Cancer Cells Invade Confined Microchannels via a Self-Directed Mesenchymal-to-Amoeboid Transition.
Cancer cell invasion through physical barriers in the extracellular matrix (ECM) requires a complex synergy of traction force against the ECM, mechanosensitive feedback, and subsequent cytoskeletal rearrangement. PDMS microchannels were used to investigate the transition from mesenchymal to amoeboid invasion in cancer cells. Migration was faster in narrow 3 μm-wide channels than in wider 10 μm channels, even in the absence of cell-binding ECM proteins. Cells permeating narrow channels exhibited blebbing and had smooth leading edge profiles, suggesting an ECM-induced transition from mesenchymal invasion to amoeboid invasion. Live cell labeling revealed a mechanosensing period in which the cell attempts mesenchymal-based migration, reorganizes its cytoskeleton, and proceeds using an amoeboid phenotype. Rho/ROCK (amoeboid) and Rac (mesenchymal) pathway inhibition revealed that amoeboid invasion through confined environments relies on both pathways in a time- and ECM-dependent manner. This demonstrates that cancer cells can dynamically modify their invasion programming to navigate physically confining matrix conditions.


Predicting intentions to continue exclusive breastfeeding for 6 months: a comparison among racial/ethnic groups.
The purpose of this study was to explore how mothers of different races/ethnicities make decisions to continue exclusive breastfeeding (EBF) for 6 months under the Theory of Planned Behavior. Participants were recruited from hospitals and WIC clinics in Central Indiana and Southern New Jersey from 2008 to 2009. Mothers (N=236: 93 non-Hispanic African American, 72 non-Hispanic white, 71 Hispanic/Latina) completed a self-administered questionnaire that measured theoretical constructs and beliefs related to their intention to practice EBF for 6 months. Intentions to continue EBF for 6 months were similar (P=0.15) across racial/ethnic groups. Significant proportions of the intention (P<0.001) were explained by the three theoretical constructs (attitude, subjective norm, and perceived behavioral control). The relative importance of each construct in predicting the intention varied by group. The most influential predictors (P<0.001) were attitude for white mothers, subjective norm for African American mothers, and perceived behavioral control for Latina mothers. Latent beliefs strongly associated with attitude in white mothers were 'bonding with the baby' and 'easy feeding.' Beliefs held by family members and the general public contributed to the subjective norm of African American mothers. Perceived behavioral control in Latina mothers was highly correlated with 'pumping breast milk'. Development of policy and intervention programs that focus on shaping strong predictors and beliefs within racial/ethnic groups could reduce disparities in EBF rates and establish EBF for 6 months as a cultural norm.


Capsular and pericapsular contributions to acetabular and femoral head perfusion.
Capsular and pericapsular vessels are believed to contribute to acetabular and femoral head perfusion, but, to our knowledge, there is no anatomic study to support this theory. The purpose of this study was to describe the vascular anatomy of the hip with particular reference to the contributions of the capsular and pericapsular vessels. Twenty fresh cadavers were dissected twenty-four hours after intra-arterial injection of colored silicone. The arteries supplying the hip were followed by careful dissection from their origins outside the pelvis to their terminal branches. Particular attention was paid to the vessels traveling between the acetabulum and the capsule and the femoral head. In all twenty specimens, the hip capsule received blood supply from the superior and inferior gluteal arteries proximally and from the medial and lateral femoral circumflex arteries distally. The contributory vessels entered the capsule peripherally and superficially. The circumflex arteries supplied the anterior capsule. The gluteal arteries supplied the posterior and posterosuperior aspect of the hip capsule, augmented by contributions from the circumflex arteries. Variable anastomoses were found between the gluteal and femoral systems on the capsular surface beneath the gluteus minimus and short hip external rotators. The medial femoral circumflex artery provided the dominant blood supply to the femoral head in eighteen specimens, and the inferior gluteal artery provided the dominant supply in two specimens. Capsular and pericapsular vessels that contribute to the blood supply of the acetabulum run on the posterior and posterolateral surface of the capsule. The dominant blood supply to the femoral head comes from vessels that approach the joint posteriorly and penetrate the joint near the femoral attachment of the capsule.


Mutations in NLRP7 are associated with diploid biparental hydatidiform moles, but not androgenetic complete moles.
NLRP7 (NALP7) has been identified as the major gene involved in the inherited predisposition to recurrent molar pregnancies, a rare recessive condition in which affected individuals have complete hydatidiform moles of diploid biparental origin (BiCHM). The role of NLRP7 in other types of molar pregnancy and reproductive wastage has not been conclusively demonstrated. The purpose of this study was to clarify this by identifying NLRP7 variation in two clinically well-defined groups of patients: women with recurrent BiCHM, and women with three or more recurrent complete hydatidiform moles of proven androgenetic origin (AnCHM). Fluorescent microsatellite genotyping of molar tissue was used to establish a diagnosis of recurrent BiCHM (four novel cases) or recurrent AnCHM (nine women with multiple CHM). These two groups were subsequently screened for mutations in NLRP7 using DNA sequencing. Additional screening for non-pathological variants was performed in 21 previously published cases of recurrent BiCHM. Taqman genotyping was used to determine the frequency of novel NLRP7 variants in two control cohorts of Caucasian and Asian women with no adverse reproductive outcomes. Of the four novel cases with recurrent BiCHM, two were homozygous for mutations in NLRP7 while one was a compound heterozygote for a nonsense mutation and a pathological variant. No NLRP7 mutations or pathological variants were identified in the fourth case. None of the women with AnCHM carried any mutations or pathological variants of NLRP7. A single case of AnCHM was found to be heterozygous for a novel variant (R413Q). NLRP7 mutations do not represent a major cause of AnCHM.


Escherichia coli common pilus (ECP) targets arabinosyl residues in plant cell walls to mediate adhesion to fresh produce plants.
Outbreaks of verotoxigenic Escherichia coli are often associated with fresh produce. However, the molecular basis to adherence is unknown beyond ionic lipid-flagellum interactions in plant cell membranes. We demonstrate that arabinans present in different constituents of plant cell walls are targeted for adherence by E. coli common pilus (ECP; or meningitis-associated and temperature-regulated (Mat) fimbriae) for E. coli serotypes O157:H7 and O18:K1:H7. l-Arabinose is a common constituent of plant cell wall that is rarely found in other organisms, whereas ECP is widespread in E. coli and other environmental enteric species. ECP bound to oligosaccharides of at least arabinotriose or longer in a glycan array, plant cell wall pectic polysaccharides, and plant glycoproteins. Recognition overlapped with the antibody LM13, which binds arabinanase-sensitive pectic epitopes, and showed a preferential affinity for (1→5)-α-linked l-arabinosyl residues and longer chains of arabinan as demonstrated with the use of arabinan-degrading enzymes. Functional adherence in planta was mediated by the adhesin EcpD in combination with the structural subunit, EcpA, and expression was demonstrated with an ecpR-GFP fusion and ECP antibodies. Spinach was found to be enriched for ECP/LM13 targets compared with lettuce. Specific recognition of arabinosyl residues may help explain the persistence of E. coli in the wider environment and association of verotoxigenic E. coli with some fresh produce plants by exploitation of a glycan found only in plant, not animal, cells.


Automated assessment of FDG-PET for differential diagnosis in patients with neurodegenerative disorders.
To review literature until November 2015 and reach a consensus on whether automatic semi-quantification of brain FDG-PET is useful in the clinical setting for neurodegenerative disorders. A literature search was conducted in Medline, Embase, and Google Scholar. Papers were selected with a lower limit of 30 patients (no limits with autopsy confirmation). Consensus recommendations were developed through a Delphi procedure, based on the expertise of panelists, who were also informed about the availability and quality of evidence, assessed by an independent methodology team. Critical outcomes were available in nine among the 17 papers initially selected. Only three papers performed a direct comparison between visual and automated assessment and quantified the incremental value provided by the latter. Sensitivity between visual and automatic analysis is similar but automatic assessment generally improves specificity and marginally accuracy. Also, automated assessment increases diagnostic confidence. As expected, performance of visual analysis is reported to depend on the expertise of readers. Tools for semi-quantitative evaluation are recommended to assist the nuclear medicine physician in reporting brain FDG-PET pattern in neurodegenerative conditions. However, heterogeneity, complexity, and drawbacks of these tools should be known by users to avoid misinterpretation. Head-to-head comparisons and an effort to harmonize procedures are encouraged.


