Development and evaluation of an anti-rabies virus phosphoprotein-specific monoclonal antibody for detection of rabies neutralizing antibodies using RFFIT.
Rabies is a major public health problem with a fatality rate close to 100%; however, complete prevention can be achieved through pre- or post-exposure prophylaxis. The rapid fluorescent focus inhibition test (RFFIT) is one of the recommended testing methods to determine the production of neutralizing antibodies after vaccination. Here, we report the development of a new monoclonal antibody (mAb) designed to react specifically with Rabies virus (RABV) phosphoprotein (P protein), and the evaluation of its applicability to the RFFIT and its effectiveness as a diagnostic reagent for human rabies. The mAb KGH P 16B8 was produced to target the P protein of the Korean KGH RABV strain. An indirect immunofluorescence assay (IFA) was conducted to detect various strains of RABV in various cell lines. Alexa-conjugated KGH P 16B8 (16B8-Alexa) was developed for the RFFIT. The IFA test could detect RABV up to a 1:2,500 dilution, with a detection limit comparable to that of a commercial diagnostic reagent. The sensitivity, specificity, positive predictive value, and negative predictive value of the RFFIT using 16B8-Alexa in 414 clinical specimens were 98.67%, 99.47%, 99.55%, and 98.42%, respectively. The results of the RFFIT with 16B8-Alexa were strongly correlated with those obtained using an existing commercial diagnostic reagent (r = 0.995, p<0.001). The mAb developed in this study shows high sensitivity and specificity, confirming its clinical utility with the RFFIT to measure the rabies neutralizing antibody titer and establish a diagnosis in human. Thus, 16B8-Alexa is expected to serve as an alternative diagnostic reagent that is widely accessible, with potentially broad applications beyond those of the RFFIT in Korea. Further studies with 16B8-Alexa should provide insight into the immunological mechanism of the P protein of Korean RABV.


Microglassification™: a novel technique for protein dehydration.
The dehydration of biologics is commonly employed to achieve solid-dose formulation and enhanced stability during long-term preservation. We have developed a novel process, Microglassification™, which can rapidly and controllably dehydrate protein solutions into solid amorphous microspheres at room temperature. Single bovine serum albumin (BSA) microdroplets were suspended in pentanol or decanol using a micropipette, and the dynamic changes in droplet dissolution were observed in real-time and correlated to protein's water of hydration, medium's water activity, and microsphere protein concentration. Microglassification™ was also carried out at bulk scale, and changes in BSA secondary structure were analyzed by Fourier transform infrared spectroscopy and fluorescence spectroscopy; multimer formation was detected by native gel electrophoresis. BSA concentration in the microsphere increased with solvent exposure time and decreasing water activity. Image analysis at single particle and bulk scale showed the formation of solid BSA microspheres with a maximum protein concentration of 1147 ± 32 mg/mL. The native BSA samples were dehydrated to approximately 450 waters per BSA, which is well below monolayer coverage of 1282 waters per BSA. The secondary structure of Microglassified™ BSA reverted to native-like conformation upon rehydration with only minor irreversible aggregation (2.7%). Results of the study establish the efficacy of the Microglassification™ for the successful dehydration of biologics.


Tizanidine versus baclofen in the treatment of spasticity in patients with multiple sclerosis.
Tizanidine (Sirdalud) was compared to baclofen (Lioresal) in a randomized, double-blind, cross-over trial. Each medication was introduced over a three week titration period and then maintained at the highest tolerated dose for five weeks. The two treatment phases were separated by a one week drug withdrawal and a two week washout period. Sixty-six patients entered the trial and forty-eight completed both treatment phases. At the end of the trial, neurologists and physiotherapists thought that baclofen was superior on the basis of perceived efficacy and tolerance (p less than or equal to 0.05). Although the efficacy of tizanidine or baclofen was judged as good to excellent by 24 and 39% of patients respectively, this difference was not statistically significant. Muscle weakness was the most common adverse effect. This was significantly more troublesome in patients treated with baclofen. Somnolence and xerostomia were more common in patients treated with tizanidine. Both baclofen and tizanidine appear to be useful adjuncts in the treatment of spasticity in patients with multiple sclerosis. Preference of either drug is tempered principally by side-effects.


On the proof and disproof of natural product stereostructures: characterization and analysis of a twenty-eight member stereoisomer library of murisolins and their Mosher ester derivatives.
Characterizing a stereoisomer library of 28 of the 64 possible isomers of the acetogenin murisolin, including 24 of the 32 possible diastereomers, provides a complete picture of the spectra of this class of molecules. Remarkably, each of the 32 diastereomers exhibits one of only six sets of substantially identical (1)H NMR spectra under standard conditions. These spectra follow directly from a local symmetry analysis of the dihydroxy-THF fragment of the molecule and provide no information about the configuration about the hydroxybutenolide. Eighteen tris-Mosher ester derivatives of library members have been made, and their spectra were analyzed to give a complete picture of the usefulness of chiral derivatives. The tris-Mosher esters of the 64 isomers of murisolin will exhibit 40 sets of spectra: 16 isomers have unique spectra whereas 24 isomers share an identical spectrum with one other isomer. This identity occurs even though the pairs of compounds were already diastereomers (not enantiomers) before the derivatization. The complete set of spectra allows any murisolin or closely related compound to be narrowed to one or two structures by simple matching and without recourse to assignment and subtraction of resonances. The structure of murisolin was proved to be the 4R,15R,16R,19R,20R,34S isomer, whereas the assignment of 16,19-cis-murisolin as RRRSSS was changed to the RSSRRS diastereomer and murisolin A is suggested to be RSRRRS.


What is remembered? Role of attention on the encoding and retrieval of hippocampal representations.
The hippocampus is critically involved in storing explicit memory such as memory for space. A defining feature of explicit memory storage is that it requires attention both for encoding and retrieval. Whereas, a great deal is now known about the mechanisms of storage, the mechanisms whereby attention modulates the encoding and retrieval of space and other hippocampus-dependent memory representations are not known. In this review we discuss recent studies, including our own, which show on the cellular level that attention is critical for the stabilization of spatial and reward-associated odour representations. Our findings support the view that in the hippocampus attention selects the reference frame for task-relevant information. This mechanism is in part mediated by dopamine acting through D1/D5 receptors and involves an increase in neuronal synchronization in the gamma band frequency. We propose that synchronous activity leads to enhancements in synaptic strength that mediate the stabilization of hippocampal representations.


Functional serotonin-2B receptors are expressed by a teratocarcinoma-derived cell line during serotoninergic differentiation.
Among immortalized teratocarcinoma-derived cells, the clone 1C11 is a committed precursor of the neuronal lineage. On day 2 of its serotoninergic differentiation, this clone expresses only one subtype of serotonin [5-hydroxytryptamine (5-HT)] receptor, which is functionally coupled to phosphatidylinositol hydrolysis. The identity of these receptors was established by comparing their properties with those of 5-HT2B receptors expressed by LMTK- fibroblasts stably transfected with the recently cloned murine cDNA NP75 (LM5 cells). In both cell types, the analysis of (+/-)-1-(2,5-dimethoxy-4-[125I]iodophenyl)- 2-aminopropane HCl ([125I]DOI) binding revealed the presence of a single class of sites, the affinity of which was 1 order of magnitude lower than that reported for 5-HT2A receptors. In 1C11 cells differentiated for 2 days, as well as in LM5 cells, DOI binding was decreased by nonhydrolyzable analogs of GTP, indicating that the 5-HT2B receptor is functionally coupled to a G protein. The DOI-induced increase of phosphoinositide hydrolysis, which was correlated with both GTPase activity and binding data, is mediated by a Gq protein. This work demonstrates that the 5-HT2B receptor is functionally expressed before complete serotoninergic differentiation of 1C11 cells. The inducible 1C11 clone thus provides an in vitro model to investigate the possible role of the 5-HT2B receptor in the expression of the serotoninergic phenotype.


Mattress double anchor footprint repair: a novel, arthroscopic rotator cuff repair technique.
In an effort to increase the immediate strength of a rotator cuff repair and to simulate the standard open reconstruction with its effective suture fixation, we have developed a novel technique for suture anchor reconstruction of the rotator cuff. The technique, termed mattress double anchor (MDA), is simple and adaptable. It makes use of 2 suture anchors that are placed independently and then connected by a suture loop. The technique produces a repair construct that distributes the stress across 2 anchors. The method also restores a large surface area for healing between the rotator cuff and the tuberosity.


Congruency in defining critical thinking by nurse educators and non-nurse scholars.
This study investigated nurse educators' definition of the concept critical thinking. A sample of 201 baccalaureate nurse educators in midwest nursing programs completed a questionnaire identifying their perception of critical-thinking skills and characteristics, and their agreement with non-nurse critical-thinking experts on items often considered to be critical thinking. This study found that nurse educators agreed with non-nurse critical-thinking experts on the skills and dispositions; however, significant differences were found between nurse educators and non-nurse experts regarding concepts related to critical thinking. Nurse educators were more likely to identify researching, problem-solving, decision-making, and planning as critical thinking. Despite their assertion otherwise, it is apparent from this study that nurse educators have a different perception of critical thinking than scholars in other disciplines. This study suggests that practice disciplines such as nursing may perceive critical thinking differently than educators in nonpractice disciplines.


