles for detecting EGFR-T790M using COBAS ver2. Repeated tests with different samples and different methods may improve accuracy of T790M detection and will lead to the maximum benefit for the patient.


Correlation between midthigh low-density muscle and insulin resistance in obese nondiabetic patients in Korea.
We investigated the link between lipid-rich skeletal muscle, namely low-density muscle, and insulin resistance in Korea. Abdominal adipose tissue areas and midthigh skeletal muscle areas of 75 obese nondiabetic subjects (23 men, 52 women; mean age +/- SD, 41.9 +/- 14.1 years) were measured by computed tomography (CT). The midthigh skeletal muscle areas were subdivided into low-density muscle (0 to +30 Hounsfield units) and normal-density muscle (+31 to +100 Hounsfield units). The homeostasis model assessment (HOMA) score was calculated to assess whole-body insulin sensitivity. The abdominal visceral fat area and the midthigh low-density muscle area were found to be well correlated with the HOMA score (r = 0.471, P < 0.01 and r = 0.513, P < 0.01, respectively). The correlation between low-density muscle area and insulin resistance persisted after adjusting for BMI or total body fat mass (r = 0.451, P < 0.01 and r = 0.522, P < 0.01, respectively) and even after adjusting for abdominal visceral fat area (r = 0.399, P < 0.01). The midthigh low-density muscle area seems to be a reliable determinant of insulin resistance in Korean obese nondiabetic patients.


Anticoagulant Activity of Naja nigricollis Venom Is Mediated by Phospholipase A2 Toxins and Inhibited by Varespladib.
Bites from elapid snakes typically result in neurotoxic symptoms in snakebite victims. Neurotoxins are, therefore, often the focus of research relating to understanding the pathogenesis of elapid bites. However, recent evidence suggests that some elapid snake venoms contain anticoagulant toxins which may help neurotoxic components spread more rapidly. This study examines the effects of venom from the West African black-necked spitting cobra (Naja nigricollis) on blood coagulation and identifies potential coagulopathic toxins. An integrated RPLC-MS methodology, coupled with nanofractionation, was first used to separate venom components, followed by MS, proteomics and coagulopathic bioassays. Coagulation assays were performed on both crude and nanofractionated N. nigricollis venom toxins as well as PLA2s and 3FTx purified from the venom. Assays were then repeated with the addition of either the phospholipase A2 inhibitor varespladib or the snake venom metalloproteinase inhibitor marimastat to assess whether either toxin inhibitor is capable of neutralizing coagulopathic venom activity. Subsequent proteomic analysis was performed on nanofractionated bioactive venom toxins using tryptic digestion followed by nanoLC-MS/MS measurements, which were then identified using Swiss-Prot and species-specific database searches. Varespladib, but not marimastat, was found to significantly reduce the anticoagulant activity of N. nigricollis venom and MS and proteomics analyses confirmed that the anticoagulant venom components mostly consisted of PLA2 proteins. We, therefore, conclude that PLA2s are the most likely candidates responsible for anticoagulant effects stimulated by N. nigricollis venom.


Predominant contribution of the G protein-mediated mechanism to NaF-induced vascular contractions in diabetic rats: association with an increased level of G(qalpha) expression.
The purpose of this study was to determine the mechanism responsible for alterations in NaF-induced contractions of blood vessels from streptozotocin-induced diabetic rats. In the presence of AlCl(3), NaF (>/=7.5 mM) produced significantly greater contractions in diabetic aorta and mesenteric artery compared with age-matched controls. Pretreatment with 1 microM nifedipine eliminated the enhanced contractile responses of diabetic vessels to NaF, resulting in no difference in the magnitude of NaF-induced contractions between control and diabetic vessels. In the presence of 100 microM deferoxamine, an Al(3+) chelator, NaF-induced contractions of diabetic vessels were markedly attenuated, whereas only the responses to lower concentrations of NaF were reduced in control vessels. No significant difference was found in the peak amplitude of transient contractions induced by 10 microM cyclopiazonic acid between control and diabetic vessels. The addition of 10 microM okadaic acid produced attenuated contractions in diabetic vessels. These findings indicate no involvement of the inhibitory effects of NaF on endoplasmic reticular Ca(2+)-pump ATPase and protein phosphatases in the genesis of the enhanced responsiveness of diabetic vessels to NaF. Western blot analysis showed a 2.5-fold increase in the expression of G(qalpha) in diabetic aortic membranes. In contrast, the G(ialpha) level was modestly decreased and the G(salpha) and G(betagamma) levels were unchanged in diabetes. The present results suggest that enhanced vascular contractions to NaF in diabetes is attributed predominantly to a G protein-mediated Ca(2+) channel activation that results from markedly increased G(qalpha) expression in vascular tissues under this pathological state.


