A novel protein kinase family in Plasmodium falciparum is differentially transcribed and secreted to various cellular compartments of the host cell.
Processes at the surface of Plasmodium falciparum-infected erythrocytes such as antigenic variation and cytoadhesion may be modulated by active signalling between host and parasite. Potential candidates for this role include the putative kinases of the FIKK family. The novel Apicomplexa-specific FIKK gene has expanded in P. falciparum to 20 sequence-related members distributed between 11 chromosomes. Specific antibodies raised against different members indicated that most FIKK proteins locate to punctate foci in the erythrocyte cytoplasm that colocalized with Maurer's clefts proteins. One FIKK member dissociates at the trophozoite stage from the Maurer's clefts and relocates with the erythrocyte cytoskeleton. Another FIKK protein, despite having a PEXEL motif, remains located within the parasite. FIKK proteins possess the essential residues for phosphotransferase activity. We show that protein kinase activity was detected in immunoprecipitates obtained with two anti-FIKK antibodies. Quantitative PCR analysis revealed differential gene transcription of the FIKK paralogues in asexual blood stages parasites. We observed significant changes in the transcription pattern between parasites with different adhesion phenotypes. Our data suggest a role of FIKK proteins in the remodelling of the erythrocyte surface and reveal the existence of an adaptable parasite system able to sense intra- and possibly extracellular changes.


Cost effectiveness of dabrafenib as a first-line treatment in patients with BRAF V600 mutation-positive unresectable or metastatic melanoma in Canada.
To evaluate the cost effectiveness of dabrafenib versus dacarbazine and vemurafenib as first-line treatments in patients with BRAF V600 mutation-positive unresectable or metastatic melanoma from a Canadian healthcare system perspective. A partitioned-survival analysis model with three mutually exclusive health states (pre-progression, post-progression, and dead) was used. The proportion of patients in each state was calculated using survival distributions for progression-free and overall survival derived from pivotal trials of dabrafenib and vemurafenib. For each treatment, expected progression-free, post-progression, overall, and quality-adjusted life-years (QALYs), and costs were calculated. Costs were based on list prices, a clinician survey, and published sources. A 5-year time horizon was used in the base case. Costs (in 2012 Canadian dollars [CA$]) and QALYs were discounted at 5% annually. Deterministic and probabilistic sensitivity analyses were conducted. Dabrafenib was estimated to yield 0.2055 more QALYs at higher cost than dacarbazine. The incremental cost-effectiveness ratio was CA$363,136/QALY. In probabilistic sensitivity analyses, at a threshold of CA$200,000/QALY, there was an 8.2% probability that dabrafenib is cost effective versus dacarbazine. In deterministic sensitivity analyses, cost effectiveness was sensitive to survival distributions, utilities, and time horizon, with the hazard ratio for overall survival for dabrafenib versus dacarbazine being the most sensitive parameter. Assuming a class effect for efficacy of BRAF inhibitors, dabrafenib was dominant versus vemurafenib (less costly, equally effective), reflecting its assumed lower daily cost. Assuming no class effect, dabrafenib yielded 0.0486 more QALYs than vemurafenib. At a threshold of CA$200,000/QALY, dabrafenib is unlikely to be cost effective compared with dacarbazine. It is not possible to make reliable conclusions regarding the relative cost effectiveness of dabrafenib versus vemurafenib based on available information.


Using ultrasound to quantify tongue shape and movement characteristics.
Objective : Previous experimental studies have demonstrated abnormal lingual articulatory patterns characterizing cleft palate speech. Most articulatory information to date has been collected using electropalatography, which records the location and size of tongue-palate contact but not the tongue shape. The latter type of data can be provided by ultrasound. The present paper aims to describe ultrasound tongue imaging as a potential tool for quantitative analysis of tongue function in speakers with cleft palate. A description of the ultrasound technique as applied to analyzing tongue movements is given, followed by the requirements for quantitative analysis. Several measures are described, and example calculations are provided. Measures : Two measures aim to quantify overuse of tongue dorsum in cleft palate articulations. Crucially for potential clinical applications, these measures do not require head-to-transducer stabilization because both are based on a single tongue curve. The other three measures compare sets of tongue curves, with the aim to quantify the dynamics of tongue displacement, token-to-token variability in tongue position, and the extent of separation between tongue curves for different speech sounds. Conclusions : All measures can be used to compare tongue function in speakers with cleft palate before and after therapy, as well as to assess their performance against that in typical speakers and to help in selecting more effective treatments.


The British Columbia Healthy Connections Project: findings on socioeconomic disadvantage in early pregnancy.
Maternal exposure to socioeconomic disadvantage increases the risk of child injuries and subsequent child developmental and mental health problems - particularly for young mothers. To inform early intervention planning, this research therefore aimed to describe the health and social adversities experienced by a cohort of girls and young women in early pregnancy in British Columbia (BC), Canada. Participants were recruited for the BC Healthy Connections Project (BCHCP), a randomized controlled trial examining the effectiveness of Nurse-Family Partnership, a home visitation program, in improving child and maternal outcomes. Baseline data were collected from 739 participants on trial entry. Participants were selected on the basis of preparing to parent for the first time and experiencing socioeconomic disadvantage. Analyses involved descriptive statistics and age-group comparisons. Most participants reported having low income (84%), having limited education (52%) and being single (91%) at trial entry. Beyond these eligibility criteria, other health and social adversities included: housing instability (52%); severe anxiety or depression (47%); other diagnosed mental disorders (22%); prenatal nicotine and cannabis use (27 and 21%); physical health problems (20%); child maltreatment when younger (56%); and intimate partner violence recently (50%). As well, few (29%) had received income assistance entitlements. More than two thirds (70%) were experiencing four or more forms of adversity. Age-group differences were observed for cognitive functioning, being single, low income, limited education, psychological distress and service use (p-value ≤0.05). This cohort was selected on the basis of socioeconomic disadvantage. Yet all participants were experiencing substantial added adversities - at higher rates than other Canadians. Furthermore, despite Canada's public programs, these pregnant girls and young women were not being adequately reached by social services. Our study adds new data to inform early intervention planning, suggesting that unacceptably high levels of socioeconomic disadvantage exist for some young British Columbians. Therefore greater health and social supports and services are warranted for these young mothers and their children. Registered August 24, 2012 with ClinicalTrials.gov Identifier: NCT01672060 . Active not recruiting.


Colocalization of taurine- and cysteine sulfinic acid decarboxylase-like immunoreactivity in the hippocampus of the rat.
It is proposed that taurine is an inhibitory neurotransmitter/neuromodulator in the CNS. The present study localized taurine-containing neurons within the rat hippocampus with the use of a monoclonal antibody against conjugated taurine (Tau2) in conjunction with an antiserum against cysteine sulfinic acid decarboxylase (CSADC), a synthesizing enzyme for taurine. Taurine-like immunoreactivity Tau-LI) and CSADC-LI were colocalized in neurons of the dentate gyrus, CA1(/CA2), CA3, and CA4. Of all the cells examined, pyramidal basket cells within the granule cell layer of the dentate gyrus were most intensely stained with both Tau2 and CSADC. Granule cells were also double-labeled with Tau-LI and CSADC-LI. Cell nuclei and dendrites in the CA1 region stained more intensely with Tau2 than somata. CSADC-LI was colocalized with Tau-LI within these neurons. Light staining with both Tau2 and the CSADC antiserum was inconsistently present in CA3 and CA4 neurons and was found to be highly dependent on the type of fixation and delay to fixation. Tau-LI was more consistently present in increased numbers of neurons in CA3 when glutaraldehyde was added to the paraformaldehyde fixative solution. Hippocampi which were immersion-fixed in paraformaldehyde following a 0-, 6-, or 24-hour postmortem delay exhibited a lack of Tau2 staining in the CA3 region in the majority of animals studied, similar to some paraformaldehyde perfusion-fixed rats. These studies suggest that taurine was present in the majority of neurons within the major cell layers of the rat hippocampus, but Tau-LI was more easily lost from neurons in the CA3 region following delay to fixation. The localization of Tau-LI in excitatory neurons such as granule cells and pyramidal cells is not consistent with its proposed inhibitory transmitter role. However, the prominent Tau2 staining in dendrites of the CA1 region provides anatomical support for the hypothesis that taurine may be released from dendrites in the CA1 region and may function as a neuromodulator of calcium flux in these pyramidal neurons.


Absorption-coefficient-determination method for particulate materials.
A method is presented for determining the optical absorption coefficient, or the imaginary refractive index, of particulate material that has been collected from aerosols or hydrosols by means of filtration. The method, based on the Kubelka-Munk theory of diffuse reflectance, is nondestructive and requires no other knowledge of the sample than the amount present, the specific gravity, and an estimate of the real index of refraction. The theoretical development of the method is discussed along with an analysis of photometric and gravimetric errors. We test the method by comparing results obtained for powdered didymium glass with measurements made before the glass was crushed. An example of the method's application to the determination of the absorption coefficient of atmospheric dust at UV, visible, and near-IR wavelengths is also presented.


Structure of receptive fields in area 3b of primary somatosensory cortex in the alert monkey.
We investigated the two-dimensional structure of area 3b neuronal receptive fields (RFs) in three alert monkeys. Three hundred thirty neurons with RFs on the distal fingerpads were studied with scanned, random dot stimuli. Each neuron was stimulated continuously for 14 min, yielding 20,000 response data points. Excitatory and inhibitory components of each RF were determined with a modified linear regression algorithm. Analyses assessing goodness-of-fit, repeatability, and generality of the RFs were developed. Two hundred forty-seven neurons yielded highly repeatable RF estimates, and most RFs accounted for a large fraction of the explainable response of each neuron. Although the area 3b RF structures appeared to be continuously distributed, certain structural generalities were apparent. Most RFs (94%) contained a single, central region of excitation and one or more regions of inhibition located on one, two, three, or all four sides of the excitatory center. The shape, area, and strength of excitatory and inhibitory RF regions ranged widely. Half the RFs contained almost evenly balanced excitation and inhibition. The findings indicate that area 3b neurons act as local spatiotemporal filters that are maximally excited by the presence of particular stimulus features. We believe that form and texture perception are based on high-level representations and that area 3b is an intermediate stage in the processes leading to these representations. Two possibilities are considered: (1) that these high-level representations are basically somatotopic and that area 3b neurons amplify some features and suppress others, or (2) that these representations are highly transformed and that area 3b effects a step in the transformation.