Is chronic hepatitis C virus infection a risk factor for breast cancer?
To evaluate the prevalence of breast tumors in adult females with chronic hepatitis C virus (HCV) infection. Prospective, single-center study, based on female outpatients consulting in a liver unit, for 1 year. The study group included females with present and/or past history of chronic infection by HCV. Patients with spontaneous recovery were excluded. Chronic hepatitis had been proved by liver biopsy in the majority of cases and/or biological markers of inflammation and fibrosis. The control group included female patients with other well documented chronic liver diseases: chronic hepatitis B, alcoholic liver disease, autoimmune hepatitis, hemochromatosis, non alcoholic liver disease, chronic cholangitis. Participating patients were prospectively questioned during consultation about past breast history and follow-up by mammography. Breast carcinoma was recorded in 17/294 patients with HCV infection (5.8%, 95% CI: 3.1-8.4) vs 5/107 control patients (4.7%, 95% CI: 0.67-8.67). Benign tumors of the breast (mastosis, nodules, cysts) were recorded in 75/294 patients with HCV infection (25.5%, 95% CI: 20.5-30.5) vs 21/107 (19.6%, 95% CI: 12.1-27.1) in the control group. No lesion was noted in 202 patients with HCV (68.7%, 95% CI: 63.4-74) vs 81 control patients (75.7%, 95% CI: 67.6-83.8). Despite a trend to an increased prevalence in the group with HCV infection, the difference was not significant compared to the control group (P = NS). In patients over 40 years, the results were, respectively, as follows: breast cancer associated with HCV: 17/266 patients (6.3%, 95% CI: 3.4-9.3) vs 5/95 patients (5.2%, 95% CI: 0.7-9.7) in the control group; benign breast tumors: 72/266 patients with HCV infection (27%, 95% CI: 21.7-32.4) vs 18/95 patients (18.9%, 95% CI: 11-26.8) in the control group; no breast lesion 177/266 (66.5%, 95% CI: 60.9-72.2) in patients with HCV infection vs 72/95 (75.7%, 95% CI: 67.1-84.4) in the control group. The differences were not significant (P = NS). These results suggest that chronic HCV infection is not a strong promoter of breast carcinoma in adult females of any age.


Genetics of Cerebellar and Neocortical Expansion in Anthropoid Primates: A Comparative Approach.
What adaptive changes in brain structure and function underpin the evolution of increased cognitive performance in humans and our close relatives? Identifying the genetic basis of brain evolution has become a major tool in answering this question. Numerous cases of positive selection, altered gene expression or gene duplication have been identified that may contribute to the evolution of the neocortex, which is widely assumed to play a predominant role in cognitive evolution. However, the components of the neocortex co-evolve with other functionally interdependent regions of the brain, most notably in the cerebellum. The cerebellum is linked to a range of cognitive tasks and expanded rapidly during hominoid evolution. Here we present data that suggest that, across anthropoid primates, protein-coding genes with known roles in cerebellum development were just as likely to be targeted by selection as genes linked to cortical development. Indeed, based on currently available gene ontology data, protein-coding genes with known roles in cerebellum development are more likely to have evolved adaptively during hominoid evolution. This is consistent with phenotypic data suggesting an accelerated rate of cerebellar expansion in apes that is beyond that predicted from scaling with the neocortex in other primates. Finally, we present evidence that the strength of selection on specific genes is associated with variation in the volume of either the neocortex or the cerebellum, but not both. This result provides preliminary evidence that co-variation between these brain components during anthropoid evolution may be at least partly regulated by selection on independent loci, a conclusion that is consistent with recent intraspecific genetic analyses and a mosaic model of brain evolution that predicts adaptive evolution of brain structure.


Quantum chemical study of low temperature oxidation mechanism of dibenzofuran.
A density functional theory (DFT) study of the reaction of dibenzofuranyl radical with oxygen molecule has been made. The geometries, energies, and vibrational frequencies of the reactant, transition states, intermediates, and products have been calculated at the B3LYP/6-311+G(3df,2p)//B3LYP/6-31G(d) level of theory. The initial reaction of dibenzofuran (DBF) with molecular oxygen results in the formation of the 1-dibenzofuranylperoxy radical. The stability of this adduct toward decomposition at low to intermediate temperatures results in it undergoing several possible rearrangements. The lowest energy pathway with a barrier of 24.2 kcal/mol involves a rearrangement to the 1,1-dioxadibenzofuran radical. The next lowest energy pathway involves fission of the O-O linkage whose reaction energy was found to be 37.6 kcal/mol. Transition state theory (TST) calculations indicate that the lowest energy pathway should predominate at temperatures up to about 1200 K. Two other unimolecular reaction pathways with barriers of 45.5 and 91.1 kcal/mol have also been discovered. The latter pathway leads to the formation of a para-quinone (dibenzofuran quinone) which has been detected experimentally in the low-temperature oxidation of DBF [Marquaire, P. M.; Worner, R.; Rambaud, P.; Baronnet, F. Organohalogen Compd. 1999, 40, 519]. Our quantum calculations, however, do not support this latter pathway to quinone formation. Instead, the quinone is most probably formed as a consequence of recombination of the 1-dibenzofuranyloxy radical (produced by peroxy fission) with an O atom in the para position. Each of the unimolecular reaction pathways have been subjected to detailed quantum chemical investigation and transition states and intermediates leading to the final products (principally CO, CO2, and C2H2 with traces of benzofuran and benzene) have been identified. For certain stable intermediates, their possible reactions with molecular oxygen have been further investigated quantum chemically. The present work therefore presents a detailed quantum chemical investigation of the reaction pathways in the low-temperature oxidation mechanism of DBF. Since the dibenzofuran moiety is present in the polychlorinated DBFs, our conclusions should be generally applicable to this family of compounds.


A new approach to the classification and management of airways diseases: identification of treatable traits.
This review outlines a new, personalized approach for the classification and management of airway diseases. The current approach to airways disease is, we believe, no longer fit for purpose. It is impractical, overgeneralizes complex and heterogeneous conditions and results in management that is imprecise and outcomes that are worse than they could be. Importantly, the assumptions we make when applying a diagnostic label have impeded new drug discovery and will continue to do so unless we change our approach. This review suggests a new mechanism-based approach where the emphasis is on identification of key causal mechanisms and targeted intervention with treatment based on possession of the relevant mechanism rather than an arbitrary label. We highlight several treatable traits and suggest how they can be identified and managed in different healthcare settings.


Multicentric Castleman's disease associated with rheumatoid arthritis: a possible role of hepatitis B antigen.
A patient with seropositive rheumatoid arthritis and a carrier of hepatitis B surface antigen developed angiofollicular hyperplasia (multicentric Castleman's disease). The hepatitis B virus and the rheumatoid factor may have had a role in the aetiology of this lymphatic disorder. The development of Castleman's disease in association with these factors may provide another clue supporting the reactive nature of this disease.


A consensus tree approach for reconstructing human evolutionary history and detecting population substructure.
The random accumulation of variations in the human genome over time implicitly encodes a history of how human populations have arisen, dispersed, and intermixed since we emerged as a species. Reconstructing that history is a challenging computational and statistical problem but has important applications both to basic research and to the discovery of genotype-phenotype correlations. We present a novel approach to inferring human evolutionary history from genetic variation data. We use the idea of consensus trees, a technique generally used to reconcile species trees from divergent gene trees, adapting it to the problem of finding robust relationships within a set of intraspecies phylogenies derived from local regions of the genome. Validation on both simulated and real data shows the method to be effective in recapitulating known true structure of the data closely matching our best current understanding of human evolutionary history. Additional comparison with results of leading methods for the problem of population substructure assignment verifies that our method provides comparable accuracy in identifying meaningful population subgroups in addition to inferring relationships among them. The consensus tree approach thus provides a promising new model for the robust inference of substructure and ancestry from large-scale genetic variation data.


A high-fat, ketogenic diet causes hepatic insulin resistance in mice, despite increasing energy expenditure and preventing weight gain.
Low-carbohydrate, high-fat ketogenic diets (KD) have been suggested to be more effective in promoting weight loss than conventional caloric restriction, whereas their effect on hepatic glucose and lipid metabolism and the mechanisms by which they may promote weight loss remain controversial. The aim of this study was to explore the role of KD on liver and muscle insulin sensitivity, hepatic lipid metabolism, energy expenditure, and food intake. Using hyperinsulinemic-euglycemic clamps, we studied insulin action in mice fed a KD or regular chow (RC). Body composition was assessed by ¹H magnetic resonance spectroscopy. Despite being 15% lighter (P < 0.001) than RC-fed mice because of a 17% increase in energy expenditure (P < 0.001), KD-fed mice manifested severe hepatic insulin resistance, as reflected by decreased suppression (0% vs. 100% in RC-fed mice, P < 0.01) of endogenous glucose production during the clamp. Hepatic insulin resistance could be attributed to a 350% increase in hepatic diacylglycerol content (P < 0.001), resulting in increased activation of PKCε (P < 0.05) and decreased insulin receptor substrate-2 tyrosine phosphorylation (P < 0.01). Food intake was 56% (P < 0.001) lower in KD-fed mice, despite similar caloric intake, and could partly be attributed to a more than threefold increase (P < 0.05) in plasma N-acylphosphatidylethanolamine concentrations. In conclusion, despite preventing weight gain in mice, KD induces hepatic insulin resistance secondary to increased hepatic diacylglycerol content. Given the key role of nonalcoholic fatty liver disease in the development of type 2 diabetes and the widespread use of KD for the treatment of obesity, these results may have potentially important clinical implications.


Prevalence of symptoms of severe asthma and allergies in Irish school children: an ISAAC protocol study, 1995-2007.
Childhood asthma is a recurring health burden and symptoms of severe asthma in children are also emerging as a health and economic issue. This study examined changing patterns in symptoms of severe asthma and allergies (ever eczema and hay fever), using the Irish International Study of Asthma and Allergies in Childhood (ISAAC) protocol. ISAAC is a cross-sectional self-administered questionnaire survey of randomly selected representative post-primary schools. Children aged 13-14 years were studied: 2,670 (in 1995), 2,273 (in 1998), 2,892 (in 2002-2003), and 2,805 (in 2007). Generalized linear modelling using Poisson distribution was employed to compute adjusted prevalence ratios (PR). A 39% significant increase in symptoms of severe asthma was estimated in 2007 relative to the baseline year 1995 (adjusted PR: 1.39 [95% CI: 1.14-1.69]) increasing from 12% in 1995 to 15.3% in 2007. Opposite trends were observed for allergies, showing a decline in 2007, with an initial rise. The potential explanations for such a complex disease pattern whose aetiological hypothesis is still evolving are speculative. Changing environmental factors may be a factor, for instance, an improvement in both outdoor and indoor air quality further reinforcing the hygiene hypothesis but obesity as a disease modifier must also be considered.


