Gadolinium Chelate Safety in Pregnancy: Barely Detectable Gadolinium Levels in the Juvenile Nonhuman Primate after in Utero Exposure.
Purpose To determine whether gadolinium remains in juvenile nonhuman primate tissue after maternal exposure to intravenous gadoteridol during pregnancy. Materials and Methods Gravid rhesus macaques and their offspring (n = 10) were maintained, as approved by the institutional animal care and utilization committee. They were prospectively studied as part of a pre-existing ongoing research protocol to evaluate the effects of maternal malnutrition on placental and fetal development. On gestational days 85 and 135, they underwent placental magnetic resonance imaging after intravenous gadoteridol administration. Amniocentesis was performed on day 135 prior to administration of the second dose of gadoteridol. After delivery, the offspring were followed for 7 months. Tissue samples from eight different organs and from blood were harvested from each juvenile macaque. Gadolinium levels were measured by using inductively coupled plasma mass spectrometry. Results Gadolinium concentration in the amniotic fluid was 0.028 × 10-5 %ID/g (percentage injected dose per gram of tissue) 50 days after administration of one gadoteridol dose. Gadolinium was most consistently detected in the femur (mean, 2.5 × 10-5 %ID/g; range, [0.81-4.1] × 10-5 %ID/g) and liver (mean, 0.15 × 10-5 %ID/g; range, [0-0.26] × 10-5 %ID/g). Levels were undetectable in the remaining sampled tissues, with the exception of one juvenile skin sample (0.07 × 10-5 %ID/g), one juvenile spleen sample (0.039 × 10-5 %ID/g), and one juvenile brain (0.095 × 10-5 %ID/g) and kidney (0.13 × 10-5 %ID/g) sample. Conclusion The presence of gadoteridol in the amniotic fluid after maternal injection enables confirmation that it crosses the placenta. Extremely low levels of gadolinium are found in juvenile macaque tissues after in utero exposure to two doses of gadoteridol, indicating that a very small amount of gadolinium persists after delivery. © RSNA, 2017.


A global assessment of civil registration and vital statistics systems: monitoring data quality and progress.
Increasing demand for better quality data and more investment to strengthen civil registration and vital statistics (CRVS) systems will require increased emphasis on objective, comparable, cost-effective monitoring and assessment methods to measure progress. We apply a composite index (the vital statistics performance index [VSPI]) to assess the performance of CRVS systems in 148 countries or territories during 1980-2012 and classify them into five distinct performance categories, ranging from rudimentary (with scores close to zero) to satisfactory (with scores close to one), with a mean VSPI score since 2005 of 0·61 (SD 0·31). As expected, the best performing systems were mostly in the European region, the Americas, and Australasia, with only two countries from east Asia and Latin America. Most low-scoring countries were in the African or Asian regions. Globally, only modest progress has been made since 2000, with the percentage of deaths registered increasing from 36% to 38%, and the percentage of children aged under 5 years whose birth has been registered increasing from 58% to 65%. However, several individual countries have made substantial improvements to their CRVS systems in the past 30 years by capturing more deaths and improving accuracy of cause-of-death information. Future monitoring of the effects of CRVS strengthening will greatly benefit from application of a metric like the VSPI, which is objective, costless to compute, and able to identify components of the system that make the largest contributions to good or poor performance.


Chronic infection of Toxoplasma gondii downregulates miR-132 expression in multiple brain regions in a sex-dependent manner.
MicroRNA-132 (miR-132) has been demonstrated to affect multiple neuronal functions and its dysregulation is linked to several neurological disorders. We previously showed that acute Toxoplasma gondii infection induces miR-132 expression both in vitro and in vivo. To investigate the impact of chronic infection on miR-132, we infected mice with T. gondii PRU strain and performed assessment 5 months later in six brain regions (cortex, hypothalamus, striatum, cerebellum, olfactory bulb and hippocampus) by qPCR. We found that while acute infection of T. gondii increases the expression of miR-132, chronic infection has the opposite effect. The effect varied amongst different regions of the brain and presented in a sex-dependent manner, with females exhibiting more susceptibility than males. MiR-132 and brain-derived neurotrophic factor (BDNF, an inducer of miR-132) were not co-varies in the brain areas of infected mice. T. gondii DNA/RNA was found in all tested brain regions and a selective tropism towards the hippocampus, based on bradyzoite density, was observed in both males and females. However, the expressions of miR-132 or BDNF were poorly reflected by the density of T. gondii in brain areas. Our findings highlight the importance of investigating the miR-132-mediated neuronal function in mice infected with T. gondii.


The limb field mesoderm determines initial limb bud anteroposterior asymmetry and budding independent of sonic hedgehog or apical ectodermal gene expressions.
We have analyzed the pattern of expression of several genes implicated in limb initiation and outgrowth using limbless chicken embryos. We demonstrate that the expressions of the apical ridge associated genes, Fgf-8, Fgf-4, Bmp-2 and Bmp-4, are undetectable in limbless limb bud ectoderm; however, FGF2 protein is present in the limb bud ectoderm. Shh expression is undetectable in limbless limb bud mesoderm. Nevertheless, limbless limb bud mesoderm shows polarization manifested by the asymmetric expression of Hoxd-11, -12 and -13, Wnt-5a and Bmp-4 genes. The posterior limbless limb bud mesoderm, although not actually expressing Shh, is competent to express it if supplied with exogenous FGF or transplanted to a normal apical ridge environment, providing further evidence of mesodermal asymmetry. Exogenous FGF applied to limbless limb buds permits further growth and determination of recognizable skeletal elements, without the development of an apical ridge. However, the cells competent to express Shh do so at reduced levels; nevertheless, Bmp-2 is then rapidly expressed in the posterior limbless mesoderm. limbless limb buds appear as bi-dorsal structures, as the entire limb bud ectoderm expresses Wnt-7a, a marker for dorsal limb bud ectoderm; the ectoderm fails to express En-1, a marker of ventral ectoderm. As expected, C-Lmx1, which is downstream of Wnt-7a, is expressed in the entire limbless limb bud mesoderm. We conclude that anteroposterior polarity is established in the initial limb bud prior to Shh expression, apical ridge gene expression or dorsal-ventral asymmetry. We propose that the initial pattern of gene expressions in the emergent limb bud is established by axial influences on the limb field. These permit the bud to emerge with asymmetric gene expression before Shh and the apical ridge appear. We report that expression of Fgf-8 by the limb ectoderm is not required for the initiation of the limb bud. The gene expressions in the pre-ridge limb bud mesoderm, as in the limb bud itself, are unstable without stimulation from the apical ridge and the polarizing region (Shh) after budding is initiated. We propose that the defect in limbless limb buds is the lack of a dorsal-ventral interface in the limb bud ectoderm where the apical ridge induction signal would be received and an apical ridge formed. These observations provide evidence for the hypothesis that the dorsal-ventral ectoderm interface is a precondition for apical ridge formation.


Cellular vacuoles induced by Mycoplasma pneumoniae CARDS toxin originate from Rab9-associated compartments.
Recently, we identified an ADP-ribosylating and vacuolating cytotoxin in Mycoplasma pneumoniae designated Community Acquired Respiratory Distress Syndrome (CARDS) toxin. In this study we show that vacuoles induced by recombinant CARDS (rCARDS) toxin are acidic and derive from the endocytic pathway as determined by the uptake of neutral red and the fluid-phase marker, Lucifer yellow, respectively. Also, we demonstrate that the formation of rCARDS toxin-associated cytoplasmic vacuoles is inhibited by the vacuolar ATPase inhibitor, bafilomycin A1, and the ionophore, monensin. To examine the ontogeny of these vacuoles, we analyzed the distribution of endosomal and lysosomal membrane markers during vacuole formation and observed the enrichment of the late endosomal GTPase, Rab9, around rCARDS toxin-induced vacuoles. Immunogold-labeled Rab9 and overexpression of green fluorescent-tagged Rab9 further confirmed vacuolar association. The late endosomal- and lysosomal-associated membrane proteins, LAMP1 and LAMP2, also localized to the vacuolar membranes, while the late endosomal protein, Rab7, and early endosomal markers, Rab5 and EEA1, were excluded. HeLa cells expressing dominant-negative (DN) Rab9 exhibited markedly reduced vacuole formation in the presence of rCARDS toxin, in contrast to cells expressing DN-Rab7, highlighting the importance of Rab9 function in rCARDS toxin-induced vacuolation. Our findings reveal the unique Rab9-association with rCARDS toxin-induced vacuoles and its possible relationship to the characteristic histopathology that accompanies M. pneumoniae infection.


Graded activation in rabbit mesotubarium smooth muscle.
The outer margin of the mesotubarium superius, an accessory ligament from the femal rabbit reproductive system, contains a long and slender bundle of smooth muscle fibers well aligned with the long axis of the tissue. Very little connective tissue is present. Massive alternating-current electrical field stimuli (variable in frequency, amplitude, and duration) applied to isolated mesotubaria from mature, nonpregnant animals produced contractions in which isometric force (P) and its first derivative (dP/dt) could be continuously graded; an optimum existed for stimulus frequency and amplitude, but not for its duration. Twitchlike contractions could not be produced. Isometric contractions in which P was plotted against dP/dt to generate a phase-plane trajectory were used to predict the ultimate (time = infinity) force (Po) in graded contractions; this value agreed with the Po derived from isotonic force-velocity curves fitted to the Hill equation. Quick stretches applied during the rise of contractile force revealed a slow onset of the ability to bear the Po force.


A new method for closure of small to medium-size nasoseptal perforations.
This article describes a new method of repairing nasoseptal perforations. The surgical procedure involves rotating a composite transposition flap of mucosa and cartilage anchored at the posterior margin of the perforation. The focus of the repair is to preserve vascularity to the flap and thereby enhance the survival of the cartilage and mucosa. This method appears to be most suitable for repairing small to medium-size perforations. Early success has been promising and warrants a trial by others.


First demonstration of long-haul transmission using silicon microring modulators.
We report error-free long-haul transmission of optical data modulated using a silicon microring resonator electro-optic modulator with modulation rates up to 12.5 Gb/s. Using bit-error-rate and power penalty characterizations, we evaluate the performance of this device with varying modulation rates, and perform a comparative analysis using a commercial electro-optic modulator. We then experimentally measure the signal integrity degradation of the high-speed optical data with increasing propagation distances, induced chromatic dispersions, and bandwidth-distance products, showing error-free transmission for propagation distances up to 80 km. These results confirm the functional ubiquity of this silicon modulator, establishing the potential role of silicon photonic interconnects for chip-scale high-performance computing systems and memory access networks, optically-interconnected data centers, as well as high-performance telecommunication networks spanning large distances.