Effects of person-centered care at the organisational-level for people with dementia. A systematic review.
The aim of the systematic review was to determine the effectiveness of organizational-level person-centered care for people living with dementia in relation to their quality of life, mood, neuropsychiatric symptoms and function. ALOIS, the Cochrane Dementia and Cognitive Improvement Group Specialised Register databases, were searched up to June 2018 using the terms dementia OR cognitive impairment OR Alzheimer AND non-pharmacological AND personhood OR person-centered care. Reviewed studies included randomized controlled trials (RCTs), cluster-randomized trials (CRTs) and quasi-experimental studies that compared outcomes of person-centered care and usual (non-person-centered) care, for people with a diagnosis of dementia. The search yielded 12 eligible studies with a total of 2599 people living with dementia in long-term care homes, 600 receiving hospital care and 293 living in extra-care community housing. Random-effects models were used to pool adjusted risk ratios and standard mean differences from all studies; the findings were assessed followed the PRISMA guidelines and GRADE criteria. Statistical heterogeneity was assessed using the I2 method and Chi2 P value; studies with low statistical heterogeneity were analyzed using a random-effects model with restricted maximum likelihood estimation in R. Analyses of pre/post data within 12 months identified: a significant effect for quality of life (standardized mean difference (SMD) 0.16 and 95% CI 0.03 to 0.28; studies = 6; I2 = 22%); non-significant effects for neuropsychiatric symptoms (SMD 0.06, 95% CI -0.08 to 0.19; studies = 4; I2 = 0%) and well-being (SMD 0.15, 95% CI -0.15 to 0.45; studies = 4; I2 = 77%); and no effects for agitation (SMD -0.05 (95% CI -0.17 to -0.07; studies 5; I2 = 0%) and depression (SMD -0.06 and 95% CI -0.27 to 0.15, studies = 5; I2 = 53%). The evidence from this review recommends implementation of person-centered care at the organizational-level to support the quality of life of people with living with dementia.


Xeno-Free Cultivation of Mesenchymal Stem Cells From the Corneal Stroma.
The human cornea has recently been described as a source of corneal stroma-derived mesenchymal stem cells (hMSCs). In vitro expansion of these cells involves basal medium supplemented with fetal bovine serum (FBS). As animal-derived serum can confer a risk of disease transmission and can be subject to considerable lot-to-lot variability, it does not comply with newer Good Manufacturing Practice (GMP)-required animal component-free culture protocols for clinical translation. This study investigated animal-free alternatives to FBS for cultivation of human corneal stromal MSCs. Proliferative capacity was studied for cultures supplemented with different concentrations (2.5%, 5%, and 10%) of FBS, human AB serum, human platelet lysate (HPL), and XerumFree. Unsupplemented basal medium was used as a control. The expression of specific hMSC markers (CD73+, CD90+, CD105+, CD19-, CD34-, CD79α-, CD11b-, CD14-, CD45-, and HLA-DR-) and trilineage differentiation (adipogenesis, osteogenesis, and chondrogenesis) were compared for the two outperforming supplements: 10% FBS and HPL. HPL is the only consistent non-xeno supplement where hMSC cultures show significantly higher proliferation than the 10% FBS-supplemented cultures. Both FBS- and HPL-supplemented hMSC cultures showed plastic adherence and trilineage differentiation, and no significant differences were found in the expression of the hMSC marker panel. No significant differences in stemness were detected between FBS and HPL cultures. We conclude that HPL is the best supplement for expansion of human corneal stromal MSCs. HPL significantly outperforms human AB serum, the chemically defined XerumFree, and even the gold standard, FBS. The xeno-free nature of HPL additionally confers preferred standing for use in GMP-regulated clinical trials using human corneal stromal MSCs.


Effects of tomato and soy on serum adipokine concentrations in postmenopausal women at increased breast cancer risk: a cross-over dietary intervention trial.
Breast cancer risk among postmenopausal women increases as body mass index increases. Practical preventive methods to reduce risk of breast cancer are lacking. Few studies have investigated the effects of carotenoids and isoflavones on circulating adipokines in postmenopausal women. The aim was to examine the effects of lycopene- and isoflavone-rich diets on serum adipokines. This was a 26-week, two-arm, longitudinal crossover trial. Participants were recruited from clinics at The Ohio State University Comprehensive Cancer Center. Seventy postmenopausal women at increased breast cancer risk participated in the study. The mean age and body mass index of participants was 57.2 years and 30.0 kg/m(2), respectively; the study was comprised of 81.4% whites. The interventions included 10 weeks of consumption of a tomato-based diet (≥25 mg lycopene daily) and 10 weeks of consumption of a soy-based diet (≥40 g of soy protein daily), with a 2-week washout in between. Changes in serum adiponectin, leptin, and the adiponectin to leptin ratio were examined for each intervention through linear mixed models, with ratio estimates corresponding to postintervention adipokine concentrations relative to preintervention concentrations. After the tomato intervention, among all women, adiponectin concentration increased (ratio 1.09, 95% confidence interval (CI) 1.00-1.18), with a stronger effect observed among nonobese women (ratio 1.13, 95% CI 1.02-1.25). After the soy intervention, adiponectin decreased overall (ratio 0.91, 95% CI 0.84-0.97), with a larger reduction observed among nonobese women (ratio 0.89, 95% CI 0.81-0.98). Overall, no significant changes in leptin or the adiponectin to leptin ratio were observed after either intervention. Increasing dietary consumption of tomato-based foods may beneficially increase serum adiponectin concentrations among postmenopausal women at increased breast cancer risk, especially those who are not obese. Additional studies are essential to confirm these effects and to elucidate the specific mechanisms that may make phytonutrients found in tomatoes practical as breast cancer chemopreventive agents.


Young women's contraceptive microbicide preferences: associations with contraceptive behavior and sexual relationship characteristics.
In time, microbicides may provide women with dual prevention against pregnancy and STDs. Although several microbicide dimensions have been evaluated, little is known about women's preferences for contraceptive microbicides and correlates of these preferences. Acceptability of a hypothetical contraceptive microbicide cream or jelly was examined among a -clinic-based sample of 266 women in Indianapolis from 2004 (when participants were aged 14-22) to 2008. Group conjoint analyses and individual conjoint analyses were used to compare preferences with respect to four microbicide -dimensions: contraceptive ability, efficacy in relation to condoms, timing of use and texture. Pearson's product moment correlations were used to examine the relationship between preferences for a contraceptive microbicide and selected characteristics of the women. Overall, the top-rated microbicide dimensions were efficacy in relation to that of condoms and contraceptive ability (importance scores, 40.0 and 35.4 out of 100.0, respectively). When all dimension levels were compared, contraceptive ability was the most strongly preferred (part-worth utility score, 8.9), and lower efficacy than that of -condoms was the least strongly preferred (-11.9). Preference for contraceptive microbicides was positively -associated with current contraceptive use, sexual agency, partner communication, commitment to avoiding pregnancy and -perceived partner agreement about avoiding pregnancy (coefficients, 0.07-0.18). It was negatively associated with current or past nonuse of contraceptives, seeking pregnancy and perceived partner agreement about seeking -pregnancy (-0.08 to -0.14). Microbicides with dual prevention properties may be attractive to young women. Microbicide development and subsequent clinical trials should incorporate contraceptive microbicides.