The application of GBS markers for extending the dense genetic map of rye (Secale cereale L.) and the localization of the Rfc1 gene restoring male fertility in plants with the C source of sterility-inducing cytoplasm.
Genotyping by sequencing (GBS) is an efficient method of genotyping in numerous plant species. One of the crucial steps toward the application of GBS markers in crop improvement is anchoring them on particular chromosomes. In rye (Secale cereale L.), chromosomal localization of GBS markers has not yet been reported. In this paper, the application of GBS markers generated by the DArTseq platform for extending the high-density map of rye is presented. Additionally, their application is used for the localization of the Rfc1 gene that restores male fertility in plants with the C source of sterility-inducing cytoplasm. The total number of markers anchored on the current version of the map is 19,081, of which 18,132 were obtained from the DArTseq platform. Numerous markers co-segregated within the studied mapping population, so, finally, only 3397 unique positions were located on the map of all seven rye chromosomes. The total length of the map is 1593 cM and the average distance between markers is 0.47 cM. In spite of the resolution of the map being not very high, it should be a useful tool for further studies of the Secale cereale genome because of the presence on this map of numerous GBS markers anchored for the first time on rye chromosomes. The Rfc1 gene was located on high-density maps of the long arm of the 4R chromosome obtained for two mapping populations. Genetic maps were composed of DArT, DArTseq, and PCR-based markers. Consistent mapping results were obtained and DArTs tightly linked to the Rfc1 gene were successfully applied for the development of six new PCR-based markers useful in marker-assisted selection.


A biophysical perspective of understanding nanoparticles at large.
In this article we present a biophysical perspective that describes the fate of nanoparticles in both the aqueous phase and in living systems. Specifically, we show the correlations between the physicochemistry of fullerenes and their uptake, translocation, transformation, transport, and biodistribution in mammalian and plant systems, at the molecular, cellular, and whole organism level. In addition to fullerenes and their structural derivatives, we describe the biological and environmental implications and applications of the condensed matter of carbon nanotubes and quantum dots, and the soft condensed matter of plastic and dendrimers. The main purpose of this article is to demonstrate the vast opportunities and unique advantages of applying experimental and simulation biophysics to the nascent research field of understanding nanoparticles at large.


Transcript expression profiling in two contrasting cultivars and molecular cloning of a SKP-1 like gene, a component of SCF-ubiquitin proteasome system from mungbean Vigna radiate L.
Protein degradation and turnover under various environmental stresses is basically regulated by ubiquitin-proteasome system (UPS), of which SKP1 is a very essential component. Isolation and cloning of an identified potential stress responsive candidate gene SKP1, was successfully done for the first time to fathom the role of SKP1 in drought tolerance at genetic level in drought tolerant mungbean cultivar Pratap, which was screened after a detailed physio-biochemical screening amongst seven popular mungbean cultivars. The cloned gene SKP1 (accession number KX881912) is 550 bp in length, encodes 114 amino acids. It shows high sequence homology with SKP1 from Zea mays (NP_001148633). The protein expression of isolated SKP1 was confirmed by GUS fused expression using a Histochemical assay under control as well as under drought stress. Further, up-regulation in relative expression level of SKP1 in different plant parts under drought stress confirmed its utility as a potential drought responsive candidate gene certainly demanding extensive genetic research for further incorporation in breeding programs. Moreover, the structure of VrSKP1 (Vigna radiata SKP1) has been modelled, validated and an Essential Dynamics (ED) was done on the Molecular Dynamics (MD) simulation trajectories for filtering large-scale concerted motions. Free-energy calculations on the ED revealed a complex free-energy landscape (FEL) implying the conformational diversity of the modelled VrSPK1 protein.


Automatic Localization of Cochlear Implant Electrode Contacts in CT.
Determining the exact location of cochlear implant (CI) electrode contacts after implantation is important, as it helps quantifying the relation between CI positioning and hearing outcome. Unfortunately, localization of individual contacts can be difficult, because the spacing between the electrode contacts is near the spatial resolution limit of high-resolution clinical computed tomography (CT) scanners. This study introduces and examines a simple, automatic method for the localization of intracochlear electrode contacts. CI geometric specifications may provide the prior knowledge that is essential to accurately estimate contact positions, even though individual contacts may not be visibly resolved. The prior knowledge in CI geometry is used to accurately estimate intracochlear electrode contact positions in high-resolution CT scans of seven adult patients implanted with a CI (Cochlear Ltd.). The automatically detected electrode contact locations were verified against locations marked by two experienced observers. The interobserver errors and the errors between the averaged locations and the automatically detected locations were calculated. The estimated contact positions were transformed to a cylindrical cochlear coordinate system, according to an international consensus, in which the insertion angles and the radius and elevation were measured. The linear correlation of the automatically detected electrode contact positions with the manually detected locations was high (R = 0.98 for the radius, and R = 1.00 for the insertion angle). The errors in radius and in insertion angle between the automatically detected locations and the manually detected locations were 0.12 mm and 1.7°. These errors were comparable to the interobserver errors. Geometrical measurements were in line with what is usually found in human cochleae. The mean insertion angle of the most apical electrode was 410° (range: 316° to 503°). The mean radius of the electrode contacts in the first turn of the cochlear spiral was 3.0 mm, and the mean radius of the remainder in the second turn was 1.7 mm. With implant geometry as prior knowledge, automatic analysis of high-resolution CT scans enables accurate localization of CI electrode contacts. The output of this method can be used to study the effect of CI positioning on hearing outcomes in more detail.


The Ross procedure offers excellent survival compared with mechanical aortic valve replacement in a real-world setting.
The ideal prosthesis for young patients requiring aortic valve replacement has not been defined to date. Although the Ross procedure provides excellent survival, its application is still limited. We compared the long-term survival after the Ross procedure with mechanical aortic valve replacement. All consecutive Ross procedures and mechanical aortic valve replacements performed between 1991 and 2008 at a single centre were analysed. Only adult patients between 18 and 50 years of age were included in the study. Survival and valve-related complications were evaluated. Furthermore, survival was compared with the age- and sex-matched Austrian population. A total of 159 Ross patients and 173 mechanical valve patients were included. The cumulative survival for the Ross procedure was significantly better, with survival rates of 96, 94 and 93% at 5, 10 and 15 years, respectively, in comparison to 90, 84 and 75% (P < 0.01) for patients with mechanical valves. A Cox regression analysis including patients' age, gender and valve type revealed age and the type of aortic valve replacement as independent significant factors influencing survival (for age, hazard ratio = 1.1, 95% confidence interval = 1.0-1.1, P = 0.03; and for valve type, hazard ratio = 2.6, 95% confidence interval = 1.2-5.8, P = 0.02). The observed survival was comparable to the expected standard survival for the Ross group but was significantly reduced in the mechanical valve group. In a real-world setting, the Ross procedure is associated with a long-term survival benefit in young adults in comparison to mechanical aortic valve replacement.


Effects of long-term use Topiramate on fertility and growth parameter in adult male rats.
A case control study was conducted to assess the effects of long-term ingestion of Topiramate on fertility, body and reproductive organ weight and level of sex hormones in Sprague-Dawley male rats. Ten adult male rats were exposed to Topiramate diets at a concentration of 100 mg /kg for 60 days. Another ten control male rats received vehicle (distilled water). After 24 hours of the last dose, animals were autopsied under light anesthesia. Several parameters including, body and reproductive organ weight, sperm count and motility, serum testosterone, FSH, levels of cholesterol, triglyceride, SGOT and SGPT were all measured. Assessment of pregnancies in females mixed with tested males was also measured. The results of histological, histometrical and biochemical profiles were compared to that of the control group, and the significance of these results was measured using student's ''t'' and Chi- square tests. Long-term ingestion of Topiramate for 60 days caused a significant decrease in spermatogenesis in seminiferous tubules of the testes. Sperm motility and density were also significantly reduced in cauda epididymides and testes of the treated group. The body weights and weight of reproductive organs (testes, epididymides, ventral prostate and seminal vesicle) were decreased considerably. Hormonal assay also showed significant decrease in testosterone levels. Testicular cell population dynamics also demonstrated a decrease in the number of both primary and secondary spermatocytes and spermatids in the treatment group. The number of female rats impregnated by male rats on long-term Topiramate diet had decreased. The number of implantations and the number of viable fetuses were also notably decreased in female rats impregnated by male rats ingested Topiramate. Histometry of reproductive organs confirmed these results. these results confirm that the long-term Topiramate ingestion produces adverse effects on fertility and reproductive system in adult male rat.


Early goal-directed therapy for severe sepsis and septic shock: A living systematic review.
Studies and meta-analyses conflict regarding the use of early goal-directed therapy (EGDT) for septic shock. We sought to clarify the conflict by performing a living systematic review and meta-regression. We performed a meta-analysis and explored heterogeneity with meta-regression. We conformed with the Preferred Reporting Items for Systematic Reviews and Meta-Analyses checklist and qualified strength of evidence with a Grading of Recommendations Assessment, Development and Evaluation profile. Overall, EGDT did not significantly reduce mortality compared with usual care (relative risk, 0.85; 95% confidence interval, 0.67-1.08); however, heterogeneity was substantial (I2=64%; 95% confidence interval, 12%-85%). Illness severity did not correlate with mortality reduction; however, there were significant correlations with control rate mortality and the strategy employed by the control group. Benefit was confined to trials with a control mortality greater than 35%. Compared with monitoring of lactate clearance and central venous pressure, EGDT mortality was higher. The benefit of EGDT is evident in populations with high mortality, in line with reported global mortality rates. In settings with low mortality the recent trials challenge the need for 6-hour goals; however, most patients in these trials met 3-hour resuscitation goals as defined by the Surviving Sepsis Campaign. In settings with higher mortality, EGDT or normalization of lactate/central venous pressure may be viable therapeutic options.


Choose to move: The motivational impact of autonomy support on motor learning.
Numerous studies in the motor learning domain have demonstrated learning advantages of self-controlled practice relative to yoked conditions. In separate lines of evidence in the social-psychological literature, findings show that providing participants with task-relevant autonomy support or minor incidental choices can result in superior outcomes when compared with conditions that thwart autonomy or do not offer choice. We hypothesized that motor learning could be enhanced by providing learners with choices - even if those choices are unrelated to task performance. In Experiment 1, two groups of participants practiced a golf putting task. While one group (the choice group) was able to select the color of golf balls (white, yellow, or orange) to be used in each upcoming block of 10 trials, participants in the second group (the yoked group) were provided with the same colored golf balls their choice-group counterparts had chosen. The results of a 24-h delayed retention test indicated significantly greater putting accuracy for the choice compared with the yoked group. Experiment 2 went one step further by asking choice group participants for their preferences regarding two issues unrelated to the practice task (balancing on a stabilometer): (1) which of two subsequent tasks (coincident timing or hand dynamometry) they wanted to perform and (2) which of two prints of paintings by Renoir they thought the investigator should hang on the laboratory wall. Yoked group participants were simply informed about which task they would perform afterwards and of which painting the experimenter would put on the wall. Balance learning was significantly more effective in the choice group on a retention test. Thus, self-controlled practice conditions can influence motor learning without providing task-relevant information, content, or strategic learning advantages. Self-controlled effects in motor learning may be motivational in nature, attributable to satisfaction of fundamental autonomy needs.