Metabolomics in premature labor: a novel approach to identify patients at risk for preterm delivery.
Biomarkers for preterm labor (PTL) and delivery can be discovered through the analysis of the transcriptome (transcriptomics) and protein composition (proteomics). Characterization of the global changes in low-molecular weight compounds which constitute the 'metabolic network' of cells (metabolome) is now possible by using a 'metabolomics' approach. Metabolomic profiling has special advantages over transcriptomics and proteomics since the metabolic network is downstream from gene expression and protein synthesis, and thus more closely reflects cell activity at a functional level. This study was conducted to determine if metabolomic profiling of the amniotic fluid can identify women with spontaneous PTL at risk for preterm delivery, regardless of the presence or absence of intraamniotic infection/inflammation (IAI). Two retrospective cross-sectional studies were conducted, including three groups of pregnant women with spontaneous PTL and intact membranes: (1) PTL who delivered at term; (2) PTL without IAI who delivered preterm; and (3) PTL with IAI who delivered preterm. The first was an exploratory study that included 16, 19, and 20 patients in groups 1, 2, and 3, respectively. The second study included 40, 33, and 40 patients in groups 1, 2, and 3, respectively. Amniotic fluid metabolic profiling was performed by combining chemical separation (with gas and liquid chromatography) and mass spectrometry. Compounds were identified using authentic standards. The data were analyzed using discriminant analysis for the first study and Random Forest for the second. (1) In the first study, metabolomic profiling of the amniotic fluid was able to identify patients as belonging to the correct clinical group with an overall 96.3% (53/55) accuracy; 15 of 16 patients with PTL who delivered at term were correctly classified; all patients with PTL without IAI who delivered preterm neonates were correctly identified as such (19/19), while 19/20 patients with PTL and IAI were correctly classified. (2) In the second study, metabolomic profiling was able to identify patients as belonging to the correct clinical group with an accuracy of 88.5% (100/113); 39 of 40 patients with PTL who delivered at term were correctly classified; 29 of 33 patients with PTL without IAI who delivered preterm neonates were correctly classified. Among patients with PTL and IAI, 32/40 were correctly classified. The metabolites responsible for the classification of patients in different clinical groups were identified. A preliminary draft of the human amniotic fluid metabolome was generated and found to contain products of the intermediate metabolism of mammalian cells and xenobiotic compounds (e.g. bacterial products and Salicylamide). Among patients with spontaneous PTL with intact membranes, metabolic profiling of the amniotic fluid can be used to assess the risk of preterm delivery in the presence or absence of infection/inflammation.


Associating child sexual abuse with child victimization in China.
To provide a comprehensive profile of the prevalence of child sexual abuse as well as other forms of child victimization in China and to examine the associations between child sexual abuse, demographic factors, and other forms of child victimization. Using a 2-staged stratified sampling procedure, we recruited a total of 18,341 students in grades 9-12 (girls 46.7%, mean age 15.86 years) from 150 randomly sampled schools during November 2009 through July 2010 in 6 Chinese cities. We assessed the students' demographic background and their experience of child sexual abuse and other forms of victimization. The independent effect on child sexual abuse of each demographic factor and form of child victimization was examined after controlling for other variables. The overall lifetime and preceding-year prevalence of child sexual abuse was 8.0% and 6.4%, respectively. Boys were more likely to report child sexual abuse than were girls. Apart from having experienced other forms of child victimization, several characteristics were associated with greater risk of child sexual abuse: being a boy; being older; having sibling(s); having divorced, separated, or widowed parents; or having an unemployed father. This study provides reliable estimates of child victimization to facilitate resource allocation in health care settings in China. The strong associations between child sexual abuse and other forms of child victimization warrant screening for additional forms of child victimization once any one of them has been identified.


Cytological grading of breast carcinoma on fine needle aspirates and its relation with histological grading.
Grading of breast carcinoma on fine needle aspiration cytology (FNAC) is beneficial for selecting patients for neoadjuvant chemotherapy. To grade the breast carcinoma on FNAC using Robinson grading system and to assess the concordance of cytological grading (CG) with histological grading (HG) using Elston-Ellis modification of Scarff-Bloom-Richardson grading system. The study was conducted for 1-year, comprising of 50 female patients attending outpatient departments (OPD) as well as admitted in various surgical wards of a teaching hospital, diagnosed as breast carcinoma. FNAC smears were stained with May-Grunwald-Giemsa and Papanicolaou (Pap) stains and CG was done using Robinson system on Pap stained smears. The results were compared with HG system after resection of tumors. Of 50 cases, 14 (28%) cases were graded as grade I, 24 (48%) grade II, and 12 (24%) grade III by CG, whereas 9 (18%), 28 (56%) and 13 (26%) cases were graded as grade I, II and III by HG. The result showed overall 72% concordance of CG with HG, with grade II and grade III showing highest degree of concordance (83.33%), which is comparable to previous studies. Kappa measurement showed a higher degree of agreement in high-grade tumors compared with low-grade tumors (0.73 in grade III, 0.53 in grade II and 0.39 in grade I). Cytological grading is comparable to HG in majority of cases. Because neoadjuvant chemotherapy is becoming increasingly popular as primary treatment modality of breast cancer, CG could be a useful parameter in selecting the mode of therapy and predicting tumor behavior.


Label-free quantitative proteomics reveals regulation of interferon-induced protein with tetratricopeptide repeats 3 (IFIT3) and 5'-3'-exoribonuclease 2 (XRN2) during respiratory syncytial virus infection.
ABSTRACT: A large quantitative study was carried out to compare the proteome of respiratory syncytial virus (RSV) infected versus uninfected cells in order to determine novel pathways regulated during viral infection. RSV infected and mock-infected HEp2 cells were lysed and proteins separated by preparative isoelectric focussing using offgel fractionation. Following tryptic digestion, purified peptides were characterized using label-free quantitative expression profiling by nano-ultra performance liquid chromatography coupled to electrospray ionisation mass spectrometry with collision energy ramping for all-ion fragmentation (UPLC-MSE). A total of 1352 unique cellular proteins were identified and their abundance compared between infected and non-infected cells. Ingenuity pathway analysis revealed regulation of several central cellular metabolic and signalling pathways during infection. Selected proteins that were found regulated in RSV infected cells were screened by quantitative real-time PCR for their regulation on the transcriptional level. Synthesis of interferon-induced protein with tetratricopeptide repeats 3 (IFIT3) and 5'-3'-exoribonuclease 2 (XRN2) mRNAs were found to be highly induced upon RSV infection in a time dependent manner. Accordingly, IFIT3 protein levels accumulated during the time course of infection. In contrast, little variation was observed in XRN2 protein levels, but different forms were present in infected versus non-infected cells. This suggests a role of these proteins in viral infection, and analysis of their function will shed further light on mechanisms of RNA virus replication and the host cell defence machinery.


Unusual phenolic compounds contribute to ecophysiological performance in the purple-colored green alga zygogonium ericetorum (zygnematophyceae, streptophyta) from a high-alpine habitat.
The filamentous green alga Zygogonium ericetorum (Zygnematophyceae, Streptophyta) was collected in a high-alpine rivulet in Tyrol, Austria. Two different morphotypes of this alga were found: a purple morph with a visible purple vacuolar content and a green morph lacking this coloration. These morphotypes were compared with respect to their secondary metabolites, ultrastructure, and ecophysiological properties. Colorimetric tests with aqueous extracts of the purple morph indicated the presence of soluble compounds such as phenolics and hydrolyzable tannins. High-performance liquid chromatography-screening showed that Z. ericetorum contained several large phenolic peaks with absorption maxima at ∼280 nm and sometimes with minor maxima at ∼380 nm. Such compounds are uncommon for freshwater green microalgae, and could contribute to protect the organism against increased UV and visible (VIS) irradiation. The purple Z. ericetorum contained larger amounts (per dry weight) of the putative phenolic substances than the green morph; exposure to irradiation may be a key factor for accumulation of these phenolic compounds. Transmission electron microscopy of the purple morph showed massive vacuolization with homogenous medium electron-dense content in the cell periphery, which possibly contains the secondary compounds. In contrast, the green morph had smaller, electron-translucent vacuoles. The ecophysiological data on photosynthesis and desiccation tolerance indicated that increasing photon fluence densities led to much higher relative electron transport rates (rETR) in the purple than in the green morph. These data suggest that the secondary metabolites in the purple morph are important for light acclimation in high-alpine habitats. However, the green morph recovered better after 4 d of rehydration following desiccation stress.


Targeting 14-3-3ε-CDC25A interactions to trigger apoptotic cell death in skin cancer.
Non-melanoma skin cancer is the most common form of cancer worldwide. We previously documented an anti-apoptotic role for CDC25A in cutaneous squamous cell carcinoma (SCC), an activity dependent on its association with 14-3-3 proteins. We hypothesized that targeting CDC25A-14-3-3ε interactions may be an effective strategy for inducing skin cancer cell apoptosis. Co-immunoprecipitation revealed that CDC25A associated with 14-3-3ε, 14-3-3γ and 14-3-3ζ in SCC cells but not normal keratinocytes. 14-3-3ε and CDC25A activated Akt/BAD/Survivin pro-survival signaling. To target the interaction of 14-3-3ε with CDC25A for cancer therapy, we developed two novel phospho-peptides, pS and pT, corresponding to each of the 14-3-3 binding sites of CDC25A, to specifically interfere with 14-3-3ε binding to CDC25A. Peptides pT (IC50 = 22.1 μM), and pS (IC50 = 29 μM) induced SCC cell death and blocked 14-3-3ε binding to CDC25A. pS or pT treatment of SCC xenografts increased apoptotic cell death and decreased pro-survival P-Akt (S473) and Survivin, demonstrating the effectiveness of the peptides in vivo. These findings lay a framework for the further development of peptides to target 14-3-3ε-CDC25A interactions for skin cancer treatment.


3D chromatin architecture and transcription regulation in cancer.
Chromatin has distinct three-dimensional (3D) architectures important in key biological processes, such as cell cycle, replication, differentiation, and transcription regulation. In turn, aberrant 3D structures play a vital role in developing abnormalities and diseases such as cancer. This review discusses key 3D chromatin structures (topologically associating domain, lamina-associated domain, and enhancer-promoter interactions) and corresponding structural protein elements mediating 3D chromatin interactions [CCCTC-binding factor, polycomb group protein, cohesin, and Brother of the Regulator of Imprinted Sites (BORIS) protein] with a highlight of their associations with cancer. We also summarise the recent development of technologies and bioinformatics approaches to study the 3D chromatin interactions in gene expression regulation, including crosslinking and proximity ligation methods in the bulk cell population (ChIA-PET and HiChIP) or single-molecule resolution (ChIA-drop), and methods other than proximity ligation, such as GAM, SPRITE, and super-resolution microscopy techniques.