Electrochemistry-mass spectrometry unveils the formation of reactive triclocarban metabolites.
Triclocarban (3,4,4'-trichlorocarbanilide, TCC) is a widely used antibacterial agent in personal care products and is frequently detected as an environmental pollutant in waste waters and surface waters. In this study, we report novel reactive metabolites potentially formed during biotransformation of TCC. The oxidative metabolism of TCC has been predicted using an electrochemical cell coupled online to liquid chromatography and electrospray ionization mass spectrometry. The electrochemical oxidation unveils the fact that hydroxylated metabolites of TCC may form reactive quinone imines. Moreover, a so-far unknown dechlorinated and hydroxylated TCC metabolite has been identified. The results were confirmed by in vitro studies with human and rat liver microsomes. The reactivity of the newly discovered quinone imines was demonstrated by their covalent binding to glutathione and macromolecules, using β-lactoglobulin A as a model protein. The results regarding the capability of the electrochemical cell to mimic the oxidative metabolism of TCC are discussed. Moreover, the occurrence of reactive metabolites is compared with findings from earlier in vivo studies and their relevance in vivo is argued.


The N-terminal propeptide of the precursor to sporamin acts as a vacuole-targeting signal even at the C terminus of the mature part in tobacco cells.
An asparagine-proline-isoleucine-arginine-leucine (NPIRL) and its related sequences in the N-terminal propeptides (NTPP) of several plant vacuolar proteins, including that of sporamin from sweet potato (SPO) function as vacuole-targeting determinants in a manner that is distinct from the vacuole-targeting determinant in the CTPPs of other plant vacuolar proteins. When the mutant precursor to sporamin, SPO-NTPP (in which NTPP was moved to the C terminus of the mature part), was expressed in tobacco (Nicotiana tabacum) cells, the pro-form was efficiently targeted to the vacuole and the NTPP was cleaved off. Unlike the results obtained with the wild-type precursor, substitution of the NPIRL sequence in the C-terminally located NTPP to asparagine-proline-glycine-arginine-leucine in the SPO-isoleucine-28-to-glycine mutant resulted in missorting of less than 20% of the pro-form to the medium. Unlike the vacuolar transport of SPO-NTPP, the vacuolar transport of SPO-isoleucine-28-to-glycine was strongly inhibited by 33 microM wortmannin, which is similar to the C-terminal propeptide-mediated vacuolar transport. These results suggest that the vacuole-targeting function of the NPIRL sequence is not strictly dependent on its location at the N terminus of a protein and that the C-terminally located mutant NTPP acquired some physicochemical properties of the C-terminal vacuole-targeting sequence.


Environmental pollution: a tangible risk for NAFLD pathogenesis.
The liver is crucial for human life, and the health of this organ often mirrors the health of the individual. The liver can be the target of several diseases, the most prevalent of which, as a consequence of development and changes in human lifestyles, is the nonalcoholic fatty liver disease (NAFLD). NAFLD is a multifactorial disease that embraces many histo-pathologic conditions and is highly linked to metabolic derangements. Technological progress and industrialization have also had the consequence of releasing pollutants in the environment, for instance pesticides or solvents, as well as by-products of discharge, such as the particulate matter. In the last decade, a growing body of evidence has emerged, shedding light on the potential impact of environmental pollutants on liver health and, in particular, on NAFLD occurrence. These contaminants have a great steatogenic potential and need to be considered as tangible NAFLD risk factors. There is an urgent need for a deeper comprehension of their molecular mechanisms of action, as well as for new lines of intervention to reduce their worldwide diffusion. This review wishes to sensitize the community to the effects of several environmental pollutants on liver health.


Clinical outcomes of patients infected with HIV through use of injected drugs compared to patients infected through sexual transmission: late presentation, delayed anti-retroviral treatment and higher mortality.
To compare patients who acquired HIV infection through use of injected drugs (HIV-IDU) with patients who acquired HIV by sexual transmission (HIV-ST) in terms of late presentation (LP), delay in anti-retroviral treatment (ART) initiation, virological and immunological response to ART, mortality and progression to AIDS. Prospective multi-centre cohort study of HIV-infected subjects naive to ART at entry (Cohort of the Spanish HIV Research Network: CoRIS). Thirty-one centres from the Spanish public health-care system. A total of 9355 patients were included (1064 HIV-IDU and 8291 HIV-ST) during 2004-13. We compared LP (defined as presentation for care with a CD4 cell count < 350/μl and/or AIDS-defining illness), delayed ART initiation (defined as initiating treatment more than 6 months after the date when treatment was indicated by the guidelines, or not initiating treatment at all when it was indicated), virological and immunological response to ART (defined as viral load < 50 HIV-1 RNA copies/ml and a CD4 count increase of at least 100 cells/μl, respectively, after 1 year of treatment), mortality and progression to AIDS in HIV-IDU and HIV-ST. Compared with HIV-ST, HIV-IDU had higher risk of LP [odds ratio (OR) = 1.76; 95% confidence interval (CI) = 1.41-2.18], delayed ART initiation (OR 1.87; 95% CI = 1.46-2.40) and higher mortality [hazard ratio (HR) = 1.43; 95% CI = 1.03-2.01] and risk of progression to AIDS [subhazard ratio (SHR) = 1.68; 95% CI = 1.29-2.18]. Virological suppression due to ART was lower in HIV-IDU than in patients with HIV-ST only among patients without hepatitis C virus (HCV) infection [adjusted OR (aOR) = 0.59; 95% CI = 0.36-0.95]; among patients with HCV infection, virological suppression due to ART did not show significant differences between HIV-IDU and HIV-ST. There were no significant differences in immunological response after adjusting by HCV (aOR = 0.74; 95% CI = 0.52-1.06). In Spain, patients who acquire HIV infection through use of injected drugs appear to have a higher risk of late presentation, delayed initiation of anti-retroviral treatment and progression to AIDS and death than patients who acquire HIV by sexual transmission.


Chondroitin sulfate A-adhering Plasmodium falciparum-infected erythrocytes express functionally important antibody epitopes shared by multiple variants.
Acquired protection from Plasmodium falciparum placental malaria, a major cause of maternal, fetal, and infant morbidity, is mediated by IgG specific for the P. falciparum erythrocyte membrane protein 1 variant VAR2CSA. This protein enables adhesion of P. falciparum-infected erythrocytes to chondroitin sulfate A in the intervillous space. Although interclonal variation of the var2csa gene is lower than that among var genes in general, VAR2CSA-specific Abs appear to target mainly polymorphic epitopes. This has raised doubts about the feasibility of VAR2CSA-based vaccines. We used eight human monoclonal IgG Abs from affinity-matured memory B cells of P. falciparum-exposed women to study interclonal variation and functional importance of Ab epitopes among placental and peripheral parasites from East and West Africa. Most placental P. falciparum isolates were labeled by several mAbs, whereas peripheral isolates from children were essentially nonreactive. The mAb reactivity of peripheral isolates from pregnant women indicated that some were placental, whereas others had alternative sequestration foci. Most of the mAbs were comparable in their reactivity with bound infected erythrocytes (IEs) and recombinant VAR2CSA and interfered with IE and/or VAR2CSA binding to chondroitin sulfate A. Pair-wise mAb combinations were more inhibitory than single mAbs, and all of the mAbs together was the most efficient combination. Each mAb could opsonize IEs for phagocytosis, and a combination of the eight mAbs caused phagocytosis similar to that of plasma IgG-opsonized IEs. We conclude that functionally important Ab epitopes are shared by the majority of polymorphic VAR2CSA variants, which supports the feasibility of VAR2CSA-based vaccines against placental malaria.


Host cytoskeleton in respiratory syncytial virus assembly and budding.
Respiratory syncytial virus (RSV) is one of the major pathogens responsible for lower respiratory tract infections (LRTI) in young children, the elderly, and the immunosuppressed. Currently, there are no antiviral drugs or vaccines available that effectively target RSV infections, proving a significant challenge in regards to prevention and treatment. An in-depth understanding of the host-virus interactions that underlie assembly and budding would inform new targets for antiviral development.Current research suggests that the polymerised form of actin, the filamentous or F-actin, plays a role in RSV assembly and budding. Treatment with cytochalasin D, which disrupts F-actin, has been shown to inhibit virus release. In addition, the actin cytoskeleton has been shown to interact with the RSV matrix (M) protein, which plays a central role in RSV assembly. For this reason, the interaction between these two components is hypothesised to facilitate the movement of viral components in the cytoplasm and to the budding site. Despite increases in our knowledge of RSV assembly and budding, M-actin interactions are not well understood. In this review, we discuss the current literature on the role of actin cytoskeleton during assembly and budding of RSV with the aim to integrate disparate studies to build a hypothetical model of the various molecular interactions between actin and RSV M protein that facilitate RSV assembly and budding.


Comparative Assessment of Procedure Cost and Outcomes Between Guidewire and Crossing Device Strategies to Cross Peripheral Artery Chronic Total Occlusions.
The aim of this study was to assess actual procedural costs and outcomes comparing wire-catheter and dedicated chronic total occlusion (CTO) device strategies to cross peripheral artery CTOs. Peripheral artery CTO interventions are frequently performed, but there are limited data on actual procedural costs and outcomes comparing wire-catheter and dedicated CTO devices. The XLPAD (Excellence in Peripheral Artery Disease Intervention) registry (NCT01904851) was accessed to retrospectively compare cost and 30-day and 12-month outcomes of wire-catheter and crossing device strategies for treatment of infrainguinal peripheral artery CTO. Of all 3,234 treated lesions, 42% (n = 1,362) were CTOs in 1,006 unique patients. Wire-catheter approaches were used in 82% of CTOs, whereas dedicated CTO devices were used in 18% (p < 0.0001). CTO crossing device use was associated with significantly higher technical success (74% vs. 65%; p < 0.0001) and mean procedure cost ($7,800.09 vs. $4,973.24; p < 0.0001). Because 12-month repeat revascularization (11.3% vs. 17.2%; p = 0.02) and amputation rates (2.8% vs. 8.5%; p = 0.002) in the CTO crossing device arm were lower compared with the wire-catheter group, the net cost for an initial CTO crossing device strategy was $423.80 per procedure. An initial wire-catheter approach to cross a peripheral artery CTO is most frequently adopted. The use of dedicated CTO crossing devices provides significantly higher technical success and lower reintervention and amputation rates, at a net cost of $423.80 per procedure at 12 months.