Serological prevalence and persistence of high-risk human papillomavirus infection among women in Santiago, Chile.
Human papillomavirus (HPV) serology is a main factor for designing vaccination programs and surveillance strategies; nevertheless, there are few reports of HPV seroprevalence in the general population, especially in Latin America. This study aimed to describe high-risk HPV serological prevalence, persistence, and association with concurrent cervical infection, in Chilean women. 1021 women from the general population, aged 15-85 years, were studied in 2001 of whom 600 were reexamined in 2006. The assessments at both time points included cervical HPV DNA testing, HPV antibody testing, cervical cytology and a sociodemographic/behavioral questionnaire. HPV DNA and antibodies against L1 protein of types 16, 18, 31, 33, 35, 45, 52, and 58 were assessed by reverse line blot and multiplex serology, respectively. Seropositivity was high at both baseline (43.2%) and follow-up (50.2%) and increased with age (p < 0.001); corresponding DNA prevalences were 6.7% and 8.7%. DNA and seroprevalence were associated at baseline (p = 0.01 for any HPV). Early age at first sexual intercourse and having had two or more sexual partners were independently associated with seropositivity. Most (82.0%) initially seropositive women remained seropositive at follow-up; 21.6% of initially seronegative women seroconverted, reaching 17.5% among women older than 60 years of age. ASCUS or worse cytology was correlated with HPV DNA positivity but not with HPV seropositivity. HPV seroprevalence studies are a useful tool for learning about the dynamics of HPV infection in a community. This study contributes to understanding the natural history of HPV infection and provides a baseline assessment before the incorporation of HPV vaccination into a national program.


Expression of c-FLIP in malignant melanoma, and its relationship with the clinicopathological features of the disease.
Numerous molecular events have been associated with the development of malignant melanoma (MM). The cellular FLICE-like inhibitory protein (c-FLIP) was originally identified as an inhibitor of death-receptor signalling through competition with FADD-like interleukin-1β-converting enzyme (FLICE; also known as caspase-8) for recruitment to the FAS-associated death domain (FADD), and it has been suggested recently that c-FLIP is required for the survival and proliferation of various cell types. Aim. To investigate the relationship of c-FLIP expression with the clinicopathological features of MM. Immunohistochemical staining with anti-c-FLIP antibody was performed on tissue samples taken from 77 MMs and 20 naevi. The proliferative cells were visualized by staining with Ki-67 antibody. Annexin V-propidium iodide labelling, a marker for chromatin condensation, was performed to observe the rate of cell apoptosis by flow cytometry. Expression of c-FLIP was increased in MM tissue compared with the matched pigmented naevi. The c-FLIP expression was significantly associated with the histological type and Clark level of MM, and correlated strongly with the Ki-67 labelling index. Downregulation of c-FLIP might increase apoptosis in MM cell lines. c-FLIP might play an important role in the obtaining of aggressive biological behaviours and be useful in predicting prognosis of patients with MM.


Cancer-associated fibroblast-derived annexin A6+ extracellular vesicles support pancreatic cancer aggressiveness.
The intratumoral microenvironment, or stroma, is of major importance in the pathobiology of pancreatic ductal adenocarcinoma (PDA), and specific conditions in the stroma may promote increased cancer aggressiveness. We hypothesized that this heterogeneous and evolving compartment drastically influences tumor cell abilities, which in turn influences PDA aggressiveness through crosstalk that is mediated by extracellular vesicles (EVs). Here, we have analyzed the PDA proteomic stromal signature and identified a contribution of the annexin A6/LDL receptor-related protein 1/thrombospondin 1 (ANXA6/LRP1/TSP1) complex in tumor cell crosstalk. Formation of the ANXA6/LRP1/TSP1 complex was restricted to cancer-associated fibroblasts (CAFs) and required physiopathologic culture conditions that improved tumor cell survival and migration. Increased PDA aggressiveness was dependent on tumor cell-mediated uptake of CAF-derived ANXA6+ EVs carrying the ANXA6/LRP1/TSP1 complex. Depletion of ANXA6 in CAFs impaired complex formation and subsequently impaired PDA and metastasis occurrence, while injection of CAF-derived ANXA6+ EVs enhanced tumorigenesis. We found that the presence of ANXA6+ EVs in serum was restricted to PDA patients and represents a potential biomarker for PDA grade. These findings suggest that CAF-tumor cell crosstalk supported by ANXA6+ EVs is predictive of PDA aggressiveness, highlighting a therapeutic target and potential biomarker for PDA.


Tracking Microscope Performance: A Workflow to Compare Point Spread Function Evaluations Over Time.
Routine system checks are essential for supervising the performance of an advanced light microscope. Recording and evaluating the point spread function (PSF) of a given system provides information about the resolution and imaging. We compared the performance of fluorescent and gold beads for PSF recordings. We then combined the open-source evaluation software PSFj with a newly developed KNIME pipeline named PSFtracker to create a standardized workflow to track a system's performance over several measurements and thus over long time periods. PSFtracker produces example images of recorded PSFs, plots full-width-half-maximum (FWHM) measurements over time and creates an html file which embeds the images and plots, together with a table of results. Changes of the PSF over time are thus easily spotted, either in FWHM plots or in the time series of bead images which allows recognition of aberrations in the shape of the PSF. The html file, viewed in a local browser or uploaded on the web, therefore provides intuitive visualization of the state of the PSF over time. In addition, uploading of the html file on the web allows other microscopists to compare such data with their own.


Comparison of methods for analysing cluster randomized trials: an example involving a factorial design.
Studies involving clustering effects are common, but there is little consistency in their analysis. Various analytical methods were compared for a factorial cluster randomized trial (CRT) of two primary care-based interventions designed to increase breast screening attendance. Three cluster-level and five individual-level options were compared in respect of log odds ratios of attendance and their standard errors (SE), for the two intervention effects and their interaction. Cluster-level analyses comprised: (C1) unweighted regression of practice log odds; (C2) regression of log odds weighted by their inverse variance; (C3) random-effects meta-regression of log odds with practice as a random effect. Individual-level analyses comprised: (I1) standard logistic regression ignoring clustering; (I2) robust SE; (I3) generalized estimating equations; (I4) random-effects logistic regression; (I5) Bayesian random-effects logistic regression. Adjustments for stratification and baseline variables were investigated. As expected, method I1 was highly anti-conservative. The other, valid, methods exhibited considerable differences in parameter estimates and standard errors, even between the various random-effects methods based on the same statistical model. Method I4 was particularly sensitive to between-cluster variation and was computationally stable only after controlling for baseline uptake. Commonly used methods for the analysis of CRT can give divergent results. Simulation studies are needed to compare results from different methods in situations typical of cluster trials but when the true model parameters are known.


A threonine turnstile defines a dynamic amphiphilic binding motif in the AAA ATPase p97 allosteric binding site.
The turnstile motion of two neighboring threonines sets up a dynamic side chain interplay that can accommodate both polar and apolar ligands in a small molecule allosteric protein binding site. A computational model based on SAR data and both X-ray and cryo-EM structures of the AAA ATPase p97 was used to analyze the effects of paired threonines at the inhibitor site. Specifically, the Thr side chain hydroxyl groups form a hydrogen bonding network that readily accommodates small, highly polar ligand substituents. Conversely, diametric rotation of the χ1 torsion by 150-180° orients the side chain β-methyl groups into the binding cleft, creating a hydrophobic pocket that can accommodate small, apolar substituents. This motif was found to be critical for rationalizing the affinities of a structurally focused set of inhibitors of p97 covering a > 2000-fold variation in potencies, with a preference for either small-highly polar or small-apolar groups. The threonine turnstile motif was further validated by a PDB search that identified analogous binding modes in ligand interactions in PKB, as well as by an analysis of NMR structures demonstrating additional gear-like interactions between adjacent Thr pairs. Combined, these data suggest that the threonine turnstile motif may be a general feature of interest in protein binding pockets.


