Epidemic surveillance using an electronic medical record: an empiric approach to performance improvement.
Electronic medical records (EMR) form a rich repository of information that could benefit public health. We asked how structured and free-text narrative EMR data should be combined to improve epidemic surveillance for acute respiratory infections (ARI). Eight previously characterized ARI case detection algorithms (CDA) were applied to historical EMR entries to create authentic time series of daily ARI case counts (background). An epidemic model simulated influenza cases (injection). From the time of the injection, cluster-detection statistics were applied daily on paired background+injection (combined) and background-only time series. This cycle was then repeated with the injection shifted to each week of the evaluation year. We computed: a) the time from injection to the first statistical alarm uniquely found in the combined dataset (Detection Delay); b) how often alarms originated in the background-only dataset (false-alarm rate, or FAR); and c) the number of cases found within these false alarms (Caseload). For each CDA, we plotted the Detection Delay as a function of FAR or Caseload, over a broad range of alarm thresholds. CDAs that combined text analyses seeking ARI symptoms in clinical notes with provider-assigned diagnostic codes in order to maximize the precision rather than the sensitivity of case-detection lowered Detection Delay at any given FAR or Caseload. An empiric approach can guide the integration of EMR data into case-detection methods that improve both the timeliness and efficiency of epidemic detection.


Effects of yoga interventions on pain and pain-associated disability: a meta-analysis.
We searched databases for controlled clinical studies, and performed a meta-analysis on the effectiveness of yoga interventions on pain and associated disability. Five randomized studies reported single-blinding and had a higher methodological quality; 7 studies were randomized but not blinded and had moderate quality; and 4 nonrandomized studies had low quality. In 6 studies, yoga was used to treat patients with back pain; in 2 studies to treat rheumatoid arthritis; in 2 studies to treat patients with headache/migraine; and 6 studies enrolled individuals for other indications. All studies reported positive effects in favor of the yoga interventions. With respect to pain, a random effect meta-analysis estimated the overall treatment effect at SMD = -.74 (CI: -.97; -.52, P < .0001), and an overall treatment effect at SMD = -.79 (CI: -1.02; -.56, P < .0001) for pain-related disability. Despite some limitations, there is evidence that yoga may be useful for several pain-associated disorders. Moreover, there are hints that even short-term interventions might be effective. Nevertheless, large-scale further studies have to identify which patients may benefit from the respective interventions. This meta-analysis suggests that yoga is a useful supplementary approach with moderate effect sizes on pain and associated disability.


Estimating the burden of antimicrobial resistance in Malawi: protocol for a prospective observational study of the morbidity, mortality and economic cost of third-generation cephalosporin resistant bloodstream infection.
Introduction: Antimicrobial resistance (AMR) is a global public health concern, but the problems are context specific, with each county or setting facing differing challenges. In sub-Saharan Africa, third-generation cephalosporin resistant Enterobacterales (3GCR-E) are of particular concern, given the widespread reliance on ceftriaxone for treatment of severe infection in this setting. In Malawi, despite rising prevalence of 3GCR-E, the health-impact of these infections has not been described. This study is designed to estimate attributable mortality, morbidity and economic cost of 3GCR-E bloodstream infection (BSI) in a large, urban hospital. Methods: This study will investigate the burden of AMR by recruiting a a prospective longitudinal cohort of patients who have bloodstream infection with 3GCR-E, at Queen Elizabeth Central Hospital, Blantyre, Malawi. Patients whose blood culture is positive for either third-generation cephalosporin susceptible (3GC-S) or third-generation resistant (3GC-R) Enterobacterales will be enrolled and provide clinical and healthcare economic data. Patients will be followed throughout their hospital stay and to 6-months post discharge. The primary outcomes for the study are mortality and morbidity from 3GCR-E. Healthcare economic outcomes will be assessed by comparing healthcare provider costs, indirect patient costs and health-related quality of life outcomes in patients with 3GC-S and 3GC-R BSI. Based on our observation that some patients with clinical suspicion of sepsis and 3GC-R BSI are surviving without an effective antibiotic, we review each patient prospectively and classify what role the isolated bacteria is playing in the patient's clinical presentation. Each BSI episode will be classified into the following categories: definite Gram-negative sepsis, probable Gram-negative sepsis, transient or occult bacteraemia, or contaminated blood culture. These classifications will be incorporated into our analysis. Ethics and dissemination: The study protocol has been approved by the Malawi College of Medicine Research Ethics Committee and by the Liverpool School of Tropical Medicine Research Ethics committee.


Dissociation of nitric oxide synthase immunoreactivity and NADPH-diaphorase enzyme activity in rat pituitary.
The relationship between nitric oxide synthase (NOS) immunocytochemistry and NADPH-diaphorase (NADPH-d) histochemistry was investigated in the anterior and posterior pituitary of ovariectomized rats. NADPH-d activity was present throughout the posterior pituitary but could not be detected in the anterior pituitary. By contrast, an antibody raised against whole recombinant rat neuronal NOS revealed strongly NOS-immunoreactive gonadotrophs and folliculo-stellate cells scattered throughout the anterior pituitary in addition to immunoreactive fibres in the posterior pituitary. The study demonstrates that NADPH-d histochemistry is not always coincident with NOS and provides evidence for an as yet uncharacterized subtype of NOS in the rat anterior pituitary.


The role of medical necessity and cost-effectiveness in making medical decisions.
The term "medical necessity" is used ubiquitously in health care, but its meaning and implementation vary substantially among providers, payers, and patients. This ambiguity has led some to suggest that cost-effectiveness be used as a basis for decision rules. This paper presents an analytical framework that is familiar to clinicians and shows that medical necessity and cost-effectiveness do not provide deterministic rules for clinical decision making. First, 2 x 2 tables are used to show the tradeoff between the sensitivity and specificity of decision rules. Then, the example of asymptomatic abdominal aortic aneurysm is used to show that these tradeoffs can be seen as a continuum of decision rules on a receiver-operating characteristic curve. Society can therefore choose a decision threshold on the basis of medical necessity that optimizes the number of lives saved or any other desired outcome, but the tradeoff between sensitivity and specificity cannot be avoided. Applying cost-effectiveness criteria may change the decision threshold because cost-effectiveness itself involves inherent tradeoffs that create additional ambiguity for clinical decisions. The conclusion is that decision rules based on medical necessity or cost-effectiveness should not be considered deterministic. Rather, decision rules are useful when they make assumptions explicit and specify tradeoffs so that clinicians, patients, and payers can make better decisions.


The mutational landscape of lethal castration-resistant prostate cancer.
Characterization of the prostate cancer transcriptome and genome has identified chromosomal rearrangements and copy number gains and losses, including ETS gene family fusions, PTEN loss and androgen receptor (AR) amplification, which drive prostate cancer development and progression to lethal, metastatic castration-resistant prostate cancer (CRPC). However, less is known about the role of mutations. Here we sequenced the exomes of 50 lethal, heavily pre-treated metastatic CRPCs obtained at rapid autopsy (including three different foci from the same patient) and 11 treatment-naive, high-grade localized prostate cancers. We identified low overall mutation rates even in heavily treated CRPCs (2.00 per megabase) and confirmed the monoclonal origin of lethal CRPC. Integrating exome copy number analysis identified disruptions of CHD1 that define a subtype of ETS gene family fusion-negative prostate cancer. Similarly, we demonstrate that ETS2, which is deleted in approximately one-third of CRPCs (commonly through TMPRSS2:ERG fusions), is also deregulated through mutation. Furthermore, we identified recurrent mutations in multiple chromatin- and histone-modifying genes, including MLL2 (mutated in 8.6% of prostate cancers), and demonstrate interaction of the MLL complex with the AR, which is required for AR-mediated signalling. We also identified novel recurrent mutations in the AR collaborating factor FOXA1, which is mutated in 5 of 147 (3.4%) prostate cancers (both untreated localized prostate cancer and CRPC), and showed that mutated FOXA1 represses androgen signalling and increases tumour growth. Proteins that physically interact with the AR, such as the ERG gene fusion product, FOXA1, MLL2, UTX (also known as KDM6A) and ASXL1 were found to be mutated in CRPC. In summary, we describe the mutational landscape of a heavily treated metastatic cancer, identify novel mechanisms of AR signalling deregulated in prostate cancer, and prioritize candidates for future study.


Dental Stem Cells and their Applications in Dental Tissue Engineering.
Tooth loss or absence is a common condition that can be caused by various pathological circumstances. The replacement of the missing tooth is important for medical and aesthetic reasons. Recently, scientists focus on tooth tissue engineering, as a potential treatment, beyond the existing prosthetic methods. Tooth engineering is a promising new therapeutic approach that seeks to replace the missing tooth with a bioengineered one or to restore the damaged dental tissue. Its main tool is the stem cells that are seeded on the surface of biomaterials (scaffolds), in order to create a biocomplex. Several populations of mesenchymal stem cells are found in the tooth. These different cell types are categorized according to their location in the tooth and they demonstrate slightly different features. It appears that the dental stem cells isolated from the dental pulp and the periodontal ligament are the most powerful cells for tooth engineering. Additional research needs to be performed in order to address the problem of finding a suitable source of epithelial stem cells, which are important for the regeneration of the enamel. Nevertheless, the results of the existing studies are encouraging and strongly support the belief that tooth engineering can offer hope to people suffering from dental problems or tooth loss.


Web-based parenting skills to reduce behavior problems following abusive head trauma: a pilot study.
Pediatric abusive head trauma causes significant cognitive and behavioral morbidity, yet very few post-acute interventions exist to facilitate long-term recovery. To meet the needs of this vulnerable population, we piloted a web-based intervention with live coaching designed to improve positive parenting and child behavior. The efficacy of this parenting skills intervention was compared with access to Internet resources on brain injury. Participants included seven families (four randomized to the parenting intervention and three randomized to receive Internet resources). Parenting skills were observed and child behavior was rated at baseline and intervention completion. At completion, parents who received the parenting skills intervention showed significantly more positive parenting behaviors and fewer undesirable behaviors during play than parents who received access to Internet resources. Additionally, during play, children in the parenting skills intervention group were more compliant following parent commands than children in the Internet resources group. Lastly, parents who received the parenting intervention reported less intense oppositional and conduct behavior problems in their children post-intervention than did parents in the Internet resources group. These findings provide preliminary evidence for the use of this web-based positive parenting skills intervention to improve parenting skills and child behavior following abusive head trauma.