Ten Lessons for Good Practice for the INHERIT Triple Win: Health, Equity, and Environmental Sustainability.
The world's challenges of climate change, damage to ecosystems, and social and health inequalities require changes in human behaviours at every level of organisation, among governments, business, communities, and individuals. An important question is how behaviour change can be enabled and supported at the scale and speed required. The research reported in this paper describes important lessons for good practice in changing contexts to modify behaviours for a triple win for health, equity and environmental sustainability. Authors synthesised learning from qualitative, quantitative and cost benefit evaluations of 15 case studies conducted in 12 countries in Europe. The case studies address ways of living (green spaces and energy efficient housing), moving (active transport) and consuming (healthy and sustainable diets) that support the triple win. Ten lessons for good practice were identified. These include bringing a triple win mindset to policy and practice in planning interventions, with potential to improve environmental sustainability, health and equity at the same time. The lessons for good practice are intended to support governmental and non-governmental actors, practitioners and researchers planning to work across sectors to achieve mutual benefits for health and environmental sustainability and in particular to benefit poorer and more socio-economically disadvantaged groups.


Leucine carboxyl methyltransferase-1 is necessary for normal progression through mitosis in mammalian cells.
Protein phosphatase 2A (PP2A) is a multifunctional phosphatase that plays important roles in many cellular processes including regulation of cell cycle and apoptosis. Because PP2A is involved in so many diverse processes, it is highly regulated by both non-covalent and covalent mechanisms that are still being defined. In this study we have investigated the importance of leucine carboxyl methyltransferase-1 (LCMT-1) for PP2A methylation and cell function. We show that reduction of LCMT-1 protein levels by small hairpin RNAs causes up to a 70% reduction in PP2A methylation in HeLa cells, indicating that LCMT-1 is the major mammalian PP2A methyltransferase. In addition, LCMT-1 knockdown reduced the formation of PP2A heterotrimers containing the Balpha regulatory subunit and, in a subset of the cells, induced apoptosis, characterized by caspase activation, nuclear condensation/fragmentation, and membrane blebbing. Knockdown of the PP2A Balpha regulatory subunit induced a similar amount of apoptosis, suggesting that LCMT-1 induces apoptosis in part by disrupting the formation of PP2A(BalphaAC) heterotrimers. Treatment with a pan-caspase inhibitor partially rescued cells from apoptosis induced by LCMT-1 or Balpha knockdown. LCMT-1 knockdown cells and Balpha knockdown cells were more sensitive to the spindle-targeting drug nocodazole, suggesting that LCMT-1 and Balpha are important for spindle checkpoint. Treatment of LCMT-1 and Balpha knockdown cells with thymidine dramatically reduced cell death, presumably by blocking progression through mitosis. Consistent with these results, homozygous gene trap knock-out of LCMT-1 in mice resulted in embryonic lethality. Collectively, our results indicate that LCMT-1 is important for normal progression through mitosis and cell survival and is essential for embryonic development in mice.


Oral Pathology in Forensic Investigation.
Forensic odontology is the subdiscipline of dentistry which analyses dental evidence in the interest of justice. Oral pathology is the subdiscipline of dentistry that deals with the pathology affecting the oral and maxillofacial regions. This subdiscipline is utilized for identification through oral and maxillofacial pathologies with associated syndromes, enamel rod patterns, sex determination using exfoliative cytology, identification from occlusal morphology of teeth, and deoxyribonucleic acid profiling from teeth. This subdiscipline is also utilized for age estimation studies which include Gustafson's method, incremental lines of Retzius, perikymata, natal line formation in teeth, neonatal line, racemization of collagen in dentin, cemental incremental lines, thickness of the cementum, and translucency of dentin. Even though the expertise of an oral pathologist is not taken in forensic investigations, this paper aims to discuss the role of oral pathology in forensic investigation.


Orally administered allyl sulfides from garlic ameliorate murine colitis.
Inflammatory bowel disease (IBD) is an incurable disease which affects millions of people. Garlic (Allium sativum) preparations have been traditionally employed for the treatment of diseases affecting the digestive tract. Here, we have investigated the effect of diallyl sulfide (DAS) and diallyl disulfide (DADS), two garlic-derived sulfur compounds, on intestinal inflammation in vivo as well as in intestinal isolated cells. Colitis was induced in mice by intracolonic administration of dinitrobenzenesulfonic acid. Intestinal damage was assessed by evaluating colon weight/colon length ratio and by histology. Murine intestinal epithelial cells stimulated with IFN-γ were used to evaluate the possible in vitro DAS and DADS anti-inflammatory effects. DAS and DADS, given for two consecutive days after DNBS administration, reduced inflammation and damage. In IFN-γ-stimulated intestinal epithelial cells, DADS reduced IP-10 and IL-6 levels, while DAS inhibited nitric oxide production and STAT-1 expression. DAS and DADS exert therapeutic effects in the DNBS model of colitis. The actions of these compounds on the production of IP-10, IL-6, hydrogen sulfide or nitric oxide and on the expression of STAT-1 observed in intestinal cells stimulated with IFN-γ, might explain the protective action of DAS and DADS in experimental IBD.


Stimulus-response compatibility tests of implicit preference for food and body image to identify people at risk for disordered eating: a validation study.
The aim of this study was to incorporate implicit measures of relevant social cognition into eating disorder research. Fifty-three females diagnosed with an eating disorder (ED), and 41 at-risk females were recruited via ED support websites, along with 23 healthy females for comparison. Computerised online tests assessing subconscious normative ideal body image (IBI-BIAT) and personalised self-identification body image (PBI-BIAT) associations and food preferences (FP-AAT) were administered, followed by the modified version of the Eating Disorder Examination Questionnaire (EDE-Q). Anthropometric data, age, need for social approval, self-reported measures of self-esteem, normative perception and body image satisfaction were recorded. Self-reported diagnosed ED status was corroborated with BMI and EDE-Q. Diagnostic performance of the implicit measures was assessed with ROC analysis. Those diagnosed with ED showed significantly stronger automatic preferences for and self-identification with thin body image, compared to healthy females, but no differences were found in food preferences. The IBI-BIAT showed better diagnostic power than PBI-BIAT, correctly classifying 87% of the diagnosed participants. No correlation was found between IBI-BIAT and the explicit measures. The results suggest that the underlying subconscious social cognitive factors of pathological eating are linked to body image, not to food items per se.


Paroxysmal nocturnal hemoglobinuria: deficiency in factor H-like functions of the abnormal erythrocytes.
Erythrocytes from patients with paroxysmal nocturnal hemoglobinuria (PNH) contained a subpopulation that lacked membrane-associated Factor H-like activity present on normal human erythrocytes. Initial deposition of C3b on the erythrocytes was effected using a fluid phase C3 convertase. The cells were then treated with fluorescein-labeled C3 and the cell-bound C3 convertase. Analysis utilizing the fluorescence-activated cell sorter revealed two distinct cell populations, one of which was highly fluorescent, indicating a large number of C3b molecules per cell. Only this population (43%) was susceptible to lysis (44%) when exposed to acidified serum before C3b deposition. The less fluorescent population resembled normal human erythrocytes and was not affected by prior treatment with acidified serum. Since C3b deposition occurred almost exclusively on the complement-sensitive cells in the PNH erythrocyte population, these cells could be examined for the Factor H-like regulatory activities without prior isolation. These functions include enhancement of inactivation of erythrocyte-bound C3b by Factor I and acceleration of the decay of erythrocyte-bound C3 convertase, C3b,Bb. It was found that C3b on PNH erythrocytes was 100-fold less susceptible to inactivation by Factor I than C3b on normal human erythrocytes. The half-life at 22 degrees C of C3b,Bb on PNH erythrocytes was threefold greater than on normal human erythrocytes and similar to that of the enzyme bound to particles that do not possess Factor H-like activity. These observations suggest that the abnormal susceptibility of PNH erythrocytes to lysis by complement is due to a functional deficiency in one or more of the Factor H-like proteins present on normal human erythrocytes.


Sodium channel dysfunction in intractable childhood epilepsy with generalized tonic-clonic seizures.
Mutations in SCN1A, the gene encoding the brain voltage-gated sodium channel alpha(1) subunit (Na(V)1.1), are associated with genetic forms of epilepsy, including generalized epilepsy with febrile seizures plus (GEFS+ type 2), severe myoclonic epilepsy of infancy (SMEI) and related conditions. Several missense SCN1A mutations have been identified in probands affected by the syndrome of intractable childhood epilepsy with generalized tonic-clonic seizures (ICEGTC), which bears similarity to SMEI. To test whether ICEGTC arises from molecular mechanisms similar to those involved in SMEI, we characterized eight ICEGTC missense mutations by whole-cell patch clamp recording of recombinant human SCN1A heterologously expressed in cultured mammalian cells. Two mutations (G979R and T1709I) were non-functional. The remaining alleles (T808S, V983A, N1011I, V1611F, P1632S and F1808L) exhibited measurable sodium current, but had heterogeneous biophysical phenotypes. Mutant channels exhibited lower (V983A, N1011I and F1808L), greater (T808S) or similar (V1611F and P1632S) peak sodium current densities compared with wild-type (WT) SCN1A. Three mutations (V1611F, P1632S and F1808L) displayed hyperpolarized conductance-voltage relationships, while V983A exhibited a strong depolarizing shift in the voltage dependence of activation. All mutants except T808S had hyperpolarized shifts in the voltage dependence of steady-state channel availability. Three mutants (V1611F, P1632S and F1808L) exhibited persistent sodium current ranging from approximately 1-3% of peak current amplitude that was significantly greater than WT-SCN1A. Several mutants had impaired slow inactivation, with V983A showing the most prominent effect. Finally, all of the functional alleles exhibited reduced use-dependent channel inhibition. In summary, SCN1A mutations associated with ICEGTC result in a wide spectrum of biophysical defects, including mild-to-moderate gating impairments, shifted voltage dependence and reduced use dependence. The constellation of biophysical abnormalities for some mutants is distinct from those previously observed for GEFS+ and SMEI, suggesting possible, but complex, genotype-phenotype correlations.