Mutation in the CYP21B gene (Ile-172----Asn) causes steroid 21-hydroxylase deficiency.
Steroid 21-hydroxylase deficiency is the most common cause of congenital adrenal hyperplasia. It results from a deficiency in a specific cytochrome P450, P450c21 (P450XXIA). The gene encoding this protein (CYP21B) and a closely linked pseudogene (CYP21A) are located in the HLA complex on chromosome 6p. Many mutant alleles are associated with deletions of CYP21B; we report the cloning and characterization of a nondeleted mutant CYP21B gene. This mutant gene is expressed on transfection into mouse Y1 adrenal cells, producing mRNA levels similar to those seen after transfection of the normal CYP21B gene. In codon 172 of the mutant gene, the normal codon ATC, encoding isoleucine, has been changed to AAC, encoding asparagine. This mutation is normally present in the CYP21A pseudogene, so that it may have been transferred to the mutant CYP21B gene by gene conversion. Hybridization of oligonucleotide probes corresponding to this and two other mutations normally present in CYP21A demonstrated that 4 out of 20 patients carried the codon 172 mutation; in one of these patients, the mutation was present as part of a larger gene conversion involving at least exons 3-6. Gene conversion may be a frequent cause of 21-hydroxylase deficiency alleles due to the presence of six chi-like sequences (GCTGGGG) in the CYP21 genes and the close proximity of the CYP21A pseudogene, which has several potentially deleterious mutations.


Preparation of molecular imprinted microspheres based on inorganic-organic co-functional monomer for miniaturized solid-phase extraction of fluoroquinolones in milk.
An inorganic-organic co-functional monomer, methacrylic acid-vinyltriethoxysilan (MAA-VTES) was designed for the synthesis of molecularly imprinted microspheres (MIMs). By virtue of the aqueous suspension polymerization and dummy template (pazufloxacin), the obtained MAA-VTES based MIMs exhibited good recognition and selectivity to fluoroquinolones (FQs), and were successfully applied as selective sorbents of a miniaturized home-made solid phase extraction device for the determination of ofloxacin (OFL), lomefloxacin (LOM) and ciprofloxacin (CIP) in milk samples. Under the optimum conditions of the miniaturized molecularly imprinted solid phase extraction (mini-MISPE) coupled with liquid chromatography-ultraviolet detector (LC-UV), good linearities were obtained for three FQs in a range of 0.2-20.0μgmL(-1) and the average recoveries at three spiked levels were ranged from 87.2% to 106.1% with the relative standard deviation (RSD) less than 5.4%. The presented co-functional monomer based mini-MISPE-LC-UV protocol introduced the rigidity and flexibility of inorganic silicon materials, exhibited excellent extraction performance towards targets, and could be potentially applied to the determination of FQs in milk samples.


Can uroflowmetry patterns in women be reliably interpreted?
Our aim was to investigate the proposition that uroflowmetry patterns can be reliably interpreted and correspond with specific urodynamic diagnoses. Uroflowmetry traces from 129 women with diagnoses of either genuine stress incontinence or detrusor instability were interpreted by four physicians with a minimum of 6 months experience in urogynecology. To test intraobserver variability, the traces were classified a second time 8 weeks later. Inter- and intraobserver variability was calculated by kappa analysis. There was marked intra- and interobserver variability in classification of traces, but no evidence of a correlation between urodynamic diagnosis and uroflowmetry pattern. Neither peak flow, total voided volume nor rate of acceleration of flow correlated with diagnosis. Although flow rates are important in predicting possible problems following surgery for stress incontinence, there is no evidence that flow patterns can be used as a screening test for specific urodynamic diagnoses.


Gli3 coordinates three-dimensional patterning and growth of the tectum and cerebellum by integrating Shh and Fgf8 signaling.
The coordination of anterior-posterior (AP) and dorsal-ventral (DV) patterning of the mesencephalon (mes) and rhombomere 1 (r1) is instrumental for the development of three distinct brain structures: the tectum and cerebellum dorsally and the tegmentum ventrally. Patterning of the mes/r1 is primarily mediated by signaling molecules secreted from two organizers: sonic hedgehog (Shh) from the floor plate (DV) and Fgf8 from the isthmus (AP). Gli3, a zinc-finger transcription factor in the Shh signaling pathway, has been implicated in regulating Fgf8 expression and is therefore a potential candidate for coordinating the action of the two organizers. By inactivating mouse Gli3 at successive embryonic time points in vivo, we uncovered the extent and the underlying mechanism of Gli3 function in the mes/r1. We demonstrate that before E9.0, Gli3 is required for establishing a distinct posterior tectum, isthmus and cerebellum, but does not play a role in the development of the tegmentum. Between E9.0 and E11.0, Gli3 continues to be required for isthmus and cerebellum development, but primarily for defining the cerebellar foliation pattern. We show that Gli3 regulates patterning of the isthmus and cerebellar anlage by confining Fgf8 expression to the isthmus, and attenuates growth of dorsal r1 (before E11.0) and the dorsal mes and isthmus (beyond E11.0) through regulation of cell proliferation and viability. In conclusion, our results show that Gli3 is essential for the coordinated three-dimensional patterning and growth of the dorsal mes/r1.