Removal of endocrine disrupting compounds from wastewater using polymer particles.
This study evaluated the use of particles of molecularly imprinted and non-imprinted polymers (MIP and NIP) as a wastewater treatment method for endocrine disrupting compounds (EDCs). MIP and NIP remove EDCs through adsorption and therefore do not result in the formation of partially degraded products. The results show that both MIP and NIP particles are effective for removal of EDCs, and NIP have the advantage of not being as compound-specific as the MIP and hence can remove a diverse range of compounds including 17-β-estradiol (E2), atrazine, bisphenol A, and diethylstilbestrol. Removal of E2 from wastewater was also tested to determine the effectiveness of NIP in the presence of interfering substances and natural organic matter. Removal of E2 from wastewater samples was high and increased with increasing NIP. NIP represent an effective way of removing a wide variety of EDCs from wastewater.


Structural analysis of phosphorylation-associated interactions of human MCC with Scribble PDZ domains.
Scribble is a crucial adaptor protein that plays a pivotal role during establishment and control of cell polarity, impacting many physiological processes ranging from cell migration to immunity and organization of tissue architecture. Scribble harbours a leucine-rich repeat domain and four PDZ domains that mediate most of Scribble's interactions with other proteins. It has become increasingly clear that post-translational modifications substantially impact Scribble-ligand interactions, with phosphorylation being a major modulator of binding to Scribble. To better understand how Scribble PDZ domains direct cell polarity signalling and how phosphorylation impacts this process, we investigated human Scribble interactions with MCC (Mutated in Colorectal Cancer). We systematically evaluated the ability of all four individual Scribble PDZ domains to bind the PDZ-binding motif (PBM) of MCC as well as MCC phosphorylated at the -1 Ser position. We show that Scribble PDZ1 and PDZ3 are the major interactors with MCC, and that modifications to Ser at the -1 position in the MCC PBM only has a minor effect on binding to Scribble PDZ domains. We then examined the structural basis for these observations by determining the crystal structures of Scribble PDZ1 domain bound to both the unphosphorylated MCC PBM as well as phosphorylated MCC. Our structures indicated that phospho-Ser at the -1 position in MCC is not involved in major contacts with Scribble PDZ1, and in conjunction with our affinity measurements suggest that the impact of phosphorylation at the -1 position of MCC must extend beyond a simple modulation of the affinity for Scribble PDZ domains.


Mi-2 complex couples DNA methylation to chromatin remodelling and histone deacetylation.
Methylation of DNA at the dinucleotide CpG is essential for mammalian development and is correlated with stable transcriptional silencing. This transcriptional silencing has recently been linked at a molecular level to histone deacetylation through the demonstration of a physical association between histone deacetylases and the methyl CpG-binding protein MeCP2 (refs 4,5). We previously purified a histone deacetylase complex from Xenopus laevis egg extracts that consists of six subunits, including an Rpd3-like deacetylase, the RbA p48/p46 histone-binding protein and the nucleosome-stimulated ATPase Mi-2 (ref. 6). Similar species were subsequently isolated from human cell lines, implying functional conservation across evolution. This complex represents the most abundant form of deacetylase in amphibian eggs and cultured mammalian cells. Here we identify the remaining three subunits of this enzyme complex. One of them binds specifically to methylated DNA in vitro and molecular cloning reveals a similarity to a known methyl CpG-binding protein. Our data substantiate the mechanistic link between DNA methylation, histone deacetylation and transcriptional silencing.


Transcriptional architecture and chromatin landscape of the core circadian clock in mammals.
The mammalian circadian clock involves a transcriptional feed back loop in which CLOCK and BMAL1 activate the Period and Cryptochrome genes, which then feedback and repress their own transcription. We have interrogated the transcriptional architecture of the circadian transcriptional regulatory loop on a genome scale in mouse liver and find a stereotyped, time-dependent pattern of transcription factor binding, RNA polymerase II (RNAPII) recruitment, RNA expression, and chromatin states. We find that the circadian transcriptional cycle of the clock consists of three distinct phases: a poised state, a coordinated de novo transcriptional activation state, and a repressed state. Only 22% of messenger RNA (mRNA) cycling genes are driven by de novo transcription, suggesting that both transcriptional and posttranscriptional mechanisms underlie the mammalian circadian clock. We also find that circadian modulation of RNAPII recruitment and chromatin remodeling occurs on a genome-wide scale far greater than that seen previously by gene expression profiling.


Activation of methionine synthase by insulin-like growth factor-1 and dopamine: a target for neurodevelopmental toxins and thimerosal.
Methylation events play a critical role in the ability of growth factors to promote normal development. Neurodevelopmental toxins, such as ethanol and heavy metals, interrupt growth factor signaling, raising the possibility that they might exert adverse effects on methylation. We found that insulin-like growth factor-1 (IGF-1)- and dopamine-stimulated methionine synthase (MS) activity and folate-dependent methylation of phospholipids in SH-SY5Y human neuroblastoma cells, via a PI3-kinase- and MAP-kinase-dependent mechanism. The stimulation of this pathway increased DNA methylation, while its inhibition increased methylation-sensitive gene expression. Ethanol potently interfered with IGF-1 activation of MS and blocked its effect on DNA methylation, whereas it did not inhibit the effects of dopamine. Metal ions potently affected IGF-1 and dopamine-stimulated MS activity, as well as folate-dependent phospholipid methylation: Cu(2+) promoted enzyme activity and methylation, while Cu(+), Pb(2+), Hg(2+) and Al(3+) were inhibitory. The ethylmercury-containing preservative thimerosal inhibited both IGF-1- and dopamine-stimulated methylation with an IC(50) of 1 nM and eliminated MS activity. Our findings outline a novel growth factor signaling pathway that regulates MS activity and thereby modulates methylation reactions, including DNA methylation. The potent inhibition of this pathway by ethanol, lead, mercury, aluminum and thimerosal suggests that it may be an important target of neurodevelopmental toxins.


Effects of strength training on bone mineral density: hormonal and bone turnover relationships.
The effects of a 16-wk strength-training program on bone mineral density (BMD) was assessed by dual-energy X-ray absorptiometry in 21 men [age 61 +/- 1 (SE) yr]. Sixteen men (age 59 +/- 2 yr) served as control subjects. To investigate the possible hormonal relationships underlying the effects on BMD, serum concentrations of growth hormone, insulin-like growth factor I, and testosterone were determined before and after training. In addition, osteocalcin and skeletal alkaline phosphatase (markers of bone formation) and tartrate-resistant acid phosphatase (a marker of bone resorption) were measured before and after training to assess bone turnover. The training program resulted in a 2.8 +/- 0.6% increase in femoral neck BMD (1.004 +/- 0.037 vs. 1.031 +/- 0.037 g/cm2; P < 0.001). However, there were no significant changes in total body, anterioposterior spine, lateral spine, Ward's triangle, or greater trochanter BMD. Moreover, there were no significant changes in growth hormone, insulin-like growth factor I, testosterone, osteocalcin, or skeletal alkaline phosphatase. There were no changes in the control group. Thus, strength training can increase femoral neck BMD, and this effect does not appear to be accompanied by changes in anabolic hormones or markers of bone formation and resorption.


CCP1, a Tubulin Deglutamylase, Increases Survival of Rodent Spinal Cord Neurons following Glutamate-Induced Excitotoxicity.
Microtubules (MTs) are cytoskeletal elements that provide structural support and act as roadways for intracellular transport in cells. MTs are also needed for neurons to extend and maintain long axons and dendrites that establish connectivity to transmit information through the nervous system. Therefore, in neurons, the ability to independently regulate cytoskeletal stability and MT-based transport in different cellular compartments is essential. Posttranslational modification of MTs is one mechanism by which neurons regulate the cytoskeleton. The carboxypeptidase CCP1 negatively regulates posttranslational polyglutamylation of MTs. In mammals, loss of CCP1, and the resulting hyperglutamylation of MTs, causes neurodegeneration. It has also long been known that CCP1 expression is activated by neuronal injury; however, whether CCP1 plays a neuroprotective role after injury is unknown. Using shRNA-mediated knock-down of CCP1 in embryonic rat spinal cord cultures, we demonstrate that CCP1 protects spinal cord neurons from excitotoxic death. Unexpectedly, excitotoxic injury reduced CCP1 expression in our system. We previously demonstrated that the CCP1 homolog in Caenorhabditis elegans is important for maintenance of neuronal cilia. Although cilia enhance neuronal survival in some contexts, it is not yet clear whether CCP1 maintains cilia in mammalian spinal cord neurons. We found that knock-down of CCP1 did not result in loss or shortening of cilia in cultured spinal cord neurons, suggesting that its effect on survival of excitotoxicity is independent of cilia. Our results support the idea that enzyme regulators of MT polyglutamylation might be therapeutically targeted to prevent excitotoxic death after spinal cord injuries.


Green tea, phytic acid, and inositol in combination reduced the incidence of azoxymethane-induced colon tumors in Fisher 344 male rats.
Experimental as well as epidemiologic studies in human populations provide evidence that consumption of phytochemicals reduces the incidence of degenerative diseases. Green tea (GT) catechins are known for their antioxidative potential. Phytic acid (PA) also acts as a natural antioxidant and may have numerous health benefits. This experiment was designed to investigate the inhibitory effects of combinations of 1% and 2% GT, PA, and inositol (I) in reducing the incidence of azoxymethane-induced colon tumors in Fisher 344 male rats. After an acclimatization period of 1 week, nine groups of rats (15 rats per group) were initially assigned to consume AIN 93 G diet and later AIN 93 M diet after 20 weeks of age. Treatments were given in drinking water. All rats received azoxymethane injections (16 mg/kg of body weight) subcutaneously at 7 and 8 weeks of age. Rats were killed at 45 weeks of age by CO(2) euthanasia. Tumor incidence (93.76%) and the number of tumors per tumor-bearing rat ratio (2.25) were significantly (P<.05) higher in the control group compared with treatment groups. Glutathione S-transferase activity was significantly (P<.05) higher in rats fed combinations of 2% GT+PA+I and GT+PA (33.25 ± 1.23 and 29.83 ± 1.10 μmol/mL, respectively) compared with other groups. These findings suggest that the synergistic effect of the 2% level of GT, PA, and I may reduce the incidence of colon tumors and therefore have potential as a chemopreventive agent.


A gigantic shark from the lower cretaceous duck creek formation of Texas.
Three large lamniform shark vertebrae are described from the Lower Cretaceous of Texas. We interpret these fossils as belonging to a single individual with a calculated total body length of 6.3 m. This large individual compares favorably to another shark specimen from the roughly contemporaneous Kiowa Shale of Kansas. Neither specimen was recovered with associated teeth, making confident identification of the species impossible. However, both formations share a similar shark fauna, with Leptostyrax macrorhiza being the largest of the common lamniform sharks. Regardless of its actual identification, this new specimen provides further evidence that large-bodied lamniform sharks had evolved prior to the Late Cretaceous.