Association of Visit-to-Visit Variability of Systolic Blood Pressure With Cardiovascular Disease and Mortality in Primary Care Chinese Patients With Type 2 Diabetes-A Retrospective Population-Based Cohort Study.
This study aimed to evaluate the impact of visit-to-visit variability (VVV) of systolic blood pressure (SBP) on cardiovascular disease (CVD) and mortality among primary care Chinese patients with type 2 diabetes mellitus (T2DM). A retrospective cohort study was conducted in 124,105 Chinese adult primary care patients with T2DM and without prior diagnosed CVD from August 2008 to December 2009. The VVV of SBP was evaluated using SDs of SBP over 24 months. The risks of CVD and all-cause mortality associated with variability in SBP were evaluated using Cox proportional hazards regression. Subgroup analysis was conducted by the stratification of age, sex, duration of diabetes, the presence of chronic kidney disease, baseline SBP and trend, and the number and class of antihypertensive drugs. A positive linear relationship between the VVV of SBP and the first incidence of CVD and all-cause mortality was identified over a median follow-up time of 39.5 months. Patients with a low SD of SBP of <5 mmHg had the lowest risks of CVD and all-cause mortality, and patients with an SD of SBP of ≥10 mmHg had significantly higher risks. For every 1 SD increase in the SD of SBP, the risks of CVD, all-cause mortality, and the composite of both events increased by 2.9% (95% CI 2.4-3.4%), 4.0% (95% CI 3.5-4.6%), and 3.4% (95% CI 3.0-3.8%), respectively. A direct linear relationship was also observed in all selected subgroups. SBP variability, irrespective of the mean SBP level, is a potential predictor for the development of CVD and all-cause mortality in patients with diabetes. In addition to monitoring BP targets for their patients with diabetes, clinicians should also remain vigilant about the visit-to-visit fluctuation of BP.


Automated medical avatar animation for warfighter mission simulation.
Virtual representations of human internal anatomy are important for military applications such as protective equipment design, injury severity prediction, thermal analysis, and physiological simulations. High-fidelity volumetric models based on imaging data are typically in static postures and difficult to use in simulations of realistic mission scenarios. This study aimed to investigate a hybrid approach to reposition medical avatars that preserves internal anatomy but allows rapid repositioning of full three-dimensional (3D) meshes. A software framework that accepts a medical avatar in a 3D tetrahedral mesh format representing 72 organs and tissues with an articulated skeleton was developed. The skeleton is automatically resized and associated to the avatar using rigging and skinning algorithms inspired by computer animation techniques. Military relevant motions were used for animations. A motion retargeting algorithm was implemented to apply animation to avatars of various sizes, and a motion blending algorithm was implemented to smoothly transition between movements. These algorithms were incorporated into a path generation tool that accepts initial, intermediate, and final coordinates of a multisegment action along with the specific motion for each segment to synthesize a realistic compound set of movements comprising the animation. The developed pipeline for dynamic repositioning of medical avatars was demonstrated. Various complex motions were automatically animated. Retargeting was demonstrated on models of varying sizes. Movements along a path were animated to demonstrate smooth motion transitions. Animation of the full 3D avatar mesh ran in real time on a standard desktop personal computer. The repositioning algorithm successfully preserved the shape and volume of rigid structures such as bone. The developed software leverages techniques from various disciplines to create a hybrid approach enabling real-time 3D mesh repositioning appropriate for use in simulated military missions using avatars containing a complete anatomy representation. The workflow is largely automated, enabling rapid evaluation of new mission scenarios.


Insulin needs after CD3-antibody therapy in new-onset type 1 diabetes.
Type 1 diabetes mellitus is a T-cell-mediated autoimmune disease that leads to a major loss of insulin-secreting beta cells. The further decline of beta-cell function after clinical onset might be prevented by treatment with CD3 monoclonal antibodies, as suggested by the results of a phase 1 study. To provide proof of this therapeutic principle at the metabolic level, we initiated a phase 2 placebo-controlled trial with a humanized antibody, an aglycosylated human IgG1 antibody directed against CD3 (ChAglyCD3). In a multicenter study, 80 patients with new-onset type 1 diabetes were randomly assigned to receive placebo or ChAglyCD3 for six consecutive days. Patients were followed for 18 months, during which their daily insulin needs and residual beta-cell function were assessed according to glucose-clamp-induced C-peptide release before and after the administration of glucagon. At 6, 12, and 18 months, residual beta-cell function was better maintained with ChAglyCD3 than with placebo. The insulin dose increased in the placebo group but not in the ChAglyCD3 group. This effect of ChAglyCD3 was most pronounced among patients with initial residual beta-cell function at or above the 50th percentile of the 80 patients. In this subgroup, the mean insulin dose at 18 months was 0.22 IU per kilogram of body weight per day with ChAglyCD3, as compared with 0.61 IU per kilogram with placebo (P<0.001). In this subgroup, 12 of 16 patients who received ChAglyCD3 (75 percent) received minimal doses of insulin (< or =0.25 IU per kilogram per day) as compared with none of the 21 patients who received placebo. Administration of ChAglyCD3 was associated with a moderate "flu-like" syndrome and transient symptoms of Epstein-Barr viral mononucleosis. Short-term treatment with CD3 antibody preserves residual beta-cell function for at least 18 months in patients with recent-onset type 1 diabetes.


Patellofemoral Pain in Adolescence and Adulthood: Same Same, but Different?
The mainstay of patellofemoral pain (PFP) treatment is exercise therapy, often in combination with adjunct treatments such as patient education, orthoses, patella taping and stretching, making the intervention multimodal in nature. The vast majority of randomised controlled trials among patients with PFP have investigated the effect of treatment among adults (>18 years of age). So, while systematic reviews and meta-analyses provide evidence-based recommendations for treating PFP, these recommendations are largely based upon the trials in adults. In the present article, we have summarised the findings on the efficacy of multimodal treatment (predominantly exercise) from the three largest trials concerning patients with PFP, focusing on the long-term success-rate 1 year after receiving multimodal treatment, and with a particular focus on the success rate across the different age groups, including both adolescents, young adults and adults. The results of this paper show that there appears to be a difference in the success rate between adolescents and adults, despite providing similar exercise treatment and having similar exercise compliance. While PFP may present in a similar fashion in adolescence and adults, it may not be the same underlying condition or stage, and different treatments may be required. Collectively, this highlights the importance of increasing our understanding of the underlying pathology, pain mechanisms and why treatment may-or may not-work in adolescents and adults with PFP.


Conventional and Single-Molecule Targeted Sequencing Method for Specific Variant Detection in IKBKG while Bypassing the IKBKGP1 Pseudogene.
In addition to Sanger sequencing, next-generation sequencing of gene panels and exomes has emerged as a standard diagnostic tool in many laboratories. However, these captures can miss regions, have poor efficiency, or capture pseudogenes, which hamper proper diagnoses. One such example is the primary immunodeficiency-associated gene IKBKG. Its pseudogene IKBKGP1 makes traditional capture methods aspecific. We therefore developed a long-range PCR method to efficiently target IKBKG, as well as two associated genes (IRAK4 and MYD88), while bypassing the IKBKGP1 pseudogene. Sequencing accuracy was evaluated using both conventional short-read technology and a newer long-read, single-molecule sequencer. Different mapping and variant calling options were evaluated in their capability to bypass the pseudogene using both sequencing platforms. Based on these evaluations, we determined a robust diagnostic application for unambiguous sequencing and variant calling in IKBKG, IRAK4, and MYD88. This method allows rapid identification of selected primary immunodeficiency diseases in patients suffering from life-threatening invasive pyogenic bacterial infections.


The definition and classification of pneumonia.
Following the publication of a volume of Pneumonia focused on diagnosis, the journal's Editorial Board members debated the definition and classification of pneumonia and came to a consensus on the need to revise both of these. The problem with our current approach to the classification of pneumonia is twofold: (i) it results in widespread empirical, and often unnecessary, use of antimicrobials that contributes to pathogen resistance; and (ii) it contributes to heterogeneity among the groups of subjects compared in research, causing misclassification bias and mixtures of effects that threaten internal validity. After outlining the problem of classification, this commentary describes the strengths and weaknesses of a range of systems for the classification of pneumonia. The commentary then calls for debate to generate consensus classifications in the field, proposing a working definition and way forward focusing on the following three points: (i) pneumonia should be defined as an acute infection of the lung parenchyma by various pathogens, excluding the condition of bronchiolitis; (ii) defining pneumonia as a group of specific (co)infections with different characteristics is an ideal that currently has limited use, because the identification of aetiologic organisms in individuals is often not possible (however, the benefits of classifying pneumonia into specific, more homogenous phenotypes should be carefully considered when designing research studies); and (iii) investigation of more homogenous pneumonia groupings is achievable and is likely to yield more rapid advances in the field.


An Environment-Wide Association Study (EWAS) on type 2 diabetes mellitus.
Type 2 Diabetes (T2D) and other chronic diseases are caused by a complex combination of many genetic and environmental factors. Few methods are available to comprehensively associate specific physical environmental factors with disease. We conducted a pilot Environmental-Wide Association Study (EWAS), in which epidemiological data are comprehensively and systematically interpreted in a manner analogous to a Genome Wide Association Study (GWAS). We performed multiple cross-sectional analyses associating 266 unique environmental factors with clinical status for T2D defined by fasting blood sugar (FBG) concentration > or =126 mg/dL. We utilized available Centers for Disease Control (CDC) National Health and Nutrition Examination Survey (NHANES) cohorts from years 1999 to 2006. Within cohort sample numbers ranged from 503 to 3,318. Logistic regression models were adjusted for age, sex, body mass index (BMI), ethnicity, and an estimate of socioeconomic status (SES). As in GWAS, multiple comparisons were controlled and significant findings were validated with other cohorts. We discovered significant associations for the pesticide-derivative heptachlor epoxide (adjusted OR in three combined cohorts of 1.7 for a 1 SD change in exposure amount; p<0.001), and the vitamin gamma-tocopherol (adjusted OR 1.5; p<0.001). Higher concentrations of polychlorinated biphenyls (PCBs) such as PCB170 (adjusted OR 2.2; p<0.001) were also found. Protective factors associated with T2D included beta-carotenes (adjusted OR 0.6; p<0.001). Despite difficulty in ascertaining causality, the potential for novel factors of large effect associated with T2D justify the use of EWAS to create hypotheses regarding the broad contribution of the environment to disease. Even in this study based on prior collected epidemiological measures, environmental factors can be found with effect sizes comparable to the best loci yet found by GWAS.