Motor imagery during action observation enhances imitation of everyday rhythmical actions in children with and without developmental coordination disorder.
Children with developmental coordination disorder (DCD) exhibit deficits both in imitation and motor imagery (MI) compared to typically developing children. Combined action observation and motor imagery (AO + MI) instructions can however enhance automatic imitation in both groups. In the present study we investigated the effects of AO + MI instructions on intentional imitation in children both with (n = 13) and without DCD (n = 12). On each trial participants observed and/or imagined before executing a familiar rhythmical pantomime action. These target actions were either habitually fast (tooth brushing or window wiping) or habitually slow (paint brushing or face washing), in the vertical or horizontal plane. Within each habitual speed, the target action speed was subtly manipulated across trials (fast vs. slow). Instruction condition was manipulated across three blocks of 16 trials: (1) observe before imitating the target action; (2) observe then imagine the action before imitating; (3) observe while imagining the same action before imitating (AO + MI). Kinematic analyses revealed typically developing children imitated the observed cycle times significantly better than children with DCD. A main effect of instruction showed imitation improved for AO + MI compared to the other two instructions. Within-group analyses found a significant advantage in DCD for AO + MI compared to observe then imagine. In typically developing children, imitation was significantly enhanced for AO + MI compared to observe then imitate. Combined AO + MI instructions therefore represent a promising new approach to refining performance of everyday rhythmical actions in children both with and without DCD, with implications for movement therapy and sports training.


TCL1A Single-Nucleotide Polymorphisms and Estrogen-Mediated Toll-Like Receptor-MYD88-Dependent Nuclear Factor-κB Activation: Single-Nucleotide Polymorphism- and Selective Estrogen Receptor Modulator-Dependent Modification of Inflammation and Immune Response.
In a previous genome-wide association study (GWAS) for musculoskeletal adverse events during aromatase inhibitor therapy for breast cancer, we reported that single nucleotide polymorphisms (SNPs) near the TCL1A gene were associated with this adverse drug reaction. Functional genomic studies showed that TCL1A expression was induced by estradiol, but only in cells with the variant sequence for the top GWAS SNP (rs11849538), a SNP that created a functional estrogen response element. In addition, TCL1A genotype influenced the downstream expression of a series of cytokines and chemokines and had a striking effect on nuclear factor κB (NF-κB) transcriptional activity. Furthermore, this SNP-dependent regulation could be reversed by selective ER modulators (SERMs). The present study was designed to pursue mechanisms underlying TCL1A SNP-mediated, estrogen-dependent NF-κB activation. Functional genomic studies were performed using a panel of 300 lymphoblastoid cell lines for which we had generated genome-wide SNP and gene expression data. It is known that toll-like receptors (TLRs) can regulate NF-κB signaling by a process that requires the adaptor protein MYD88. We found that TLR2, TLR7, TLR9, and TLR10 expression, as well as that of MYD88, could be modulated by TCL1A in a SNP and estrogen-dependent fashion and that these effects were reversed in the presence of SERMs. Furthermore, MYD88 inhibition blocked the TCL1A SNP and estrogen-dependent NF-κB activation, as well as protein-protein interaction between TCL1A and MYD88. These observations greatly expand the range of pathways influenced by TCL1A genotype and raise the possibility that this estrogen- and SNP-dependent regulation might be altered pharmacologically by SERMs.


Compact portable biosensor for arsenic detection in aqueous samples with Escherichia coli bioreporter cells.
We present a compact portable biosensor to measure arsenic As(III) concentrations in water using Escherichia coli bioreporter cells. Escherichia coli expresses green fluorescent protein in a linearly dependent manner as a function of the arsenic concentration (between 0 and 100 μg/L). The device accommodates a small polydimethylsiloxane microfluidic chip that holds the agarose-encapsulated bacteria, and a complete optical illumination/collection/detection system for automated quantitative fluorescence measurements. The device is capable of sampling water autonomously, controlling the whole measurement, storing and transmitting data over GSM networks. We demonstrate highly reproducible measurements of arsenic in drinking water at 10 and 50 μg/L within 100 and 80 min, respectively.


The Indian pediatric HIV epidemic: a systematic review.
Despite an estimated 70,000 Indian children living with HIV infection, little is known about India's pediatric HIV epidemic. Generalizations about epidemiology, natural history, and treatment outcomes from other resource-limited settings (RLS) may be inaccurate for several biologic and social reasons. A review of the Indian literature is needed to optimize country-specific HIV management and examine these generalizations. MEDLINE and EMBASE were searched for articles published in English by November 2007 on HIV-infected, Indian children (0-18 years) that detailed epidemiology, natural history, or treatment. Articles with original, extractable data were selected and summarized using descriptive statistics. Of 370 citations, 58 studies were included in this review (median study size 24 children). Significant heterogeneity was noted among the studies. HIV infection was reported nearly twice as often in males (male/female ratio 1.9) and diagnosed earlier (4.7 years) than in other RLS. Over 2% of hospitalized children were reported to be HIV-infected. The reported mortality among HIV-infected newborns of 22% at 18 months was lower than other RLS. Improved anthropometrics were the only consistently reported and comparable benefit of short-term HAART to other RLS. Review of the Indian literature yielded potentially unique epidemiology and natural history compared to other RLS. However, important questions about the accuracy and representativeness of the Indian data limit its generalizability and comparability. Targeted interventions to curb India's pediatric HIV epidemic require urgent clarification of these findings. If such differences truly exist, management guidelines should be tailored accordingly.