Comprehensive analysis of full genome sequence and Bd-milRNA/target mRNAs to discover the mechanism of hypovirulence in Botryosphaeria dothidea strains on pear infection with BdCV1 and BdPV1.
Pear ring rot disease, mainly caused by Botryosphaeria dothidea, is widespread in most pear and apple-growing regions. Mycoviruses are used for biocontrol, especially in fruit tree disease. BdCV1 (Botryosphaeria dothidea chrysovirus 1) and BdPV1 (Botryosphaeria dothidea partitivirus 1) influence the biological characteristics of B. dothidea strains. BdCV1 is a potential candidate for the control of fungal disease. Therefore, it is vital to explore interactions between B. dothidea and mycovirus to clarify the pathogenic mechanisms of B. dothidea and hypovirulence of B. dothidea in pear. A high-quality full-length genome sequence of the B. dothidea LW-Hubei isolate was obtained using Single Molecule Real-Time sequencing. It has high repeat sequence with 9.3% and DNA methylation existence in the genome. The 46.34 Mb genomes contained 14,091 predicted genes, which of 13,135 were annotated. B. dothidea was predicted to express 3833 secreted proteins. In bioinformatics analysis, 351 CAZy members, 552 transporters, 128 kinases, and 1096 proteins associated with plant-host interaction (PHI) were identified. RNA-silencing components including two endoribonuclease Dicer, four argonaute (Ago) and three RNA-dependent RNA polymerase (RdRp) molecules were identified and expressed in response to mycovirus infection. Horizontal transfer of the LW-C and LW-P strains indicated that BdCV1 induced host gene silencing in LW-C to suppress BdPV1 transmission. To investigate the role of RNA-silencing in B. dothidea defense, we constructed four small RNA libraries and sequenced B. dothidea micro-like RNAs (Bd-milRNAs) produced in response to BdCV1 and BdPV1 infection. Among these, 167 conserved and 68 candidate novel Bd-milRNAs were identified, of which 161 conserved and 20 novel Bd-milRNA were differentially expressed. WEGO analysis revealed involvement of the differentially expressed Bd-milRNA-targeted genes in metabolic process, catalytic activity, cell process and response to stress or stimulus. BdCV1 had a greater effect on the phenotype, virulence, conidiomata, vertical and horizontal transmission ability, and mycelia cellular structure biological characteristics of B. dothidea strains than BdPV1 and virus-free strains. The results obtained in this study indicate that mycovirus regulates biological processes in B. dothidea through the combined interaction of antiviral defense mediated by RNA-silencing and milRNA-mediated regulation of target gene mRNA expression.


SemiSPECT: a small-animal single-photon emission computed tomography (SPECT) imager based on eight cadmium zinc telluride (CZT) detector arrays.
The first full single-photon emission computed tomography (SPECT) imager to exploit eight compact high-intrinsic-resolution cadmium zinc telluride (CZT) detectors, called SemiSPECT, has been completed. Each detector consists of a CZT crystal and a customized application-specific integrated circuit (ASIC). The CZT crystal is a 2.7 cm x 2.7 cm x -0.2 cm slab with a continuous top electrode and a bottom electrode patterned into a 64 x 64 pixel array by photolithography. The ASIC is attached to the bottom of the CZT crystal by indium-bump bonding. A bias voltage of -180 V is applied to the continuous electrode. The eight detectors are arranged in an octagonal lead-shielded ring. Each pinhole in the eight-pinhole aperture placed at the center of the ring is matched to each individual detector array. An object is imaged onto each detector through a pinhole, and each detector is operated independently with list-mode acquisition. The imaging subject can be rotated about a vertical axis to obtain additional angular projections. The performance of SemiSPECT was characterized using 99mTc. When a 0.5 mm diameter pinhole is used, the spatial resolution on each axis is about 1.4 mm as estimated by the Fourier crosstalk matrix, which provides an algorithm-independent average resolution over the field of view. The energy resolution achieved by summing neighboring pixel signals in a 3 x 3 window is about 10% full-width-at-half-maximum of the photopeak. The overall system sensitivity is about 0.5 x 10(-4) with the energy window of +/-10% from the photopeak. Line-phantom images are presented to visualize the spatial resolution provided by SemiSPECT, and images of bone, myocardium, and human tumor xenografts in mice demonstrate the feasibility of preclinical small-animal studies with SemiSPECT.


Identification of a novel human Rho protein with unusual properties: GTPase deficiency and in vivo farnesylation.
We have identified a human Rho protein, RhoE, which has unusual structural and biochemical properties that suggest a novel mechanism of regulation. Within a region that is highly conserved among small GTPases, RhoE contains amino acid differences specifically at three positions that confer oncogenicity to Ras (12, 59, and 61). As predicted by these substitutions, which impair GTP hydrolysis in Ras, RhoE binds GTP but lacks intrinsic GTPase activity and is resistant to Rho-specific GTPase-activating proteins. Replacing all three positions in RhoE with conventional amino acids completely restores GTPase activity. In vivo, RhoE is found exclusively in the GTP-bound form, suggesting that unlike previously characterized small GTPases, RhoE may be normally maintained in an activated state. Thus, amino acid changes in Ras that are selected during tumorigenesis have evolved naturally in this Rho protein and have similar consequences for catalytic function. All previously described Rho family proteins are modified by geranylgeranylation, a lipid attachment required for proper membrane localization. In contrast, the carboxy-terminal sequence of RhoE predicts that, like Ras proteins, RhoE is normally farnesylated. Indeed, we have found that RhoE in farnesylated in vivo and that this modification is required for association with the plasma membrane and with an unidentified cellular structure that may play a role in adhesion. Thus, two unusual structural features of this novel Rho protein suggest a striking evolutionary divergence from the Rho family of GTPases.


Symptom profiles in children with ADHD: effects of comorbidity and gender.
To examine ratings and objective measures of attention-deficit/hyperactivity disorder (ADHD) symptoms to assess whether ADHD children with and without comorbid conditions have equally high levels of core symptoms and whether symptom profiles differ as a function of comorbidity and gender. Four hundred ninety-eight children from the NIMH Collaborative Multisite Multimodal Treatment Study of Children With Attention-Deficit/Hyperactivity Disorder (MTA) were divided into comorbid groups based on the parent Diagnostic Interview Schedule for Children and assessed via parents' and teachers' Swanson, Nolan, and Pelham (SNAP) ratings and a continuous performance test (CPT). Comorbidity and gender effects were examined using analyses of covariance controlled for age and site. CPT inattention, impulsivity, and dyscontrol errors were high in all ADHD groups. Children with ADHD + oppositional defiant or conduct disorder were rated as more impulsive than inattentive, while children with ADHD + anxiety disorders (ANX) were relatively more inattentive than impulsive. Girls were less impaired than boys on most ratings and several CPT indices, particularly impulsivity, and girls with ADHD + ANX made fewer CPT impulsivity errors than girls with ADHD-only. Children with ADHD have high levels of core symptoms as measured by rating scales and CPT, irrespective of comorbidity. However, there are important differences in symptomatology as a function of comorbidity and gender.


A comparison of the performance of microsatellite and methylation urine analysis for predicting the recurrence of urothelial cell carcinoma, and definition of a set of markers by Bayesian network analysis.
To compare the potential of two diagnostic methods for detecting recurrence of urothelial cell carcinoma (UCC) of the bladder, by (i) detecting alterations in microsatellite DNA markers and loss of heterozygosity (LOH), and (ii) detecting aberrant gene hypermethylation, as UCC has a high recurrence rate in the urinary tract and the disease can invade muscle if new tumours are overlooked. Over 1 year, urine samples were retrieved from 40 patients already diagnosed with bladder UCC (30 pTa, two pTis, eight pT1). Samples were collected 6 months after bladder tumour resection, during the follow-up schedule. We used samples to analyse nine microsatellite markers and the methylation status of 11 gene promoters. Receiver operating characteristic curves were generated and Bayesian statistics used to create an interaction network between recurrence and the biomarkers. During the study, 15 of the 40 patients (38%) had a tumour recurrence and 14 were identified by cystoscopy (reference method). Overall, microsatellite markers (area under curve, AUC 0.819, 95% confidence interval, CI, 0.677-0.961) had better performance characteristics than promoter hypermethylation (AUC 0.448, 0.259-0.637) for detecting recurrence. A marker panel of IFNA, MBP, ACTBP2, D9S162 and of RASSF1A, and WIF1 generated a higher diagnostic accuracy of 86% (AUC 0.92, 0.772-0.981). Microsatellite markers have better performance characteristics than promoter hypermethylation for detecting UCC recurrence. These data support the further development of a combination of only six markers from both methods in urinary DNA. Once validated, it could be used routinely during the follow-up for the early detection and surveillance of UCC from the lower and upper urinary tract.