Diagnosis of early stage knee osteoarthritis based on early clinical course: data from the CHECK cohort.
Early diagnosis of knee osteoarthritis (OA) is important in managing this disease, but such an early diagnostic tool is still lacking in clinical practice. The purpose of this study was to develop diagnostic models for early stage knee OA based on the first 2-year clinical course after the patient's initial presentation in primary care and to identify whether these course factors had additive discriminative value over baseline factors. We extracted eligible patients' clinical and radiographic data from the CHECK cohort and formed the first 2-year course factors according to the factors' changes over the 2 years. Clinical expert consensus-based diagnosis, which was made via evaluating patients' 5- to 10-year follow-up data, was used as the outcome factor. Four models were developed: model 1, included clinical course factors only; model 2, included clinical and radiographic course factors; model 3, clinical baseline factors + clinical course factors; and model 4, clinical and radiographic baseline factors + clinical and radiographic course factors. All the models were built by a generalized estimating equation with a backward selection method. Area under the receiver operating characteristic curve (AUC) and its 95% confidence interval (CI) were calculated for assessing model discrimination. Delong's method compared AUCs. Seven hundred sixty-one patients with 1185 symptomatic knees were included in this study. Thirty-seven percent knees were diagnosed as OA at follow-up. Model 1 contained 6 clinical course factors; model 2: 6 clinical and 3 radiographic course factors; model 3: 6 baseline clinical factors combined with 5 clinical course factors; and model 4: 4 clinical and 1 radiographic baseline factors combined with 5 clinical and 3 radiographic course factors. Model discriminations are as follows: model 1, AUC 0.70 (95% CI 0.67-0.74); model 2, 0.74 (95% CI 0.71-0.77); model 3, 0.77 (95% CI 0.74-0.80); and model 4, 0.80 (95% CI 0.77-0.82). AUCs of model 3 and model 4 were slightly but significantly higher than corresponding baseline-factor models (model 3 0.77 vs 0.75, p = 0.031; model 4 0.80 vs 0.76, p = 0.003). Four diagnostic models were developed with "fair" to "good" discriminations. First 2-year course factors had additive discriminative value over baseline factors.


Early metabolic response to chemoradiotherapy by interim FDG PET/CT is associated with better overall survival and histological response in esophageal cancers.
Early assessment of response to neoadjuvant chemoradiotherapy (CRT) is crucial in determining the most suitable treatment strategy in locally advanced oesophageal cancer (LAEC). We evaluated the impact of early metabolic response during CRT on overall survival (OS) and histological response. Patients with biopsy-proven oesophageal carcinoma underwent FDG PET/CT with evaluation of the standardized uptake value (SUV) before any treatment and during CRT after 20 Gy. 116 patients (Male: 66.4%, Median age: 63; squamous cell carcinomas (SCC): 70%) met inclusion criteria. Median OS was 21.7 months. There was a significant positive correlation between interim metabolic response and OS. In multivariate analysis, only metabolic response using the 50% cut-off value remained significantly associated with OS (IC95% = 0.28-0.73; p = 0.001). In this statistical analysis, surgery (p = 0.007) and T stage (p = 0.023) were also correlated with OS. There was a significant correlation between early metabolic response and local recurrence (Chi-squared test p = 0.0001). Early metabolic response using FDG PET/CT is associated with better OS, disease-free survival, local control and pathological response in patients treated by CRT for LAEC.


A novel molecularly imprinted method with computational simulation for the affinity isolation and knockout of baicalein from Scutellaria baicalensis.
A novel molecularly imprinted polymer (MIP) was synthesized by precipitation polymerization with baicalein (BAI) as the template and used as solid-phase extraction (SPE) adsorbent, aiming at the affinity isolation and selective knockout of BAI from Scutellaria baicalensis Georgi (SB). We used computational simulation to predict the optimal functional monomer, polymerization solvent and molar ratio of template to functional monomer. Characterization and performance tests revealed that MIP exhibited uniform spherical morphology, rapid binding kinetics, and higher adsorption capacity for BAI compared with nonimprinted polymer (NIP). The application of MIP in SPE coupled with high-performance liquid chromatography to extract BAI from SB showed excellent recovery (94.3%) and purity (97.0%). Not only the single BAI compound, but also the BAI-removed SB extract was obtained by one-step process. This new method is useful for isolation and knockout of key bioactive compounds from herbal medicines.


Differential-phase-shift quantum secret sharing.
A quantum secret sharing (QSS) protocol based on a differential-phase-shift scheme is proposed, which quantum mechanically provides a full secret key to one party and partial keys to two other parties. A weak coherent pulse train is utilized instead of individual photons as in conventional schemes. Compared with previous QSS protocols, the present one features a simple setup, is suitable for fiber transmission, and offers the possibility for a high key creation rate. An experiment is also carried out to demonstrate the operation.


Generation of constitutive and inducible trans-sialylation dominant-negative phenotypes in Trypanosoma brucei and Trypanosoma cruzi.
Trans-sialylation is a unique enzymatic process that is restricted to some trypanosome species. By expressing developmentally regulated trans-sialidases, these protozoan parasites cleave sialic acids from host glycoconjugates and transfer them to acceptors on their own cell surfaces. The biological function of this process is not understood, but trans-sialylation is expected to be important in the invasion of mammalian cells by Trypanosoma cruzi and the survival of Trypanosoma brucei within its insect vector. Since a conventional gene knockout approach was precluded, we developed a dominant-negative strategy, in which fusion proteins consisting of a bacterial sialidase and trypanosome proteins were expressed in T.brucei and T.cruzi. The strong recombinant sialidase activity shifted the reaction equilibrium from sialic acid transfer to hydrolysis, in this way creating a sialic-acid-negative phenotype. Taking advantage of a recently introduced inducible expression system, we were able to control the expression of sialidase fusion proteins in T.brucei. Reversion of the sialic-acid-negative state to wild-type sialylation was accomplished by selective inhibition of the foreign sialidase, leaving the parasite trans-sialidase unaffected. Both desialylation and resialylation of trypanosomes was rapidly achieved. Our results show that neither T.brucei nor T.cruzi require sialic acids for survival in vitro, ruling out the involvement of sialylation in cell surface integrity. The versatile system introduced here will allow a detailed in vivo study of the role of trans-sialylation during the trypanosome infection cycle. Furthermore, cell-surface sialic acids are implicated in a multitude of (patho-) biochemical processes in other organisms. The quantitative and qualitative manipulation of cell surface sialic acids, by expressing of counteracting enzymes, constitutes a novel approach with potentially broad applications in glycobiology.


Microbial diversity and biogeochemistry of the Guaymas Basin deep-sea hydrothermal plume.
Hydrothermal plumes are hot spots of microbial biogeochemistry in the deep ocean, yet little is known about the diversity or ecology of microorganisms inhabiting plumes. Recent biogeochemical evidence shows that Mn(II) oxidation in the Guaymas Basin (GB) hydrothermal plume is microbially mediated and suggests that the plume microbial community is distinct from deep-sea communities. Here we use a molecular approach to compare microbial diversity in the GB plume and in background deep seawater communities, and cultivation to identify Mn(II)-oxidizing bacteria from plumes and sediments. Despite dramatic differences in Mn(II) oxidation rates between plumes and background seawater, microbial diversity and membership were remarkably similar. All bacterial clone libraries were dominated by Gammaproteobacteria and archaeal clone libraries were dominated by Crenarchaeota. Two lineages, both phylogenetically related to methanotrophs and/or methylotrophs, were consistently over-represented in the plume. Eight Mn(II)-oxidizing bacteria were isolated, but none of these or previously identified Mn(II) oxidizers were abundant in clone libraries. Taken together with Mn(II) oxidation rates measured in laboratory cultures and in the field, these results suggest that Mn(II) oxidation in the GB hydrothermal plume is mediated by genome-level dynamics (gene content and/or expression) of microorganisms that are indigenous and abundant in the deep sea but have yet to be unidentified as Mn(II) oxidizers.


Synchronicity of frequently sampled thyrotropin (TSH) and leptin concentrations in healthy adults and leptin-deficient subjects: evidence for possible partial TSH regulation by leptin in humans.
Leptin signals the status of energy reserves to the brain. Leptin stimulates biosynthesis of TRH in vitro and influences the activity of the hypothalamic-pituitary-thyroid axis in vivo in rodents. Because blood levels of both leptin and TSH display diurnal variation with a distinct nocturnal rise, we sought to determine whether a relationship exists between fluctuations in circulating leptin and TSH. We measured serum leptin and TSH levels every 7 min for 24 h in five healthy men and found that both leptin and TSH levels are highly organized and pulsatile. A similar pattern of leptin and TSH rhythms was observed, with TSH and leptin levels reaching a nadir in late morning and a peak in the early morning hours. Importantly, cosinor analysis on the absolute leptin and TSH levels revealed a statistically significant fit for a 24-h period and the two hormones showed similar probabilities of rhythm and superimposable peak values. Furthermore, this study shows a strong positive Pearson correlation between the 24-h patterns of variability of leptin and TSH in healthy subjects. Finally, the ultradian fluctuations in leptin levels showed pattern synchrony with those of TSH as determined by cross-correlation analysis, by cross-approximate enthropy and Bayessian analysis applied independently. To further explore whether these associations could reflect an underlying regulation of TSH secretion by leptin, we also studied frequently sampled leptin and TSH levels in four brothers, members of a family with leptin deficiency (one normal homozygote, two heterozygotes, and one leptin-deficient homozygote). Leptin levels of the homozygous leptin-deficient subject are detectable but bioinactive, and the rhythm of his TSH is disorganized. 24-h pattern of leptin and TSH variability in the heterozygous subjects, although significantly correlated, showed a weaker correlation compared with the strong correlation in the normal subjects. These data are consistent with the possibility that leptin may regulate TSH pulsatility and circadian rhythmicity, but interventional studies are needed to definitively prove whether leptin regulates the minute-to-minute oscillations and ultradian rhythm of TSH levels.