Reagent Controlled Stereoselective Assembly of α-(1,3)-Glucans.
Pre-activation based glycosylations have become a very powerful tool in the assembly of oligosaccharides and the use of nucleophilic additives allows for the in situ generation of reactive intermediates with tailored reactivity. We here use a glycosylation strategy that is based on the use of per-benzylated imidate building blocks for the fully stereoselective construction of a spacer equipped Aspergillus fumigatus α-1,3-octaglucan. We have used the trimethylsilyl iodide (TMSI)-triphenylphosphine oxide (Ph3P=O) for the stereoselective installation of an azidopropanol spacer and triflic acid (TfOH)-dimethyl formamide (DMF) enabled glycosylations for the coupling reactions with the secondary glucosyl C-3-alcohols. An operationally simple in situ activation coupling procedure is introduced and used for the final glycosylation events towards the octasaccharide.


Screening of compounds interacting with HIV-1 proteinase using optical biosensor technology.
A high-resolution optical biosensor assay for screening of low-molecular-weight compounds, using an immobilized protein target, has been developed. HIV-1 proteinase was immobilized on the sensor surface by direct amine coupling and a variety of inhibitors and noninteracting reference drugs were applied to the sensor surface in a continuous flow of buffer. The procedure did not require intrinsic reporter groups, substrates, inhibitors, or other ligands for detection. By using a reference protein, the signal could be corrected for the relatively large background signal caused by differences in dimethyl sulfoxide concentration between running and sample buffers. Substances binding with high affinity (Ki in nM range) required efficient regeneration of the sensor surface and washing of the injection system between sample cycles to get consistent results. Analysis was simplified by using report points, extracted during both association and dissociation phases, and a simple graphical display of data. The optimized assay could correctly distinguish HIV-1 inhibitors from other compounds in a randomized series, indicate differences in their interaction kinetics, and reveal artifacts due to nonspecific signals, incomplete regeneration, or carryover. The method is expected to be generally applicable to secondary screening of low-molecular-weight compound libraries with proteins as targets.


Pathogen susceptibility and fitness costs explain variation in immune priming across natural populations of flour beetles.
In many insects, individuals primed with low doses of pathogens early in life have higher survival after exposure to the same pathogen later in life. Yet, our understanding of the evolutionary and ecological history of priming of immune response in natural insect populations is limited. Previous work demonstrated population-, sex- and stage-specific variation in the survival benefit of priming response in flour beetles (Tribolium castaneum) infected with their natural pathogen Bacillus thuringiensis. However, the evolutionary forces responsible for this natural variation remained unclear. In the present work, we tested whether the strength of the priming response (measured as the survival benefit after priming and subsequent infection, relative to unprimed controls) was associated with multiple fitness parameters and immune components across 10 flour beetle populations collected from different locations in India. Our results suggest two major selective pressures that may explain the observed inter-population variation in priming: (a) Basal pathogen susceptibility - populations that were more susceptible to infection produced a stronger priming response, and (b) Short-term early reproductive success - populations where primed females produced more offspring early in life (measured over 2 days) had lower survival benefit (measured over 120 days), suggesting a potential trade-off between early reproduction and priming response. However, the negative association between survival and reproduction is limited to priming and infection in adults, but not in larvae. While other components of beetle fitness (starvation resistance and larval development) and immune function (haemolymph antibacterial activity and antimicrobial quinone secretion) also varied widely across populations, none of them was correlated with the variation in priming responses across populations. Our work is the first systematic empirical demonstration of multiple selective pressures that may govern the evolution of immune priming in the wild. We hope that this motivates further experiments to establish the role of pathogen-imposed selection and fitness costs in the evolution of priming in natural insect populations.


Preparative and Kinetic Analysis of β-1,4- and β-1,3-Glucan Phosphorylases Informs Access to Human Milk Oligosaccharide Fragments and Analogues Thereof.
The enzymatic synthesis of oligosaccharides depends on the availability of suitable enzymes, which remains a limitation. Without recourse to enzyme engineering or evolution approaches, herein we demonstrate the ability of wild-type cellodextrin phosphorylase (CDP: β-1,4-glucan linkage-dependent) and laminaridextrin phosphorylase (Pro_7066: β-1,3-glucan linkage-dependent) to tolerate a number of sugar-1- phosphate substrates, albeit with reduced kinetic efficiency. In spite of catalytic efficiencies of <1 % of the natural reactions, we demonstrate the utility of given phosphorylase-sugar phosphate pairs to access new-to-nature fragments of human milk oligosaccharides, or analogues thereof, in multi-milligram quantities.