Crosstalk between cAMP-dependent kinase and MAP kinase through a protein tyrosine phosphatase.
The haematopoietic protein tyrosine phosphatase (HePTP) is a negative regulator of the MAP kinases Erk1, Erk2 and p38. HePTP binds to these kinases through a kinase-interaction motif (KIM) in its non-catalytic amino terminus and inactivates them by dephosphorylating the critical phosphorylated tyrosine residue in their activation loop. Here we show that cyclic-AMP-dependent protein kinase (PKA) phosphorylates serine residue 23 in the KIM of HePTP in vitro and in intact cells. This modification reduces binding of MAP kinases to the KIM, an effect that is prevented by mutation of serine 23 to alanine. The PKA-mediated release of MAP kinase from HePTP is sufficient to activate the kinase and to induce transcription from the c-fos promoter. Expression of a HePTP serine-23-to-alanine mutant inhibits MAP-kinase dissociation and activation and induction of transcription from the c-fos promoter. We conclude that HePTP not only controls the activity of MAP kinases, but also mediates crosstalk between the cAMP system and the MAP-kinase cascade.


Cellular differentiation state modulates the mRNA export activity of SR proteins.
SR proteins function in nuclear pre-mRNA processing, mRNA export, and translation. To investigate their cellular dynamics, we developed a quantitative assay, which detects differences in nucleocytoplasmic shuttling among seven canonical SR protein family members. As expected, SRSF2 and SRSF5 shuttle poorly in HeLa cells but surprisingly display considerable shuttling in pluripotent murine P19 cells. Combining individual-resolution cross-linking and immunoprecipitation (iCLIP) and mass spectrometry, we show that elevated arginine methylation of SRSF5 and lower phosphorylation levels of cobound SRSF2 enhance shuttling of SRSF5 in P19 cells by modulating protein-protein and protein-RNA interactions. Moreover, SRSF5 is bound to pluripotency-specific transcripts such as Lin28a and Pou5f1/Oct4 in the cytoplasm. SRSF5 depletion reduces and overexpression increases their cytoplasmic mRNA levels, suggesting that enhanced mRNA export by SRSF5 is required for the expression of pluripotency factors. Remarkably, neural differentiation of P19 cells leads to dramatically reduced SRSF5 shuttling. Our findings indicate that posttranslational modification of SR proteins underlies the regulation of their mRNA export activities and distinguishes pluripotent from differentiated cells.


Expression of cell growth and bone specific genes at single cell resolution during development of bone tissue-like organization in primary osteoblast cultures.
Primary cultures of calvarial derived normal diploid osteoblasts undergo a developmental expression of genes reflecting growth, extracellular matrix maturation, and mineralization during development of multilayered nodules having a bone tissue-like organization. Scanning electron microscopy of the developing cultures indicates the transition from the uniform distribution of cuboidal osteoblasts to multilayered nodules of smaller cells with a pronounced orientation of perinodular cells towards the apex of the nodule. Ultrastructural analysis of the nodule by transmission electron microscopy indicates that the deposition of mineral is confined to the extracellular matrix where cells appear more osteocytic. The cell body contains rough endoplasmic reticulum and golgi, while these intracellular organelles are not present in the developing cellular processes. To understand the regulation of temporally expressed genes requires an understanding of which genes are selectively expressed on a single cell basis as the bone tissue-like organization develops. In situ hybridization analysis using 35S labelled histone gene probes, together with 3H-thymidine labelling and autoradiography, indicate that greater than 98% of the pre-confluent osteoblasts are proliferating. By two weeks, both the foci of multilayered cells and internodular cell regions have down-regulated cell growth associated genes. Post-proliferatively, but not earlier, initial expression of both osteocalcin and osteopontin are restricted to the multilayered nodules where all cells exhibit expression. While total mRNA levels for osteopontin and osteocalcin are coordinately upregulated with an increase in mineral deposition, in situ hybridization has revealed that expression of osteocalcin and osteopontin occurs predominantly in cells associated with the developing nodules. In contrast, proliferating rat osteosarcoma cells (ROS 17/2.8) concomitantly express histone H4, along with osteopontin and osteocalcin. These in situ analyses of gene expression during osteoblast growth and differentiation at the single cell level establish that a population of proliferating calvarial-derived cells subsequently expresses osteopontin and osteocalcin in cells developing into multilayered nodules with a tissue-like organization.


Differential regulation of fixN-reiterated genes in Rhizobium etli by a novel fixL-fixK cascade.
Among the complexities in the regulation of nitrogen fixation in the Rhizobiaceae are reiteration of regulatory components as well as variant roles for each component between species. For Rhizobium etli CFN42, we reported that the symbiotic plasmid (pCFN42d) contains a key regulatory gene (fixKd) and genes for a symbiotic cytochrome oxidase (fixNOQPd). Here we discuss the occurrence of reiteration of these genes (fixKf and fixNOQPf) and the finding of an unusual fixL homolog on a plasmid previously considered cryptic (pCFN42f). The structure of the deduced FixL polypeptide is suggestive of a fusion of the receiver and transmitter modules of a two-component regulatory system as described in R. leguminosarum bv. viciae VF39. Gene fusion analysis, coupled with mutation of each regulatory element, revealed that free-living expression of FixKf was dependent fully on FixL. In contrast, synthesis of FixKd was not detected under the conditions tested. The FixKf protein is needed for microaerobic expression of both fixN reiterations, whereas the FixKd protein appears to be dispensable. Interestingly, expression of the fixN reiterations exhibits a differential dependence for FixL, where transcription of fixNf was suppressed in the absence of FixL but expression of fixNd still showed significant levels. This suggests the existence of a FixL-independent mechanism for expression of the fixNd reiteration. Surprisingly, mutations in fixL, fixKd, or fixKf (either singly or in combination) did not alter symbiotic effectiveness. A mutation in fixNd (but not in fixNf) was, however, severely affected, indicating a differential role for these reiterations in nitrogen fixation.


The maximum rate of mammal evolution.
How fast can a mammal evolve from the size of a mouse to the size of an elephant? Achieving such a large transformation calls for major biological reorganization. Thus, the speed at which this occurs has important implications for extensive faunal changes, including adaptive radiations and recovery from mass extinctions. To quantify the pace of large-scale evolution we developed a metric, clade maximum rate, which represents the maximum evolutionary rate of a trait within a clade. We applied this metric to body mass evolution in mammals over the last 70 million years, during which multiple large evolutionary transitions occurred in oceans and on continents and islands. Our computations suggest that it took a minimum of 1.6, 5.1, and 10 million generations for terrestrial mammal mass to increase 100-, and 1,000-, and 5,000-fold, respectively. Values for whales were down to half the length (i.e., 1.1, 3, and 5 million generations), perhaps due to the reduced mechanical constraints of living in an aquatic environment. When differences in generation time are considered, we find an exponential increase in maximum mammal body mass during the 35 million years following the Cretaceous-Paleogene (K-Pg) extinction event. Our results also indicate a basic asymmetry in macroevolution: very large decreases (such as extreme insular dwarfism) can happen at more than 10 times the rate of increases. Our findings allow more rigorous comparisons of microevolutionary and macroevolutionary patterns and processes.


Multiple-integrating sphere spectrophotometer for measuring absolute spectral reflectance and transmittance.
A spectroreflectometer/transmissometer is described that permits determination of absolute optical characteristics in the 300-2600-nm wavelength region (which is essentially the complete solar spectrum). The uniqueness of the instrument derives from use of three rapidly interchangeable 20-cm (8-in.) integrating spheres to measure (1) absolute hemispherical spectral reflectance as a function of angles of incidence from -40 to +40 degrees employing an Edwards-type integrating sphere with a center-mounted sample [using small 2.5-cm (1-in.) diam specimens], (2) absolute hemispherical and absolute diffuse spectral reflectance at an angle of incidence of 20 degrees employing a sphere with a wall-mounted sample (for large specimens) and a screened detector, and (3) absolute hemispherical and absolute directional (near-normal exitance) transmittance employing a complete integrating sphere with the only ports being for the sample and reference beams. Data are presented that demonstrate the ability to measure the spectral reflectance of nonmirror surfaces to an absolute accuracy of 0.995 (an uncertainty of +/-0.005 reflectance units) in both reflectance spheres and of highly specular mirrors to an absolute accuracy of 0.993 (an uncertainty of +/-0.007 reflectance units). Spectral transmittance can be measured to an absolute accuracy of better than 0.995 (an uncertainty of +/-0.005 transmittance units).


Primate Brain Anatomy: New Volumetric MRI Measurements for Neuroanatomical Studies.
Since the publication of the primate brain volumetric dataset of Stephan and colleagues in the early 1980s, no major new comparative datasets covering multiple brain regions and a large number of primate species have become available. However, technological and other advances in the last two decades, particularly magnetic resonance imaging (MRI) and the creation of institutions devoted to the collection and preservation of rare brain specimens, provide opportunities to rectify this situation. Here, we present a new dataset including brain region volumetric measurements of 39 species, including 20 species not previously available in the literature, with measurements of 16 brain areas. These volumes were extracted from MRI of 46 brains of 38 species from the Netherlands Institute of Neuroscience Primate Brain Bank, scanned at high resolution with a 9.4-T scanner, plus a further 7 donated MRI of 4 primate species. Partial measurements were made on an additional 8 brains of 5 species. We make the dataset and MRI scans available online in the hope that they will be of value to researchers conducting comparative studies of primate evolution.