Bartonella strains in small mammals from Dhaka, Bangladesh, related to Bartonella in America and Europe.
Ecological and bacteriologic observations of small mammals captured in Dhaka, Bangladesh, indicated that Bartonella infections occurred in high prevalence among lesser bandicoot rats (Bandicota bengalensis), black rats (Rattus rattus), and house shrews (Suncus murinus). Sequence analysis of the citrate synthase gene of Bartonella isolates showed that small mammals in Bangladesh harbored a diverse assemblage of strains. Some cultures were genetically related to Bartonella elizabethae, a species identified from a human patient in the United States. Sequences of some other cultures from Bandicota and Rattus rats were identical to sequences of cultures from domestic rats in France, Portugal, and the United States. The finding of Bartonella species in a high proportion of the mammalian samples from Dhaka suggests the need to study whether these agents might be responsible for human cases of febrile illness of unknown etiology in Bangladesh and elsewhere in south Asia.


Comparison of antireflection surfaces based on two-dimensional binary gratings and thin-film coatings.
A comparison of antireflection surfaces based on the two-dimensional binary gratings and thin-film coatings is presented. First, a two-dimensional hybrid binary grating is proposed and analyzed by use of a vector-based implementation of the rigorous coupled-wave analysis method. The optimum parameters of the structure are determined and the effects that changing them have on spectral characteristics of the structure are studied. Then this structure is compared with multilayer thin-film antireflection filters. These filters are designed by genetic algorithm and needle methods, which are powerful methods for multilayer filter design. The comparison results show that the sensitivity of the grating to changes in the incident wavelength is high. However, a reflectance of the order of 10(-3)% at the design wavelength can be achieved. The sensitivity of designed antireflection thin-film filters to wavelength changes is lower, however, and the minimum achievable reflectance is higher.


Racial/ethnic differences in surgical outcomes in veterans following knee or hip arthroplasty.
The utilization of joint arthroplasty for knee or hip osteoarthritis varies markedly by patient race/ethnicity. Because of concerns about surgical risk, black patients are less willing to consider this treatment. There are few published race/ethnicity-specific data on joint arthroplasty outcomes. The present study was undertaken to examine racial/ethnic differences in mortality and morbidity following elective knee or hip arthroplasty. Using information from the Veterans Administration National Surgical Quality Improvement Program database, data on 12,108 patients who underwent knee arthroplasty and 6,703 patients who underwent hip arthroplasty over a 5-year period were analyzed. Racial/ethnic differences were determined using prospectively collected data on patient characteristics, procedures, and short-term outcomes. The main outcome measures were risk-adjusted 30-day mortality and complication rates. Adjusted rates of both non-infection-related and infection-related complications after knee arthroplasty were higher among black patients compared with white patients (relative risk [RR] 1.50, 95% confidence interval [95% CI] 1.08-2.10 and RR 1.42, 95% CI 1.06-1.90, respectively). Hispanic patients had a significantly higher risk of infection-related complications after knee arthroplasty (RR 1.64, 95% CI 1.08-2.49) relative to otherwise similar white patients. Race/ethnicity was not significantly associated with the risk of non-infection-related complications (RR 0.97, 95% CI 0.68-1.38 in blacks; RR 1.18, 95% CI 0.60-2.30 in Hispanics) or infection-related complications (RR 1.27, 95% CI 0.91-1.78 in blacks; RR 1.22, 95% CI 0.63-2.36 in Hispanics) after hip arthroplasty. The overall 30-day mortality was 0.6% following knee arthroplasty and 0.7% following hip arthroplasty, with no significant differences by race/ethnicity observed for either procedure. Although absolute risks of complication are low, our findings indicate that, after adjustment, black patients have significantly higher rates of infection-related and non-infection-related complications following knee arthroplasty, compared with white patients. In addition, adjusted rates of infection-related complications after knee arthroplasty are higher in Hispanic patients than in white patients. Such differences between ethnic groups are not seen following hip arthroplasty. These groups do not appear to differ significantly in terms of post-arthroplasty mortality rates.


Screening for severe anaemia in pregnancy in Kenya, using pallor examination and self-reported morbidity.
Severe anaemia in pregnancy is an important preventable cause of maternal and perinatal morbidity and mortality. Different methods of screening for severe anaemia in pregnancy were evaluated in a 2-phased study conducted in Kilifi, Kenya. In phase 1 (in 1994/95), pallor testing was evaluated alone and in addition to raised respiratory/pulse rates: 1787 pregnant women were examined by one of 2 midwives. Sensitivities for detecting severe anaemia (haemoglobin < 7 g/dL) were 62% and 69% and specificities 87% and 77%, respectively for each of the midwives. Addition of high pulse rate increased sensitivity to 77% and 81%, but specificity reduced to 60% and 51%, respectively. In phase 2, following qualitative in-depth work, a screening questionnaire was developed. An algorithm based on screening questions had 80% sensitivity and 40% specificity. Midwife pallor-assessment was conducted following the screening questionnaire. In this phase (conducted in 1997), the midwife performed very highly in detecting severe anaemia, achieving sensitivity of 84% and specificity of 92%. Spending a few minutes asking women questions may have improved the ability to interpret pallor findings. This study demonstrates the value of pallor testing and raises alternative approaches to improving it.


Calcium depletion in frog muscle tubules: the decline of calcium current under maintained depolarization.
1. Ca2+ currents in frog skeletal muscle fibres were studied with a voltage-clamp technique. Under membrane depolarization maintained for several seconds, Ca2+ current was found to decline with time constants of 0.2-2 sec when [Ca2+]o = 10 mM. 2. Ca2+ currents are diminished by nifedipine, D-600, tetracaine and Ni2+. 3. When peak current is diminished by making the membrane potential positive, by block with drugs or by substituting the relatively less permeant Mn2+ for Ca2+ then the rate of decline is diminished also. When peak current is increased by recording at relatively negative membrane potentials or by substituting for Ca2+ the more permeant ions Ba2+ or Sr2+, then the rate of decline is increased in proportion. Evidently, the size of the current determines the rate of decline. 4. Decline of current is greatly slowed in isotonic Ca2+ saline or when the [Ca2+]o is buffered by the organic anion malate. These findings indicate that the decline of current arises from Ca2+ depletion in an extracellular compartment, most probably the transverse tubules. On this basis, an analysis of Ca2+ current decline and recovery leads to the following conclusions. 5. Ca2+ current flows almost entirely across the membranes of the transverse tubules. 6. After allowing for the tortuosity of the tubular network, the apparent diffusion coefficient for Ca2+ in the transverse tubules is about 2.6 X 10(-6) cm2/sec, three times less than the diffusion coefficient for K+ in the transverse tubules and about three times less than the diffusion coefficient for Ca2+ in free solution. 7. The transverse tubule lumen does not appear to have a large Ca2+-buffering capacity in the millimolar range. At [Ca2+]o = 10 mM, the tubule lumen binds less than 0.6 dissociable Ca2+ ions for every free ion.


Sub-Saharan Africa preparedness and response to the COVID-19 pandemic: A perspective of early career African scientists.
Emerging highly transmissible viral infections such as SARS-CoV-2 pose a significant global threat to human health and the economy. Since its first appearance in December 2019 in the city of Wuhan, Hubei province, China, SARS-CoV-2 infection has quickly spread across the globe, with the first case reported on the African continent, in Egypt on February 14 th, 2020. Although the global number of COVID-19 infections has increased exponentially since the beginning of the pandemic, the number of new infections and deaths recorded in African countries have been relatively modest, suggesting slower transmission dynamics of the virus on the continent, a lower case fatality rate, or simply a lack of testing or reliable data. Notably, there is no significant increase in unexplained pneumonias or deaths on the continent which could possibly indicate the effectiveness of interventions introduced by several African governments. However, there has not yet been a comprehensive assessment of sub-Saharan Africa's (SSA) preparedness and response to the COVID-19 pandemic that may have contributed to prevent an uncontrolled outbreak so far. As a group of early career scientists and the next generation of African scientific leaders with experience of working in medical and diverse health research fields in both SSA and resource-rich countries, we present a unique perspective on the current public health interventions to fight COVID-19 in Africa. Our perspective is based on extensive review of the available scientific publications, official technical reports and announcements released by governmental and non-governmental health organizations as well as from our personal experiences as workers on the COVID-19 battlefield in SSA. We documented public health interventions implemented in seven SSA countries including Uganda, Kenya, Rwanda, Cameroon, Zambia, South Africa and Botswana, the existing gaps and the important components of disease control that may strengthen SSA response to future outbreaks.