TLR Adaptor Protein MYD88 Mediates Sensitivity to HDAC Inhibitors via a Cytokine-Dependent Mechanism.
Histone deacetylase (HDAC) inhibitors have proven useful therapeutic agents for certain hematologic cancers. However, HDAC inhibition causes diverse cellular outcomes, and identification of cancer-relevant pathways within these outcomes remains unresolved. In this study, we utilized an unbiased loss-of-function screen and identified the Toll-like receptor (TLR) adaptor protein MYD88 as a key regulator of the antiproliferative effects of HDAC inhibition. High expression of MYD88 exhibited increased sensitivity to HDAC inhibitors; conversely, low expression coincided with reduced sensitivity. MYD88-dependent TLR signaling controlled cytokine levels, which then acted via an extracellular mechanism to maintain cell proliferation and sensitize cells to HDAC inhibition. MYD88 activity was directly regulated through lysine acetylation and was deacetylated by HDAC6. MYD88 was a component of a wider acetylation signature in the ABC subgroup of diffuse large B-cell lymphoma, and one of the most frequent mutations in MYD88, L265P, conferred increased cell sensitivity to HDAC inhibitors. Our study defines acetylation of MYD88, which, by regulating TLR-dependent signaling to cytokine genes, influences the antiproliferative effects of HDAC inhibitors. Our results provide a possible explanation for the sensitivity of malignancies of hematologic origin to HDAC inhibitor-based therapy. Cancer Res; 76(23); 6975-87. ©2016 AACR.


Undergraduate physiotherapy students' basic wheelchair provision knowledge: a pilot study in two universities in Colombia.
Purpose: Access to an appropriate wheelchair is a human right. Only between 5-15% of people who need a wheelchair have access to one. One of the key barriers to access is the lack of appropriately trained rehabilitation professionals. The objective of this study was to evaluate basic manual wheelchair provision knowledge in final-year physiotherapy undergraduate students in two programs in Colombia.Materials and methods: Students took the International Society of Wheelchair Professionals Wheelchair Service Provision - Basic Test which was administered online and in Spanish. The minimum score to pass the test is 70%; it assesses seven domains: Assessment; Prescription; Products; Fitting; User training; Follow-up, maintenance, and repairs; and Process.Results and conclusions: One-hundred sixteen students took the test and no one passed the test. The highest median domain scores were in Assessment and Process while the lowest were in Fitting and Products. The limitations of this study include that this sample does not represent all physiotherapy programmes or students in Colombia, there may be potential errors in the Spanish translation of the outcome measure, and students encountered Internet connectivity issues during the test that may have impacted their scores. Immediate interventions are required to improve teaching and students' learning outcomes related to basic manual wheelchair provision in these two programs. This study may serve as a foundation for future regional or national studies that assess the situation of wheelchair provision training in rehabilitation programs that will inform improvement actions. This manuscript is also available in Spanish as Supplemental Material.Implications for rehabilitationThis study indicates that students' current knowledge on basic appropriate manual wheelchair provision from two physiotherapy programs in Colombia is insufficient. Students' knowledge does not align with the minimum guidelines recommended for wheelchair service provision by the World Health Organization.Objectively identifying the gap in knowledge in rehabilitation trainees (i.e., physiotherapy students) is a strategy to promote the inclusion of assistive technology related content in formal academic training.The need to include formal training of appropriate wheelchair provision persists and without this training, people with disabilities who require a wheelchair for mobility will continue to face barriers to full participation in society.


Cross center single-cell RNA sequencing study of the immune microenvironment in rapid progressing multiple myeloma.
Despite advancements in understanding the pathophysiology of Multiple Myeloma (MM), the cause of rapid progressing disease in a subset of patients is still unclear. MM's progression is facilitated by complex interactions with the surrounding bone marrow (BM) cells, forming a microenvironment that supports tumor growth and drug resistance. Understanding the immune microenvironment is key to identifying factors that promote rapid progression of MM. To accomplish this, we performed a multi-center single-cell RNA sequencing (scRNA-seq) study on 102,207 cells from 48 CD138- BM samples collected at the time of disease diagnosis from 18 patients with either rapid progressing (progression-free survival (PFS) < 18 months) or non-progressing (PFS > 4 years) disease. Comparative analysis of data from three centers demonstrated similar transcriptome profiles and cell type distributions, indicating subtle technical variation in scRNA-seq, opening avenues for an expanded multicenter trial. Rapid progressors depicted significantly higher enrichment of GZMK+ and TIGIT+ exhausted CD8+ T-cells (P = 0.022) along with decreased expression of cytolytic markers (PRF1, GZMB, GNLY). We also observed a significantly higher enrichment of M2 tolerogenic macrophages in rapid progressors and activation of pro-proliferative signaling pathways, such as BAFF, CCL, and IL16. On the other hand, non-progressive patients depicted higher enrichment for immature B Cells (i.e., Pre/Pro B cells), with elevated expression for markers of B cell development (IGLL1, SOX4, DNTT). This multi-center study identifies the enrichment of various pro-tumorigenic cell populations and pathways in those with rapid progressing disease and further validates the robustness of scRNA-seq data generated at different study centers.