Generalized scale-change models for recurrent event processes under informative censoring.
Two major challenges arise in regression analyses of recurrent event data: first, popular existing models, such as the Cox proportional rates model, may not fully capture the covariate effects on the underlying recurrent event process; second, the censoring time remains informative about the risk of experiencing recurrent events after accounting for covariates. We tackle both challenges by a general class of semiparametric scale-change models that allow a scale-change covariate effect as well as a multiplicative covariate effect. The proposed model is flexible and includes several existing models as special cases, such as the popular proportional rates model, the accelerated mean model, and the accelerated rate model. Moreover, it accommodates informative censoring through a subject-level latent frailty whose distribution is left unspecified. A robust estimation procedure which requires neither a parametric assumption on the distribution of the frailty nor a Poisson assumption on the recurrent event process is proposed to estimate the model parameters. The asymptotic properties of the resulting estimator are established, with the asymptotic variance estimated from a novel resampling approach. As a byproduct, the structure of the model provides a model selection approach among the submodels via hypothesis testing of model parameters. Numerical studies show that the proposed estimator and the model selection procedure perform well under both noninformative and informative censoring scenarios. The methods are applied to data from two transplant cohorts to study the risk of infections after transplantation.


Isolation and analysis of the genetic diversity of repertoires of VSG expression site containing telomeres from Trypanosoma brucei gambiense, T. b. brucei and T. equiperdum.
African trypanosomes (including Trypanosoma brucei) are unicellular parasites which multiply in the mammalian bloodstream. T. brucei has about twenty telomeric bloodstream form Variant Surface Glycoprotein (VSG) expression sites (BESs), of which one is expressed at a time in a mutually exclusive fashion. BESs are polycistronic transcription units, containing a variety of families of expression site associated genes (ESAGs) in addition to the telomeric VSG. These polymorphic ESAG families are thought to play a role in parasite-host adaptation, and it has been proposed that ESAG diversity might be related to host range. Analysis of the genetic diversity of these telomeric gene families has been confounded by the underrepresentation of telomeric sequences in standard libraries. We have previously developed a method to selectively isolate sets of trypanosome BES containing telomeres using Transformation associated recombination (TAR) cloning in yeast. Here we describe the isolation of repertoires of BES containing telomeres from three trypanosome subspecies: Trypanosoma brucei gambiense DAL 972 (causative agent of West-African trypanosomiasis), T. b. brucei EATRO 2340 (a nonhuman infective strain) and T. equiperdum STIB 818 (which causes a sexually transmitted disease in equines). We have sequenced and analysed the genetic diversity at four BES loci (BES promoter region, ESAG6, ESAG5 and ESAG2) from these three trypanosome BES repertoires. With the exception of ESAG2, the BES sequence repertoires derived from T. b. gambiense are both less diverse than and nearly reciprocally monophyletic relative to those from T. b. brucei and T. equiperdum. Furthermore, although we find evidence for adaptive evolution in all three ESAG repertoires in T. b. brucei and T. equiperdum, only ESAG2 appears to be under diversifying selection in T. b. gambiense. This low level of variation in the T. b. gambiense BES sequence repertoires is consistent both with the relatively narrow host range of this subspecies and its apparent long-term clonality. However, our data does not show a clear correlation between size of trypanosome host range and either number of BESs or extent of ESAG genetic diversity.


Cytochrome P450 2B6 (CYP2B6) and constitutive androstane receptor (CAR) polymorphisms are associated with early discontinuation of efavirenz-containing regimens.
Cytochrome P450 2B6 (CYP2B6) is responsible for the metabolic clearance of efavirenz and single nucleotide polymorphisms (SNPs) in the CYP2B6 gene are associated with efavirenz pharmacokinetics. Since the constitutive androstane receptor (CAR) and the pregnane X receptor (PXR) correlate with CYP2B6 in liver, and a CAR polymorphism (rs2307424) and smoking correlate with efavirenz plasma concentrations, we investigated their association with early (<3 months) discontinuation of efavirenz therapy. Three hundred and seventy-three patients initiating therapy with an efavirenz-based regimen were included (278 white patients and 95 black patients; 293 male). DNA was extracted from whole blood and genotyping for CYP2B6 (516G → T, rs3745274), CAR (540C → T, rs2307424) and PXR (44477T → C, rs1523130; 63396C → T, rs2472677; and 69789A → G, rs763645) was conducted. Binary logistic regression using the backwards method was employed to assess the influence of SNPs and demographics on early discontinuation. Of the 373 patients, 131 withdrew from therapy within the first 3 months. Black ethnicity [odds ratio (OR) = 0.27; P = 0.0001], CYP2B6 516TT (OR = 2.81; P = 0.006), CAR rs2307424 CC (OR = 1.92; P = 0.007) and smoking status (OR = 0.45; P = 0.002) were associated with discontinuation within 3 months. These data indicate that genetic variability in CYP2B6 and CAR contributes to early treatment discontinuation for efavirenz-based antiretroviral regimens. Further studies are now required to define the clinical utility of these associations.


Feasibility and effectiveness of integrating provider-initiated testing and counselling within family planning services in Kenya.
To assess an intervention for increasing access to and use of HIV testing among family planning clients through provider-initiated testing and counselling for HIV. Two versions of the intervention were prospectively compared using a prepost intervention only design. Health facilities were purposively selected and family planning consultations randomly selected. Twenty-three public-sector hospitals, health centres and dispensaries in two districts of Central Province, Kenya. One group of 28 family planning providers were trained in the integrated family planning-HIV counselling intervention and in providing HIV testing and counselling to family planning clients requesting a test during the consultation and another group of 47 family planning providers were trained in the intervention and in referring clients interested in an HIV test. Samples of family planning clients willing to be observed and interviewed were randomly selected (538 preintervention, 520 postintervention) and their informed consent obtained to observe their consultation. All family planning providers were trained in an algorithm that integrates HIV/sexually transmitted infection prevention counselling, including offering HIV testing and counselling, with family planning counselling. Clients choosing to be tested were either referred or tested during the consultation by a trained family planning provider. The proportion of family planning clients with whom HIV testing was discussed; the proportion offered HIV testing; and the proportion choosing to have a test. The proportion of consultations in which HIV prevention counselling was provided and HIV testing offered increased significantly. The proportion of clients requesting an HIV test increased from 1 to 26%; approximately one third of these had never been tested previously. Provider-initiated testing and counselling is feasible and acceptable in family planning services, does not adversely affect the quality of the family planning consultation and increases access to and use of HIV testing in a population who would benefit from knowing their status.


Response prediction in chronic hepatitis C by assessment of IP-10 and IL28B-related single nucleotide polymorphisms.
High baseline levels of IP-10 predict a slower first phase decline in HCV RNA and a poor outcome following interferon/ribavirin therapy in patients with chronic hepatitis C. Several recent studies report that single nucleotide polymorphisms (SNPs) adjacent to IL28B predict spontaneous resolution of HCV infection and outcome of treatment among HCV genotype 1 infected patients. In the present study, we correlated the occurrence of variants at three such SNPs (rs12979860, rs12980275, and rs8099917) with pretreatment plasma IP-10 and HCV RNA throughout therapy within a phase III treatment trial (HCV-DITTO) involving 253 Caucasian patients. The favorable SNP variants (CC, AA, and TT, respectively) were associated with lower baseline IP-10 (P = 0.02, P = 0.01, P = 0.04) and were less common among HCV genotype 1 infected patients than genotype 2/3 (P<0.0001, P<0.0001, and P = 0.01). Patients carrying favorable SNP genotypes had higher baseline viral load than those carrying unfavorable variants (P = 0.0013, P = 0.029, P = 0.0004 respectively). Among HCV genotype 1 infected carriers of the favorable C, A, or T alleles, IP-10 below 150 pg/mL significantly predicted a more pronounced reduction of HCV RNA from day 0 to 4 (first phase decline), which translated into increased rates of RVR (62%, 53%, and 39%) and SVR (85%, 76%, and 75% respectively) among homozygous carriers with baseline IP-10 below 150 pg/mL. In multivariate analyses of genotype 1-infected patients, baseline IP-10 and C genotype at rs12979860 independently predicted the first phase viral decline and RVR, which in turn independently predicted SVR. Concomitant assessment of pretreatment IP-10 and IL28B-related SNPs augments the prediction of the first phase decline in HCV RNA, RVR, and final therapeutic outcome.


Interfacial anchor properties of tryptophan residues in transmembrane peptides can dominate over hydrophobic matching effects in peptide-lipid interactions.
Membrane model systems consisting of phosphatidylcholines and hydrophobic alpha-helical peptides with tryptophan flanking residues, a characteristic motif for transmembrane protein segments, were used to investigate the contribution of tryptophans to peptide-lipid interactions. Peptides of different lengths and with the flanking tryptophans at different positions in the sequence were incorporated in relatively thick or thin lipid bilayers. The organization of the systems was assessed by NMR methods and by hydrogen/deuterium exchange in combination with mass spectrometry. Previously, it was found that relatively short peptides induce nonlamellar phases and that relatively long analogues order the lipid acyl chains in response to peptide-bilayer mismatch. Here it is shown that these effects do not correlate with the total hydrophobic peptide length, but instead with the length of the stretch between the flanking tryptophan residues. The tryptophan indole ring was consistently found to be positioned near the lipid carbonyl moieties, regardless of the peptide-lipid combination, as indicated by magic angle spinning NMR measurements. These observations suggest that the lipid adaptations are not primarily directed to avoid a peptide-lipid hydrophobic mismatch, but instead to prevent displacement of the tryptophan side chains from the polar-apolar interface. In contrast, long lysine-flanked analogues fully associate with a bilayer without significant lipid adaptations, and hydrogen/deuterium exchange experiments indicate that this is achieved by simply exposing more (hydrophobic) residues to the lipid headgroup region. The results highlight the specific properties that are imposed on transmembrane protein segments by flanking tryptophan residues.