Prevention of esophageal cancer: the nutrition intervention trials in Linxian, China. Linxian Nutrition Intervention Trials Study Group.
In Linxian China, the esophageal/gastric cardia cancer mortality rates are among the highest in the world. There is suspicion that the population's chronic deficiencies of multiple micronutrients are etiologically involved. We conducted two randomized, placebo-controlled nutrition intervention trials to test the effects of vitamin and mineral supplements in lowering the rates of esophageal/gastric cancer. In the first trial, the dysplasia trial, 3318 adults with a cytological diagnosis of esophageal dysplasia received daily supplementation with 26 vitamins and minerals in doses typically 2-3 times the United States Recommended Daily Allowances, or placebos, for 6 years. The second trial, the general population trial, involved 29,584 adults and used a one-half replicate of a 2(4) factorial experimental design which tested the effects of four combinations of nutrients: A, retinol and zinc; B, riboflavin and niacin; C, vitamin C and molybdenum; and D, beta-carotene, vitamin E, and selenium. Doses for these daily supplements ranged from 1 to 2 times the United States Recommended Daily Allowances, and the different vitamin/mineral combinations or placebos were taken for a period of 5.25 years. As part of the general population trial, and end-of-intervention endoscopy survey was carried out in a small (1.3%) sample of subjects to see if supplementation affected the prevalence of dysplasia and early cancer. Herein we review the methods of these trials and the results of the endoscopic survey. Fifteen esophageal and 16 gastric cancers were identified in endoscopic biopsies from the 391 subjects evaluated from two villages, and nearly all were asymptomatic. No significant reductions in the prevalence of esophageal or gastric dysplasia or cancer were seen with any of the four supplement groups. However, the prevalence of gastric cancer among participants receiving retinol and zinc was 62% lower than those not receiving those supplements (P = 0.09), while participants receiving beta-carotene, vitamin E, and selenium had a 42% reduction in esophageal cancer prevalence (0.34). We have reported separately that cancer mortality over the entire 5.25-year period was significantly reduced among those receiving beta-carotene, vitamin E, and selenium. The findings from the overall trial and the endoscopic sample offer a hopeful sign and should encourage additional studies with these agents in larger numbers of subjects.


Ordinal-level phylogenomics of the arthropod class Diplopoda (millipedes) based on an analysis of 221 nuclear protein-coding loci generated using next-generation sequence analyses.
The ancient and diverse, yet understudied arthropod class Diplopoda, the millipedes, has a muddled taxonomic history. Despite having a cosmopolitan distribution and a number of unique and interesting characteristics, the group has received relatively little attention; interest in millipede systematics is low compared to taxa of comparable diversity. The existing classification of the group comprises 16 orders. Past attempts to reconstruct millipede phylogenies have suffered from a paucity of characters and included too few taxa to confidently resolve relationships and make formal nomenclatural changes. Herein, we reconstruct an ordinal-level phylogeny for the class Diplopoda using the largest character set ever assembled for the group. Transcriptomic sequences were obtained from exemplar taxa representing much of the diversity of millipede orders using second-generation (i.e., next-generation or high-throughput) sequencing. These data were subject to rigorous orthology selection and phylogenetic dataset optimization and then used to reconstruct phylogenies employing Bayesian inference and maximum likelihood optimality criteria. Ancestral reconstructions of sperm transfer appendage development (gonopods), presence of lateral defense secretion pores (ozopores), and presence of spinnerets were considered. The timings of major millipede lineage divergence points were estimated. The resulting phylogeny differed from the existing classifications in a number of fundamental ways. Our phylogeny includes a grouping that has never been described (Juliformia+Merocheta+Stemmiulida), and the ancestral reconstructions suggest caution with respect to using spinnerets as a unifying characteristic for the Nematophora. Our results are shown to have significantly stronger support than previous hypotheses given our data. Our efforts represent the first step toward obtaining a well-supported and robust phylogeny of the Diplopoda that can be used to answer many questions concerning the evolution of this ancient and diverse animal group.


Extracellular matrix protein CCN1 limits oncolytic efficacy in glioma.
Oncolytic viral therapy has been explored widely as an option for glioma treatment but its effectiveness has remained limited. Cysteine rich 61 (CCN1) is an extracellular matrix (ECM) protein elevated in cancer cells that modulates their adhesion and migration by binding cell surface receptors. In this study, we examined a hypothesized role for CCN1 in limiting the efficacy of oncolytic viral therapy for glioma, based on evidence of CCN1 induction that occurs in this setting. Strikingly, we found that exogenous CCN1 in glioma ECM orchestrated a cellular antiviral response that reduced viral replication and limited cytolytic efficacy. Gene expression profiling and real-time PCR analysis revealed a significant induction of type-I interferon responsive genes in response to CCN1 exposure. This induction was accompanied by activation of the Jak/Stat signaling pathway, consistent with induction of an innate antiviral cellular response. Both effects were mediated by the binding of CCN1 to the cell surface integrin α6β1, activating its signaling and leading to rapid secretion of interferon-α, which was essential for the innate antiviral effect. Together, our findings reveal how an integrin signaling pathway mediates activation of a type-I antiviral interferon response that can limit the efficacy of oncolytic viral therapy. Furthermore, they suggest therapeutic interventions to inhibit CCN1-integrin α6 interactions to sensitize gliomas to viral oncolysis.


Nodulosis in systemic onset juvenile idiopathic arthritis: an uncommon event with spontaneous resolution.
Accelerated nodulosis is a rare complication of methotrexate therapy in juvenile idiopathic arthritis. When nodulosis does occur in patients with juvenile idiopathic arthritis on methotrexate, it is almost always seen in patients with polyarthritis with rheumatoid factor seropositivity, but only occasionally in polyarthritis patients who are rheumatoid factor negative. It has been described previously in only one patient with systemic arthritis. In this study, we describe three patients with systemic arthritis, all of whom developed nodulosis while receiving methotrexate. Interestingly, it was not necessary to discontinue methotrexate in any of these patients. In fact, methotrexate dose was escalated without consequences and with complete resolution of nodules. This observation suggests that nodulosis occurring in patients with systemic arthritis already on methotrexate may not be because of methotrexate itself, but may be a component of the disease process. The other likely possibility given the fact that nodulosis occurs with other immunomodulatory agents and is not specifically related to methotrexate is that immunomodulatory agents in general precipitate the development of nodulosis. Thus, we propose that the new terminology "immunomodulatory agents induced nodulosis" rather than "methotrexate-induced nodulosis" be used in the literature.


Color reproduction from noisy CFA data of single sensor digital cameras.
Single sensor digital color still/video cameras capture images using a color filter array (CFA) and require color interpolation (demosaicking) to reconstruct full color images. The color reproduction has to combat sensor noises which are channel dependent. If untreated in demosaicking, sensor noises can cause color artifacts that are hard to remove later by a separate denoising process, because the demosaicking process complicates the noise characteristics by blending noises of different color channels. This paper presents a joint demosaicking-denoising approach to overcome this difficulty. The color image is restored from noisy mosaic data in two steps. First, the difference signals of color channels are estimated by linear minimum mean square-error estimation. This process exploits both spectral and spatial correlations to simultaneously suppress sensor noise and interpolation error. With the estimated difference signals, the full resolution green channel is recovered. The second step involves in a wavelet-based denoising process to remove the CFA channel-dependent noises from the reconstructed green channel. The red and blue channels are subsequently recovered. Simulated and real CFA mosaic data are used to evaluate the performance of the proposed joint demosaicking-denoising scheme and compare it with many recently developed sophisticated demosaicking and denoising schemes.


Developing a Dissemination Model to Improve Intervention Reach among West Virginia Youth Smokers.
The not-on-tobacco program is an evidence-based teen smoking cessation program adopted by the American Lung Association (ALA). Although widely disseminated nationally via ALA Master Trainers, in recent years, adoption and implementation of the N-O-T program in West Virginia (WV) has slowed. WV, unfortunately, has one of the highest smoking rates in the US. Although it is a goal of public health science, dissemination of evidence-based interventions is woefully understudied. The present manuscript reviews a theoretical model of dissemination of the not-on-tobacco program in WV. Based on social marketing, diffusion of innovations, and social cognitive theories, the nine-phase model incorporates elements of infrastructure development, accountability, training, delivery, incentives, and communication. The model components as well as preliminary lessons learned from initial implementation are discussed.