Estimating climate resilience for conservation across geophysical settings.
Conservationists need methods to conserve biological diversity while allowing species and communities to rearrange in response to a changing climate. We developed and tested such a method for northeastern North America that we based on physical features associated with ecological diversity and site resilience to climate change. We comprehensively mapped 30 distinct geophysical settings based on geology and elevation. Within each geophysical setting, we identified sites that were both connected by natural cover and that had relatively more microclimates indicated by diverse topography and elevation gradients. We did this by scoring every 405 ha hexagon in the region for these two characteristics and selecting those that scored >SD 0.5 above the mean combined score for each setting. We hypothesized that these high-scoring sites had the greatest resilience to climate change, and we compared them with sites selected by The Nature Conservancy for their high-quality rare species populations and natural community occurrences. High-scoring sites captured significantly more of the biodiversity sites than expected by chance (p < 0.0001): 75% of the 414 target species, 49% of the 4592 target species locations, and 53% of the 2170 target community locations. Calcareous bedrock, coarse sand, and fine silt settings scored markedly lower for estimated resilience and had low levels of permanent land protection (average 7%). Because our method identifies-for every geophysical setting-sites that are the most likely to retain species and functions longer under a changing climate, it reveals natural strongholds for future conservation that would also capture substantial existing biodiversity and correct the bias in current secured lands.


The use of Resonant X-ray Emission Spectroscopy (RXES) for the electronic analysis of metal complexes and their interactions with biomolecules.
This review presents a new application of Resonant X-ray Emission Spectroscopy (RXES) to study the mechanism of action of metal containing anticancer derivatives and in particular platinum in situ and in vivo. The technique is an example of a photon-in photon-out X-ray spectroscopic approach, which enables chemical speciation of drugs to be determined and therefore to derive action mechanisms, and to determine drug binding rates under physiological conditions and therapeutic concentrations. This is made feasible due to the atomic specificity and high penetration depth of RXES. The review presents examples of the three main types of information that can be obtained by RXES and establishes an experimental protocol to perfect the measurements within cells.


Exploring the longitudinal stability of the CSQ24 in a back pain population.
The CSQ24 is a shortened version of the Coping Strategies Questionnaire and includes 24 items and four factors-Catastrophizing, Diversion, Reinterpreting, and Cognitive Coping. The factor structure of the CSQ has been a matter for debate for some time. This study aimed to explore the stability of the factor structure of the CSQ24 using repeated measurements in a back pain population at assessment, after physiotherapy treatment, and at 6 and 12 months after treatment. A consecutive sample of 194 adults with chronic low back pain was recruited over 24 months. They completed the CSQ24, with other measures of pain, disability and mood, on 4 occasions. Ninety-six participants satisfactorily completed and returned both 6- and 12-month longitudinal follow-up questionnaires. Exploratory Factor Analysis showed, at each time point, a 4-factor structure of Catastrophizing, Cognitive Coping, Diversion, and Reinterpreting. Within each factor, the factor scores at each time point were significantly correlated and showed good internal reliability. This stability was demonstrated against a background of significant change in measures of pain, disability, and mood. These results provide support for the 4-factor structure of the CSQ24 and its longitudinal stability. Argument is provided for the need to explore the longitudinal stability of related tools in populations that undergo change as a result of treatment.


Lithium-ion storage properties of a micro/nanosheet-like NaV6O15 anode in aqueous solution.
The micromorphologies of NaV6O15 materials synthesized using the hydrothermal method and calcination comprise staggered micro/nanosheet-like shapes. These materials can act as anode active materials in aqueous rechargeable lithium-ion batteries (ARLBs). NaV6O15 has a charge transfer resistance of several ohms in the ARLB, which is an order of magnitude smaller than in traditional lithium-ion batteries. The lithium-ion diffusion coefficients at 0.15, -0.21 and -0.65 V vs. saturated calomel electrode (SCE) were calculated as 7.3 × 10-9, 3 × 10-12, 7.6 × 10-11 cm2 s-1, respectively, from the Warburg spectrum, which were three orders of magnitude larger than in conventional lithium-ion batteries. Furthermore, NaV6O15 materials used in this ARLB have a high energy density of about 134.9 Wh kg-1 at a power of 640 W kg-1. In contrast to traditional LIB behavior, we found that the capacity retention and coulombic efficiency of the as-synthesized materials increased with the increasing ARLB discharge current density, which showed potential for this new ARLB system to be applied in the field of large-scale energy storage and power source devices.