The challenges and potential solutions of achieving meaningful consent amongst research participants in northern Thailand: a qualitative study.
Achieving meaningful consent can be challenging, particularly in contexts of diminished literacy, yet is a vital part of participant protection in global health research. We explored the challenges and potential solutions of achieving meaningful consent through a qualitative study in a predominantly hill tribe ethnic minority population in northern Thailand, a culturally distinctive population with low literacy. Semi-structured interviews were conducted with 37 respondents who had participated in scrub typhus clinical research, their family members, researchers and other key informants. A thematic analysis was conducted. Our analysis identified four interrelated themes surrounding participants' ability to give consent: varying degrees of research understanding, limitations of using informal translators, issues impacting decisions to join research, and voluntariness of consent. Suggestions for achieving more meaningful consent included the use of formal translators and community engagement with research populations. Participant's agency in decision making to join research should be supported, but research information needs to be communicated to potential participants in a way that they can understand. We found that improved understanding about the study and its potential benefits and harms goes beyond literacy or translation and requires attention to social and cultural factors.


Molecular genetic analysis of FNR-dependent promoters.
In enteric bacteria, the expression of many genes encoding various anaerobic electron transfer functions is controlled by FNR, the product of the autoregulated fnr gene. FNR is structurally and functionally homologous to CAP, the catabolite gene activator protein, and increased FNR production strongly stimulates transcription of its target genes. By analysis of RNA produced in vivo the promoters of four FNR-dependent genes were localized and shown to display a common arrangement. A 22bp dyad symmetry was found about 30 nucleotides upstream of the transcriptional startpoints and a similar sequence was shown to overlap the site of transcription initiation in the negatively controlled fnr gene. The consensus sequence for the half site recognized by FNR (AAA-TTGAT) is only slightly different from that of CAP (AA-TGTGA). Studies with two mutant frd promoters from Escherichia coli, displaying altered regulation and FNR response, provided additional evidence for recognition of this sequence by FNR.


Deep profiling of apoptotic pathways with mass cytometry identifies a synergistic drug combination for killing myeloma cells.
Multiple myeloma is an incurable and fatal cancer of immunoglobulin-secreting plasma cells. Most conventional therapies aim to induce apoptosis in myeloma cells but resistance to these drugs often arises and drives relapse. In this study, we sought to identify the best adjunct targets to kill myeloma cells resistant to conventional therapies using deep profiling by mass cytometry (CyTOF). We validated probes to simultaneously detect 26 regulators of cell death, mitosis, cell signaling, and cancer-related pathways at the single-cell level following treatment of myeloma cells with dexamethasone or bortezomib. Time-resolved visualization algorithms and machine learning random forest models (RFMs) delineated putative cell death trajectories and a hierarchy of parameters that specified myeloma cell survival versus apoptosis following treatment. Among these parameters, increased amounts of phosphorylated cAMP response element-binding protein (CREB) and the pro-survival protein, MCL-1, were defining features of cells surviving drug treatment. Importantly, the RFM prediction that the combination of an MCL-1 inhibitor with dexamethasone would elicit potent, synergistic killing of myeloma cells was validated in other cell lines, in vivo preclinical models and primary myeloma samples from patients. Furthermore, CyTOF analysis of patient bone marrow cells clearly identified myeloma cells and their key cell survival features. This study demonstrates the utility of CyTOF profiling at the single-cell level to identify clinically relevant drug combinations and tracking of patient responses for future clinical trials.


Identifying stochastic oscillations in single-cell live imaging time series using Gaussian processes.
Multiple biological processes are driven by oscillatory gene expression at different time scales. Pulsatile dynamics are thought to be widespread, and single-cell live imaging of gene expression has lead to a surge of dynamic, possibly oscillatory, data for different gene networks. However, the regulation of gene expression at the level of an individual cell involves reactions between finite numbers of molecules, and this can result in inherent randomness in expression dynamics, which blurs the boundaries between aperiodic fluctuations and noisy oscillators. This underlies a new challenge to the experimentalist because neither intuition nor pre-existing methods work well for identifying oscillatory activity in noisy biological time series. Thus, there is an acute need for an objective statistical method for classifying whether an experimentally derived noisy time series is periodic. Here, we present a new data analysis method that combines mechanistic stochastic modelling with the powerful methods of non-parametric regression with Gaussian processes. Our method can distinguish oscillatory gene expression from random fluctuations of non-oscillatory expression in single-cell time series, despite peak-to-peak variability in period and amplitude of single-cell oscillations. We show that our method outperforms the Lomb-Scargle periodogram in successfully classifying cells as oscillatory or non-oscillatory in data simulated from a simple genetic oscillator model and in experimental data. Analysis of bioluminescent live-cell imaging shows a significantly greater number of oscillatory cells when luciferase is driven by a Hes1 promoter (10/19), which has previously been reported to oscillate, than the constitutive MoMuLV 5' LTR (MMLV) promoter (0/25). The method can be applied to data from any gene network to both quantify the proportion of oscillating cells within a population and to measure the period and quality of oscillations. It is publicly available as a MATLAB package.


Effect of Game Design, Goal Type, and Player Numbers on the Physiological and Physical Demands of Hurling-Specific Small-Sided Games.
The current study examined the effect that game design modification, goal type, and player numbers on the running performance and physiological demands of small-sided hurling games (SSG). Forty-eight hurling players (age, 25.5 ± 3.2 years; height, 178.9 ± 3.2 cm; body mass, 78.5 ± 4.5 kg) performed 4 types of SSG (possession [P], normal play [NP], regular goals [RG] and small goals [SG]) in 4-a-side, 5-a-side, and 6-a-side formats. Heart rate (Polar Electro Oy) and global positioning system technology (VX Sport, 4-Hz, Lower Hutt) were used to analyze the physical and physiological differences between SSG. Total distance (m), high-speed running distance (m) (≥17 km·h), very-high speed running distance (≥22 km·h) (m), peak and mean velocity (km·h) were analyzed as an indicator of the physical demands of play. The 4-a-side SSG independent of game design and goal type resulted in a significantly higher relative exercise intensity compared with 5-a-side (mean change: 6 ± 2%; p = 0.001; d = 1.9 ± 0.2; large) and 6-a-side SSG independent of game design or goal type (mean change: 12 ± 2%; p = 0.001; d = 2.9 ± 0.8; very large). The 4-a-side SG (619 ± 106-m [419-735-m]) resulted in the highest distance when compared with all PP (mean change: 141 ± 9 m; p = 0.05; d = 1.9 ± 0.3; moderate) and RG (mean change: 119 ± 39 m; p = 0.004; d = 2.1 ± 0.8; large). Similar trends were observed for 5-a-side and 6-a-side games with SG resulting in increased total running performance. In conclusion, the current observations reveal that 4-a-side NP, SG, and RG have the highest physiological demands with 4-a-side SG having increased running performance in contrast to other game design and goal-type games. Furthermore, independent of game design and goal type, 4-a-side SSG show increased relative intensity compared with 5-a-side and 6-a-side SSG.


Clinical follow-up method for frontal sinus obliteration with bioactive glass S53P4.
A clinical follow-up method was developed to investigate the behavior of a massive amount of bioactive glass S53P4 (BG) clinically used in frontal sinus obliteration. Two sizes of granules (0.63-0.8 mm or 0.8-1.0 mm) in 16 separate BG amounts, weight 25 g, were tested both in simulated body fluid (SBF) and in a buffer containing tris-hydroxymethyl aminomethane citric acid (TRIS-c.a) in standard conditions. The dissolution of silicon (Si) and phosphate (P) was detected with direct current plasma atom emission spectroscopy (DCP-AES) monthly up to 6 months. The BG masses were scanned both wet in the solutions and dried by computer tomography (CT), and the scans were analyzed by Region of Interest (ROI) technique. Calcium phosphate (CaP)- and silica (Si)-gel-layers were studied by scanning electron microscopy (SEM) at 1, 3, and 6 months. Cumulative loss of Si and P was stronger in TRIS-c.a than in SBF (p < 0.0001), and it was higher with smaller than with larger granules in both solutions (p < 0.0001). This was shown correspondingly by the decrease of Hounsfield units (HUs) in ROI analysis (p < 0.0001). The level of HUs was lower with dried than with wet BG (p < 0.0001). The results were compared for clinical ROI analysis of patients with obliterated frontal sinuses up to 48 months and they were parallel. The follow-up method seems to indirectly reveal the behavior of BG and the healing process in the obliterated cavity.


Increased T cell glucose uptake reflects acute rejection in lung grafts.
Although T cells are required for acute lung rejection, other graft-infiltrating cells such as neutrophils accumulate in allografts and are also high glucose utilizers. Positron emission tomography (PET) with the glucose probe [(18)F]fluorodeoxyglucose ([(18)F]FDG) has been employed to image solid organ acute rejection, but the sources of glucose utilization remain undefined. Using a mouse model of orthotopic lung transplantation, we analyzed glucose probe uptake in the grafts of syngeneic and allogeneic recipients with or without immunosuppression treatment. Pulmonary microPET scans demonstrated significantly higher [(18)F]FDG uptake in rejecting allografts when compared to transplanted lungs of either immunosuppressed or syngeneic recipients. [(18)F]FDG uptake was also markedly attenuated following T cell depletion therapy in lung recipients with ongoing acute rejection. Flow cytometric analysis using the fluorescent deoxyglucose analog 2-NBDG revealed that T cells, and in particular CD8(+) T cells, were the largest glucose utilizers in acutely rejecting lung grafts followed by neutrophils and antigen-presenting cells. These data indicate that imaging modalities tailored toward assessing T cell metabolism may be useful in identifying acute rejection in lung recipients.


Whey protein but not soy protein supplementation alters body weight and composition in free-living overweight and obese adults.
A double-blind, randomized clinical trial was conducted to determine the effect of consumption of supplemental whey protein (WP), soy protein (SP), and an isoenergetic amount of carbohydrate (CHO) on body weight and composition in free-living overweight and obese but otherwise healthy participants. Ninety overweight and obese participants were randomly assigned to 1 of 3 treatment groups for 23 wk: 1) WP; 2) SP (each providing ~56 g/d of protein and 1670 kJ/d); or 3) an isoenergetic amount of CHO. Supplements were consumed as a beverage twice daily. Participants were provided no dietary advice and continued to consume their free-choice diets. Participants' body weight and composition data were obtained monthly. Dietary intake was determined by 24-h dietary recalls collected every 10 d. After 23 wk, body weight and composition did not differ between the groups consuming the SP and WP or between SP and CHO; however, body weight and fat mass of the group consuming the WP were lower by 1.8 kg (P < 0.006) and 2.3 kg (P < 0.005), respectively, than the group consuming CHO. Lean body mass did not differ among any of the groups. Waist circumference was smaller in the participants consuming WP than in the other groups (P < 0.05). Fasting ghrelin was lower in participants consuming WP compared with SP or CHO. Through yet-unknown mechanisms, different sources of dietary protein may differentially facilitate weight loss and affect body composition. Dietary recommendations, especially those that emphasize the role of dietary protein in facilitating weight change, should also address the demonstrated clinical potential of supplemental WP.