Red-Green Tactical Lighting Is Preferred for Suturing Wounds in a Simulated Night Environment.
Delivering medical care in nighttime conditions is challenging, as 25% of Special Operations medical Operators have reported that problems with lighting contributed to poor casualty outcomes. Red light is often used in nighttime operations but makes blood detection difficult and diminishes depth perception and visual acuity. Red-green combination lighting may be superior for differentiating blood from tissue and other fluids but had not been tested versus red-only or green-only lighting for combat-related medical procedures, such as wound suturing. Dark-adapted medical resident physicians (N = 24) sutured 6cm long, 3cm deep, full-thickness lacerations in deceased swine under red-only, green-only, and red-green lighting provided by a tactical flashlight using a randomized within-subjects design. Time to suture completion, suture quality, user ratings, and user preference data were contrasted at p < .05. This study was approved by Naval Medical Center Portsmouth IRB. Suture completion time and suture quality were similar across all lighting conditions. Participants rated red-green lighting as significantly easier for identifying blood, identifying instruments, and performing suturing (p < .01). Red-green lighting was preferred by 83% of participants compared to 8% each for red-only and green-only (p < .001). Pending further study under tactical conditions, red-green lighting is tentatively recommended for treating battlefield wounds in low-light environments.


Quantitative trait loci affecting methamphetamine responses in BXD recombinant inbred mouse strains.
Individual differences in most behavioral and pharmacological responses to abused drugs are dependent on both genetic and environmental factors. The genetic influences on the complex phenotypes related to drug abuse have been difficult to study using classical genetic analyses. Quantitative trait locus (QTL) mapping is a method that has been used successfully to examine genetic contributions to some of these traits by correlating allelic variation in polymorphic genetic markers of known chromosomal location with variation in drug-response phenotypes. We evaluated several behavioral responses to multiple doses of methamphetamine (METH) in C57BL/6J (B6), DBA/2J (D2), and 25 of their recombinant inbred (BXD RI) strains. Stereotyped chewing, horizontal home cage activity, and changes in body temperature after 0, 4, 8, or 16 mg/kg METH, as well as stereotyped climbing behavior after 16 mg/kg METH, were examined. Associations (p < 0.01) between METH sensitivity and allelic status at multiple microsatellite genetic markers were subsequently determined for each response. QTLs were provisionally identified for each phenotype, some unique to a particular behavior and others that appeared to influence multiple phenotypes. Candidate genes suggested by these analyses included several that mapped near genes relevant for the neurotransmitters acetylcholine and glutamate. The locations of QTLs provisionally identified by this analysis were compared with QTLs hypothesized in other studies to influence methamphetamine- and cocaine-related phenotypes. In several instances, QTLs appeared to overlap, which is consistent with idea that common neural substrates underlie some responses to psychostimulants.


Myoepithelial cell staining patterns of papillary breast lesions: from intraductal papillomas to invasive papillary carcinomas.
We evaluated 25 intraductal papillomas and 18 papillary carcinomas (invasive, 4; micropapillary ductal carcinoma in situ [DCIS], 5; cases originally classified as intracystic/intraductal papillary carcinoma, 9) by calponin, smooth muscle myosin heavy chain (SMM-HC), and p63 immunostains. Calponin, SMM-HC, and p63 labeled myoepithelial cells (MECs) in all intraductal papillomas and all micropapillary DCIS cases. The invasive papillary carcinoma cases were uniformly negative for all stains. The 9 cases originally diagnosed as intracystic/intraductal papillary carcinoma showed more variable results, with identification of an MEC layer in only 4 cases. Comparison of staining of MECs by these 3 stains showed that calponin was more sensitive and intense than SMM-HC; however, there was cross-reactivity with myofibroblastic cells. Staining with p63 was discontinuous, making interpretation of an intact myoepithelial layer difficult. Of 9 cases originally classified as intraductal papillary carcinoma, 5 showed absence of a basal MEC layer by immunohistochemical analysis. The lack of a basal MEC layer in these cases suggests a spectrum of progression from in situ to invasive disease and might help explain distant metastases from previously reported "intraductal papillary carcinoma."


Efficacy and safety of canagliflozin when used in conjunction with incretin-mimetic therapy in patients with type 2 diabetes.
To assess the efficacy and safety of canagliflozin, a sodium glucose co-transporter 2 (SGLT2) inhibitor, in patients with type 2 diabetes enrolled in the CANagliflozin cardioVascular Assessment Study (CANVAS) who were on an incretin mimetic [dipeptidyl peptidase-4 (DPP-4) inhibitor or glucagon-like peptide-1 (GLP-1) receptor agonist]. CANVAS is a double-blind, placebo-controlled study that randomized participants to canagliflozin 100 or 300 mg or placebo added to routine therapy. The present post hoc analysis assessed the efficacy and safety of canagliflozin 100 and 300 mg compared with placebo in subsets of patients from CANVAS who were taking background DPP-4 inhibitors or GLP-1 receptor agonists with or without other antihyperglycaemic agents at week 18. Of the 4330 patients in CANVAS, 316 were taking DPP-4 inhibitors and 95 were taking GLP-1 receptor agonists. At 18 weeks, canagliflozin 100 and 300 mg provided larger placebo-subtracted reductions in glycated haemoglobin (HbA1c) in patients taking DPP-4 inhibitors [-0.56% (95% confidence interval [CI]: -0.77, -0.35), and -0.75% (95% CI: -0.95, -0.54), respectively] and GLP-1 receptor agonists [-1.00% (95% CI: -1.35, -0.65), and -1.06% (95% CI: -1.43, -0.69), respectively]. Body weight and blood pressure (BP) reductions were seen with canagliflozin versus placebo in both subsets. Higher incidences of genital mycotic infections and osmotic diuresis-related adverse events (AEs) were seen with canagliflozin compared with placebo. The incidence of hypoglycaemia was numerically higher with canagliflozin versus placebo; nearly all events occurred in patients on background insulin or insulin secretagogues. In patients on background incretin mimetics, canagliflozin improved HbA1c, body weight and BP, with an increased incidence of AEs related to SGLT2 inhibition.


Pseudoknots in prion protein mRNAs confirmed by comparative sequence analysis and pattern searching.
The human prion gene contains five copies of a 24 nt repeat that is highly conserved among species. An analysis of folding free energies of the human prion mRNA, in particular in the repeat region, suggested biased codon selection and the presence of RNA patterns. In particular, pseudoknots, similar to the one predicted by Wills in the human prion mRNA, were identified in the repeat region of all available prion mRNAs available in GenBank, but not those of birds and the red slider turtle. An alignment of these mRNAs, which share low sequence homology, shows several co-variations that maintain the pseudoknot pattern. The presence of pseudoknots in yeast Sup35p and Rnq1 suggests acquisition in the prokaryotic era. Computer generated three-dimensional structures of the human prion pseudoknot highlight protein and RNA interaction domains, which suggest a possible effect in prion protein translation. The role of pseudoknots in prion diseases is discussed as individuals with extra copies of the 24 nt repeat develop the familial form of Creutzfeldt-Jakob disease.


Activation of the interferon response by human cytomegalovirus occurs via cytoplasmic double-stranded DNA but not glycoprotein B.
In vitro infection of cells with the betaherpesvirus human cytomegalovirus (HCMV) stimulates an innate immune response characterized by phosphorylation of the transcription factor interferon regulatory factor 3 (IRF3) and subsequent expression of IRF3-dependent genes. While previous work suggests that HCMV envelope glycoprotein B is responsible for initiating this reaction, the signaling pathways stimulated by virus infection that lead to IRF3 phosphorylation have largely been uncharacterized. Recently, we identified Z DNA binding protein 1 (ZBP1), a sensor of cytoplasmic DNA, as an essential protein for this response. We now describe a human fibroblast cell line exhibiting a recessive defect that results in the absence of activation of IRF3 following treatment with HCMV but not Sendai virus or double-stranded RNA. In addition, we show that while exposure of these cells to soluble HCMV glycoprotein B is capable of triggering IRF3-dependent gene transcription, transfection of the cells with double-stranded DNA is not. Furthermore, we show that overexpression of ZBP1 in these cells reestablishes their ability to secrete interferon in response to HCMV and that multiple ZBP1 transcriptional variants exist in both wild-type and mutant cells. These results have two major implications for the understanding of innate immune stimulation by HCMV. First, they demonstrate that HCMV glycoprotein B is not the essential molecular pattern that induces an IRF3-dependent innate immune response. Second, IRF3-terminal signaling triggered by HCMV particles closely resembles that which is activated by cytoplasmic double-stranded DNA.


Association of serum ferritin levels with the risk of stomach cancer.
A group of 5908 men provided serum samples during their study examination from 1967 to 1970. After a surveillance period of over 20 years, 121 incident cases of tissue-confirmed gastric cancer were identified. Their stored sera and those of 121 matched controls from the study population were tested for serum ferritin and transferrin levels. Because of the suggested effects of previous thawing on the serum results, detailed data analyses were limited to the 46 cases and matched controls whose sera were never thawed before this study. The mean serum levels (In ng/ml) were 5.26 for the 46 gastric cancer cases and 5.68 for their controls (P < 0.01). For serum transferrin, the mean levels (mg/dl) were 249.8 for cases and 254.1 for controls (P = 0.53). The inverse association with serum ferritin, which reflect total iron body stores, was stronger for the 21 cases diagnosed within 15 years of examination (P = 0.02) than for the 25 cases diagnosed after 15 years (P = 0.15). The limitations of the study and the implications of its findings are discussed.


Food vacuole-associated lipid bodies and heterogeneous lipid environments in the malaria parasite, Plasmodium falciparum.
The malaria parasite Plasmodium falciparum induces a sixfold increase in the phospholipid content of infected erythrocytes during its intraerythrocytic growth. We have characterized the lipid environments in parasitized erythrocyte using the hydrophobic probe, Nile Red. Spectral imaging with a confocal microscope revealed heterogeneous lipid environments in parasite-infected erythrocytes. An insight into the nature of these environments was gained by comparing these spectra with those of triacylglycerol/phospholipid emulsions and phospholipid membranes. Using this approach, we identified a population of intensely stained particles of a few hundred nanometers in size that are closely associated with the digestive vacuole of the parasite and appear to be composed of neutral lipids. Electron microscopy and isolation of food vacuoles confirmed the size of these particles and their intimate association respectively. Lipid analysis suggests that these neutral lipid bodies are composed of di- and triacylgycerols and may represent storage organelles for lipid intermediates that are generated during digestion of phospholipids in the food vacuole. Mono-, di- and triacylglycerol suspensions promote beta-haematin formation, suggesting that these neutral lipid bodies, or their precursors, may also be involved in haem detoxification. We also characterized other compartments of the infected erythrocyte that were stained less intensely with the Nile Red probe. Both the erythrocyte membrane and the parasite membrane network exhibit red shifts compared with the neutral lipid bodies that are consistent with cholesterol-rich and cholesterol-poor membranes respectively. Ratiometric imaging revealed more subtle variations in the lipid environments within the parasite membrane network.