The impact on healthcare, policy and practice from 36 multi-project research programmes: findings from two reviews.
We sought to analyse the impacts found, and the methods used, in a series of assessments of programmes and portfolios of health research consisting of multiple projects. We analysed a sample of 36 impact studies of multi-project research programmes, selected from a wider sample of impact studies included in two narrative systematic reviews published in 2007 and 2016. We included impact studies in which the individual projects in a programme had been assessed for wider impact, especially on policy or practice, and where findings had been described in such a way that allowed them to be collated and compared. Included programmes were highly diverse in terms of location (11 different countries plus two multi-country ones), number of component projects (8 to 178), nature of the programme, research field, mode of funding, time between completion and impact assessment, methods used to assess impact, and level of impact identified. Thirty-one studies reported on policy impact, 17 on clinician behaviour or informing clinical practice, three on a combined category such as policy and clinician impact, and 12 on wider elements of impact (health gain, patient benefit, improved care or other benefits to the healthcare system). In those multi-programme projects that assessed the respective categories, the percentage of projects that reported some impact was policy 35% (range 5-100%), practice 32% (10-69%), combined category 64% (60-67%), and health gain/health services 27% (6-48%). Variations in levels of impact achieved partly reflected differences in the types of programme, levels of collaboration with users, and methods and timing of impact assessment. Most commonly, principal investigators were surveyed; some studies involved desk research and some interviews with investigators and/or stakeholders. Most studies used a conceptual framework such as the Payback Framework. One study attempted to assess the monetary value of a research programme's health gain. The widespread impact reported for some multi-project programmes, including needs-led and collaborative ones, could potentially be used to promote further research funding. Moves towards greater standardisation of assessment methods could address existing inconsistencies and better inform strategic decisions about research investment; however, unresolved issues about such moves remain.


Genes required for the engulfment of cell corpses during programmed cell death in Caenorhabditis elegans.
After programmed cell death, a cell corpse is engulfed and quickly degraded by a neighboring cell. For degradation to occur, engulfing cells must recognize, phagocytose and digest the corpses of dying cells. Previously, three genes were known to be involved in eliminating cell corpses in the nematode Caenorhabditis elegans: ced-1, ced-2 and nuc-1. We have identified five new genes that play a role in this process: ced-5, ced-6, ced-7, ced-8 and ced-10. Electron microscopic studies reveal that mutations in each of these genes prevent engulfment, indicating that these genes are needed either for the recognition of corpses by other cells or for the initiation of phagocytosis. Based upon our study of double mutants, these genes can be divided into two sets. Animals with mutations in only one of these sets of genes have relatively few unengulfed cell corpses. By contrast, animals with mutations in both sets of genes have many unengulfed corpses. These observations suggest that these two sets of genes are involved in distinct and partially redundant processes that act in the engulfment of cell corpses.


The right way to self-fuse bi- and terpyrenyls to afford graphenic cutouts.
In this work, we subject bi- and terpyrenyls to selective fusion for formation of extended polycyclic aromatic hydrocarbons (PAHs). Connecting the pyrene units at 4-4'- or 1-4'-positions led to smooth formation of extended PAHs, achieved via cyclodehydrogenation. This is far more difficult if pyrene is coupled in the 1,1'-position.


Social-epidemiological study for evaluation of water supply and sanitation systems of low-income urban community in Dhaka, Bangladesh.
This study aims at quantification of health losses, considering social and environmental factors. Morbidity and mortality cases of diarrhoea for children under five years old were used to estimate the disability adjusted life years (DALYs) lost for the target households in low-income communities in Dhaka, Bangladesh. Water supply facilities and sanitation systems, along with hygiene practices and their health outcomes, were studied at community level. Demographic, socio-economic and socio-cultural aspects were also studied to support the research findings and give a better understanding of the local conditions. The four selected communities, Ward 60 (W60), Ward 61 (W61), Ward 62 (W62) and Ward 65 (W65), all had different existing urban services such as water supply, sanitation, garbage management and drainage facilities. All of these services existed in W62, but W60 did not have any of the services; W61 had sanitation and drainage coverage, whereas W65 had only a water supply facility. The results conclusively showed that, compared with the null (absence of services) scenario (W60), the other three scenarios (W61, W62 and W65) showed a substantial decrease of diarrhoea (1.219, 1.284 and 2.052 DALYs/household/year, respectively) reported for children under five years old. Besides urban services, other socio-economic characteristics might also influence the prevalence of diseases.


International Psychogeriatric Association consensus statement on defining and measuring treatment benefits in dementia.
Current symptomatic treatments for dementia have only modest efficacy. Assessing meaningful benefits in this variably progressive syndrome is complex and difficult. This consensus statement was generated by an international group representing caregivers, organizations and professionals with expertise in dementia. We recommend the statement of clear, pre-defined diagnostic and severity criteria and outcome measures, which include functional and executive capacity, in treatment trials. Outcomes can include effects on people with dementia (PWD) with regard to cognition, behavioral and psychological symptoms, quality of life, global assessments, and activities of daily living, and must be tailored to the education and culture of the participants. Outcomes can also appropriately encompass effects on caregivers. New instruments may be needed, as validation of many existing measures has been in relatively homogenous populations. Treatment goals can be to prevent dementia emerging, or in those with established dementia to slow deterioration, and to postpone, stabilize or improve symptoms. Comparisons between treatment groups should be on the basis of clinically relevant measures with both risk and benefit reported for all participants regardless of whether or not they continue to receive treatment throughout the trial. Characterization of any groups that respond well to treatment has been unsuccessful to date, but may be facilitated in the future by measurement of putative biomarkers. Despite considerable recent progress and several 'candidate' biomarkers, none is yet satisfactory for determining diagnosis, severity, progression or prediction of response. To provide meaningful data, economic analyses should use up-to-date, country-specific data. Health economic measures should be incorporated as secondary outcomes in all Phase 3 trials since health systems are concerned with cost-effectiveness as well as clinical outcome. Health utility measures are not, however, validated satisfactorily in dementia, thus calling into question previous health economic analyses. While current drugs appear to reduce the amount of family caregiver time required by PWD, these costs fall in the main on older individuals who often exert little political leverage, rather than on society at large. Traditionally, elderly people have been marginalized in the political process. The growth in the older population across the world, and their potential for increasing political empowerment may lead to a radical re-evaluation of the economics of treatment in dementia.