Zika Virus Outbreak in Haiti in 2014: Molecular and Clinical Data.
Zika virus (ZIKV), first isolated in Uganda in 1947, is currently spreading rapidly through South America and the Caribbean. In Brazil, infection has been linked with microcephaly and other serious complications, leading to declaration of a public health emergency of international concern; however, there currently are only limited data on the virus (and its possible sources and manifestations) in the Caribbean. From May, 2014-February, 2015, in conjunction with studies of chikungunya (CHIKV) and dengue (DENV) virus infections, blood samples were collected from children in the Gressier/Leogane region of Haiti who presented to a school clinic with undifferentiated febrile illness. Samples were initially screened by RT-PCR for CHIKV and DENV, with samples negative in these assays further screened by viral culture. Of 177 samples screened, three were positive for ZIKV, confirmed by viral sequencing; DENV-1 was also identified in culture from one of the three positive case patients. Patients were from two different schools and 3 different towns, with all three cases occurring within a single week, consistent with the occurrence of an outbreak in the region. Phylogenetic analysis of known full genome viral sequences demonstrated a close relationship with ZIKV from Brazil; additional analysis of the NS5 gene, for which more sequences are currently available, showed the Haitian strains clustering within a monophyletic clade distinct from Brazilian, Puerto Rican and Guatemalan sequences, with all part of a larger clade including isolates from Easter Island. Phylogeography also clarified that at least three major African sub-lineages exist, and confirmed that the South American epidemic is most likely to have originated from an initial ZIKV introduction from French Polynesia into Easter Island, and then to the remainder of the Americas. ZIKV epidemics in South America, as well as in Africa, show complex dissemination patterns. The virus appears to have been circulating in Haiti prior to the first reported cases in Brazil. Factors contributing to transmission and the possible linkage of this early Haitian outbreak with microcephaly remain to be determined.


[Preeclampsia and thrombin generation test].
Acquiring new information to allow prediction of the development of diseases associated with impaired coagulation. Design effective preventive measures most serious diseases (TEN) in the fields of gynecology and obstetrics. For pregnant women with preeclampsia, hypertension compared with women with normal pregnancies could lead to increased thrombin generation due to the synergistic effect of thrombotic risk factors. Based on the results and found statistically significant differences between the groups among pregnant can select for a higher risk of developing deep vein thrombosis. This risk group could then greatly benefit from more stringent follow-up and possible preventive treatment prophylactic doses of LMWH in reducing maternal and perinatal morbidity and mortality. Prospective study. Department of Obstetrics and Gynecology, University Hospital Olomouc. In early pregnancy - during pregnancy standard samples (up to the end of the first trimester) patients venous blood was sampled and they completed information questionnaire. A second sampling was carried out between 24 to 28 week, the third sample and between 36th to 40th week. Obtained blood samples were subsequently processed in the coagulation laboratory Hemato-Oncology Clinic and Olomouc. The blood samples were investigated protein C and S, antithrombin, FVIII level, FII, Leiden, and plasma endothelial microparticles, and lupus anticoagulant and APC resistance standardized methodologies. Thrombin generation was determined thrombin generation test. Thrombin generation was measured fully automatically using a kit (Technothrombin TGA, Technoclone, Vienna, Austria) and analyzer Ceveron Alpha (Technoclone, Vienna, Austria) with fully automatic analysis software. As the main parameter is evaluated by the maximum thrombin generation, at the same time, however, was also detected in the total amount of thrombin and the time until the beginning of the formation of thrombin. In the period 2008-2011 were analyzed blood samples of 303 healthy pregnant women. 215 women, ie 71% were nuliparas, 60 women, ie 19.8% were primiparas, 28 women, 9,2% were secundiparas. The average age of pregnant women was 28.6 years(± 3.8 years). The average maternal weight at the beginning of pregnancy was 63.6 kg (± 7.8 kg). Of the 303 women in 18 (6%) developed slight to moderate degree of preeclampsia or HELLP syndrome with varying severity of clinical manifestations. 20 mothers (6.6%) gave birth prematurely terminated before 37 week of pregnancy. 3 pregnancies (0.9%) were discontinued due to genetic indication for fetal birth defect. The complete study protocol (sampling in all three trimesters) thus completed 280 pregnancies. Of the three evaluated, parameter Lag time, ETP and peak we observed significant differences when comparing physiological pregnancies and pregnancies with preeclampsia (Table 3 and Figure 5-7), the statistical level of p < 0.01. In pregnancies with chronic hypertension, these differences were not significant. Comparison of 18 pregnancies, in which the III. trimester developed preeclampsia with other pregnant with physiological pregnancy did not show statistically significant differences in I. and II. trimester. The results suggest the activation of coagulation through the late stages of pregnancy. Results are influenced by strong clinical variability of disease. In severe and early preeclampsia this activation and significant differences begin much earlier. We demonstrated significantly higher activation of thrombin generation in women with preeclampsia [10]. Changes in preeclampsia are characterized by increased generation of thrombin in plasma. This fact may explain the partial success of the clinical use of aspirin in preeclampsia. In the third trimester, during the manifestation of the disease, patients with preeclampsia have significantly higher ETP compared to patients with a normal pregnancies. Pregnant women with chronic hypertension also show a slight increase in the activation of thrombin. However, these results are not statistically significant. Examination of coagulation in the first and second trimester in women who later developed preeclampsia, showed no statistically significant differences and thus can not be used in this case as predictive, but only as a diagnostic test.


Motor Memory: Revealing Conditioned Action Tendencies Using Transcranial Magnetic Stimulation.
Action tendencies can be elicited by motivationally salient stimuli (e.g., appetitive rewards) or objects that support utilization behaviors. These action tendencies can benefit behavioral performance through speeded RTs in response tasks and improve detection accuracy in attentional capture tasks. However, action tendencies can be counterproductive when goals change (e.g., refraining from junk foods or abstaining from alcohol). Maintaining control over cue-elicited action tendencies is therefore critical for successful behavior modification. To better understand this relationship, we used transcranial magnetic stimulation to investigate the neural signatures of action tendencies in the presence of previously trained response cues. Participants were presented with a continuous letter stream and instructed to respond quickly to two target letters using two different response keys. Following this training phase, the target letters were embedded in a new task (test phase), and we applied transcranial magnetic stimulation to the motor cortex and measured motor evoked potentials as an index of corticospinal excitability (CSE). We found that CSE could be potentiated by a former response cue trained within a single experimental session, even when participants were instructed to withhold responses during the test phase. Critically, attention to the previously trained response cue was required to elicit the primed modulation in CSE, and successful control of this activity was accompanied by CSE suppression. These findings suggest that well-trained response cues can come to prime a conditioned action tendency and provide a model for understanding how the implementation of cognitive control can override action automaticity.


Human nucleolus organizers on nonhomologous chromosomes can share the same ribosomal gene variants.
The distributions of three human ribosomal gene polymorphisms among individual chromosomes containing nucleolus organizers were analyzed by using mouse--human hybrid cells. Different nucleolus organizers can contain the same variant, suggesting the occurrence of genetic exchanges among ribosomal gene clusters on nonhomologous chromosomes. Such exchanges appear to occur less frequently in mice. This difference is discussed in terms of the nucleolar organization and chromosomal location of ribosomal gene clusters in humans and mice.


Cellular contribution of bone graft to fusion.
Although a number of studies have examined the fate of graft-derived cells during the process of fusion, there remains no consensus regarding their exact contribution to bone formation within the fusion mass. We developed two chimeric mouse isograft fusion models that allowed us to track the fate of graft cells within the host fusion bed. Cortical/cancellous bone graft (1:1 ratio of pelvic to vertebral body bone) from male mice was placed between (a) the tibia and fibula or (b) the coccygeal spine transverse processes of syngeneic female hosts. Both models were characterized histologically and histochemically. Graft-derived cells were then identified by fluorescent in situ hybridization for Y-chromosome sequences present in only the graft (male) cells. When the fusion mass was healing but not yet fused (at 1 and 2 weeks), numerous graft-derived cells were observed throughout the fusion site. The predominant graft-derived cell types included chondrocytes, osteoblasts, and fibroblasts. Chondrocytes arose from precursor cells in the graft de novo. as cartilage was not transplanted during the surgical procedure. By the time a mature fusion mass had formed (at 6 weeks), graft-derived cells persisted as osteocytes within the cortical rim surrounding the fusion mass. These osteocytes likely differentiated from graft-derived precursors that had directly formed bone, because transplanted osteocytes within cortical bone graft fragments were noted to rarely survive even at 1 and 2 weeks. Collectively, our results demonstrate for the first time that bone graft contributes cells that, in conjunction with host cells, directly form bone within the fusion mass during all phases of fusion rather than just the early phases.


Differences in Establishment of Persistence of Vaccine and Wild Type Rubella Viruses in Fetal Endothelial Cells.
Both wild type (WT) and vaccine rubella virus (RV) can pass through the placenta to infect a human fetus, but only wtRV routinely causes pathology. To investigate possible reasons for this, we compared establishment of persistence of wtRV and RA27/3 vaccine strains in fetal endothelial cells. We showed that yields of RA27/3 and wtRV were similar after the first round of replication, but then only vaccine-infected cultures went through a crisis characterized by partial cell loss and gradual decline of virus titer followed by recovery and establishment of persistent cultures with low levels of RA27/3 secretion. We compared various steps of virus replication, but we were unable to identify changes, which might explain the 2-log difference in RA27/3 and wtRV yields in persistently infected cultures. Whole genome sequencing did not reveal selection of virus variants in either the wtRV or RA27/3 cultures. Quantitative single-cell analysis of RV replication by in situ hybridization detected, on average, 1-4 copies of negative-strand RNA and ~50 copies of positive-strand genomic RNA in cells infected with both vaccine and WT viruses. The distinct characteristics of RA27/3 replication were the presence of large amounts of negative-strand RV RNA and RV dsRNA at the beginning of the crisis and the accumulation of high amounts of genomic RNA in a subpopulation of infected cells during crisis and persistence. These results suggest that RA27/3 can persist in fetal endothelial cells, but the characteristics of persistence and mechanisms for the establishment and maintenance of persistence are different from wtRV.