Fibroblast growth factor 21 reduces the severity of cerulein-induced pancreatitis in mice.
Fibroblast growth factor 21 (FGF21) acts as a hormonal regulator during fasting and is involved in lipid metabolism. Fgf21 gene expression is regulated by peroxisome proliferator-activated receptor (PPAR)-dependent pathways, which are enhanced during pancreatitis. Therefore, the aim of this study was to investigate FGF21's role in pancreatic injury. Fgf21 expression was quantified during cerulein-induced pancreatitis (CIP) or following mechanical or thapsigargin-induced stress through Northern blot analysis, in situ hybridization, and quantitative reverse transcription polymerase chain reaction. FGF21 protein was quantified by Western blot analysis. Isolated acinar cells or AR42J acinar cells were treated with recombinant FGF21 protein, and extracellular regulated kinase 1/2 activation was examined. The severity of CIP was compared between wild-type mice and mice overexpressing FGF21 (FGF21Tg) or harboring a targeted deletion of Fgf21 (Fgf21(-/-)). Acinar cell Fgf21 expression markedly increased during CIP and following injury in vitro. Purified FGF21 activated the extracellular regulated kinase 1/2 pathway in pancreatic acinar cells. The severity of CIP is inversely correlated to FGF21 expression because FGF21Tg mice exhibited decreased serum amylase and decreased pancreatic stellate cell activation, whereas Fgf21(-/-) mice had increased serum amylase and tissue damage. The expression of Fgf21 was also inversely correlated to expression of Early growth response 1, a proinflammatory and profibrotic transcription factor. These studies suggest a novel function for Fgf21 as an immediate response gene protecting pancreatic acini from overt damage.


Social and endocrine correlates of immune function in meerkats: implications for the immunocompetence handicap hypothesis.
Social status can mediate effects on the immune system, with profound consequences for individual health; nevertheless, most investigators of status-related disparities in free-ranging animals have used faecal parasite burdens to proxy immune function in the males of male-dominant species. We instead use direct measures of innate immune function (complement and natural antibodies) to examine status-related immunocompetence in both sexes of a female-dominant species. The meerkat is a unique model for such a study because it is a cooperatively breeding species in which status-related differences are extreme, evident in reproductive skew, morphology, behaviour, communication and physiology, including that dominant females naturally express the greatest total androgen (androstenedione plus testosterone) concentrations. We found that, relative to subordinates, dominant animals had reduced serum bacteria-killing abilities; also, relative to subordinate females, dominant females had reduced haemolytic complement activities. Irrespective of an individual's sex or social status, androstenedione concentrations (but not body condition, age or reproductive activity) negatively predicted concurrent immunocompetence. Thus, dominant meerkats of both sexes are immunocompromised. Moreover, in female meerkats, androstenedione perhaps acting directly or via local conversion, may exert a double-edged effect of promoting dominance and reproductive success at the cost of increased parasitism and reduced immune function. Given the prominent signalling of dominance in female meerkats, these findings may relate to the immunocompetence handicap hypothesis (ICHH); however, our data would suggest that the endocrine mechanism underlying the ICHH need not be mediated solely by testosterone and might explain trade-offs in females, as well as in males.


Characteristics of educational software use in 106 clinical laboratories.
The University of Washington, Seattle, has developed educational software for clinical laboratories. We used a 32-question survey to study software implementation. Of 106 clinical laboratories (response rate, 60%) that purchased the software and completed the survey, 89 laboratories (84%) that reported using the software formed the basis for the study. The most common software users were laboratory personnel, followed by medical technologist or medical laboratory technician students, residents, and medical students; the mean (SD) number of personnel categories using the software per laboratory was 1.8 (0.8). The most common reasons for use were initial instruction, cross-training, and competency assessment. The most frequent setting for software use was an area where laboratory testing occurred, followed by a dedicated training location, a location chosen by the employee, a classroom, and a distance learning mode. On a scale of 1 (poor) to 5 (excellent), the average satisfaction rating as an instructional tool was 4.4 and as a competency assessment tool, 4.2. Compared with laboratories in hospitals with 400 beds or fewer, laboratories in hospitals with more than 400 beds used the software for more categories of users (P = .008), had a higher proportion of laboratories using it for residents (P = .003), and had a higher proportion of laboratories with dedicated training areas (P = .02).


Endovascular Thrombus Removal for Acute Iliofemoral Deep Vein Thrombosis.
The ATTRACT trial (Acute Venous Thrombosis: Thrombus Removal with Adjunctive Catheter-Directed Thrombolysis) previously reported that pharmacomechanical catheter-directed thrombolysis (PCDT) did not prevent postthrombotic syndrome (PTS) in patients with acute proximal deep vein thrombosis. In the current analysis, we examine the effect of PCDT in ATTRACT patients with iliofemoral deep vein thrombosis. Within a large multicenter randomized trial, 391 patients with acute deep vein thrombosis involving the iliac or common femoral veins were randomized to PCDT with anticoagulation versus anticoagulation alone (No-PCDT) and were followed for 24 months to compare short-term and long-term outcomes. Between 6 and 24 months, there was no difference in the occurrence of PTS (Villalta scale ≥5 or ulcer: 49% PCDT versus 51% No-PCDT; risk ratio, 0.95; 95% CI, 0.78-1.15; P=0.59). PCDT led to reduced PTS severity as shown by lower mean Villalta and Venous Clinical Severity Scores ( P<0.01 for comparisons at 6, 12, 18, and 24 months), and fewer patients with moderate-or-severe PTS (Villalta scale ≥10 or ulcer: 18% versus 28%; risk ratio, 0.65; 95% CI, 0.45-0.94; P=0.021) or severe PTS (Villalta scale ≥15 or ulcer: 8.7% versus 15%; risk ratio, 0.57; 95% CI, 0.32-1.01; P=0.048; and Venous Clinical Severity Score ≥8: 6.6% versus 14%; risk ratio, 0.46; 95% CI, 0.24-0.87; P=0.013). From baseline, PCDT led to greater reduction in leg pain and swelling ( P<0.01 for comparisons at 10 and 30 days) and greater improvement in venous disease-specific quality of life (Venous Insufficiency Epidemiological and Economic Study Quality of Life unit difference 5.6 through 24 months, P=0.029), but no difference in generic quality of life ( P>0.2 for comparisons of SF-36 mental and physical component summary scores through 24 months). In patients having PCDT versus No-PCDT, major bleeding within 10 days occurred in 1.5% versus 0.5% ( P=0.32), and recurrent venous thromboembolism over 24 months was observed in 13% versus 9.2% ( P=0.21). In patients with acute iliofemoral deep vein thrombosis, PCDT did not influence the occurrence of PTS or recurrent venous thromboembolism. However, PCDT significantly reduced early leg symptoms and, over 24 months, reduced PTS severity scores, reduced the proportion of patients who developed moderate-or-severe PTS, and resulted in greater improvement in venous disease-specific quality of life. URL: https://www.clinicaltrials.gov . Unique identifier: NCT00790335.


[Infant botulism – why honey should be avoided for children up to one year].
Infant botulism - why honey should be avoided for children up to one year Infant botulism means that Clostridium botulinum colonize and produce toxin in the infant gut, usually during the first year of life. Illness severity varies widely and the incidence may be under-estimated. Infant botulism should be considered in cases of acute muscle weakness or floppiness in infants, especially when accompanied by constipation or feeding difficulties. Respiratory failure and need for mechanical ventilation is common, but full recovery is gradually obtained. Diagnosis is based on stool culture and toxin detection in stool. Botulinum spores are frequently present in honey, which should consequently be avoided for infants.


The effect of pathogen-specific clinical mastitis on the lactation curve for somatic cell count.
Data from 274 Dutch herds recording clinical mastitis (CM) over an 18-mo period were used to investigate the effect of pathogen-specific CM on the lactation curve for somatic cell count (SCC). Analyzed pathogens were Staphylococcus aureus, coagulase-negative staphylococci, Escherichia coli, Streptococcus dysgalactiae, Streptococcus uberis, other streptococci, and the culture-negative samples. The dataset contained 178,754 test-day records on SCC, recorded in 26,411 lactations of 21,525 cows of different parities. In lactations without both clinical and subclinical mastitis, SCC was high shortly after parturition, decreased to a minimum at 50 days in milk (DIM), and increased slowly toward the end of the lactation. Effects of CM on lactation curves for SCC differed among the pathogens isolated. Before a case of clinical E. coli mastitis occurred, SCC was close to the SCC of lactations without both clinical and subclinical mastitis, and after the case of CM had occurred, SCC returned rather quickly to a low level again. Similar curves were found for lactations with cases of CM associated with culture-negative samples. Before a case of clinical Staph. aureus mastitis occurred, average SCC was already high, and it remained high after the occurrence. Effects of CM associated with Strep. dysgalactiae, Strep. uberis, and other streptococci on the lactation curve for SCC were comparable. They showed a continuous increase in SCC until the case of pathogen-specific CM occurred, and afterwards SCC stayed at a higher level. Using SCC test-day records, these typical characteristics of each pathogen may be used to find more effective indicators of CM.


Clade I-Associated Mpox Cases Associated with Sexual Contact, the Democratic Republic of the Congo.
We report a cluster of clade I monkeypox virus infections linked to sexual contact in the Democratic Republic of the Congo. Case investigations resulted in 5 reverse transcription PCR-confirmed infections; genome sequencing suggest they belonged to the same transmission chain. This finding demonstrates that mpox transmission through sexual contact extends beyond clade IIb.


Lactate oxidation in human skeletal muscle mitochondria.
Lactate is an important intermediate metabolite in human bioenergetics and is oxidized in many different tissues including the heart, brain, kidney, adipose tissue, liver, and skeletal muscle. The mechanism(s) explaining the metabolism of lactate in these tissues, however, remains unclear. Here, we analyze the ability of skeletal muscle to respire lactate by using an in situ mitochondrial preparation that leaves the native tubular reticulum and subcellular interactions of the organelle unaltered. Skeletal muscle biopsies were obtained from vastus lateralis muscle in 16 human subjects. Samples were chemically permeabilized with saponin, which selectively perforates the sarcolemma and facilitates the loss of cytosolic content without altering mitochondrial membranes, structure, and subcellular interactions. High-resolution respirometry was performed on permeabilized muscle biopsy preparations. By use of four separate and specific substrate titration protocols, the respirometric analysis revealed that mitochondria were capable of oxidizing lactate in the absence of exogenous LDH. The titration of lactate and NAD(+) into the respiration medium stimulated respiration (P ≤ 0.003). The addition of exogenous LDH failed to increase lactate-stimulated respiration (P = 1.0). The results further demonstrate that human skeletal muscle mitochondria cannot directly oxidize lactate within the mitochondrial matrix. Alternately, these data support previous claims that lactate is converted to pyruvate within the mitochondrial intermembrane space with the pyruvate subsequently taken into the mitochondrial matrix where it enters the TCA cycle and is ultimately oxidized.