Studies of metabolism of round spermatids: glucose as unfavorable substrate.
The exposure of spermatids to glucose in the absence of pyruvate and lactate resulted in an extremely low energy charge. The adenosine 5'-triphosphate (ATP) level rapidly declined and the fructose 1,6-bisphosphate (FBP) and triose levels increased. These changes were prevented by the addition of pyruvate or lactate. The levels of ATP and FBP were inversely correlated. In cells exposed to glucose, FBP did not flow appreciably through the step of glyceraldehyde 3-phosphate dehydrogenase (GA3PDH). The lactate level did not change. However, when pyruvate or lactate was administered to cells exposed to glucose, the FBP level declined rapidly. This drop was accompanied by a commensurate increase in lactate. In these cells, pyruvate transport was suppressed, and the pyruvate taken up by these cells was mostly oxidized in the tricarboxylic acid (TCA) cycle without its being reduced to lactate. In this case, the ATP level increased, but to a level still lower than existed before exposure to glucose. Furthermore, when kinetic studies on the activity of 6-phosphofructokinase (PFK) were carried out, PFK appeared to be fully activated at intracellular levels of fructose 6-phosphate, ATP and adenosine 5'-monophosphate (AMP). These results indicate that the rate of glucose metabolism in glycolysis depends heavily on the energy charge. In cells exposed to glucose, the sugar does not flow appreciably through the glycolytic pathway due to inhibition of GA3PDH. Moreover, the ATP level cannot be recovered fully from the lowest level by the addition of pyruvate or lactate.(ABSTRACT TRUNCATED AT 250 WORDS)


Natural-scene statistics predict how the figure-ground cue of convexity affects human depth perception.
The shape of the contour separating two regions strongly influences judgments of which region is "figure" and which is "ground." Convexity and other figure-ground cues are generally assumed to indicate only which region is nearer, but nothing about how much the regions are separated in depth. To determine the depth information conveyed by convexity, we examined natural scenes and found that depth steps across surfaces with convex silhouettes are likely to be larger than steps across surfaces with concave silhouettes. In a psychophysical experiment, we found that humans exploit this correlation. For a given binocular disparity, observers perceived more depth when the near surface's silhouette was convex rather than concave. We estimated the depth distributions observers used in making those judgments: they were similar to the natural-scene distributions. Our findings show that convexity should be reclassified as a metric depth cue. They also suggest that the dichotomy between metric and nonmetric depth cues is false and that the depth information provided many cues should be evaluated with respect to natural-scene statistics. Finally, the findings provide an explanation for why figure-ground cues modulate the responses of disparity-sensitive cells in visual cortex.


A critical review of brand and generic alendronate for the treatment of osteoporosis.
Compare in vitro and in vivo characteristics and clinical outcomes of brand and generic alendronate. Relevant search terms were input into Medline ("alendronate" AND "generic" up to August 5, 2013) and any abstracts deemed possibly relevant selected for full paper review and abstraction. Multicentre, randomized, placebo-controlled Phase III clinical trials of substantial size and duration have established the anti-fracture efficacy and safety of brand amino-bisphosphonates. For regulatory approval, generic versions of brand drugs need to demonstrate bioequivalence in young, healthy volunteers and have similar dissolution times. While the potency and amount of active drug within generic formulations must be identical to the brand, differences are permitted in the excipients. Significant differences in tablet disintegration time among different versions of generic and brand alendronate have been reported. Rapidly disintegrating alendronate pills may increase oesophageal bioadhesion and adverse event risk. Oesophageal-bound alendronate or slow disintegrating alendronate tablets may be made inert and ineffective by subsequently ingested food or drink. Investigations have reported a lower persistence to therapy with generic brands of alendronate as compared to brand bisphosphonates and patients switched from brand to generic alendronate have increased adverse event rates and losses in bone mineral density. Numerous differences exist between brand and generic alendronate including: disintegration time, bioadhesion to the oesophagus, patient persistence to therapy, adverse event incidence, and maintenance of bone mineral density. Generic forms of alendronate warrant closer clinical study before they are ascribed the clinical effectiveness and tolerability of brand alendronate.


Human nonspecific suppressive lymphokines.
Since the term "lymphokine" first appeared in print over 20 years ago, a tremendous number of these soluble mediators of the immune system have been described. Within the past few years, many human nonspecific suppressive lymphokines have been identified. This review discusses the historical basis of immunologic suppression and suppressor factors. Later reports describing suppressive human lymphokines are then grouped into four categories: primarily stimulatory lymphokines that also mediate certain suppressive activities, suppressive lymphokines produced during altered states of immunity, suppressive lymphokines produced by exogenously stimulated lymphocytes, and suppressive lymphokines produced by unstimulated lymphocytes. Recent work I have been involved in focusing on the human suppressive lymphokine soluble suppressor factor (SSF) is also discussed.


Themis sets the signal threshold for positive and negative selection in T-cell development.
Development of a self-tolerant T-cell receptor (TCR) repertoire with the potential to recognize the universe of infectious agents depends on proper regulation of TCR signalling. The repertoire is whittled down during T-cell development in the thymus by the ability of quasi-randomly generated TCRs to interact with self-peptides presented by major histocompatibility complex (MHC) proteins. Low-affinity TCR interactions with self-MHC proteins generate weak signals that initiate 'positive selection', causing maturation of CD4- or CD8αβ-expressing 'single-positive' thymocytes from CD4(+)CD8αβ(+) 'double-positive' precursors. These develop into mature naive T cells of the secondary lymphoid organs. TCR interaction with high-affinity agonist self-ligands results in 'negative selection' by activation-induced apoptosis or 'agonist selection' of functionally differentiated self-antigen-experienced T cells. Here we show that positive selection is enabled by the ability of the T-cell-specific protein Themis to specifically attenuate TCR signal strength via SHP1 recruitment and activation in response to low- but not high-affinity TCR engagement. Themis acts as an analog-to-digital converter translating graded TCR affinity into clear-cut selection outcome. By dampening mild TCR signals Themis increases the affinity threshold for activation, enabling positive selection of T cells with a naive phenotype in response to low-affinity self-antigens.


Albumin-based nanoconjugates for targeted delivery of therapeutic oligonucleotides.
Nanoparticle-based delivery has become an important strategy to advance siRNA and antisense oligonucleotides into clinical reality. However, limited biodistribution of nanoparticles and the toxicity of some nanocarriers restrict the wider application of this strategy. To address these issues we aimed to construct oligonucleotide delivery systems which are non-cytotoxic and smaller than typical nanoparticles. Thus, a morpholino oligonucleotide was conjugated to a tumor-targeting RGD peptide, and then, multiple RGD-oligo conjugates were linked to a single molecule of human serum albumin via a reductively responsive linkage. The resultant nanoconjugates showed uniform and monodispersed size distribution with a diameter of 13 nm. A single nanoconjugate molecule contains 15 oligonucleotides as well as 15 targeting ligands on the surface of albumin. The nanoparticle demonstrated 61-fold enhancement in receptor-specific cellular delivery of oligonucleotides in integrin-expressing tumor cells compared to the non-targeted control nanoconjugates and were able to robustly enhance functional activity of the oligonucleotide at low nanomolar concentrations without causing cytotoxicity. Due to their small size, the targeted nanoconjugates could penetrate deeply and distribute throughout 3-D tumor spheroids, whereas the conventional nanoparticles with sizes over 300 nm could only deliver to the cells on the surface of the tumor spheroids. As a result of their greater cellular delivery, smaller size, and lack of cytotoxicity compared to conventional nanoparticles, the multivalent nanoconjugates may provide an effective tool for targeting oligonucleotides to tumors and other diseased tissues.


The topology of higher-order complexes associated with brain hubs in human connectomes.
Higher-order connectivity in complex systems described by simplexes of different orders provides a geometry for simplex-based dynamical variables and interactions. Simplicial complexes that constitute a functional geometry of the human connectome can be crucial for the brain complex dynamics. In this context, the best-connected brain areas, designated as hub nodes, play a central role in supporting integrated brain function. Here, we study the structure of simplicial complexes attached to eight global hubs in the female and male connectomes and identify the core networks among the affected brain regions. These eight hubs (Putamen, Caudate, Hippocampus and Thalamus-Proper in the left and right cerebral hemisphere) are the highest-ranking according to their topological dimension, defined as the number of simplexes of all orders in which the node participates. Furthermore, we analyse the weight-dependent heterogeneity of simplexes. We demonstrate changes in the structure of identified core networks and topological entropy when the threshold weight is gradually increased. These results highlight the role of higher-order interactions in human brain networks and provide additional evidence for (dis)similarity between the female and male connectomes.


Genetic risk score analysis indicates migraine with and without comorbid depression are genetically different disorders.
Migraine and major depressive disorder (MDD) are comorbid, moderately heritable and to some extent influenced by the same genes. In a previous paper, we suggested the possibility of causality (one trait causing the other) underlying this comorbidity. We present a new application of polygenic (genetic risk) score analysis to investigate the mechanisms underlying the genetic overlap of migraine and MDD. Genetic risk scores were constructed based on data from two discovery samples in which genome-wide association analyses (GWA) were performed for migraine and MDD, respectively. The Australian Twin Migraine GWA study (N = 6,350) included 2,825 migraine cases and 3,525 controls, 805 of whom met the diagnostic criteria for MDD. The RADIANT GWA study (N = 3,230) included 1,636 MDD cases and 1,594 controls. Genetic risk scores for migraine and for MDD were used to predict pure and comorbid forms of migraine and MDD in an independent Dutch target sample (NTR-NESDA, N = 2,966), which included 1,476 MDD cases and 1,058 migraine cases (723 of these individuals had both disorders concurrently). The observed patterns of prediction suggest that the 'pure' forms of migraine and MDD are genetically distinct disorders. The subgroup of individuals with comorbid MDD and migraine were genetically most similar to MDD patients. These results indicate that in at least a subset of migraine patients with MDD, migraine may be a symptom or consequence of MDD.


Near- to far-field transformation in the aperiodic Fourier modal method.
This paper addresses the task of obtaining the far-field spectrum for a finite structure given the near-field calculated by the aperiodic Fourier modal method in contrast-field formulation (AFMM-CFF). The AFMM-CFF efficiently calculates the solution to Maxwell's equations for a finite structure by truncating the computational domain with perfectly matched layers (PMLs). However, this limits the far-field solution to a narrow strip between the PMLs. The Green's function for layered media is used to extend the solution over the whole super- and substrate. The approach is validated by applying it to the problem of scattering from a cylinder for which the analytical solution is available. Moreover, a numerical study is conducted on the accuracy of the approximate far-field computed with the super-cell Fourier modal method by using the AFMM-CFF with near- to far-field transformation as a reference.