White Paper: Yoga Therapy and Pain-How Yoga Therapy Serves in Comprehensive Integrative Pain Management, and How It Can Do More.
This paper examines the role of yoga therapy in comprehensive integrative pain management (CIPM). The pain crisis is described, and how yoga therapists can contribute to its solution is explained. Yoga therapy can be an essential component of the multidisciplinary undertaking that will be required to improve patient outcomes and alter the trajectory of the global public health crisis constituted by an epidemic of poorly understood and inadequately addressed pain. Additional context and evidence are presented to document the effectiveness of yoga therapy interventions to support people living with pain. The white paper concludes by listing recommendations to providers, consumers, payers, and legislators, who together can address systemic and structural barriers to CIPM, as well as suggestions for enabling the yoga therapy profession to more fully participate in these solutions.


Should symptomatic menopausal women be offered hormone therapy?
Many physicians remain uncertain about prescribing hormone therapy for symptomatic women at the onset of menopause. The American Society for Reproductive Medicine (ASRM) convened a multidisciplinary group of healthcare providers to discuss the efficacy and risks of hormone therapy for symptomatic women, and to determine whether it would be appropriate to treat women at the onset of menopause who were complaining of menopausal symptoms. Numerous controlled clinical trials consistently demonstrate that hormone therapy, administered via oral, transdermal, or vaginal routes, is the most effective treatment for vasomotor symptoms. Topical vaginal formulations of hormone therapy should be preferred when prescribing solely for the treatment of symptoms of vulvar and vaginal atrophy. Data from the Women's Health Initiative indicate that the overall attributable risk of invasive breast cancer in women receiving estrogen plus progestin was 8 more cases per 10,000 women-years. No increased risk for invasive breast cancer was detected for women who never used hormone therapy in the past or for those receiving estrogen only. Hormone therapy is not effective for the treatment of cardiovascular disease and that the risk of cardiovascular disease with hormone therapy is principally in older women who are considerably postmenopause. Healthy symptomatic women should be offered the option of hormone therapy for menopausal symptoms. Symptom relief with hormone therapy for many younger women (at the onset of menopause) with menopausal symptoms outweighs the risks and may provide an overall improvement in quality of life. Hormone therapy should be individualized for symptomatic women. This involves tailoring the regimen and dose to individual needs.


The brominated flame retardants TBP-AE and TBP-DBPE antagonize the chicken androgen receptor and act as potential endocrine disrupters in chicken LMH cells.
Increased exposure of birds to endocrine disrupting compounds has resulted in developmental and reproductive dysfunctions. We have recently identified the flame retardants, allyl-2,4,6-tribromophenyl ether (TBP-AE), 2-3-dibromopropyl-2,4,6-tribromophenyl ether (TBP-DBPE) and the TBP-DBPE metabolite 2-bromoallyl-2,4,6-tribromophenyl ether (TBP-BAE) as antagonists to both the human androgen receptor (AR) and the zebrafish AR. In the present study, we aimed at determining whether these compounds also interact with the chicken AR. In silico modeling studies showed that TBP-AE, TBP-BAE and TBP-DBPE were able to dock into to the chicken AR ligand-binding pocket. In vitro transfection assays revealed that all three brominated compounds acted as chicken AR antagonists, inhibiting testosterone induced AR activation. In addition, qRT-PCR studies confirmed that they act as AR antagonists and demonstrated that they also alter gene expression patterns of apoptotic, anti-apoptotic, drug metabolizing and amino acid transporter genes. These studies, using chicken LMH cells, suggest that TBP-AE, TBP-BAE and TBP-DBPE are potential endocrine disrupters in chicken.


Causal influences in the human brain during face discrimination: a short-window directed transfer function approach.
In this letter, we considered the application of parametric spectral analysis, namely a short-window directed transfer function (DTF) approach, to multichannel electroencephalography (EEG) data during a face discrimination task. We identified causal influences between occipitoparietal and centrofrontal electrode sites, the timing of which corresponded to previously reported EEG face-selective components. More importantly we present evidence that there are both feedforward and feedback influences, a finding that is in direct contrast to current computational models of perceptual discrimination and decision making which tend to favor a purely feedforward processing scheme.


Enhanced presynaptic neurotransmitter release in the anterior cingulate cortex of mice with chronic pain.
The anterior cingulate cortex (ACC) is a forebrain structure known for its roles in learning and memory. Recent studies show that painful stimuli activate the prefrontal cortex and that brain chemistry is altered in this area in patients with chronic pain. Components of the CNS that are involved in pain transmission and modulation, from the spinal cord to the ACC, are very plastic and undergo rapid and long-term changes after injury. Patients suffering from chronic pain often complain of memory and concentration difficulties, but little is known about the neural circuitry underlying these deficits. To address this question, we analyzed synaptic transmission in the ACC from mice with chronic pain induced by hindpaw injection of complete Freund's adjuvant (CFA). In vitro whole-cell patch-clamp recordings revealed a significant enhancement in neurotransmitter release probability in ACC synapses from mice with chronic pain. Trace fear memory, which requires sustained attention and the activity of the ACC, was impaired in CFA-injected mice. Using knock-out mice, we found that calmodulin-stimulated adenylyl cyclases, AC1 and/or AC8, were crucial in mediating the long-lasting enhanced presynaptic transmitter release in the ACC of mice with chronic pain. Our findings provide strong evidence that presynaptic alterations caused by peripheral inflammation contribute to memory impairments after injury.


Prevalence and determinants of hyperuricemia in middle-aged, urban Chinese men.
Hyperuricemia is associated with metabolic syndrome and has emerged as a marker for both type 2 diabetes and cardiovascular disease. We estimated the prevalence and lifestyle risk factors of hyperuricemia in middle-aged, urban Chinese men. The study included 3,978 urban Chinese men 40-74 years of age from a population-based cohort study, the Shanghai Men's Health Study, who were free of type 2 diabetes at baseline and had provided fasting blood samples. Uric acid concentrations were measured by the uricase method. Hyperuricemia was defined as >7.0 mg/dL. Anthropometric measurements and information on lifestyle factors and disease history were collected by in-person interviews. One quarter of the study subjects had hyperuricemia. Participants with metabolic syndrome had a higher prevalence of hyperuricemia. Body mass index (BMI), waist-to-hip ratio (WHR), waist circumference, and weight gain (since age 20) were positively associated with the prevalence of hyperuricemia. Physical activity was inversely related to the prevalence of hyperuricemia. The odds ratios for hyperuricemia for quintiles of nonoccupational physical activity were 1.00, 0.80, 0.73, 0.75, and 0.57 (P trend <0.001). Participants with hyperuricemia were less likely to be current smokers, but were more likely to drink alcohol regularly. Beer consumption was associated with higher risk of hyperuricemia compared with consumption of wine or liquor. In this representative sample of middle-aged, urban Chinese men, hyperuricemia is highly prevalent. Obesity, weight gain in adulthood, and alcohol intake were associated with a higher prevalence of hyperuricemia, whereas daily physical activity and smoking were inversely related to the prevalence of hyperuricemia.


Motivation towards dual career of European student-athletes.
The present study aimed to investigate motivations for the dual career of European student-athletes living in countries providing different educational services for elite athletes: State-centric regulation-State as sponsor/facilitator (State), National Sporting Federations/Institutes as intermediary (Federation) and Laisser Faire, no formal structures (No Structure). Therefore, the European Student-athletes' Motivation towards Sports and Academics Questionnaire (SAMSAQ-EU) was administered to 524 European student-athletes. Exploratory Factor Analysis, and Confirmatory Factor Analysis were applied to test the factor structure, and the reliability and validity of the SAMSAQ-EU, respectively. A multivariate approach was applied to verify subgroup effects (P ≤ 0.05) according to gender (i.e., female and male), age (i.e., ≤ 24 years, > 24 years), type of sport (i.e., individual sport and team sport) and competition level (i.e., national and international). Insufficient confirmatory indexes were reported for the whole European student-athlete group, whereas distinct three factor models [i.e., Student Athletic Motivation (SAM); Academic Motivation (AM); Career Athletic Motivation (CAM)] emerged, with acceptable reliability estimates, for State (SAM = 0.82; AM = 0.75; and CAM = 0.75), Federation (SAM = 0.82; AM = 0.66; and CAM = 0.87) and No Structure (SAM = 0.78; AM = 0.74; and CAM = 0.79) subgroups. Differences between subgroups were found only for competition level (P < 0.001) in relation to SAM (P = 0.001) and CAM (P < 0.001). For SAM, the highest and lowest values emerged for Federation (national, 5.1 ± 0.5; international, 5.4 ± 0.5) and State (national, 4.5 ± 0.9; international, 4.8 ± 0.7). The opposite picture emerged for CAM (Federation: national, 3.3 ± 0.7; international, 3.5 ± 0.9; State: national, 5.0 ± 0.8; international, 5.0 ± 0.9). Therefore, despite SAMSAQ-EU demonstrated to be a useful tool, results showed that European student-athletes' motivation for dual career has to be specifically investigated according to social contexts.


Prehospital ticagrelor in ST-segment elevation myocardial infarction.
The direct-acting platelet P2Y12 receptor antagonist ticagrelor can reduce the incidence of major adverse cardiovascular events when administered at hospital admission to patients with ST-segment elevation myocardial infarction (STEMI). Whether prehospital administration of ticagrelor can improve coronary reperfusion and the clinical outcome is unknown. We conducted an international, multicenter, randomized, double-blind study involving 1862 patients with ongoing STEMI of less than 6 hours' duration, comparing prehospital (in the ambulance) versus in-hospital (in the catheterization laboratory) treatment with ticagrelor. The coprimary end points were the proportion of patients who did not have a 70% or greater resolution of ST-segment elevation before percutaneous coronary intervention (PCI) and the proportion of patients who did not have Thrombolysis in Myocardial Infarction flow grade 3 in the infarct-related artery at initial angiography. Secondary end points included the rates of major adverse cardiovascular events and definite stent thrombosis at 30 days. The median time from randomization to angiography was 48 minutes, and the median time difference between the two treatment strategies was 31 minutes. The two coprimary end points did not differ significantly between the prehospital and in-hospital groups. The absence of ST-segment elevation resolution of 70% or greater after PCI (a secondary end point) was reported for 42.5% and 47.5% of the patients, respectively. The rates of major adverse cardiovascular events did not differ significantly between the two study groups. The rates of definite stent thrombosis were lower in the prehospital group than in the in-hospital group (0% vs. 0.8% in the first 24 hours; 0.2% vs. 1.2% at 30 days). Rates of major bleeding events were low and virtually identical in the two groups, regardless of the bleeding definition used. Prehospital administration of ticagrelor in patients with acute STEMI appeared to be safe but did not improve pre-PCI coronary reperfusion. (Funded by AstraZeneca; ATLANTIC ClinicalTrials.gov number, NCT01347580.).