Additional molecular support for the new chordate phylogeny.
Recent phylogenomic analyses have suggested tunicates instead of cephalochordates as the closest living relatives of vertebrates. In direct contradiction with the long accepted view of Euchordates, this new phylogenetic hypothesis for chordate evolution has been the object of some skepticism. We assembled an expanded phylogenomic dataset focused on deuterostomes. Maximum-likelihood using standard models and Bayesian phylogenetic analyses using the CAT site-heterogeneous mixture model of amino-acid replacement both provided unequivocal support for the sister-group relationship between tunicates and vertebrates (Olfactores). Chordates were recovered as monophyletic with cephalochordates as the most basal lineage. These results were robust to both gene sampling and missing data. New analyses of ribosomal rRNA also recovered Olfactores when compositional bias was alleviated. Despitethe inclusion of 25 taxa representing all major lineages, the monophyly of deuterostomes remained poorly supported. The implications of these phylogenetic results for interpreting chordate evolution are discussed in light of recent advances from evolutionary developmental biology and genomics.


The +838 C/G MT2A polymorphism, metals, and the inflammatory/immune response in carotid artery stenosis in elderly people.
Carotid artery stenosis (CS) is a well-established risk factor for stroke. Increased proinflammatory chemokines, enhanced metallothionein (MT), and altered metal homeostasis may play roles in atherosclerosis progression and plaque destabilization. MT may sequester zinc during chronic inflammation, provoke zinc deficiency, and modulate NK cell cytotoxicity. A recent investigation of older patients with diabetes and atherosclerosis showed an association between the -209 A/G MT2A polymorphism, CS, and zinc status. In this study, we evaluated the relationship between two MT2A polymorphisms (-209 and + 838 locus), metal status, and inflammatory/immune response in older patients with CS only (the CS1 group) or with CS and previous cerebrovascular episodes (transient ischemic attack or stroke) (the CS2 group). A total of 506 individuals (188 CS1, 100 CS2, and 218 healthy controls) were studied. Atherosclerotic patients (CS1 and CS2) showed increased levels of MT, MCP-1, and RANTES, reduced NK cell cytotoxicity, and altered trace element concentrations (zinc, copper, magnesium, iron). The +838 C/G MT2A polymorphism was differently distributed in CS1 and CS2 patients, who displayed the GG genotype (C-) with significantly higher frequency than elderly controls. C- carriers showed increased MCP-1 and decreased NK cell cytotoxicity, CD56+ cells, and intracellular zinc availability along with decreased zinc, copper, and magnesium content in erythrocytes and increased iron in plasma. C- carriers also showed a major incidence of soft carotid plaques. In conclusion, the +838 C/G MT2A polymorphism seems to influence inflammatory markers, zinc availability, NK cell cytotoxicity, and trace element status, all of which may promote CS development.


[Epidemiological and virological aspects Rotavirus diarrhoea in Abidjan, Côte d'Ivoire (1997-2000)].
Viruses, mainly rotaviruses are aetiological agents in more than 80% of the cases of acute diarrhoea in children. In order to determine the epidemiological characteristics and genotypes of human rotaviruses involved in gastroenteritis in diarrheic children aged from 0 to 5 years old in Abidjan, 642 specimens of stools were collected between 1997 and 2000 in the urban health centres and University Teaching Hospitals in Abidjan. The antigenic detection of rotaviruses carried out by ELISA test was followed by the antigenic (VP6 sub-groups) and molecular characterization: polyacrylamide gel electrophoresis and genetic typing. The general prevalence of Rotavirus diarrhoea was 27.9%. Among the children who were found positive, those whose age ranged from 0 to 11 months old accounted for 45.8% against 41.3% and 12.9% for those whose age ranged from 1 to 2 and 3 to 5 years old respectively proving thus the precocity of rotavirus infection. From an electrophoretypical and antigenic point of view 74.5% of 141 extracts of RNA had a "long" profile and belonged to the VP6 II sub-group against 24.8% of "short" profile belonging to sub-group I. The electrophoretypes with short profile were identified in majority in infants whose age ranged from 0 to 2 years old. Out of the P genotypes identified, the P [8] genotype (59.6%) was predominant followed by the P [6] genotype (26.2%), P [4] (2.8%) and one mosaic genotype P[6,8] which represented 11.4%. These results will need to be completed by the determination of VP7 genotypes in order to provide interesting information on rotaviruses before the introduction of anti-Rotavirus vaccines in the country.


Advances in understanding the molecular basis of frontotemporal dementia.
Frontotemporal dementia (FTD) is a clinical syndrome with a heterogeneous molecular basis. Until recently, the underlying cause was known in only a minority of cases that were associated with abnormalities of the tau protein or gene. In 2006, however, mutations in the progranulin gene were discovered as another important cause of familial FTD. That same year, TAR DNA-binding protein 43 (TDP-43) was identified as the pathological protein in the most common subtypes of FTD and amyotrophic lateral sclerosis (ALS). Since then, substantial efforts have been made to understand the functions and regulation of progranulin and TDP-43, as well as their roles in neurodegeneration. More recently, other DNA/RNA binding proteins (FET family proteins) have been identified as the pathological proteins in most of the remaining cases of FTD. In 2011, abnormal expansion of a hexanucleotide repeat in the gene C9orf72 was found to be the most common genetic cause of both FTD and ALS. All common FTD-causing genes have seemingly now been discovered and the main pathological proteins identified. In this Review, we highlight recent advances in understanding the molecular aspects of FTD, which will provide the basis for improved patient care through the development of more-targeted diagnostic tests and therapies.


Separation of organic and inorganic arsenic species by capillary electrophoresis using direct spectrophotometric detection.
Capillary electrophoresis (CE) with UV detection was used for the determination of arsenite, arsenate, monomethylarsonic acid, dimethylarsinic acid, p-aminophenylarsonic acid, 4-hydroxy-3-nitrobenzenearsonic acid, 4-nitrophenylarsonic acid, phenylarsonic acid, and phenylarsine oxide. The electrophoretic mobilities of these anionic species were determined in a 20 mM phosphate buffer in a pH range from 4 to 11, which established pH 10 as the optimum for the separation. The target analytes were then separated in a fused-silica capillary using 20 mM NaHCO(3)-Na(2)CO(3) buffer, pH 10, as electrolyte and detected at 192 nm. Both normal- and reversed electroosmotic flow (EOF) separation modes were investigated and in the latter case, poly(diallydimethylammonium chloride) (PDDAC), was used for dynamic coating of the capillary and to provide a stable and reproducible reversed EOF (relative standard deviation RSD, 0.39%). The influence of electrolyte pH and composition, applied voltage, as well as EOF reversal protocols upon the method performance criteria were investigated. The optimised method provided limits of detection for the target analytes of 1.62, 6.22, 1.45, 1.83, 0.34, 0.40, 0.40, 0.18, and 0.30 mg/L As, respectively. Linearity was obtained in the range of 0.5-40 mg/L As (for aryl compounds) and from 5-100 mg/L As (for the remaining analytes). Reproducibility of peak areas was in the range of 0.8-5.5% RSD. The method was applied to the determination of four aryl arsenic compounds used as additives in animal feed. Analytes were extracted with 40 mM hydrochloric acid - acetonitrile 4:1 v/v, and then cleaned up by passing through a C(18) solid-phase extraction cartridge before analysis by CE with detection at 200 nm. Recoveries for the four analytes were in the range of 78.8-108.3%.


[The role of Robben Island as a stock breeding station for the settlement at the Cape of Good Hope].
The universal notoriety of Robben Island as a penitentiary for political prisoners, notably in the 19th and 20th centuries, overshadows its previous historical significance established centuries ago. The Island, initially a source of seals and penguins to European mariners rounding the southern tip of Africa, and later for several other reasons, including its proximity to the Cape of Good Hope, played a pivotal role in the selection of this halfway station. The seals would provide blubber for train oil and the penguins, meat and eggs. The transhumant Peninsular Khoekhoe was to provide cattle and sheep by a barter process as before. Inconsistent access to Khoen livestock forced the Vereenigde Oost-Indische Compagnie (VOC) to consider their own breeding programmes and ultimately the establishment of Free Burgers. Van Riebeeck confirmed the suitability of Robben Island for the fattening and breeding of sheep and this island made a substantial contribution to the provision of sheep and mutton to the fleets and the local community. Khoen sheep did not do well in the Table Valley in early summer and it was expected that they would thrive on the drier island. Predators and stock theft were major problems at the Cape and neither occurred on the island. It is contended that it was unlikely that the settlement at the Cape would have occurred and succeeded without Robben Island.


Virus Coat Protein Transgenic Papaya Provides Practical Control of Papaya ringspot virus in Hawaii.
Since 1992, Papaya ringspot virus (PRSV) destroyed nearly all of the papaya hectarage in the Puna district of Hawaii, where 95% of Hawaii's papayas are grown. Two field trials to evaluate transgenic resistance (TR) were established in Puna in October 1995. One trial included the following: SunUp, a newly named homozygous transformant of Sunset; Rainbow, a hybrid of SunUp, the nontransgenic Kapoho cultivar widely grown in Puna, and 63-1, another segregating transgenic line of Sunset. The second trial was a 0.4-ha block of Rainbow, simulating a near-commercial planting. Both trials were installed within a matrix of Sunrise, a PRSV-susceptible sibling line of Sunset. The matrix served to contain and trace pollen flow from TR plants, and as a secondary inoculum source. Virus infection was first observed 3.5 months after planting. At a year, 100% of the non-TR control and 91% of the matrix plants were infected, while PRSV infection was not observed on any of the TR plants. Fruit production data of SunUp and Rainbow show that yields were at least three times higher than the industry average, while maintaining percent soluble solids above the minimum of 11% required for commercial fruit. These data suggest that transgenic SunUp and Rainbow, homozygous and hemizygous for the coat protein transgene, respectively, offer a good solution to the PRSV problem in Hawaii.