Photochemically-assisted synthesis and photophysical properties of difluoroboronated β-diketones with fused four-benzene-ring chromophores, chrysene and pyrene.
We investigated the photophysical properties of difluoroboronated β-diketones (BF2DK) with chrysene and pyrene skeletons (ChB and PyB, respectively) in solution and in the solid state. Acetylchrysenes, as the key precursors to ChBs, were photochemically prepared from the corresponding (acetylphenyl)naphthylethenes by means of a modified photocyclization method. The absorption and emission spectra of the BF2DKs were obtained in chloroform and acetonitrile, and the quantum yields and lifetimes of the fluorescence were determined. Excimeric fluorescence from PyB was absent even in highly concentrated solution. Based on the Lippert-Mataga analysis of the absorption and fluorescence features, the photophysical properties of the ChBs were discussed in comparison with those of PyB. The fluorescence states of the studied BF2DKs are shown to be of a charge-transfer character. The fluorescence quantum yields decrease with increasing the solvent polarity due to the enhanced internal conversion process. The fluorescence quantum yields in the solid state of the studied BF2DKs were determined, and it was found that PyB is fluorescent, whereas the fluorescence quantum yields of the ChBs depend on the substituted position of the chrysene moiety.


Auditory brainstem implantation in primates: lessons for human surgery and application.
We report on the surgical technique for surface electro-auditory prosthesis (EAP) implantation, pathological changes occurring at the cochlear nucleus complex (CNC), and its relation with electrical stimulation. Fourteen Macaca fascicularis were operated upon for a translabyrinthine bilateral auditory neurectomy, and simultaneous unilateral EAP implantation. Six animals were not stimulated, and the remaining eight were connected to an external active device. Stimulation was planned for 1000 hours. Biotolerance to the materials was adequate without significant reactions in the CNC surface, but an ependymal reaction. Lesions attributed to surgical trauma were also found. Two animals being stimulated could not complete the planned course due to cable break or EAP extrusion. One stimulated animal developed an asymptomatic brainstem abscess. A good knowledge of CNC topography is required to avoid surgical trauma. Externally connected devices may facilitate extrusion of the EAP or ascending infections.


Endoscopic approach to maxillofacial trauma.
Endoscopic approaches to maxillofacial trauma have included their use in temporomandibular joint arthroscopy, zygomatic arch repair, repair after orbital trauma, repair of frontal sinus fractures, and finally in the repair of subcondylar mandibular fractures. These techniques provide exciting new options for the management of facial fractures. Many of the techniques provide a steep learning curve and require specialized equipment. The approach to an orbital blowout fracture allows the use of a Caldwell-Luc approach and the use of a naturally occurring body space (the maxillary sinus) for the visualization and maneuvering of the endoscope, tools, and implants. The use of the endoscope in the management of fractures for facial structures such as mandibular subcondylar fractures, which do not have a naturally occurring body cavity, may present greater challenges when trying to obtain visualization, reduction, and fixation. When used for the treatment of subcondylar fractures, wide exposure and the use of right-angled drills and screw drivers may significantly help with this procedure. This may be considered by some surgeons as an endoscopically "assisted" procedure that can possibly be accomplished with direct visualization and the use of dental mirrors. It remains to be seen as to whether some of these diverse endoscopic applications will represent the standard of care in the future or remain a specialized technique practiced by a minority of surgeons in a few specialized centers. This article reviews various endoscopic applications to maxillofacial trauma and discusses some of the controversies of these techniques.


Longitudinal associations between micronutrient consumption and leukocyte telomere length.
There are few studies on the association between nutrient intake and telomere length, which may reflect cumulative oxidative stress and indicate biological ageing. In the present study, we evaluated longitudinal associations between the consumption of micronutrients, including antioxidant nutrients and B vitamins involved in one-carbon transfer pathways, and leukocyte telomere length (LTL). The study included 1958 middle-aged and older Korean men and women (age range at baseline: 40-69 years) from a population-based cohort. We collected dietary information at baseline using a semiquantitative food frequency questionnaire (June 2001 to January 2003) and assessed the consumption of micronutrients, including vitamins A, B1 , B2 , B3 , B6 , B9 (folate), C and E, as well as calcium, phosphorus, potassium, iron and zinc. We measured LTL using a real-time polymerase chain reaction at the 10-year follow-up examination (February 2011 to November 2012). In the multiple regression model adjusted for potential confounders, LTL was positively associated with the consumption of vitamin C (P < 0.05), folate (P = 0.05) and potassium (P = 0.05) in all participants. In the age-stratified analysis, the association between the consumption of vitamin C (P < 0.01), folate (P < 0.05) and potassium (P < 0.05) with LTL was significant only among participants aged <50 years. Our findings suggest that the earlier consumption of vitamin C, folate and potassium, which are abundant in fruits and vegetables, can delay biological ageing in middle-aged and older adults.


What is the future of training in surgery? Needs assessment of national stakeholders.
The Curriculum Committee of the American College of Surgeons-Accredited Educational Institutes conducted a need assessment to (1) identify gaps between ideal and actual practices in areas of surgical care, (2) explore educational solutions for addressing these gaps, and (3) shape a vision to advance the future of training in surgery. National stakeholders were recruited from the committee members' professional network and interviewed via telephone. Interview questions targeted areas for improving surgical patient care, optimal educational solutions for training in surgery including simulation roles, and entities that should primarily bear training costs. We performed an iterative, qualitative analysis including member checking to identify key themes. Twenty-two interviewees included state/national board representatives, risk managers, multispecialty faculty/program directors, nurses, trainees, an industry representative, and a patient. Surgeons' communication with patients, families, and team members was raised consistently by stakeholders as a way to establish clear expectations regarding pre-, peri-, and postoperative care. Other comments highlighted the surgeon's development and demonstration and maintenance of cognitive and technical skills, including surgical judgment. Stakeholders also reiterated the critical need for surgeons to engage in on-going self-assessment and professional development to identify and remediate recognized limitations. Recommended learning modalities for meeting surgeons' needs included active learning (deliberate practice, diverse patient experiences), experiential learning (simulation), and peer and mentored learning (preceptorship). This first formal needs assessment of education for surgeons points to opportunities for educational programs in patient-centered communication, learning models that match preferences of new generations of trainees, and training in interprofessional/interdisciplinary team communication and teamwork.


Clinical aspects of paediatric mastocytosis: a review of 101 cases.
Cutaneous mastocytosis (CM) is a typical presentation of mastocytosis in children. However, systemic mastocytosis may also occur in children. We tried to characterize the clinical features of childhood-onset mastocytosis and estimate the value of the SCORMA (SCORing Mastocytosis) Index and serum tryptase levels as disease severity parameters. In a survey of 101 children mastocytosis was diagnosed and classified according to World Health Organization criteria. In all the cases serum tryptase levels and the SCORMA Index were done to assess the extent and intensity of the disease. Cutaneous mastocytosis was diagnosed in 100 children; 84% of them presented maculopapular CM, 10% mastocytoma and 6% diffuse cutaneous mastocytosis. Moreover, systemic mastocytosis with bone marrow infiltration and associated with maculopapular CM was found in one case. There was a positive correlation of serum tryptase level to the SCORMA Index. Both the mean tryptase level and the mean SCORMA Index were elevated in diffuse cutaneous mastocytosis children when compared with other forms CM. A significantly higher mean tryptase level was found in children with flushing, hypotension, diarrhoea, extensive bullous lesions and osteoporosis or osteopenia. Mastocytosis in children usually has a benign course. Nevertheless, severe mediator-related symptoms and systemic involvement may appear. Therefore, a multidisciplinary approach involving careful monitoring of the serum tryptase level, SCORMA Index and the organ function is recommended. Both tryptase levels and the SCORMA Index are of a great value as disease severity parameters and they should be assessed simultaneously in all mastocytosis patients.


Using theatre in preceptorship.
Using theatre methods and techniques in post registration education has the unique advantage of facilitating learning that is based in the reality of practice yet in a safe space free from the demands and pressures of the work place. It offers compassionate insights that allow participants to reflect on others' behaviour and how their behaviours can impact on the culture in which they work. This article offers a description of how this has been implemented within an acute NHS trust as part of a preceptorship programme in three ways: using a professional theatre company, with other educators and as an individual educator.