Influence of fibrillin-1 genotype on the aortic stiffness in men.
Aortic stiffness is a predictor of cardiovascular mortality. The mechanical properties of the arterial wall depend on the connective tissue framework, with variation in fibrillin-1 and collagen I genes being associated with aortic stiffness and/or pulse pressure elevation. The aim of this study was to investigate whether variation in fibrillin-1 genotype was associated with aortic stiffness in men. The mechanical properties of the abdominal aorta of 79 healthy men (range 28-81 yr) were investigated by ultrasonographic phase-locked echo tracking. Fibrillin-1 genotype, characterized by the variable tandem repeat in intron 28, and collagen type I alpha 1 genotype characterized by the 2,064 G>T polymorphism, were determined by using DNA from peripheral blood cells. Three common fibrillin-1 genotypes, 2-2, 2-3, and 2-4, were observed in 50 (64%), 10 (13%), and 11 (14%) of the men, respectively. Those of 2-3 genotype had higher pressure strain elastic modulus and aortic stiffness compared with men of 2-2 or 2-4 genotype (P = 0.005). Pulse pressure also was increased in the 2-3 genotype (P = 0.04). There was no significant association between type 1 collagen genotype and aortic stiffness in this cohort. In conclusion, the fibrillin-1 2-3 genotype in men was associated with increased aortic stiffness and pulse pressure, indicative of an increased risk for cardiovascular disease.


Updates on attention-deficit/hyperactivity disorder and learning disorders.
The relationship of attention-deficit/hyperactivity disorder (ADHD) to learning disorders was reviewed and included reading disability, mathematics learning disability, and nonverbal learning disability. Genetic, neuroimaging, and neuropsychological functioning were examined for each disorder, along with a discussion of any existing literature when ADHD co-occurred with the disorder. All the disorders were found to frequently co-occur with ADHD. A review of the underlying neuroanatomic and neurofunctional data found specific structures that frequently co-occur in these disorders with others that are specific to the individual diagnosis. Aberrations in structure and/or function were found for the caudate, corpus callosum, and cerebellum, making these structures sensitive for the disorder but not specific. Suggestions for future research, particularly in relation to intervention, are provided.


Antioxidant and Neuroprotective Effects of Caffeine against Alzheimer's and Parkinson's Disease: Insight into the Role of Nrf-2 and A2AR Signaling.
This paper reviews the results of studies conducted on the role of caffeine in the management of different neurological disorders, such as Parkinson's disease (PD) and Alzheimer's disease (AD). To highlight the potential role of caffeine in managing different neurodegenerative diseases, we identified studies by searching PubMed, Web of Science, and Google Scholar by scrutinizing the lists of pertinent publications. According to the collected overall findings, caffeine may reduce the elevated oxidative stress; inhibit the activation of adenosine A2A, thereby regulating the accumulation of Aβ; reduce the hyperphosphorylation of tau; and reduce the accumulation of misfolded proteins, such as α-synuclein, in Alzheimer's and Parkinson's diseases. The studies have suggested that caffeine has promising protective effects against different neurodegenerative diseases and that these effects may be used to tackle the neurological diseases and/or their consequences. Here, we review the ongoing research on the role of caffeine in the management of different neurodegenerative disorders, focusing on AD and PD. The current findings suggest that caffeine produces potent antioxidant, inflammatory, and anti-apoptotic effects against different models of neurodegenerative disease, including AD, PD, and other neurodegenerative disorders. Caffeine has shown strong antagonistic effects against the adenosine A2A receptor, which is a microglial receptor, and strong agonistic effects against nuclear-related factor-2 (Nrf-2), thereby regulating the cellular homeostasis at the brain by reducing oxidative stress, neuroinflammation, regulating the accumulation of α-synuclein in PD and tau hyperphosphorylation, amyloidogenesis, and synaptic deficits in AD, which are the cardinal features of these neurodegenerative diseases.


The role of atmospheric transformations in determining environmental impacts of carbonaceous nanoparticles.
In studies that have explored the potential environmental impacts of manufactured nanomaterials, the atmosphere has largely been viewed as an inert setting that acts merely as a route for inhalation exposure. Manufactured nanomaterials will enter the atmosphere during production, use, and disposal, and rather than simply being transported, airborne nanoparticles are in fact subject to physical and chemical transformations that could modify their fate, transport, bioavailability, and toxicity once they deposit to aqueous and terrestrial ecosystems. The objective of this paper is to review the factors affecting carbonaceous nanomaterials' behavior in the environment and to show that atmospheric transformations, often overlooked, have the potential to alter nanoparticles' physical and chemical properties and thus influence their environmental fate and impact. Atmospheric processing of naturally occurring and incidental nanoparticles takes place through coagulation, condensation, and oxidation; these phenomena are expected to affect manufactured nanoparticles as well. It is likely that carbonaceous nanomaterials in the atmosphere will be oxidized, effectively functionalizing them. By influencing size, shape, and surface chemistry, atmospheric transformations have the potential to affect a variety of nanoparticle-environment interactions, including solubility, interaction with natural surfactants, deposition to porous media, and ecotoxicity. Potential directions for future research are suggested to address the current lack of information surrounding atmospheric transformations of engineered nanomaterials.


Prognostic value of fractional flow reserve: linking physiologic severity to clinical outcomes.
Fractional flow reserve (FFR) has become an established tool for guiding treatment, but its graded relationship to clinical outcomes as modulated by medical therapy versus revascularization remains unclear. The study hypothesized that FFR displays a continuous relationship between its numeric value and prognosis, such that lower FFR values confer a higher risk and therefore receive larger absolute benefits from revascularization. Meta-analysis of study- and patient-level data investigated prognosis after FFR measurement. An interaction term between FFR and revascularization status allowed for an outcomes-based threshold. A total of 9,173 (study-level) and 6,961 (patient-level) lesions were included with a median follow-up of 16 and 14 months, respectively. Clinical events increased as FFR decreased, and revascularization showed larger net benefit for lower baseline FFR values. Outcomes-derived FFR thresholds generally occurred around the range 0.75 to 0.80, although limited due to confounding by indication. FFR measured immediately after stenting also showed an inverse relationship with prognosis (hazard ratio: 0.86, 95% confidence interval: 0.80 to 0.93; p < 0.001). An FFR-assisted strategy led to revascularization roughly half as often as an anatomy-based strategy, but with 20% fewer adverse events and 10% better angina relief. FFR demonstrates a continuous and independent relationship with subsequent outcomes, modulated by medical therapy versus revascularization. Lesions with lower FFR values receive larger absolute benefits from revascularization. Measurement of FFR immediately after stenting also shows an inverse gradient of risk, likely from residual diffuse disease. An FFR-guided revascularization strategy significantly reduces events and increases freedom from angina with fewer procedures than an anatomy-based strategy.


Molecular motors: thermodynamics and the random walk.
The biochemical cycle of a molecular motor provides the essential link between its thermodynamics and kinetics. The thermodynamics of the cycle determine the motor's ability to perform mechanical work, whilst the kinetics of the cycle govern its stochastic behaviour. We concentrate here on tightly coupled, processive molecular motors, such as kinesin and myosin V, which hydrolyse one molecule of ATP per forward step. Thermodynamics require that, when such a motor pulls against a constant load f, the ratio of the forward and backward products of the rate constants for its cycle is exp [-(DeltaG + u(0)f)/kT], where -DeltaG is the free energy available from ATP hydrolysis and u(0) is the motor's step size. A hypothetical one-state motor can therefore act as a chemically driven ratchet executing a biased random walk. Treating this random walk as a diffusion problem, we calculate the forward velocity v and the diffusion coefficient D and we find that its randomness parameter r is determined solely by thermodynamics. However, real molecular motors pass through several states at each attachment site. They satisfy a modified diffusion equation that follows directly from the rate equations for the biochemical cycle and their effective diffusion coefficient is reduced to D-v(2)tau, where tau is the time-constant for the motor to reach the steady state. Hence, the randomness of multistate motors is reduced compared with the one-state case and can be used for determining tau. Our analysis therefore demonstrates the intimate relationship between the biochemical cycle, the force-velocity relation and the random motion of molecular motors.


Crocin Bleaching Assay Using Purified Di-gentiobiosyl Crocin (-crocin) from Iranian Saffron.
Crocin bleaching assay (CBA) is a new method for determination of antioxidant capacity. In CBA, addition of hydrogen to the conjugated double bonds of crocin results in reduction of crocin and increase in the absorbance at 440 nm, which is considered as a measure of antioxidant potential. Here CBA method was set up using di-gentiobiosyl crocin or α-crocin from Iranian saffron. Then, the antioxidant activity of some known antioxidants i.e. L-ascorbic acid, bilirubin, Trolox, uric acid, and some plasma samples of infants, were tested. The results were compared to that obtained by ferric reducing antioxidant power" (FRAP) as a standard method. Di-gentiobiosyl crocin was extracted and purified from Iranian saffron as previously described by us. Then, the CBA procedure using α-crocin was done in 2 different aquatic conditions, >50% or >90% of water. Results were analyzed by both the Bors method (calculating the relative rate constants= Krel) and the Tsimidou method (calculating the percent of -crocin bleaching inhibition=% Inh). OUR RESULTS INDICATED THE FOLLOWING ORDER OF ANTIOXIDANT POTENTIAL FOR THE ABOVE MENTIONED AGENTS: ascorbic acid > uric acid > Trolox. However, these results are very similar to the data reported by others, but they are strongly related to the aqueous condition. In addition, uric acid showed different properties at different concentrations; so that it showed the antioxidant activity at low concentrations but it acted as a prooxidant at higher concentrations. Bilirubin interfered with this test, possibly because its maximum absorbance is close to the crocin. The obtained data for the antioxidant capacity of the serums was comparable with FRAP assay, except for the sample that contains high bilirubin concentration. In conclusion, it seems that CBA using the main fraction of crocin (-crocin) is a simple and useful method for determination of antioxidant potential of aqueous samples. In addition, the CBA ability to distinguish the samples that contain bilirubin or high uric acid content is helpful in clinical laboratories.