Cigarette smoking control strategies of firms with small work forces in two Northeastern States.
Cigarette smoking control strategies of firms with small work forces were assessed and compared with those of larger firms. A cross-sectional telephone survey was conducted in 1990 among private employers systematically selected from a proprietary database. These firms were located in four counties of two northeastern states. Interviews were conducted with managers of 470 small (< or = 25 employees; n = 262), medium (26-50; n = 87), and larger (> 50; n = 121) firms. Interviews assessed characteristics of the firms and their cigarette smoking policies and cessation programs. Small firms differed from larger firms in several areas. They were less likely to have written policies, used fewer methods to communicate their policies, and their policies were consistently less restrictive. Small firms also offered less assistance to employees who wished to quit. The less restrictive smoking policies reported here may be relatively ineffective in protecting nonsmokers in small firms. Small firms may encounter difficulties introducing more restrictive smoking policies because of the relative closeness of employee relations, smaller work spaces, and inability to deliver smoking cessation services to employees. Methods should be developed to assist managers of smaller firms to implement stronger smoking policies and to devise ways of making cessation assistance more easily available to their employees.


Determinants of Urinary Output Response to IV Furosemide in Acute Kidney Injury: A Pharmacokinetic/Pharmacodynamic Study.
This study assessed the determinants of urinary output response to furosemide in acute kidney injury; specifically, whether the response is related to altered pharmacokinetics or pharmacodynamics. Prospective cohort. Tertiary ICU. Thirty critically ill patients with acute kidney injury without preexisting renal impairment or recent diuretic exposure. A single dose of IV furosemide. Baseline markers of intravascular volume status were obtained prior to administering furosemide. Six-hour creatinine clearance, hourly plasma/urinary furosemide concentrations, and hourly urinary output were used to assess furosemide pharmacokinetics/pharmacodynamics parameters. Of 30 patients enrolled, 11 had stage-1 (37%), nine had stage-2 (30%), and 10 had stage-3 (33%) Acute Kidney Injury Network acute kidney injury. Seventy-three percent were septic, 47% required norepinephrine, and 53% were mechanically ventilated. Urinary output doubled in 20 patients (67%) following IV furosemide. Measured creatinine clearance was strongly associated with the amount of urinary furosemide excreted and was the only reliable predictor of the urinary output after furosemide (area under the receiver-operating-characteristic curve, 0.75; 95% CI, 0.57-0.93). In addition to an altered pharmacokinetics (p < 0.01), a reduced pharmacodynamics response to furosemide also became important when creatinine clearance was reduced to less than 40 mL/min/1.73 m (p = 0.01). Acute kidney injury staging and markers of intravascular volume, including central venous pressure, brain-natriuretic-peptide concentration, and fractional urinary sodium excretion were not predictive of urinary output response to furosemide. The severity of acute kidney injury, as reflected by the measured creatinine clearance, alters both pharmacokinetics and pharmacodynamics of furosemide in acute kidney injury, and was the only reliable predictor of the urinary output response to furosemide in acute kidney injury.


Effects of antineoplaston AS2-1 against post-operative lung metastasis in orthotopically implanted colon cancer in nude rat.
We have investigated the efficacy and mechanisms of antineoplaston AS2-1 against post-operative lung metastasis following removal of implanted human colon cancer in nude rat. The influence of AS2-1 on in vitro KM12SM human colon carcinoma cell activities (growth, cell cycle, and apoptosis) was evaluated. AS2-1 was administered perorally after removal of the implanted KM12SM cecal cancer in nude rat. AS2-1 inhibited KM12SM cell proliferation through G1 cell arrest and, at a higher concentration, induction of apoptosis. AS2-1 showed significant reduction in lung metastasis at 5 weeks after cecal removal. The survival rate in the AS2-1 group was significantly higher than that in the control. TUNEL staining on the lung metastatic tumors revealed that the apoptosis index (AI) in the AS2-1 group was significantly higher. Antineoplaston AS2-1 showed an antimetastatic effect against post-operative lung metastases from colon cancer through G1 cell arrest and the subsequent induction of apoptosis.


Accurate computation of survival statistics in genome-wide studies.
A key challenge in genomics is to identify genetic variants that distinguish patients with different survival time following diagnosis or treatment. While the log-rank test is widely used for this purpose, nearly all implementations of the log-rank test rely on an asymptotic approximation that is not appropriate in many genomics applications. This is because: the two populations determined by a genetic variant may have very different sizes; and the evaluation of many possible variants demands highly accurate computation of very small p-values. We demonstrate this problem for cancer genomics data where the standard log-rank test leads to many false positive associations between somatic mutations and survival time. We develop and analyze a novel algorithm, Exact Log-rank Test (ExaLT), that accurately computes the p-value of the log-rank statistic under an exact distribution that is appropriate for any size populations. We demonstrate the advantages of ExaLT on data from published cancer genomics studies, finding significant differences from the reported p-values. We analyze somatic mutations in six cancer types from The Cancer Genome Atlas (TCGA), finding mutations with known association to survival as well as several novel associations. In contrast, standard implementations of the log-rank test report dozens-hundreds of likely false positive associations as more significant than these known associations.


Neurogenic placodes: a common front.
Neurogenic placodes are specialized regions of the embryonic ectoderm that are central to the development of the cranial sensory systems of vertebrates. These structures generate many of the sensory neurones of the head and also, in some instances, the associated sensory receptor cells. The neurogenic placodes have often been thought to share common pathways of development and to have evolved together. However, recent work has shown that this is not the case. The term 'neurogenic placodes' no longer seems to describe a cohesive group. Rather, the neurogenic placodes fall into a number of categories, and it is within each of these that the members share a common development and evolution.


The sensitivity of transbronchial biopsy for the diagnosis of acute lung rejection.
Transbronchial biopsy has become the procedure of choice for the diagnosis of acute lung rejection after transplantation, but the sensitivity of the technique in this setting remains unknown. In this study, 14 mongrel dogs underwent left lung transplantation, after which triple-drug immunosuppression was given for 5 days and then all immunosuppression was stopped. All animals had clear chest radiographs at this time. Transbronchial biopsy was performed in nine lung regions (two to six pieces of lung tissue were obtained per region, with a mean of 4.3 pieces per region) before the animals were killed 2 to 4 days later, at which time varying degrees of rejection had occurred. Rejection was graded histologically on a scale of 0 to 3 (0 = no rejection, 1 = mild rejection, 2 = moderate rejection, 3 = severe rejection) in each piece of lung tissue obtained at transbronchial biopsy. After the dogs were put to death, the true state of lung rejection was determined by histologic examination of the entire lung. We calculated the sensitivity of transbronchial biopsy with 95% confidence intervals. Five pieces of lung tissue were needed to yield a sensitivity of 92% (82%, 100%) to identify mild rejection in the entire lung with transbronchial biopsy. Three pieces of lung tissue were needed to yield a sensitivity of 92% (84%, 100%) to identify the presence of moderate to severe rejection in the entire lung (that is, rejection that requires pulse therapy) on transbronchial biopsy. These results indicate that three to five pieces of lung tissue that are suitable for diagnostic purposes obtained at transbronchial biopsy are adequate for the diagnosis of acute pulmonary rejection after lung transplantation.


Blood pressure variables and cardiovascular risk: new findings from ADVANCE.
The relative importance of various blood pressure indices on cardiovascular risk in people with type 2 diabetes mellitus has not been established. This study compares the strengths of the associations between different baseline blood pressure variables (systolic blood pressure [SBP], diastolic blood pressure [DBP], pulse pressure [PP], and mean arterial pressure) and the 4.3-year risk of major cardiovascular events in the Action in Diabetes and Vascular Disease: Preterax and Diamicron-Modified Release Controlled Evaluation Study. Mean (SD) age for the 11 140 participants was 65.8 years (6.4 years). During follow-up, 1000 major cardiovascular events, 559 major coronary events, and 468 cardiovascular deaths were recorded. After adjustment for age, sex, and treatment allocation, the hazard ratios (95% CIs) associated with 1 increment in SD for the risk of major cardiovascular events were 1.17 (1.10 to 1.24) for SBP; 1.20 (1.13 to 1.28) for PP; 1.12 (1.05 to 1.19) for mean arterial pressure; and 1.04 (0.98 to 1.11) for DBP. The areas under the receiver operating characteristic curve were slightly higher for SBP and PP compared with mean arterial pressure and DBP for major cardiovascular and coronary events. Using achieved instead of baseline blood pressure values marginally improved the effect estimates for SBP, DBP, and mean arterial pressure, with no significant differences in the areas under the receiver operating characteristic curve between models with SBP and those with PP. In conclusion, SBP and PP are the 2 best and DBP is the least effective determinant of the risk of major cardiovascular outcomes in the relatively old patients with type 2 diabetes mellitus participating in the Action in Diabetes and Vascular Disease: Preterax and Diamicron-Modified Release Controlled Evaluation Study. However, SBP may be the simplest and most useful predictor across a wider range of age groups and populations.


The challenges of being a student nurse and the advice proffered by peers.
EBN engages through a range of Online social media activities to debate issues important to nurses and nursing. EBN Opinion papers highlight and expand on these debates.


Optimization of nanotube electrode geometry in a liquid crystal media from wavefront aberrations.
This paper presents experimental optimization of number and geometry of nanotube electrodes in a liquid crystal media from wavefront aberrations for realizing nanophotonic devices. The refractive-index gradient profiles from different nanotube geometries--arrays of one, three, four, and five--were studied along with wavefront aberrations using Zernike polynomials. The optimizations help the device to make application in the areas of voltage reconfigurable microlens arrays, high-resolution displays, wavefront sensors, holograms, and phase modulators.


Serologic evidence of West Nile virus infections in wild birds captured in Germany.
To assess the risk of acquiring a West Nile virus (WNV) infection in Germany, we investigated samples from migrating and from resident birds. Because of their stay in or migration through WNV-endemic regions, these birds are at risk to become infected with WNV. Blood samples from 3,399 birds, representing 87 bird species, were collected in Germany in 2000 and in 2002-2005. Overall, 53 birds belonging to 5 species had WNV-neutralizing antibodies. Fifty-nine birds belonging to 9 species were reactive by WNV immunofluorescence assay, and 8 birds had neutralizing antibodies against Usutu virus. Because of maternal antibody transfer via egg yolk, WNV-antibody titers in white stork nestlings were generally lower than those in adults. Despite a relatively high percentage of stork nestlings with antibodies, no viral genomes were detectable by polymerase chain reaction. In Germany, the prevalence of antibodies to WNV in migrating birds wintering in Africa or southern Europe is comparatively low.