First Report of Leaf Blight Caused by Nigrospora sphaerica on Curcuma in China.
Curcuma (family Zingiberaceae) is commonly cultivated for the use of rhizomes within traditional Chinese medicines. In October 2009 and 2010, severe leaf blight was observed on Curcuma wenyujin Y.H. Chen & C. Ling (4) in fields located in Ruian, China. The area of cultivation in Ruian encompasses 90% of the production in Zhejiang Province. Disease incidence was approximately 90% of plants observed in affected fields. Early symptoms were yellow-to-brown, irregular-shaped lesions on the leaf margin or tip. After several days, lesions expanded along the mid-vein until the entire leaf was destroyed. Blighted leaves turned grayish to dark brown and withered, and severely affected plants died. Eight fungal isolates were recovered from symptomatic C. wenyujin leaves, collected from eight different fields, on potato dextrose agar (PDA). These fungal colonies were initially white, becoming light to dark gray and produced black, spherical to subspherical, single-celled conidia (14 to 17 × 12 to 15 μm), which were borne on a hyaline vesicle at the tip of the conidiophores. On the basis of these morphological features, the isolates appeared to be similar to Nigrospora sphaerica (2). Strain ZJW-1 was selected as a representative for molecular identification. Genomic DNA was extracted from the isolate, and the internal transcribed spacer (ITS) region of the ribosomal DNA (ITS1-5.8S-ITS2) was amplified using ITS1 (5'-TCCGTAGGTGAACCTGCGG-3') and ITS4 (5'-TCCTCCGCTTATTGATATGC-3') primers (3). The ITS region was further cloned and sequenced (GenBank Accession No. JF738028) and was 99% identical to N. sphaerica (GenBank Accession No. FJ478134.1). On the basis of morphological data and the ITS rDNA sequence, the isolate was determined to be N. sphaerica. Pathogenicity tests were conducted on four leaves of four C. wenyujin plants by placing agar pieces (5 mm in diameter) from 8-day-old cultures on pushpin-wounded leaves. An equal number of control plants were wounded and inoculated with noncolonized PDA agar pieces. Plants were placed in moist chambers at 25°C with a 12-h photoperiod. Brown-to-black lesions were observed on wounded leaves after 3 days and expanded to an average of 56 × 40 mm 15 days after inoculation. No symptoms developed on the control leaves. The pathogen was reisolated from the margins of necrotic tissues but not from the controls. The pathogen has been reported as a leaf pathogen on several hosts worldwide (1). To our knowledge, this is the first report of N. sphaerica as a leaf pathogen of C. wenyujin in China. Future research will focus primarily on management of this disease. References: (1) D. F. Farr and A. Y. Rossman. Fungal Databases, Systematic Mycology and Microbiology Laboratory, USDA-ARS, Retrieved from http://nt.ars-grin.gov/fungaldatabases/ , March 31, 2011. (2) E. W. Mason. Trans. Brit. Mycol. Soc. 12:152, 1927. (3) T. J. White et al. PCR Protocols: A Guide to Methods and Applications. Academic Press, San Diego, 1990. (4) J. Zhao et al. Molecules 15:7547, 2010.


Outcome instruments: rationale for their use.
The number of outcome instruments available for use in orthopaedic observational studies has increased dramatically in recent years. Properly developed and tested outcome instruments provide a very useful tool for orthopaedic research. Criteria have been proposed to assess the measurement properties and quality of health-status instruments. Unfortunately, not all instruments are developed with use of strict quality criteria. In this article, we discuss these quality criteria and provide the reader with a tool to help select the most appropriate instrument for use in an observational study. We also review the steps for future use of outcome instruments, including the standardization of their use in orthopaedic research.


Predictors of parent-child relationships that support physical activity in Mexican-American families.
Family-based physical activity (PA) interventions would benefit from research that identifies how to build support for PA among family members. This study examined the extent to which relationships of encouragement to do PA, and co-engagement in PA, exist among Mexican-American parents and children, and sought to identify individual, relational, and household factors associated with these dimensions of support. Participants were 224 Mexican-origin adults, with at least one child aged 5-20 years, participating in a larger study conducted between 2008 and 2010. In baseline surveys, adult participants enumerated the names and attributes of their family and kin; this study focuses on 455 parent-child dyads, nested in 118 households. Parental encouragement of PA in their children was found in about half of dyads, and in 20 % of dyads children encouraged parents. Encouragement relationships were highly reciprocal. Reciprocal parent-child encouragement was also positively associated with co-participation in PA; the latter found in just 17 % of dyads. Results indicated that relational, individual, and socio-cultural attributes were associated with PA support among parents and children, and provide insights into how these relationships might be fostered within Mexican-American families.


MICA polymorphism: biology and importance in cancer.
The major histocompatibility complex class I polypeptide-related sequence A gene (MICA) encodes a membrane-bound protein acting as a ligand to stimulate an activating receptor, NKG2D, expressed on the surface of essentially all human natural killer (NK), γδ T and CD8(+) αβ T cells. MICA protein is absent from most cells but can be induced by infections and oncogenic transformation and is frequently expressed in epithelial tumors. Upon binding to MICA, NKG2D activates cytolytic responses of NK and γδ T cells against infected and tumor cells expressing MICA. Therefore, membrane-bound MICA acts as a signal during the early immune response against infection or spontaneously arising tumors. On the other hand, human tumor cells spontaneously release a soluble form of MICA, causing the downregulation of NKG2D and in turn severe impairment of the antitumor immune response of NK and CD8(+) T cells. This is considered to promote tumor immune evasion and also to compromise host resistance to infections. MICA is the most polymorphic non-classical class I gene. A possible association of MICA polymorphism with genetic predisposition to different cancer types has been investigated in candidate gene-based studies. Two genome-wide association studies have identified loci in MICA that influence susceptibility to cervical neoplasia and hepatitis C virus-induced hepatocellular carcinoma, respectively. Given the current level of interest in the field of MICA gene, we discuss the genetics and biology of the MICA gene and the role of its polymorphism in cancer. Gaps in our understanding and future research needs are also discussed.


Short-term effects of ambient particles on mortality in the elderly: results from 28 cities in the APHEA2 project.
Within the framework of the APHEA2 (Air Pollution on Health: a European Approach) project, the effects of ambient particles on mortality among persons > or = 65 yrs were investigated. Daily measurements for particles with a 50% cut-off aerodynamic diameter of 10 microm (PM10) and black smoke (BS), as well as the daily number of deaths among persons > or = 65 yrs of age, from 29 European cities, have been collected. Data on other pollutants and meteorological variables, to adjust for confounding effects and data on city characteristics, to investigate potential effect modification, were also recorded. For individual city analysis, generalised additive models extending Poisson regression, using a locally weighted regression (LOESS) smoother to control for seasonal effects, were applied. To combine individual city results and explore effect modification, second stage regression models were applied. The per cent increase (95% confidence intervals), associated with a 10 microg x m(-3) increase in PM10, in the elderly daily number of deaths was 0.8%, (0.7-0.9%) and the corresponding number for BS was 0.6%, (0.5-0.8%). The effect size was modified by the long-term average levels of nitrogen dioxide (higher levels were associated with larger effects), temperature (larger effects were observed in warmer countries), and by the proportion of the elderly in each city (a larger proportion was associated with higher effects). These results indicate that ambient particles have effects on mortality among the elderly, with relative risks comparable or slightly higher than those observed for total mortality and similar effect modification patterns. The effects among the older persons are of particular importance, since the attributable number of events will be much larger, compared to the number of deaths among the younger population.


Water Availability for Shale Gas Development in Sichuan Basin, China.
Unconventional shale gas development holds promise for reducing the predominant consumption of coal and increasing the utilization of natural gas in China. While China possesses some of the most abundant technically recoverable shale gas resources in the world, water availability could still be a limiting factor for hydraulic fracturing operations, in addition to geological, infrastructural, and technological barriers. Here, we project the baseline water availability for the next 15 years in Sichuan Basin, one of the most promising shale gas basins in China. Our projection shows that continued water demand for the domestic sector in Sichuan Basin could result in high to extremely high water stress in certain areas. By simulating shale gas development and using information from current water use for hydraulic fracturing in Sichuan Basin (20,000-30,000 m(3) per well), we project that during the next decade water use for shale gas development could reach 20-30 million m(3)/year, when shale gas well development is projected to be most active. While this volume is negligible relative to the projected overall domestic water use of ∼36 billion m(3)/year, we posit that intensification of hydraulic fracturing and water use might compete with other water utilization in local water-stress areas in Sichuan Basin.


The Midlands Trial of Empirical Amiodarone versus Electrophysiology-guided Interventions and Implantable Cardioverter-defibrillators (MAVERIC): a multi-centre prospective randomised clinical trial on the secondary prevention of sudden cardiac death.
MAVERIC was a randomised clinical trial designed to test the possibility of prospectively identifying patients who would benefit most from the implantable cardioverter-defibrillator (ICD) by electrophysiology (EP) study in the context of secondary prevention of sudden cardiac death (SCD) through comparing EP-guided interventions (anti-arrhythmic drugs, coronary revascularization, and ICD) against empirical amiodarone therapy. Two hundred and fourteen survivors of sustained ventricular tachycardia (VT), ventricular fibrillation (VF) or SCD were randomized to either treatment strategy, pre-stratified for haemodynamic status at index event, and followed up for a median of 5 years. Of the 106 amiodarone arm patients, 89 (84%) received the drug and 5 (5%) received an ICD after crossing over. Of the 108 EP arm patients, 31 (29%) received an ICD, 46 (43%) received anti-arrhythmic drugs only (mainly amiodarone or sotalol) and 18 (17%) received coronary revascularization but no ICD. No significant differences in survival or arrhythmia recurrence existed between the two treatment arms after 6 years. However, ICD recipients had a lower mortality than non-ICD recipients, regardless of allocated treatment (hazard ratio=0.54, p=0.0391). Prospective selection of patients to receive the ICD by EP study did not improve survival compared with empirical amiodarone therapy among survivors of VT, VF or SCD, whereas ICD implantation improved survival regardless of allocated treatment. On this basis, routine EP study has no role in the management of such patients, who should be offered empirical ICD therapy according to the results of other secondary prevention ICD trials.