A multi-gene phylogeny of Cephalopoda supports convergent morphological evolution in association with multiple habitat shifts in the marine environment.
The marine environment is comprised of numerous divergent organisms living under similar selective pressures, often resulting in the evolution of convergent structures such as the fusiform body shape of pelagic squids, fishes, and some marine mammals. However, little is known about the frequency of, and circumstances leading to, convergent evolution in the open ocean. Here, we present a comparative study of the molluscan class Cephalopoda, a marine group known to occupy habitats from the intertidal to the deep sea. Several lineages bear features that may coincide with a benthic or pelagic existence, making this a valuable group for testing hypotheses of correlated evolution. To test for convergence and correlation, we generate the most taxonomically comprehensive multi-gene phylogeny of cephalopods to date. We then create a character matrix of habitat type and morphological characters, which we use to infer ancestral character states and test for correlation between habitat and morphology. Our study utilizes a taxonomically well-sampled phylogeny to show convergent evolution in all six morphological characters we analyzed. Three of these characters also correlate with habitat. The presence of an autogenic photophore (those relying upon autonomous enzymatic light reactions) is correlated with a pelagic habitat, while the cornea and accessory nidamental gland correlate with a benthic lifestyle. Here, we present the first statistical tests for correlation between convergent traits and habitat in cephalopods to better understand the evolutionary history of characters that are adaptive in benthic or pelagic environments, respectively. Our study supports the hypothesis that habitat has influenced convergent evolution in the marine environment: benthic organisms tend to exhibit similar characteristics that confer protection from invasion by other benthic taxa, while pelagic organisms possess features that facilitate crypsis and communication in an environment lacking physical refuges. Features that have originated multiple times in distantly related lineages are likely adaptive for the organisms inhabiting a particular environment: studying the frequency and evolutionary history of such convergent characters can increase understanding of the underlying forces driving ecological and evolutionary transitions in the marine environment.


Posterior ring apophysis separation combined with lumbar disc herniation in adults: a 10-year experience in the surgical management of 87 cases.
The association of posterior ring apophysis separation (PRAS) with lumbar disc herniation (LDH) is uncommon and represents a true subgroup of disc herniation mainly seen in the adolescent population. The objective of this study was to describe a decade of experience in the care of adult patients with PRAS with LDH, giving particular attention to its diagnosis, surgical treatment, and outcome. This retrospective study focuses on a case series of adult patients with PRAS associated with LDH who underwent surgery for lumbar disc disease in the author's neurosurgical department between 1999 and 2008. Posterior ring apophysis separation was diagnosed in 87 (5.35%) of 1625 patients surgically treated for LDH; these patients made up the PRAS group. During a 6-month period in 2005, LDH without PRAS was diagnosed in 89 consecutive patients at the same facility; these patients constituted the control group. Presenting symptoms, physical examination findings, and preoperative imaging results were obtained from medical records. Immediate operative results were assessed, as were complications, long-term outcome, and the need for repeat surgery. This study is the first to document the distinguishing features between adult patients with and those without PRAS. The difference in average age was statistically significant (p < 0.001) between the study group (36.22 years) and the control group (44.30 years), as was the incidence of male patients (86.20% vs 71.91%, p = 0.020), incidence of military patients (74.71% vs 57.30%, p = 0.015), average duration of symptoms (16.13 vs 8.4 months, p = 0.016), and incidence of reactive scoliosis (19.54% vs 4.49%, p = 0.002). The most common anatomical location of disc herniation in the PRAS group was L5-S1 (51.72%) versus L4-L5 (53.93%) in the control group (p = 0.017). In terms of previous injury, motor deficits, back and/or leg pain, lateral or central location of LDH, mean anteroposterior diameter of disc herniation, hard or soft discs, and surgical complications, there was no statistical difference between the 2 patient groups. Similarly, there was no difference in recurrence rates and clinical outcomes between the patients with or without PRAS. Posterior ring apophysis separation with LDH is probably more common in adults than is generally recognized. It must be suspected when young male patients with persistent sciatic scoliosis and no history of injury show signs of calcified LDH. Computed tomography scanning with sagittal reconstructions is the procedure of choice for diagnosing. The L5-S1 intervertebral disc level is most commonly affected, especially the superior endplate of S-1. This condition needs more extensive surgical exposure and resection to relieve the nerve impingement. The occurrence of an apophyseal lesion was not associated with recurrent disc herniation or a fair outcome.