Dual neonate vaccine platform against HIV-1 and M. tuberculosis.
Acquired immunodeficiency syndrome and tuberculosis (TB) are two of the world's most devastating diseases. The first vaccine the majority of infants born in Africa receive is Mycobacterium bovis bacillus Calmette-Guérin (BCG) as a prevention against TB. BCG protects against disseminated disease in the first 10 years of life, but provides a variable protection against pulmonary TB and enhancing boost delivered by recombinant modified vaccinia virus Ankara (rMVA) expressing antigen 85A (Ag85A) of M. tuberculosis is currently in phase IIb evaluation in African neonates. If the newborn's mother is positive for human immunodeficiency virus type 1 (HIV-1), the baby is at high risk of acquiring HIV-1 through breastfeeding. We suggested that a vaccination consisting of recombinant BCG expressing HIV-1 immunogen administered at birth followed by a boost with rMVA sharing the same immunogen could serve as a strategy for prevention of mother-to-child transmission of HIV-1 and rMVA expressing an African HIV-1-derived immunogen HIVA is currently in phase I trials in African neonates. Here, we aim to develop a dual neonate vaccine platform against HIV-1 and TB consisting of BCG.HIVA administered at birth followed by a boost with MVA.HIVA.85A. Thus, mMVA.HIVA.85A and sMVA.HIVA.85A vaccines were constructed, in which the transgene transcription is driven by either modified H5 or short synthetic promoters, respectively, and tested for immunogenicity alone and in combination with BCG.HIVA(222). mMVA.HIVA.85A was produced markerless and thus suitable for clinical manufacture. While sMVA.HIVA.85A expressed higher levels of the immunogens, it was less immunogenic than mMVA.HIVA.85A in BALB/c mice. A BCG.HIVA(222)-mMVA.HIVA.85A prime-boost regimen induced robust T cell responses to both HIV-1 and M. tuberculosis. Therefore, proof-of-principle for a dual anti-HIV-1/M. tuberculosis infant vaccine platform is established. Induction of immune responses against these pathogens soon after birth is highly desirable and may provide a basis for lifetime protection maintained by boosts later in life.


Lymphoproliferative cancer and other malignancy in patients with rheumatoid arthritis treated with azathioprine: a 20 year follow up study.
Two hundred and two patients with rheumatoid arthritis (RA) starting treatment with large doses of azathioprine (median 300 mg/day) between 1964 and 1974 were followed up until March 1984. All but one patient (99.5%) were traced from either hospital or general practice records; and death certificates, where relevant, were obtained. A comparison group of 202 patients with RA not treated with azathioprine was selected from the diagnostic index of another rheumatology unit and followed up in 1985 to assess their status retrospectively at March 1984. Each patient treated with azathioprine was matched for year of birth, year of diagnosis, sex, and serostatus with a control patient from the latter group. Four lymphoproliferative cancers occurred in the azathioprine treated group compared with two in the control group. Further analysis of these findings suggested an increased risk of lymphoma of one case per 1000 patient years of azathioprine treatment. The lymphoma rates were then compared with those expected based on the incidence in the general population. This comparison suggested a fivefold increase in the RA control group and a 10-fold increase in the azathioprine treated group. There was also an excess of patients with non-lymphoproliferative cancers, including one with myeloma in the azathioprine group (29 v 19), the excess being greater in the group with the longest duration of treatment. This significantly increased risk did not, however, persist on matched analysis, was not related to maximum daily dose, and was not site specific. These results from a possibly unique series of patients treated with high dose azathioprine give some reassurance about the magnitude of the previously postulated carcinogenic risk of such treatment in RA.


Using multiple classifiers for predicting the risk of endovascular aortic aneurysm repair re-intervention through hybrid feature selection.
Feature selection is essential in medical area; however, its process becomes complicated with the presence of censoring which is the unique character of survival analysis. Most survival feature selection methods are based on Cox's proportional hazard model, though machine learning classifiers are preferred. They are less employed in survival analysis due to censoring which prevents them from directly being used to survival data. Among the few work that employed machine learning classifiers, partial logistic artificial neural network with auto-relevance determination is a well-known method that deals with censoring and perform feature selection for survival data. However, it depends on data replication to handle censoring which leads to unbalanced and biased prediction results especially in highly censored data. Other methods cannot deal with high censoring. Therefore, in this article, a new hybrid feature selection method is proposed which presents a solution to high level censoring. It combines support vector machine, neural network, and K-nearest neighbor classifiers using simple majority voting and a new weighted majority voting method based on survival metric to construct a multiple classifier system. The new hybrid feature selection process uses multiple classifier system as a wrapper method and merges it with iterated feature ranking filter method to further reduce features. Two endovascular aortic repair datasets containing 91% censored patients collected from two centers were used to construct a multicenter study to evaluate the performance of the proposed approach. The results showed the proposed technique outperformed individual classifiers and variable selection methods based on Cox's model such as Akaike and Bayesian information criterions and least absolute shrinkage and selector operator in p values of the log-rank test, sensitivity, and concordance index. This indicates that the proposed classifier is more powerful in correctly predicting the risk of re-intervention enabling doctor in selecting patients' future follow-up plan.


Traumatic dissociation, epileptic-like phenomena, and schizophrenia.
According to recent evidence, trauma and stress are important etiological factors in pathogenesis of schizophrenia. However, the hypothetical influence of traumatic stress on epileptic-like (or kindling) phenomena in schizophrenia is at this time unclear. In order to discover the influence of trauma, 82 patients with a diagnosis of paranoid schizophrenia and 50 healthy controls were assessed for symptoms of dissociation, traumatic stress and psycho-sensory symptoms of epileptic origin. We have found significant traumatization and dissociation in patients who met the cut-off score for psychosensory epileptic-like symptoms and also significant correlations among these measures have been found. These data suggest a certain role of epileptic-like phenomena in dissociative states in schizophrenia and are in accordance with rare intracranial EEG findings which suggest a certain role of epileptiform events in schizophrenia.


Kick Control: Using the Attracting States Arising Within the Sensorimotor Loop of Self-Organized Robots as Motor Primitives.
Self-organized robots may develop attracting states within the sensorimotor loop, that is within the phase space of neural activity, body and environmental variables. Fixpoints, limit cycles and chaotic attractors correspond in this setting to a non-moving robot, to directed, and to irregular locomotion respectively. Short higher-order control commands may hence be used to kick the system from one self-organized attractor robustly into the basin of attraction of a different attractor, a concept termed here as kick control. The individual sensorimotor states serve in this context as highly compliant motor primitives. We study different implementations of kick control for the case of simulated and real-world wheeled robots, for which the dynamics of the distinct wheels is generated independently by local feedback loops. The feedback loops are mediated by rate-encoding neurons disposing exclusively of propriosensoric inputs in terms of projections of the actual rotational angle of the wheel. The changes of the neural activity are then transmitted into a rotational motion by a simulated transmission rod akin to the transmission rods used for steam locomotives. We find that the self-organized attractor landscape may be morphed both by higher-level control signals, in the spirit of kick control, and by interacting with the environment. Bumping against a wall destroys the limit cycle corresponding to forward motion, with the consequence that the dynamical variables are then attracted in phase space by the limit cycle corresponding to backward moving. The robot, which does not dispose of any distance or contact sensors, hence reverses direction autonomously.


Modified Mason type III and IV radial head fractures: results of different surgical treatments.
Fractures of the radial head account for 4% of all fractures and 33 % of all elbow fractures. Their treatment is somewhat challenging and diversified, especially in more complex fractures (type III and IV of modified Mason's classification). The aim of this study was to identify the best surgical treatment for patients having sustained these latter lesions and outline possible predictive factors of worse outcomes. Data were retrospectively collected for 63 patients affected by radial head fracture and operated between 2006 and 2014 at the University Hospital of Parma. In 34 patients open reduction and internal fixation (ORIF) was used, in 20 radial head arthroplasty (RHA) was the treatment choice and radial head resection (RHR) was done in the remaining 9. Clinical and radiographic assessments were done at a minimum follow-up of 1 year. Clinical evaluation was performed with the Mayo Elbow performance Score (MEPS). No statistical differences were observed in either type between the MEPS of affected and unaffected elbow. Multiple regression analysis showed that modified Mason IV fractures were a predictive factor of worse outcome and that an associated coronoid fracture can lead to a higher instability of elbow. Mason IV fractures treated with primary RHA are associated to better outcomes. According to this retrospective clinical study, it was not possible to identify the optimal surgical treatment for Mason type III fractures. However, RHA seems to be the preferred choice for Mason type IV fractures. These latter types of lesions are associated to worse outcomes.


Down-regulation of placental Cdc42 and Rac1 links mTORC2 inhibition to decreased trophoblast amino acid transport in human intrauterine growth restriction.
Intrauterine growth restriction (IUGR) increases the risk for perinatal complications and metabolic and cardiovascular disease later in life. The syncytiotrophoblast (ST) is the transporting epithelium of the human placenta, and decreased expression of amino acid transporter isoforms in the ST plasma membranes is believed to contribute to IUGR. Placental mechanistic target of rapamycin Complex 2 (mTORC2) signaling is inhibited in IUGR and regulates the trafficking of key amino acid transporter (AAT) isoforms to the ST plasma membrane; however, the molecular mechanisms are unknown. Cdc42 and Rac1 are Rho-GTPases that regulate actin-binding proteins, thereby modulating the structure and dynamics of the actin cytoskeleton. We hypothesized that inhibition of mTORC2 decreases AAT expression in the plasma membrane and amino acid uptake in primary human trophoblast (PHT) cells mediated by down-regulation of Cdc42 and Rac1. mTORC2, but not mTORC1, inhibition decreased the Cdc42 and Rac1 expression. Silencing of Cdc42 and Rac1 inhibited the activity of the System L and A transporters and markedly decreased the trafficking of LAT1 (System L isoform) and SNAT2 (System A isoform) to the plasma membrane. mTORC2 inhibition by silencing of rictor failed to decrease AAT following activation of Cdc42/Rac1. Placental Cdc42 and Rac1 protein expression was down-regulated in human IUGR and was positively correlated with placental mTORC2 signaling. In conclusion, mTORC2 regulates AAT trafficking in PHT cells by modulating Cdc42 and Rac1. Placental mTORC2 inhibition in human IUGR may contribute to decreased placental amino acid transfer and reduced fetal growth mediated by down-regulation of Cdc42 and Rac1.