Assessment of blood-brain barrier penetration of miltefosine used to treat a fatal case of granulomatous amebic encephalitis possibly caused by an unusual Balamuthia mandrillaris strain.
Balamuthia mandrillaris, a free-living ameba, causes rare but frequently fatal granulomatous amebic encephalitis (GAE). Few patients have survived after receiving experimental drug combinations, with or without brain lesion excisions. Some GAE survivors have been treated with a multi-drug regimen including miltefosine, an investigational anti-leishmanial agent with in vitro amebacidal activity. Miltefosine dosing for GAE has been based on leishmaniasis dosing because no data exist in humans concerning its pharmacologic distribution in the central nervous system. We describe results of limited cerebrospinal fluid (CSF) and serum drug level testing performed during clinical management of a child with fatal GAE who was treated with a multiple drug regimen including miltefosine. Brain biopsy specimens, CSF, and sera were tested for B. mandrillaris using multiple techniques, including culture, real-time polymerase chain reaction, immunohistochemical techniques, and serology. CSF and serum miltefosine levels were determined using a liquid chromatography method coupled to tandem mass spectrometry. The CSF miltefosine concentration on hospital admission day 12 was 0.4 μg/mL. The serum miltefosine concentration on day 37, about 80 h post-miltefosine treatment, was 15.3 μg/mL. These are the first results confirming some blood-brain barrier penetration by miltefosine in a human, although with low-level CSF accumulation. Further evaluation of brain parenchyma penetration is required to determine optimal miltefosine dosing for Balamuthia GAE, balanced with the drug's toxicity profile. Additionally, the Balamuthia isolate was evaluated by real-time polymerase chain reaction (PCR), demonstrating genetic variability in 18S ribosomal RNA (18S rRNA) sequences and possibly signaling the first identification of multiple Balamuthia strains with varying pathogenicities.


Resveratrol and quercetin in combination have anticancer activity in colon cancer cells and repress oncogenic microRNA-27a.
Resveratrol and quercetin (RQ) in combination (1:1 ratio) previously inhibited growth in human leukemia cells. This study investigated the anticancer activity of the same mixture in HT-29 colon cancer cells. RQ decreased the generation of reactive oxygen species (ROS) by up to 2.25-fold and increased the antioxidant capacity by up to 3-fold in HT-29 cells (3.8-60 μg/mL), whereas IC50 values for viability were 18.13, 18.73, and 11.85 μg/mL, respectively. RQ also induced caspase-3-cleavage (2-fold) and increased PARP cleavage. Specificity protein (Sp) transcription factors are overexpressed in colon and other cancers and regulate genes required for cell proliferation survival and angiogenesis. RQ treatment decreased the expression of Sp1, Sp3, and Sp4 mRNA and this was accompanied by decreased protein expression. Moreover, the Sp-dependent antiapoptotic survival gene survivin was also significantly reduced, both at mRNA and protein levels. RQ decreased microRNA-27a (miR-27a) and induced zinc finger protein ZBTB10, an Sp-repressor, suggesting that interactions of RQ with the miR-27a-ZBTB10-axis play a role in Sp downregulation. This was confirmed by transfection of cells with the specific mimic for miR-27a, which partially reversed the effects of RQ. These findings are consistent with previous studies on botanical anticancer agents in colon cancer cells.


Abnormal metabolism of arachidonic acid in chronic inflammatory bowel disease: enhanced release of leucotriene B4 from activated neutrophils.
The metabolism of endogenous arachidonic acid P(AA) was investigated in activated neutrophils from 20 patients with Crohn's disease, 20 with ulcerative colitis, and 25 healthy volunteers. 1-14C-P(AA) was incorporated into intracellular pools of phospholipids prior to activation of the cells with ionophore A23187 and analyses of released arachidonic acid metabolites by thin layer chromatography. Total release of radioactivity expressing the release of arachidonic acid and its metabolites, was equal in the experimental and control groups, which suggests a normal substrate availability. In contrast, there was a marked increase in the relative release of leucotriene B4 (LTB4) and its omega-oxidation products, 20-hydroxy-LTB4 (20-OH-LTB4) and 20-carboxy-LTB4 (20-COOH-LTB4), with LTB4 values exceeding the reference interval in seven of 20 patients with Crohn's disease, median 8.7%, and in six of 20 patients with ulcerative colitis, median 7.7%, as compared with a median of 5.3% in healthy volunteers. Furthermore, a decreased release of unmetabolised arachidonic acid, correlating inversely with the release of LTB4 in all experimental and control groups, and normal values for the production of other metabolites of arachidonic acid--for example, 5-hydroxyeicosatetraenoic acid (5-HETE) and 12-hydroxyheptadecatrienoic acid (HHT), point to an enzymatic abnormality such as increased activity of leucotriene B synthetase. An increased capacity for release of LTB4, the major pro-inflammatory metabolite of arachidonic acid lipoxygenation by polymorphonuclear leucocytes, may contribute to perpetuation of the inflammation and to tissue destruction in chronic inflammatory bowel disease. Our findings agree with previous reports of an increased release of LTB4 by the colonic mucosa in this condition.


The surveillance and diagnosis of hepatocellular carcinoma.
The increasing incidence of hepatocellular carcinoma (HCC) is becoming a considerable problem in Europe. While no national surveillance programme exists, there is increasing evidence that surveillance programmes are efficacious and may be cost-effective. The prognosis of large, symptomatic HCC is poor and only palliative treatment is available. In contrast small tumours are now amenable to several modes of treatment including liver transplantation, surgical resection and loco-regional ablation with acceptable 5 year survival rates. Therefore, the identification of small lesions through screening should prolong survival. Consequently, the European Association for the Study of the Liver (EASL) has recommended surveillance with ultrasound scans and tests for alpha fetoprotein every 6 months. Screening is now routine clinical practice in many parts of the developed world.


Psychological interventions for people with motor neuron disease: a scoping review.
Motor neuron disease (MND) is a rapidly progressive neurodegenerative condition with no known cure. MND can affect every aspect of a person's life and has been associated with a wide range of psychological difficulties, which can occur from pre-diagnosis through to the condition's later stages. However, very little research has been conducted on psychological interventions for people with MND (pwMND). This paper aimed to provide the first review specifically targeting psychological interventions in MND and offer potential directions for future research. Methods: A scoping review was carried out across five major databases (PubMed, PsycINFO, CINAHL, Academic Search Ultimate, and Cochrane Library) until 1st of March 2020. Results: From an initial return of 1278 citations, 10 papers were included in the review. These included three randomized controlled trials (RCTs), two quasi-experiments, three uncontrolled pretest-post-test designs, one single case study, and one qualitative secondary analysis. The existing studies focused on a limited number of psychological outcomes and did not take into account site of MND onset or level of depression/anxiety before intervention. Implications for clinical practice are discussed and suggestions for future research are provided. Conclusions: The literature on psychological interventions is still extremely sparse. Mindfulness-based stress reduction (MBSR) and cognitive behavioral therapy (CBT) based on the stress-coping model show promise in RCTs, but require further evaluation. The need for further development and evaluation of psychological interventions to improve the well-being of pwMND cannot be overstated, particularly as the struggle toward the discovery of an effective treatment for MND continues.


A radiolucent lesion presenting calcification histopathologically: a classical case of CEOT.
The calcifying epithelial odontogenic tumor (CEOT) is a rare benign odontogenic neoplasm that accounts for approximately 1% of all odontogenic tumors. Histologically, it shows sheets of polyhedral epithelial cells with deep eosinophilic cytoplasm and prominent nuclei. Globules of amyloid like material among tumor cells which undergoes calcification are evident. We present a case of CEOT which presented classical histopathologic picture but clinico- radiological it presented variably. Its non association with impacted tooth, no calcifications in the incisional material and finally on excision giving its classical picture of prominent intercellular bridges, hyperchromatic polyhedral cells and amyloid with calcifications made it an interesting case to report.


Integrative Analysis of Genomic and Clinical Data Reveals Intrinsic Characteristics of Bladder Urothelial Carcinoma Progression.
The progression of bladder cancer is generally a complex and dynamic process, involving a variety of biological factors. Here, we aimed to identify a set of survival-related genes that play an important role in the progression of bladder cancer and uncover their synergistic patterns. Based on the large-scale genomic profiling data and clinical information of 404 bladder cancer patients derived from The Cancer Genome Atlas (TCGA) database, we first discovered 1078 survival-related genes related to their survival states using univariate and multivariate Cox proportional hazardous regression. We then investigated the dynamic changes of the cooperative behaviors of these 1078 genes by analyzing their respective genomic features, including copy number variations, DNA methylations, somatic mutations, and microRNA regulatory networks. Our analyses showed that during the progression of bladder cancer, the biological disorder involving the identified survival-related genes can be reflected by multiple levels of abnormal gene regulation, ranging from genomic alteration to post-transcriptional dysregulation. In particular, the stage-specific co-expression networks of these genes undergo a series of structural variations. Our findings provide useful hints on understanding the underlying complex molecular mechanisms related to the evolution of bladder cancer and offer a new perspective on clinical diagnosis and treatment of bladder cancer.


Epithelial-mesenchymal transition: potential regulator of ABC transporters in tumor progression.
Epithelial-mesenchymal transition (EMT) can directly contribute to some malignant phenotypes of tumor cells including invasion, metastasis and resistance to chemotherapy. Although EMT is widely demonstrated to play a critical role in chemoresistance and metastasis, the potential signaling network between EMT and drug resistance is still unclear. The distribution of drugs in the internal and external environment of the tumor cells is tightly linked with ATP-binding cassette (ABC) transporters. Recent studies have shown that ABC transporters expression changed continuously during EMT. We believe that EMT is an important regulator of ABC transporters. In this review, we discuss how EMT regulates ABC transporters and their potential linkages. And we hope the knowledge of EMT and ABC transporters will offer more effective targets to experimental research.


Unexpected HLA haplotype sharing in dizygotic twin pairs discordant for rheumatoid arthritis.
Dizygotic twins are generally believed to be no more genetically similar than sibs born from separate pregnancies. In the present study, a panel of 93 dizygotic twin pairs discordant for rheumatoid arthritis were typed for HLA-A, -B, -Cw, and -DR antigens. HLA haplotype sharing identical by descent between the twins showed a trend towards increased sharing of both HLA haplotypes; this increased sharing was statistically significant when the female/female twin pairs were considered separately. In contrast, the pattern of HLA haplotype sharing in sib pairs (n = 128) was consistent with a 1:2:1 ratio of 2, 1, or 0 haplotypes shared. An analysis of 16 normal dizygotic twin pairs was consistent with these results raising the possibility that dizygotic twins in general are genetically more similar at the HLA complex than sibs born from separate pregnancies.