Improvements in Test-Retest Variability of Static Automated Perimetry by Censoring Results With Low Sensitivity in Retinitis Pigmentosa.
We evaluated whether omitting (censoring) points in more severely damaged visual field areas can reduce test-retest variability of static automated perimetry (SAP) in retinitis pigmentosa (RP), as variability creates a significant challenge when monitoring for changes. Cohort 1 included 27 eyes in 16 RP subjects with visual acuity (VA) ranging from 20/20 to 20/70 who completed Humphrey 10-2 size III SAP, once per visit at three visits. Cohort 2 included 15 eyes in nine RP subjects with VA ≤ 20/60 who completed Humphrey 30-2 size V SAP, twice per visit at three visits. Variability was assessed using 95% coefficient of repeatability (CR) calculations for uncensored (all threshold values and data points included) and censored data. In cohort 1, the uncensored between-visit 95% CR was 11.6 decibels (dB); censoring locations with threshold values of <8 to 20 dB resulted in 31% to 53% reductions in the 95% CR. For cohort 2, uncensored 95% CRs were 8.7 and 8.0 dB for within- and between-visit variability, respectively; censoring <8 to 17 dB resulted in 15% to 41% and 15% to 43% reductions in within-visit and between-visit 95% CRs, respectively. For both cohorts, censoring at higher values yielded slightly less variability, at the expense of discarding data from a greater number of eyes and test locations. For 20/20 to 20/70 VA tested with size III stimuli, censoring lower sensitivity values results in substantially lower test-retest variability, which may help detect true changes for locations without severe baseline loss. A rule of thumb for clinical practices using SAP to monitor RP is that longitudinal losses of >9 dB for individual test locations with initial values ≥ 9 dB are likely to be real and meaningful, as they exceed typical variability.


Network centrality of metro systems.
Whilst being hailed as the remedy to the world's ills, cities will need to adapt in the 21(st) century. In particular, the role of public transport is likely to increase significantly, and new methods and technics to better plan transit systems are in dire need. This paper examines one fundamental aspect of transit: network centrality. By applying the notion of betweenness centrality to 28 worldwide metro systems, the main goal of this paper is to study the emergence of global trends in the evolution of centrality with network size and examine several individual systems in more detail. Betweenness was notably found to consistently become more evenly distributed with size (i.e. no "winner takes all") unlike other complex network properties. Two distinct regimes were also observed that are representative of their structure. Moreover, the share of betweenness was found to decrease in a power law with size (with exponent 1 for the average node), but the share of most central nodes decreases much slower than least central nodes (0.87 vs. 2.48). Finally the betweenness of individual stations in several systems were examined, which can be useful to locate stations where passengers can be redistributed to relieve pressure from overcrowded stations. Overall, this study offers significant insights that can help planners in their task to design the systems of tomorrow, and similar undertakings can easily be imagined to other urban infrastructure systems (e.g., electricity grid, water/wastewater system, etc.) to develop more sustainable cities.


CARBAMYL PHOSPHATE SYNTHESIS IN THE EARTHWORM LUMBRICUS TERRESTRIS.
The enzymatic synthesis of citrulline from ammonia, bicarbonate, adenosine triphosphate, and L-ornithine takes place in the soluble fraction of gut tissue of the earthworm. The synthesis occurs at low ammonia concentrations, is dependent upon added N-acetyl-L-glutamate, and results in incorporation of the bicarbonate carbon into the ureido group of the citrulline molecule. Carbamyl phosphate is the intermediate in the reaction and its biosynthesis in the earthworm is mediated by a carbamyl phosphate synthetase system similar to that of ureotelic vertebrates.


Promotion of regeneration of corticospinal tract axons in rats with recombinant vascular endothelial growth factor alone and combined with adenovirus coding for this factor.
After spinal cord transection in adult rats, the axons of the corticospinal tract (CST) degenerate retrogradely and do not regenerate. This phenomenon is thought to be related to either secondary ischemia or deficiency of growth factors. To overcome the deficiency of both blood flow and growth factors, the authors added exogenous vascular endothelial growth factor (VEGF165) to the transected spinal cord either as recombinant protein alone or combined with an adenovirus coding for VEGF165. Because most growth factors are rapidly inactivated in the extracellular environment, the authors used an adenovirus coding for VEGF165 to maintain its activity for several days. In adult rats, the dorsal two thirds of the spinal cord were transected at the T-8 level. In experimental rats, either human recombinant VEGF165 or a combination of this factor and a replication-defective adenovirus coding for VEGF165 (Ad.CMV.VEGF165) was applied at the lesion site. Both recombinant VEGF165 alone and combined with Ad.CMV.VEGF165 were mixed with Matrigel, which is a reconstituted membrane basement protein extract. Control rats received Matrigel alone or Matrigel plus an adenoviral vector containing the LACZ gene (Ad.CMV.LACZ). Thirty days after spinal cord injury, the number of newly formed blood vessels was assessed in the injured area. In addition, the sensorimotor cortex was injected with anterogradely transported horseradish peroxidase (HRP) to label the CST axons in the spinal cord and to evaluate the extent of retrograde axonal degeneration and regeneration. Gene transfer was assessed using semiquantitative reverse transcription-polymerase chain reaction analysis, enzyme-linked immunosorbent assay for human VEGF and beta-galactosidase expression in injured rats treated with Matrigel plus Ad.CMV.LACZ, Matrigel plus Ad.CMV.VEGF165, and untreated injured rats. A strong gene transfer in the spinal cord tissue of adenovirus-treated rats was found from Day 3 to Day 10 postinjury, confirming infection. In the injured spinal cord area, a significant increase of blood vessels (300% over control, p < 0.005) occurred both in rats treated with recombinant VEGF165 alone and in those treated with the combination of recombinant VEGF165 and Ad.CMV.VEGF165. Also, in both of these groups of animals the retrograde degeneration of CST axons was significantly reduced compared with rats treated with Matrigel alone or Matrigel plus Ad.CMV.LACZ. Furthermore, in rats treated with recombinant VEGF165 alone or combined with Ad.CMV.VEGF165, a few HRP-labeled CST axons, which were not detectable in control rats, were seen distal to the spinal cord injury, indicating some regeneration across the injured area. These results indicate that locally applied VEGF exerts angiogenic as well as neurotrophic effects in the injured spinal cord of rats.


Cucurbitane-Type Triterpenoids from the Blood Glucose-Lowering Extracts of Coccinia indica and Momordica balsamina Fruits.
Few vegetables that are commonly consumed in India as part of diet have been claimed for their antidiabetic potential. The present study was aimed at evaluating preventive effects of cucurbit vegetables namely, Coccinia indica and Momordica balsamina belonging to family Cucurbitaceae in diabetic hyperglycemia. The fruits of M. balsamina and C. indica were extracted with chloroform and fractionated with hexane to prepare an extract rich in moderately polar components. These extracts were used for evaluating the effect of these cucurbits in nicotinamide/streptozotocin-induced type 2 diabetes. Streptozotocin-nicotinamide-induced diabetic animals were orally treated with chloroform extract of fruits (250 mg/kg BW) given daily for a week separately. Both the extracts reduced fasting blood glucose significantly (P < 0.05 versus diabetic control) when estimated on seventh day of treatments. Pretreatment with fruit extracts for 7 days also blunted the OGTT (oral glucose tolerance test) curve. Results indicated that C. indica and M. balsamina fruits possess beneficial effects in diabetes by lowering elevated blood glucose level. Six cucurbitane-type triterpenoids were isolated from bioactive extracts of C. indica ((1-3) and M. balsamina (4-6). The structures of these compounds were elucidated on the basis of spectroscopic data analysis. The study concludes that the inclusion of C. indica and M. balsamina fruits in food can be useful for newly diagnosed diabetic patients or highrisk group of population for glycemic control. "Cucurbitane-type triterpenoids from the blood glucose-lowering extracts of Coccinia indica and M. balsamina fruit" The beneficial effects of chloroform extracts of vegetal cucurbits namely C. indica (Ivy gourd) and M. balsamina (Balsam apple) in streptozotocin-nicotinamide (STZ-NA)-induced diabetic rats has been evaluated.The isolation and characterization of six cucurbitacins from bioactive extracts of C. indica (Coccinoside A, B, and C) and M. balsamina (cucurbit-5, 7-dien-3ß-ol, cucurbita-5-en-3ß-ol-3-O-ß-d-glucopyranoside, and cucurbit-5-en-3ß-ol-3-O-ß-d-glucopyranosyl-(4'→1")-O-ß-d-glucopyranoside) have been reported for the first time.The study concludes that the inclusion of C. indica and M. balsamina fruits in food can be useful for newly diagnosed diabetic patients or high risk group of population for glycemic control. Abbreviation used: C: indica (Coccinia indica), M: balsamina (Momordica balsamina), Kbr: Potassium bromide, FTIR: Fourier transform infrared spectroscopy, COSY: Corelated Spectroscopy, DEPT: Distortionless Enhancement by Polarization Transfer, DMSO: Dimethyl sulfoxide, TMS: tetramethylsilane, ESI-MS: Electrospray Ionization mass spectrometry, TLC: thin layer chromatography, STZ-NA: Streptozotocin-nicotinamide, CMC: carboxy methyl cellulose, BW: body weight, ANOVA: analysis of variance, AUC: area under curve.


MRI of long-distance water transport: a comparison of the phloem and xylem flow characteristics and dynamics in poplar, castor bean, tomato and tobacco.
We used dedicated magnetic resonance imaging (MRI) equipment and methods to study phloem and xylem transport in large potted plants. Quantitative flow profiles were obtained on a per-pixel basis, giving parameter maps of velocity, flow-conducting area and volume flow (flux). The diurnal xylem and phloem flow dynamics in poplar, castor bean, tomato and tobacco were compared. In poplar, clear diurnal differences in phloem flow profile were found, but phloem flux remained constant. In tomato, only small diurnal differences in flow profile were observed. In castor bean and tobacco, phloem flow remained unchanged. In all plants, xylem flow profiles showed large diurnal variation. Decreases in xylem flux were accompanied by a decrease in velocity and flow-conducting area. The diurnal changes in flow-conducting area of phloem and xylem could not be explained by pressure-dependent elastic changes in conduit diameter. The phloem to xylem flux ratio reflects what fraction of xylem water is used for phloem transport (Münch's counterflow). This ratio was large at night for poplar (0.19), castor bean (0.37) and tobacco (0.55), but low in tomato (0.04). The differences in phloem flow velocity between the four species, as well as within a diurnal cycle, were remarkably small (0.25-0.40 mm s(-1)). We hypothesize that upper and lower bounds for phloem flow velocity may exist: when phloem flow velocity is too high, parietal organelles may be stripped away from sieve tube walls; when sap flow is too slow or is highly variable, phloem-borne signalling could become unpredictable.