An Applicant's Residency Program Region May Influence Where They Complete Fellowship.
The primary goal of this study was to determine if an applicant's geographic region of residency was associated with where they matched for fellowship. San Francisco Match (SF Match) provided results regarding applicant data and match results from 2014-2018 for orthopaedic subspecialties except hand and shoulder and elbow. Residency programs were divided into five regions: (Northeast [NE], Southeast [SE], Midwest [MW], Southwest [SW] and West [W]). The MW region had the fewest number of fellowship positions per applicant (0.62), the W region had the most (1.7). Applicants from each region were significantly (p < 0.0001) more likely to complete fellowship in the same region where they completed residency, and there were significant (p < 0.05) differences between regions for specific subspecialties. There are imbalances in terms of the number of applicants and specific fellowship spots available in each region. This imbalance seems important considering the strong associations found between the region in which an applicant completes residency and fellowship. Level of Evidence: Level 3. (Journal of Surgical Orthopaedic Advances 30(1):030-035, 2021).


Exploring functional roles of TRPV1 intracellular domains with unstructured peptide-insertion screening.
TRPV1 is a polymodal nociceptor for diverse physical and chemical stimuli that interact with different parts of the channel protein. Recent cryo-EM studies revealed detailed channel structures, opening the door for mapping structural elements mediating activation by each stimulus. Towards this goal, here we have combined unstructured peptide-insertion screening (UPS) with electrophysiological and fluorescence recordings to explore structural and functional roles of the intracellular regions of TRPV1 in mediating various activation stimuli. We found that most of the tightly packed protein regions did not tolerate structural perturbation by UPS when tested, indicating that structural integrity of the intracellular region is critical. In agreement with previous reports, Ca2+-dependent desensitization is strongly dependent on both intracellular N- and C-terminal domains; insertions of an unstructured peptide between these domains and the transmembrane core domain nearly eliminated Ca2+-dependent desensitization. In contrast, channel activations by capsaicin, low pH, divalent cations, and even heat are mostly intact in mutant channels containing the same insertions. These observations suggest that the transmembrane core domain of TRPV1, but not the intracellular domains, is responsible for sensing these stimuli.


Effects of 10-hydroxycamptothecin, delivered from locally injectable poly(lactide-co-glycolide) microspheres, in a murine human oral squamous cell carcinoma regression model.
This study investigated whether local delivery of 10-hydroxycamptothecin provides effective inductive chemotherapy as assessed by significant tumor reduction. Established tumorigenic human oral squamous cell carcinoma cells were used for these experiments. The experimental groups were comprised of: control (blank (no drug) poly(lactide-co-glycolide) (PLGA) microspheres), intraperitoneal 10-hydroxycamptothecin delivery + blank microspheres, local bolus 10-hydroxycamptothecin + blank microspheres, and PLGA controlled-release microspheres. The 10-hydroxycamptothecin dose administered was 12 mg/kg (bolus-intraperitoneal, local) or controlled-release over 10 days. Regardless of delivery route, 10-hydroxycamptothecin significantly reduces tumor volume. However, PLGA microspheres provide significantly higher intratumor-drug concentrations (approximately 10 and 100 fold higher) relative to local bolus and intraperitoneal routes, respectively. Also, only the PLGA microspheres significantly reduced tumor weights. Camptothecin clinical applications are limited by drug inactivation at physiological pH and the need for sustained infusions. However, due to their acidic, camptothecin-stabilizing microclimate, PLGA microspheres could provide a novel delivery system for camptothecin-based induction chemotherapy.


White matter microstructural changes in adolescent anorexia nervosa including an exploratory longitudinal study.
Anorexia nervosa (AN) often begins in adolescence, however, the understanding of the underlying pathophysiology at this developmentally important age is scarce, impeding early interventions. We used diffusion tensor imaging (DTI) to investigate microstructural white matter (WM) brain changes including an experimental longitudinal follow-up. We acquired whole brain diffusion-weighted brain scans of 22 adolescent female hospitalized patients with AN at admission and nine patients longitudinally at discharge after weight rehabilitation. Patients (10-18 years) were compared to 21 typically developing controls (TD). Tract-based spatial statistics (TBSS) were applied to compare fractional anisotropy (FA) across groups and time points. Associations between average FA values of the global WM skeleton and weight as well as illness duration parameters were analyzed by multiple linear regression. We observed increased FA in bilateral frontal, parietal and temporal areas in AN patients at admission compared to TD. Higher FA of the global WM skeleton at admission was associated with faster weight loss prior to admission. Exploratory longitudinal analysis showed this FA increase to be partially normalized after weight rehabilitation. Our findings reveal a markedly different pattern of WM microstructural changes in adolescent AN compared to most previous results in adult AN. This could signify a different susceptibility and reaction to semi-starvation in the still developing brain of adolescents or a time-dependent pathomechanism differing with extend of chronicity. Higher FA at admission in adolescents with AN could point to WM fibers being packed together more closely.


Casting a Health Equity Lens on Endocrinology and Diabetes.
As endocrinologists we have focused on biological contributors to disparities in diabetes, obesity and other endocrine disorders. Given that diabetes is an exemplar health disparity condition, we, as a specialty, are also positioned to view the contributing factors and solutions more broadly. This will give us agency in contributing to health system, public health, and policy-level interventions to address the structural and institutional racism embedded in our medical and social systems. A history of unconsented medical and research experimentation on vulnerable groups and perpetuation of eugenics theory in the early 20th century have resulted in residual health care provider biases toward minority patients and patient distrust of medical systems, leading to poor quality of care. Historical discriminatory housing and lending policies resulted in racial residential segregation and neighborhoods with inadequate housing, healthy food access, and educational resources, setting the foundation for the social determinants of health (SDOH) contributing to present-day disparities. To reduce these disparities we need to ensure our health systems are implementing the National Standards for Culturally and Linguistically Appropriate Services in Health and Health Care to promote health equity. Because of racial biases inherent in our medical systems due to historical unethical practices in minority communities, health care provider training should incorporate awareness of unconscious bias, antiracism, and the value of diversity. Finally, we must also address poverty-related SDOH (eg, food and housing insecurity) by integrating social needs into medical care and using our voices to advocate for social policies that redress SDOH and restore environmental justice.


Spatial working memory deficits in GluA1 AMPA receptor subunit knockout mice reflect impaired short-term habituation: evidence for Wagner's dual-process memory model.
Genetically modified mice, lacking the GluA1 AMPA receptor subunit, are impaired on spatial working memory tasks, but display normal acquisition of spatial reference memory tasks. One explanation for this dissociation is that working memory, win-shift performance engages a GluA1-dependent, non-associative, short-term memory process through which animals choose relatively novel arms in preference to relatively familiar options. In contrast, spatial reference memory, as exemplified by the Morris water maze task, reflects a GluA1-independent, associative, long-term memory mechanism. These results can be accommodated by Wagner's dual-process model of memory in which short and long-term memory mechanisms exist in parallel and, under certain circumstances, compete with each other. According to our analysis, GluA1(-/-) mice lack short-term memory for recently experienced spatial stimuli. One consequence of this impairment is that these stimuli should remain surprising and thus be better able to form long-term associative representations. Consistent with this hypothesis, we have recently shown that long-term spatial memory for recently visited locations is enhanced in GluA1(-/-) mice, despite impairments in hippocampal synaptic plasticity. Taken together, these results support a role for GluA1-containing AMPA receptors in short-term habituation, and in modulating the intensity or perceived salience of stimuli.


Bcl-2 protein in 518A2 melanoma cells in vivo and in vitro.
Bcl-2 is an apoptotic protein that is highly expressed in advanced melanoma. Several strategies have been employed to target the expression of this protein, including G3139, an 18-mer phosphorothioate oligodeoxyribonucleotide targeted to the initiation region of the Bcl-2 mRNA. This compound has recently completed phase III global clinical evaluation, but the function of Bcl-2 as a target in melanoma has not been completely clarified. To help resolve this question, we have permanently and stably down-regulated Bcl-2 protein and mRNA expression in 518A2 cells by two different technologies and evaluated the resulting clones both in vitro and in vivo. 518A2 melanoma cells were transfected with plasmids engineered to produce either a single-stranded antisense oligonucleotide targeted to the initiation codon region of the Bcl-2 mRNA or a short hairpin RNA also targeted to the Bcl-2 mRNA. In vitro growth, the apoptotic response to G3139, and the G3139-induced release of cytochrome c from isolated mitochondria were evaluated. Cells were then xenografted into severe combined immunodeficient mice and tumor growth was measured. In vitro, down-regulation of Bcl-2 expression by either method produced no change either in the rate of growth or in sensitivity to standard cytotoxic chemotherapeutic agents. Likewise, the induction of apoptosis by G3139 was entirely Bcl-2 independent. In addition, the G3139-induced release from isolated mitochondria was also relatively independent of Bcl-2 expression. However, when xenografted into severe combined immunodeficient mice, cells with silenced Bcl-2, using either technology, either failed to grow at all or grew to tumors of low volume and then completely regressed. In contrast, control cells with "normal" levels of Bcl-2 protein expression expanded to be large, necrotic tumors. The presence of Bcl-2 protein profoundly affects the ability of 518A2 melanoma cells to grow as human tumor xenografts in severe combined immunodeficient mice. The in vivo role of Bcl-2 in melanoma cells thus differs significantly from its in vitro role, and these experiments further suggest that Bcl-2 may be an important therapeutic target even in tumors that do not contain the t14:18 translocation.