Pathogenesis of Cryptococcus neoformans in congenitally immunodeficient beige athymic mice.
Mortality after intravenous challenge with 10(4) Cryptococcus neoformans demonstrated that doubly immunodeficient beige athymic (bg/bg nu/nu) mice were more susceptible to systemic cryptococcosis than either bg/bg or nu/nu mice. Infected bg/bg nu/nu mice also had a shortened lifespan compared with their bg/bg nu/+ littermates. Beige athymic (bg/bg nu/nu) but not bg/bg nu/+mice developed cryptococcal lesions in the skin, demonstrating that C. neoformans is dermatotropic in a T-cell-deficient host. Higher numbers of C. neoformans were isolated from the lungs and spleen of infected bg/bg nu/nu than bg/bg nu/+ mice as early as day 3 after challenge, indicating that in lymphoid-rich organs, T cells can alter the course of systemic cryptococcosis early in the infection. Despite extensive abscess formation in the brains of bg/bg nu/+ mice, dissemination and growth rate of C. neoformans in the brain was similar in both genotypes. The primary histopathological feature in tissues from bg/bg nu/nu mice infected with C. neoformans consisted of foci of encapsulated yeast cells with minimal to no inflammatory response. In contrast to bg/bg nu/nu mice, bg/bg nu/+ mice mounted a vigorous inflammatory response to C. neoformans that progressed from acute to chronic inflammation. Beige athymic mice are a new animal model that will be useful in clarifying the innate and acquired immune factors important in resistance to cryptococcosis.


Auxins reverse plant male sterility caused by high temperatures.
With global warming, plant high temperature injury is becoming an increasingly serious problem. In wheat, barley, and various other commercially important crops, the early phase of anther development is especially susceptible to high temperatures. Activation of auxin biosynthesis with increased temperatures has been reported in certain plant tissues. In contrast, we here found that under high temperature conditions, endogenous auxin levels specifically decreased in the developing anthers of barley and Arabidopsis. In addition, expression of the YUCCA auxin biosynthesis genes was repressed by increasing temperatures. Application of auxin completely reversed male sterility in both plant species. These findings suggest that tissue-specific auxin reduction is the primary cause of high temperature injury, which leads to the abortion of pollen development. Thus, the application of auxin may help sustain steady yields of crops despite future climate change.


Dose-dependent regulation of cell proliferation and collagen degradation by estradiol on ligamentum flavum.
Estradiol plays an important role in the regulation of collagen metabolism. Deficiency of estradiol has been reported to be associated with the degeneration of many connective tissues. However, the association of estradiol and hypertrophy of the ligamentum flavum was seldom explored. Therefore, we studied the effects of estradiol on cultured cells from the ligamentum flavum. Primary cultures of human ligamentum flavum cells obtained from surgical specimens of 14 patients undergoing spinal surgery were used to investigate the effect of estradiol on cell proliferation and the expression of collagen, elastin, and matrix metalloproteinases. Downstream pathways of estrogen receptor underlying the regulation of metalloproteinases were also investigated. In our study, we revealed the existence of estrogen receptors on both female and male ligamentum flavum cells with a gender difference. 17β-estradiol increased early (24 hours) proliferation of ligamentum flavum cells in a dose dependent manner and the effect could not be seen when the cell density increased. Estradiol with a concentration of 10(-9) M decreased collagen levels and increased the expression of MMP-13. Adding an antagonist of PI3K downstream pathway could reverse the expression of MMP-13 caused by estradiol. The results implied estradiol regulated the expression of MMP-13 via PI3K pathway and contributed to the homeostasis of extracellular matrix in the ligamentum flavum.


Antioxidant properties of kilned and roasted malts.
Compounds possessing antioxidant activity play a crucial role in delaying or preventing lipid oxidation in foods and beverages during processing and storage. Such reactions lead to loss of product quality, especially as a consequence of off-flavor formation. The aim of this study was to determine the antioxidant activity of kilned (standard) and roasted (speciality) malts in relation to phenolic compounds, sugars, amino acids, and color [assessed as European Brewing Convention units (degrees EBC) and absorbance at 420 nm]. The concentrations of sugars and amino acids decreased with the intensity of the applied heat treatment, and this was attributed to the extent of the Maillard reaction, as well as sugar caramelization, in the highly roasted malts. Proline, followed by glutamine, was the most abundant free amino/imino acid in the malt samples, except those that were highly roasted, and maltose was the most abundant sugar in all malts. Levels of total phenolic compounds decreased with heat treatment. Catechin and ferulic acid were the most abundant phenolic compounds in the majority of the malts, and amounts were highest in the kilned samples. In highly roasted malts, degradation products of ferulic acid were identified. Antioxidant activity increased with the intensity of heating, in parallel with color formation, and was significantly higher for roasted malts compared to kilned malts. In kilned malts, phenolic compounds were the main identified contributors to antioxidant activity, with Maillard reaction products also playing a role. In roasted malts, Maillard reaction products were responsible for the majority of the antioxidant activity.


Chronic Alcohol Consumption Exacerbates the Severity of Psoriasiform Dermatitis in Mice.
A relationship between alcohol consumption and psoriasis has been reported, but it is unclear whether alcohol consumption aggravates psoriasis. Here, we studied the effect of chronic ethanol (EtOH) consumption in the murine model of Aldara-induced psoriasiform dermatitis. C57BL/6 mice received 5% EtOH in their drinking water for 10 weeks. Dermatitis was induced from weeks 9 to 10, by applying Aldara to the shaved patch of skin on the back. Inflammation was characterized by histological and transcriptomic analyses. EtOH consumption aggravated Aldara-induced dermatitis. The scales were more severe, epidermal thickening was more pronounced, and cutaneous expression of Th17-related cytokines was exacerbated. Control mice simply receiving EtOH displayed minimal cutaneous inflammation, characterized by epidermal infiltrates of T lymphocytes and the overexpression of IL-17A and the Th17-recruiting chemokine CCL20. In vitro studies showed that low concentrations of EtOH induce the expression of CCL20 by murine epidermal keratinocytes. Alcohol consumption leads to subliminar skin inflammation, which is revealed by the exacerbation of Aldara-induced experimental psoriasiform dermatitis, likely through Th17-type minimal skin inflammation. These results favor the systematic management of alcohol consumption in psoriatic patients.


Meeting physical activity recommendations is associated with health-related quality of life in women before and after Roux-en-Y gastric bypass surgery.
Meeting physical activity (PA) recommendations is positively associated with health-related quality of life (HRQoL), but it is still unclear whether PA (specifically objectively measured) is associated with HRQoL in bariatric surgery candidates, both before and after surgery. Thus, the aim of this study was to examine the cross-sectional association between meeting objectively measured PA recommendations and HRQoL before and after Roux-en-Y gastric bypass (RYGB) surgery. Sixty-six women undergoing RYGB with pre-surgery and 62 women with post-surgery valid PA and HRQoL data were included from the control group of a RCT study aiming at improving HRQoL and PA post-RYGB surgery. Measures before and 12 months after RYGB included the Short Form Health Survey (SF-36) and objectively measured PA, sedentary time (ST), and step counts with GT3X+ accelerometers. Multiple linear regression models were used to analyze the associations between PA and HRQoL. Participants who engaged in more than 150 min of moderate to vigorous PA (MVPA)/week (PA recommendations) had considerably higher SF-36 scores (HRQoL) than those who did not, both pre and 12-month post-surgery, with greatest difference in the subscale bodily pain, 15.5 (p = 0.021) higher score (higher scores means less pain) pre-surgery and a 19.7 (p = 0.004) higher score post-surgery. Higher LPA and step counts and lower ST also showed positive associations in some of the subscales of SF-36. Meeting the PA recommendations and overall engaging in more PA was associated with higher HRQoL, pre-, and post-RYGB surgery, highlighting the importance of PA both pre- and post-surgery.