Vibrodissociation of neurons from rodent brain slices to study synaptic transmission and image presynaptic terminals.
Mechanical dissociation of neurons from the central nervous system has the advantage that presynaptic boutons remain attached to the isolated neuron of interest. This allows for examination of synaptic transmission under conditions where the extracellular and postsynaptic intracellular environments can be well controlled. A vibration-based technique without the use of proteases, known as vibrodissociation, is the most popular technique for mechanical isolation. A micropipette, with the tip fire-polished to the shape of a small ball, is placed into a brain slice made from a P1-P21 rodent. The micropipette is vibrated parallel to the slice surface and lowered through the slice thickness resulting in the liberation of isolated neurons. The isolated neurons are ready for study within a few minutes of vibrodissociation. This technique has advantages over the use of primary neuronal cultures, brain slices and enzymatically isolated neurons including: rapid production of viable, relatively mature neurons suitable for electrophysiological and imaging studies; superior control of the extracellular environment free from the influence of neighboring cells; suitability for well-controlled pharmacological experiments using rapid drug application and total cell superfusion; and improved space-clamp in whole-cell recordings relative to neurons in slice or cell culture preparations. This preparation can be used to examine synaptic physiology, pharmacology, modulation and plasticity. Real-time imaging of both pre- and postsynaptic elements in the living cells and boutons is also possible using vibrodissociated neurons. Characterization of the molecular constituents of pre- and postsynaptic elements can also be achieved with immunological and imaging-based approaches.


Inhibition of Candida albicans growth by brominated furanones.
Candida albicans is the most virulent Candida species of medical importance, which presents a great threat to immunocompromised individuals such as HIV patients. Currently, there are only four classes of antifungal agents available for treating fungal infections: azoles, polyenes, pyrimidines, and echinocandins. The fast spread of multidrug resistant C. albicans strains has increased the demand for new antifungal drugs. In this study, we demonstrate the antifungal activity of brominated furanones on C. albicans. Studying the structure and activity of this class of furanones reveals that the exocyclic vinyl bromide conjugated with the carbonyl group is the most important structural element for fungal inhibition. Furthermore, gene expression analysis using DNA microarrays showed that 3 microg/mL of 4-bromo-5Z-(bromomethylene)-3-butylfuran-2-one (BF1) upregulated 32 C. albicans genes with functions of stress response, NADPH dehydrogenation, and small-molecule transport, and repressed 21 genes involved mainly in cell-wall maintenance. Interestingly, only a small overlap is observed between the gene expression changes caused by the representative brominated furanone (BF1) in this study and other antifungal drugs reported in literature. This result suggests that brominated furanones and other antifungal drugs may target different fungal proteins or genes. The existence of such new targets provides an opportunity for developing new agents to control fungal pathogens which are resistant to currently available drugs.


Optic disc measurements in full term infants.
The objectives of this study were to measure optic disc size in full term infants and to determine whether this value is influenced by sex or birth weight. Retinal images from a cohort of full term infants admitted to a tertiary perinatal centre were obtained using a retinal camera. Optic disc size was measured by carefully delineating the outline with a cursor using image analysis software. MEDLINE was then systematically searched to compare the data with other published articles. 35 images of left and right eyes from 35 infants were assessed. An image from one eye per patient was then chosen for analysis. The following results were found: mean birth weight 3050±706 g; mean gestation 38.9±1.4 weeks. Mean optic disc area was 1.26±0.23 mm(2); mean vertical diameter was 1.37±0.15 mm; and mean horizontal diameter was 1.14±0.12 mm. The vertical diameter of the optic disc was significantly longer than the horizontal diameter (p<0.0001). Birth weight and sex did not influence the size of the optic disc in term infants. There were no differences in optic disc measurements between male and female infants and between low birth weight and normal birth weight infants.


Software configurable optical test system: a computerized reverse Hartmann test.
A software configurable optical test system (SCOTS) based on the geometry of the fringe reflection or phase measuring deflectometry method was developed for rapidly, robustly, and accurately measuring large, highly aspherical shapes such as solar collectors and primary mirrors for astronomical telescopes. In addition to using phase shifting methods for data collection and reduction, we explore the test from the point view of performing traditional optical testing methods, such as Hartmann or Hartmann-Shack tests, in a reverse way. Using this concept, the slope data calculation and unwrapping in the test can also be done with centroiding and line-scanning methods. These concepts expand the test to work in more general situations where fringe illumination is not practical. Experimental results show that the test can be implemented without complex calibration for many applications by taking the geometric advantage of working near the center curvature of the test part. The results also show that the test has a large dynamic range, can achieve measurement accuracy comparable with interferometric methods, and can provide a good complement to interferometric tests in certain circumstances. A variation of this method is also useful for measuring refractive optics and optical systems. As such, SCOTS provides optical manufacturers with a new tool for performing quantitative full field system evaluation.


GCN2-dependent metabolic stress is essential for endotoxemic cytokine induction and pathology.
Activated inflammatory macrophages can express indoleamine 2,3-dioxygenase (IDO) and thus actively deplete their own tryptophan supply; however, it is not clear how amino acid depletion influences macrophage behavior in inflammatory environments. In this report, we demonstrate that the stress response kinase GCN2 promotes macrophage inflammation and mortality in a mouse model of septicemia. In vitro, enzymatic amino acid consumption enhanced sensitivity of macrophages to the Toll-like receptor 4 (TLR4) ligand lipopolysaccharide (LPS) with significantly increased interleukin 6 (IL-6) production. Tryptophan withdrawal induced the stress response proteins ATF4 and CHOP/GADD153; however, LPS stimulation rapidly enhanced expression of both proteins. Moreover, LPS-driven cytokine production under amino acid-deficient conditions was dependent on GCN2, as GCN2 knockout (GCN2KO) macrophages had a significant reduction of cytokine gene expression after LPS stimulation. To test the in vivo relevance of these findings, monocytic-lineage-specific GCN2KO mice were challenged with a lethal dose of LPS intraperitoneally (i.p.). The GCN2KO mice showed reduced inflammatory responses, with decreased IL-6 and IL-12 expression correlating with significant reduction in animal mortality. Thus, the data show that amino acid depletion stress signals (via GCN2) synergize with proinflammatory signals to potently increase innate immune responsiveness.


HMGA1 reprograms somatic cells into pluripotent stem cells by inducing stem cell transcriptional networks.
Although recent studies have identified genes expressed in human embryonic stem cells (hESCs) that induce pluripotency, the molecular underpinnings of normal stem cell function remain poorly understood. The high mobility group A1 (HMGA1) gene is highly expressed in hESCs and poorly differentiated, stem-like cancers; however, its role in these settings has been unclear. We show that HMGA1 is highly expressed in fully reprogrammed iPSCs and hESCs, with intermediate levels in ECCs and low levels in fibroblasts. When hESCs are induced to differentiate, HMGA1 decreases and parallels that of other pluripotency factors. Conversely, forced expression of HMGA1 blocks differentiation of hESCs. We also discovered that HMGA1 enhances cellular reprogramming of somatic cells to iPSCs together with the Yamanaka factors (OCT4, SOX2, KLF4, cMYC - OSKM). HMGA1 increases the number and size of iPSC colonies compared to OSKM controls. Surprisingly, there was normal differentiation in vitro and benign teratoma formation in vivo of the HMGA1-derived iPSCs. During the reprogramming process, HMGA1 induces the expression of pluripotency genes, including SOX2, LIN28, and cMYC, while knockdown of HMGA1 in hESCs results in the repression of these genes. Chromatin immunoprecipitation shows that HMGA1 binds to the promoters of these pluripotency genes in vivo. In addition, interfering with HMGA1 function using a short hairpin RNA or a dominant-negative construct blocks cellular reprogramming to a pluripotent state. Our findings demonstrate for the first time that HMGA1 enhances cellular reprogramming from a somatic cell to a fully pluripotent stem cell. These findings identify a novel role for HMGA1 as a key regulator of the stem cell state by inducing transcriptional networks that drive pluripotency. Although further studies are needed, these HMGA1 pathways could be exploited in regenerative medicine or as novel therapeutic targets for poorly differentiated, stem-like cancers.


Physical activity program for patients with dementia and their relative caregivers: randomized clinical trial in Primary Health Care (AFISDEMyF study).
The aging of the population has led to the increase of chronic diseases, especially dementia and cardiovascular diseases, and it has become necessary for their relatives to dedicate more time in caregiving.The objective in the first phase of this study is to evaluate the effectiveness of a Primary Health Care procedure to increase the physical activity of people with dementia and their relative caregivers. Also the effect on the cognitive state and cardiovascular risk will be assessed. Clinical, multicentric and randomized trial. A simple random sampling to select 134 patients diagnosed with dementia will be carried out. After contacting their relatives, his/her participation in the trial will be requested. A basal assessment will be made and the participants will be asigned to control or intervention group (1:1). The main measure will be the assessment of physical activity (podometer and 7-PAR) in patients and caregivers. In patients with dementia: ADAS-cog, functional degree and cardiovascular risk. In caregivers: cardiovascular risk, general health and quality of life. For 3 months, participants will receive instructions to do physical activity with an adapted program. This program will be designed and applied by Primary Health Care professionals in patients with dementia and their caregivers. The control group will receive regular care. An intention-to-treat analysis will be carried out by comparing the observed differences between basal, 6 and 12 months measures. Change in the mean of daily steps assessed with the podometer and 7-PAR will be the main result. If the main hypothesis is confirmed, it could be useful to improve the cognitive state of patients with dementia, as well as the cardiovascular risk of all of them. The results can be good to improve technical features of the devices that register the physical activity in the patients with dementia, and it could facilitate its commercialization. Clinical Trials.gov Identifier: NCT02044887.


Site-specific inter-strand cross-links of DNA duplexes.
We report the development of technology that allows inter-strand coupling across various positions within one turn of DNA. Four 2'-modified nucleotides were synthesized as protected phosphoramidites and incorporated into DNA oligonucleotides. The modified nucleotides contain either 5-atom or 16-atom linker components, with either amine or carboxylic acid functional groups at their termini, forming 10 or 32 atom (11 or 33 bond) linkages. Chemical coupling of the amine and carboxylate groups in designed strands resulted in the formation of an amide bond. Coupling efficiency as a function of trajectory distance between the individual linker components was examined. For those nucleotides capable of forming inter-strand cross-links (ICLs), coupling yields were found to depend on temperature, distance, and linker length, enabling several approaches that can control regioselective linkage. In the most favorable cases, the coupling yields are quantitative. Spectroscopic measurements of strands that were chemically cross-linked indicate that the global structure of the DNA duplex does not appear to be distorted from the B form after coupling. Thermal denaturing profiles of those strands were shifted to somewhat higher temperatures than those of their respective control duplexes. Thus, the robust amide ICLs formed by this approach are site-specific, do not destabilize the rest of the duplex, and only minimally perturb the secondary structure.