New records of the blacklegged tick, Ixodes scapularis (Acari: Ixodidae) in Minnesota.
The Minnesota distribution of the blacklegged tick, Ixodes scapularis, was studied during 1998 and 1999. The majority of tick collecting was done by grouse hunters, who sent in specimens collected during the fall of 1998 and 1999. I. scapularis specimens were received from 17 counties, including eight counties in northern and central Minnesota that previously had no reports of I. scapularis. This brings the number of Minnesota counties with reported populations of I. scapularis to 30, mostly in the east-central, central, and northern parts of the state.


Whole-exome sequencing identifies rare variants in STAB2 associated with venous thromboembolic disease.
Deep vein thrombosis and pulmonary embolism, collectively defined as venous thromboembolism (VTE), are the third leading cause of cardiovascular death in the United States. Common genetic variants conferring increased varying degrees of VTE risk have been identified by genome-wide association studies (GWAS). Rare mutations in the anticoagulant genes PROC, PROS1 and SERPINC1 result in perinatal lethal thrombosis in homozygotes and markedly increased VTE risk in heterozygotes. However, currently described VTE variants account for an insufficient portion of risk to be routinely used for clinical decision making. To identify new rare VTE risk variants, we performed a whole-exome study of 393 individuals with unprovoked VTE and 6114 controls. This study identified 4 genes harboring an excess number of rare damaging variants in patients with VTE: PROS1, STAB2, PROC, and SERPINC1. At STAB2, 7.8% of VTE cases and 2.4% of controls had a qualifying rare variant. In cell culture, VTE-associated variants of STAB2 had a reduced surface expression compared with reference STAB2. Common variants in STAB2 have been previously associated with plasma von Willebrand factor and coagulation factor VIII levels in GWAS, suggesting that haploinsufficiency of stabilin-2 may increase VTE risk through elevated levels of these procoagulants. In an independent cohort, we found higher von Willebrand factor levels and equivalent propeptide levels in individuals with rare STAB2 variants compared with controls. Taken together, this study demonstrates the utility of gene-based collapsing analyses to identify loci harboring an excess of rare variants with functional connections to a complex thrombotic disease.


A group ICA based framework for evaluating resting fMRI markers when disease categories are unclear: application to schizophrenia, bipolar, and schizoaffective disorders.
Schizophrenia (SZ), bipolar disorder (BP) and schizoaffective disorder (SAD) share some common symptoms, and there is still a debate about whether SAD is an independent category. To the best of our knowledge, no study has been done to differentiate these three disorders or to investigate the distinction of SAD as an independent category using fMRI data. This study is aimed to explore biomarkers from resting-state fMRI networks for differentiating these disorders and investigate the relationship among these disorders based on fMRI networks with an emphasis on SAD. Firstly, a novel group ICA method, group information guided independent component analysis (GIG-ICA), was applied to extract subject-specific brain networks from fMRI data of 20 healthy controls (HC), 20 SZ patients, 20 BP patients, 20 patients suffering from SAD with manic episodes (SADM), and 13 patients suffering from SAD with depressive episodes exclusively (SADD). Then, five-level one-way analysis of covariance and multiclass support vector machine recursive feature elimination were employed to identify discriminative regions from the networks. Subsequently, the t-distributed stochastic neighbor embedding (t-SNE) projection and the hierarchical clustering were implemented to investigate the relationship among those groups. Finally, to evaluate the generalization ability, 16 new subjects were classified based on the found regions and the trained model using original 93 subjects. Results show that the discriminative regions mainly included frontal, parietal, precuneus, cingulate, supplementary motor, cerebellar, insula and supramarginal cortices, which performed well in distinguishing different groups. SADM and SADD were the most similar to each other, although SADD had greater similarity to SZ compared to other groups, which indicates that SAD may be an independent category. BP was closer to HC compared with other psychotic disorders. In summary, resting-state fMRI brain networks extracted via GIG-ICA provide a promising potential to differentiate SZ, BP, and SAD.


Interleukin-12 (IL-12) and IL-18 synergistically induce the fungicidal activity of murine peritoneal exudate cells against Cryptococcus neoformans through production of gamma interferon by natural killer cells.
We examined the ability of interleukin-12 (IL-12) and IL-18 to induce the production of gamma interferon (IFN-gamma) and nitric oxide (NO) by murine peritoneal exudate cells (PEC) and to stimulate the growth-inhibitory activity of these cells against Cryptococcus neoformans. PEC produced IFN-gamma and NO when stimulated with a combination of IL-12 and IL-18 but little or no IFN-gamma or NO when either cytokine was used alone. PEC anticryptococcal activity was mediated by IFN-gamma and NO production, since it was completely inhibited by a neutralizing anti-IFN-gamma monoclonal antibody (MAb) and N(G)-monomethyl-L-arginine, a competitive inhibitor of NO synthesis, respectively. To identify the IFN-gamma-producing cells among PEC stimulated with IL-12 and IL-18, we depleted NK cells, gammadelta T cells, or CD4+ T cells by treating PEC with specific Abs and complement. NK cell depletion strongly suppressed IFN-gamma production and almost completely inhibited NO production and anticryptococcal activity, while depletion of other cells had no such influence. Alternatively, purified NK cells by two cycles of glass adherence and magnetic separation with anti-CD3, -CD4, -CD8, and -B220 MAbs produced a greater amount of IFN-gamma by stimulation with IL-12 and IL-18 than unseparated non-glass-adherent PEC. Our results demonstrated that IL-12 and IL-18 synergistically induced NO-dependent anticryptococcal activity of PEC by stimulating NK cells to produce IFN-gamma.


Alterations of MicroRNAs in Solid Cancers and Their Prognostic Value.
MicroRNAs (miRNAs) are evolutionarily conserved, naturally abundant, small, regulatory non-coding RNAs that inhibit gene expression at the post-transcriptional level in a sequence-specific manner. Each miRNA represses the protein expression of several coding genes in a manner proportional to the sequence complementarity with the target transcripts. MicroRNAs play key regulatory roles in organismal development and homeostasis. They control fundamental biological processes, such as stem-cell regulation and cellular metabolism, proliferation, differentiation, stress resistance, and apoptosis. Differential miRNA expression is found in malignant tumors in comparison to normal tissue counterparts. This indicates that miRNA deregulation contributes to the initiation and progression of cancer. Currently, miRNA expression signatures are being rigorously investigated in various tumor types, with the aim of developing novel, efficient biomarkers that can improve clinical management of cancer patients. This review discusses deregulated miRNAs in solid tumors, and focuses on their emerging prognostic potential.


Comparison of three techniques for calculation of target coordinates in functional stereotactic procedures.
We compared three techniques of target coordinate determination for various functional stereotactic procedures. All procedures were based on preoperative MRI with contiguous 3-mm cuts. The first technique involved determination of anatomical landmarks and fiducial markers of the stereotactic frame on the monitor screen of an MRI scanner and calculation of the target point using a series of formulas; the second technique used a Leksell tabletop localizer, and the third technique is a part of 'Stealth' stereotactic navigation software. Final coordinates for the procedure were derived from all three techniques and subsequently adjusted using intraoperative electrical macrostimulation. We found that difference between techniques was on average 0.9 +/- 0.4 mm in each of three directions, and 1.8 +/- 0.9 mm in absolute distance. There were 7 cases in which one of the techniques had a discrepancy of more than 3 mm (more than 1 MRI slice thickness) compared with the other two, indicating a potential error in coordinate determination. This difference could potentially result in inappropriate placement of the electrode, thus affecting the procedure outcome. In 6 cases, such an error apparently occurred with the first or second technique of calculation. The average number of mapping trajectories decreased from 1.8 to 1.4 since this stereotactic software became a part of operative planning. We conclude that use of computerized planning software increases the precision of target coordinate calculation and improves the accuracy of functional stereotactic procedures.


Inflammation-related cytokines and their roles in gastroenteropancreatic neuroendocrine neoplasms.
Proinflammatory counterworks are important at different stages of tumor development, particularly during invasion and metastasis. Immune cells and their signal molecules can influence all stages of tumor progression, as well as therapeutic intervention. Proinflammatory cytokines are known triggers of growth in gastroenteropancreatic neuroendocrine neoplasms (GEP-NENs). In this study, we explored the immunohistochemical expression of tumor necrosis factor alpha (TNF-α), interleukin 1 beta (IL-1β), IL-2, and IL-6 in tissues from 43 GEP-NEN patients with tumors of gastric, duodenal, ileal, appendiceal, and colonic origin. The immunohistochemical expression of TNF-α was increased in tumor groups with high proliferation rates (Ki-67; p = 0.034), as well as in those with higher tumor grades (p = 0.05). Moreover, the immunohistochemical expression of TNF-α positively correlated with death outcomes (p = 0.016). Expression of IL-6, IL-1β, and IL-2 displayed similar immunohistochemical expression patterns regardless of Ki-67, although the expression between the ILs differed. Most GEP-NENs had high levels of IL-6 and lower levels of IL-1β and IL-2. Although further comprehensive studies are required for a complete understanding of activated mechanisms in proinflammatory protumoral microenvironment of GEP-NENs, TNF-α is a potential marker in the prognosis of those tumors.


Transient ischaemic attack: clinical relevance, risk prediction and urgency of secondary prevention.
Transient ischaemic attack (TIA) is increasingly recognized as a harbinger of stroke and an important opportunity for secondary prevention. We have reviewed recent evidence on the burden of TIA and prediction and prevention of stroke in the acute phase. Although recent data on the incidence and prevalence of TIA are lacking, available data suggest that the burden of TIA is higher than previously estimated and may be expected to increase with the ageing of the population. Prospective prognostic studies have shown that the early risk of stroke after TIA is approximately 5% at 7 days and 10-15% at 90 days depending on clinical settings and study methodology. This risk can be reliably predicted by risk scores based on clinical features (the ABCD system), TIA aetiology and findings on brain imaging, although the optimal combined prognostic strategy is uncertain because the interaction between individual predictors is not established. Studies of the urgent assessment and initiation of secondary prevention in specialist centres suggest that the early risk of stroke after TIA can be reduced by up to 80%. The risk of stroke after TIA is considerable. However, recent advances have shown that this risk can be predicted for individuals and substantially reduced by appropriate secondary prevention measures.