Modeling near-edge fine structure X-ray spectra of the manganese catalytic site for water oxidation in photosystem II.
The Mn 1s near-edge absorption fine structure (NEXAFS) has been computed by means of transition-state gradient-corrected density functional theory (DFT) on four Mn(4)Ca clusters modeling the successive S(0) to S(3) steps of the oxygen-evolving complex (OEC) in photosystem II (PSII). The model clusters were obtained from a previous theoretical study where they were determined by energy minimization. They are composed of Mn(III) and Mn(IV) atoms, progressing from Mn(III)(3)Mn(IV) for S(0) to Mn(III)(2)Mn(IV)(2) for S(1) to Mn(III)Mn(IV)(3) for S(2) to Mn(IV)(4) for S(3), implying an Mn-centered oxidation during each step of the photosynthetic oxygen evolution. The DFT simulations of the Mn 1s absorption edge reproduce the experimentally measured curves quite well. By the half-height method, the theoretical IPEs are shifted by 0.93 eV for the S(0) → S(1) transition, by 1.43 eV for the S(1) → S(2) transition, and by 0.63 eV for the S(2) → S(3) transition. The inflection point energy (IPE) shifts depend strongly on the method used to determine them, and the most interesting result is that the present clusters reproduce the shift in the S(2) → S(3) transition obtained by both the half-height and second-derivative methods, thus giving strong support to the previously suggested structures and assignments.


Bioisosteric Discovery of NPA101.3, a Second-Generation RET/VEGFR2 Inhibitor Optimized for Single-Agent Polypharmacology.
RET receptor tyrosine kinase is a driver oncogene in human cancer. We recently identified the clinical drug candidate Pz-1, which targets RET and VEGFR2. A key in vivo metabolite of Pz-1 is its less active demethylated pyrazole analogue. Using bioisosteric substitution methods, here, we report the identification of NPA101.3, lacking the structural liability for demethylation. NPA101.3 showed a selective inhibitory profile and an inhibitory concentration 50 (IC50) of <0.003 μM for both RET and VEGFR2. NPA101.3 inhibited phosphorylation of all tested RET oncoproteins as well as VEGFR2 and proliferation of cells transformed by RET. Oral administration of NPA101.3 (10 mg/kg/day) completely prevented formation of tumors induced by RET/C634Y-transformed cells, while it weakened, but did not abrogate, formation of tumors induced by a control oncogene (HRAS/G12V). The balanced synchronous inhibition of both RET and VEGFR2, as well the resistance to demethylation, renders NPA101.3 a potential clinical candidate for RET-driven cancers.


Hysteresis control of epithelial-mesenchymal transition dynamics conveys a distinct program with enhanced metastatic ability.
Epithelial-mesenchymal transition (EMT) have been extensively characterized in development and cancer, and its dynamics have been modeled as a non-linear process. However, less is known about how such dynamics may affect its biological impact. Here, we use mathematical modeling and experimental analysis of the TGF-β-induced EMT to reveal a non-linear hysteretic response of E-cadherin repression tightly controlled by the strength of the miR-200s/ZEBs negative feedback loop. Hysteretic EMT conveys memory state, ensures rapid and robust cellular response and enables EMT to persist long after withdrawal of stimuli. Importantly, while both hysteretic and non-hysteretic EMT confer similar morphological changes and invasive potential of cancer cells, only hysteretic EMT enhances lung metastatic colonization efficiency. Cells that undergo hysteretic EMT differentially express subsets of stem cell and extracellular matrix related genes with significant clinical prognosis value. These findings illustrate distinct biological impact of EMT depending on the dynamics of the transition.


Detraining produces minimal changes in physical performance and hormonal variables in recreationally strength-trained men.
The object of this study was to examine changes in muscular strength, power, and resting hormonal concentrations during 6 weeks of detraining (DTR) in recreationally strength-trained men. Each subject was randomly assigned to either a DTR (n = 9) or resistance training (RT; n = 7) group after being matched for strength, body size, and training experience. Muscular strength and power testing, anthropometry, and blood sampling were performed before the experimental period (T1), after 3 weeks (T2), and after the 6-week experimental period (T3). One-repetition maximum (1RM) shoulder and bench press increased in RT at T3 (p </= 0.05), whereas no significant changes were observed in DTR. Peak power output and mean power output significantly decreased (9 and 10%) in DTR at T2. Peak torque of the elbow flexors at 90 degrees did not change in the RT group but did significantly decrease by 11.9% at T3 compared with T1 in the DTR group. Vertical jump height increased in RT at T2 but did not change in DTR. Neither group displayed any changes in 1RM squat, body mass, percent body fat, or resting concentrations of growth hormone, follicle-stimulating hormone, luteinizing hormone, sex hormone-binding globulin, testosterone, cortisol, or adrenocorticotropin. These data demonstrate that 6 weeks of resistance DTR in recreationally trained men affects power more than it does strength without any accompanying changes in resting hormonal concentrations. For the recreational weight trainer, losses in strength over 6 weeks are less of a concern compared with anaerobic power and upper arm isometric force production. Anaerobic power exercise with a high metabolic component coming from glycolysis might be of importance for reducing the impact of DTR on Wingate power performances. A minimal maintenance training program is recommended for the recreational lifter to offset any reductions in performance.


Are Birth Weight, Early Growth, and Motor Development Determinants of Physical Activity in Children and Youth? A Systematic Review and Meta-Analysis.
The purpose of this systematic review was to explore whether birth weight, early growth and motor development act as determinants of physical activity in children and youth. We performed a systematic literature search on the possible early life determinants. A meta-analysis was performed on the association between birthweight and objectively measured physical activity. We identified 9 studies examining birth weight, in which none of the studies with objectively measured physical activity observed an association between birth weight and physical activity. The meta-analysis confirmed this result (b=-3.08, 95% CI -10.20, 4.04). The 3 studies examining early growth and physical activity in youth differ in methodology and the results are inconsistent. Two studies suggest an association between earlier motor development and physical activity and sport participation in youth. This was not confirmed in a third study. Our meta-analysis suggests that birth weight is not an important determinant of physical activity in youth. Available data does not allow firm conclusions whether early growth and motor development act as determinants of physical activity in youth.


Knockdown of SVCT2 impairs in-vitro cell attachment, migration and wound healing in bone marrow stromal cells.
Bone marrow stromal cell (BMSC) adhesion and migration are fundamental to a number of pathophysiologic processes, including fracture and wound healing. Vitamin C is beneficial for bone formation, fracture repair and wound healing. However, the role of the vitamin C transporter in BMSC adhesion, migration and wound healing is not known. In this study, we knocked-down the sodium-dependent vitamin C transporter, SVCT2, the only known transporter of vitamin C in BMSCs, and performed cell adhesion, migration, in-vitro scratch wound healing and F-actin re-arrangement studies. We also investigated the role of oxidative stress on the above processes. Our results demonstrate that both oxidative stress and down-regulation of SVCT2 decreased cell attachment and spreading. A trans-well cell migration assay showed that vitamin C helped in BMSC migration and that knockdown of SVCT2 decreased cell migration. In the in-vitro scratch wound healing studies, we established that oxidative stress dose-dependently impairs wound healing. Furthermore, the supplementation of vitamin C significantly rescued the BMSCs from oxidative stress and increased wound closing. The knockdown of SVCT2 in BMSCs strikingly decreased wound healing, and supplementing with vitamin C failed to rescue cells efficiently. The knockdown of SVCT2 and induction of oxidative stress in cells produced an alteration in cytoskeletal dynamics. Signaling studies showed that oxidative stress phosphorylated members of the MAP kinase family (p38) and that vitamin C inhibited their phosphorylation. Taken together, these results indicate that both the SVCT2 transporter and oxidative stress play a vital role in BMSC attachment, migration and cytoskeletal re-arrangement. BMSC-based cell therapy and modulation of SVCT2 could lead to a novel therapeutic approach that enhances bone remodeling, fracture repair and wound healing in chronic disease conditions.


Different ecophysiological responses of freshwater fish to warming and acidification.
Future climate change scenarios predict threatening outcomes to biodiversity. Available empirical data concerning biological response of freshwater fish to climate change remains scarce. In this study, we investigated the physiological and biochemical responses of two Iberian freshwater fish species (Squalius carolitertii and the endangered S. torgalensis), inhabiting different climatic conditions, to projected future scenarios of warming (+3°C) and acidification (ΔpH=-0.4). Herein, metabolic enzyme activities of glycolytic (citrate synthase - CS, lactate dehydrogenase - LDH) and antioxidant (glutathione S-transferase, catalase and superoxide dismutase) pathways, as well as the heat shock response (HSR) and lipid peroxidation were determined. Our results show that, under current water pH, warming causes differential interspecific changes on LDH activity, increasing and decreasing its activity in S. carolitertii and in S. torgalensis, respectively. Furthermore, the synergistic effect of warming and acidification caused an increase in LDH activity of S. torgalensis, comparing with the warming condition. As for CS activity, acidification significantly decreased its activity in S. carolitertii whereas in S. torgalensis no significant effect was observed. These results suggest that S. carolitertii is more vulnerable to climate change, possibly as the result of its evolutionary acclimatization to milder climatic condition, while S. torgalensis evolved in the warmer Mediterranean climate. However, significant changes in HSR were observed under the combined warming and acidification (S. carolitertii) or under acidification (S. torgalensis). Our results underlie the importance of conducting experimental studies and address species endpoint responses under projected climate change scenarios to improve conservation strategies, and to safeguard endangered freshwater fish.


Co-creating a program for teams to maintain and reflect on their flexibility.
To prevent rigidity within teams in health care and to support teams in detecting early warning signs of decreasing flexibility, a program has been co-created in collaboration with mental healthcare teams. This program is intended to systematically monitor team behavior, and by doing so to facilitate team intervention. We aim to lay foundations for the further development of methods that can help teams to recognize and respond to processes going on under the surface. This paper introduces the program to the reader; and describes its premises and the co-creation process, leading to a program of nine steps. Then, it describes the application of the program within a team, what a team needs to use the program, and whether the nine steps are sufficient. This pilot shows that the program is a helpful framework within which teams can talk about rigidity, define indicators of their flexibility, and think about appropriate actions and interventions for maintaining or restoring their flexibility. Team ownership and the customizability of the program are important attributes. The program appears to provide a useful framework that helps a team to observe and discuss processes. Team members become aware of the indicators of their team and make their goals explicit.