Metagenome-based diversity analyses suggest a strong locality signal for bacterial communities associated with oyster aquaculture farms in Ofunato Bay.
Ofunato Bay, in Japan, is the home of buoy-and-rope-type oyster aquaculture activities. Since the oysters filter suspended materials and excrete organic matters into the seawater, bacterial communities residing in its vicinity may show dynamic changes depending on the oyster culture activities. We employed a shotgun metagenomic technique to study bacterial communities near oyster aquaculture facilities at the center of the bay (KSt. 2) and compared the results with those of two other localities far from the station, one to the northeast (innermost bay, KSt. 1) and the other to the southwest (bay entrance, KSt. 3). Seawater samples were collected every month from January to December 2015 from the surface (1 m) and deeper (8 or 10 m) layers of the three locations, and the sequentially filtered fraction on 0.2-μm membranes was sequenced on an Illumina MiSeq system. The acquired reads were uploaded to MG-RAST for KEGG functional abundance analysis, while taxonomic analyses at the phylum and genus levels were performed using MEGAN after parsing the BLAST output. Discrimination analyses were then performed using the ROC-AUC value of the cross validation, targeting the depth (shallow or deep), locality [(KSt. 1 + KSt. 2) vs. KSt 3; (KSt. 1 + KSt. 3) vs. KSt. 2 or the (KSt. 2 + KSt. 3) vs. KSt. 1] and seasonality (12 months). The matrix discrimination analysis on the adjacent 2 continuous seasons by ROC-AUC, which was based on the datasets that originated from different depths, localities and months, showed the strongest discrimination signal on the taxonomy matrix at the phylum level for the datasets from July to August compared with those from September to June, while the KEGG matrix showed the strongest signal for the datasets from March to June compared with those from July to February. Then, the locality combination was subjected to the same ROC-AUC discrimination analysis, resulting in significant differences between KSt. 2 and KSt. 1 + KSt. 3 on the KEGG matrix. These results suggest that aquaculture activities markedly affect bacterial functions.


Feasibility and safety of extended-release naltrexone treatment of opioid and alcohol use disorder in HIV clinics: a pilot/feasibility randomized trial.
HIV-infected people with substance use disorders are least likely to benefit from advances in HIV treatment. Integration of extended-release naltrexone (XR-NTX) into HIV clinics may increase engagement in the HIV care continuum by decreasing substance use. We aimed to compare (1) XR-NTX treatment initiation, (2) retention and (3) safety of XR-NTX versus treatment as usual (TAU) for treating opioid use disorder (OUD) and/or alcohol use disorder (AUD) in HIV clinics. Non-blinded randomized trial of XR-NTX versus pharmacotherapy TAU. HIV primary care clinics in Vancouver, BC, Canada and Chicago, IL, USA. Fifty-one HIV-infected patients seeking treatment for OUD (n = 16), AUD (n = 27) or both OUD and AUD (n = 8). Primary outcomes were XR-NTX initiation (receipt of first injection within 4 weeks of randomization) and retention at 16 weeks. Secondary outcomes generated point estimates for change in substance use, HIV viral suppression [HIV RNA polymerase chain reaction (pcr) < 200 copies/ml] and safety. Two-thirds (68%) of participants assigned to XR-NTX initiated treatment, and 88% of these were retained on XR-NTX at 16 weeks. In comparison, 96% of TAU participants initiated treatment, but only 50% were retained on medication at 16 weeks. Mean days of opioid use in past 30 days decreased from 17.3 to 4.1 for TAU and from 20.3 to 7.7 for XR-NTX. Mean heavy drinking days decreased from 15.6 to 5.7 for TAU and 12.5 to 2.8 for XR-NTX. Among those with OUD, HIV suppression improved from 67 to 80% for XR-NTX and 58 to 75% for TAU. XR-NTX was well tolerated, with no precipitated withdrawals and one serious injection-site reaction. Extended-release naltrexone (XR-NTX) is feasible and safe for treatment of opioid use disorder and alcohol use disorder in HIV clinics. Treatment initiation appears to be lower and retention greater for XR-NTX compared with treatment as usual (clinicaltrials.gov NCT01908062).


Autologous bilayered self-assembled skin substitutes (SASSs) as permanent grafts: a case series of 14 severely burned patients indicating clinical effectiveness.
Split-thickness skin autografts (AGs) are the standard surgical treatment for severe burn injuries. However, the treatment of patients with substantial skin loss is limited by the availability of donor sites for skin harvesting. As an alternative to skin autografts, our research group developed autologous self-assembled skin substitutes (SASSs), allowing the replacement of both dermis and epidermis in a single surgical procedure. The aim of the study was to assess the clinical outcome of the SASSs as a permanent coverage for full-thickness burn wounds. Patients were recruited through the Health Canada's Special Access Program. SASSs were grafted on debrided full-thickness wounds according to similar protocols used for AGs. The graft-take and the persistence of the SASS epithelium over time were evaluated. 14 patients received surgical care with SASSs. The mean percentage of the SASS graft-take was 98 % (standard deviation = 5) at 5 to 7 d after surgery. SASS integrity persisted over time (average follow-up time: 3.2 years), without noticeable deficiency in epidermal regeneration. Assessment of scar quality (skin elasticity, erythema, thickness) was performed on a subset of patients. Non-homogeneous pigmentation was noticed in several patients. These results indicated that the SASS allowed the successful coverage of full-thickness burns given its high graft-take, aesthetic outcome equivalent to autografting and the promotion of long-term tissue regeneration. When skin donor sites are in short supply, SASSs could be a valuable alternative to treat patients with full-thickness burns covering more than 50 % of their total body surface area.


Urokinase-type plasminogen activator is a preferred substrate of the human epithelium serine protease tryptase epsilon/PRSS22.
Tryptase epsilon is a member of the chromosome 16p13.3 family of human serine proteases that is preferentially expressed by epithelial cells. Recombinant pro-tryptase epsilon was generated to understand how the exocytosed zymogen might be activated outside of the epithelial cell, as well as to address its possible role in normal and diseased states. Using expression/site-directed mutagenesis approaches, we now show that Lys20, Cys90, and Asp92 in the protease's substrate-binding cleft regulate its enzymatic activity. We also show that Arg(-1) in the propeptide domain controls its ability to autoactivate. In vitro studies revealed that recombinant tryptase epsilon possesses a restricted substrate specificity. Once activated, tryptase epsilon cannot be inhibited effectively by the diverse array of protease inhibitors present in normal human plasma. Moreover, this epithelium protease is not highly susceptible to alpha1-antitrypsin or secretory leukocyte protease inhibitor, which are present in the lung. Recombinant tryptase epsilon could not cleave fibronectin, vitronectin, laminin, single-chain tissue-type plasminogen activator, plasminogen, or any prominent serum protein. Nevertheless, tryptase epsilon readily converted single-chain pro-urokinase-type plasminogen activator (pro-uPA/scuPA) into its mature, enzymatically active protease. Tryptase epsilon also was able to induce pro-uPA-expressing smooth muscle cells to increase their migration through a basement membrane-like extracellular matrix. The ability to activate uPA in the presence of varied protease inhibitors suggests that tryptase epsilon plays a prominent role in fibrinolysis and other uPA-dependent reactions in the lung.


Hypoxia induces accumulation of p53 protein, but activation of a G1-phase checkpoint by low-oxygen conditions is independent of p53 status.
It has been convincingly demonstrated that genotoxic stresses cause the accumulation of the tumor suppressor gene p53. One important consequence of increased p53 protein levels in response to DNA damage is the activation of a G1-phase cell cycle checkpoint. It has also been shown that G1-phase cell cycle checkpoints are activated in response to other stresses, such as lack of oxygen. Here we show that hypoxia and heat, agents that induce cellular stress primarily by inhibiting oxygen-dependent metabolism and denaturing proteins, respectively, also cause an increase in p53 protein levels. The p53 protein induced by heat is localized in the cytoplasm and forms a complex with the heat shock protein hsc70. The increase in nuclear p53 protein levels and DNA-binding activity and the induction of reporter gene constructs containing p53 binding sites following hypoxia occur in cells that are wild type for p53 but not in cells that possess mutant p53. However, unlike ionizing radiation, the accumulation of cells in G1 phase by hypoxia is not strictly dependent on wild-type p53 function. In addition, cells expressing the human papillomavirus E6 gene, which show increased degradation of p53 by ubiquitination and fail to accumulate p53 in response to DNA-damaging agents, do increase their p53 levels following heat and hypoxia. These results suggest that hypoxia is an example of a "nongenotoxic" stress which induces p53 activity by a different pathway than DNA-damaging agents.


OSCA1 mediates osmotic-stress-evoked Ca2+ increases vital for osmosensing in Arabidopsis.
Water is crucial to plant growth and development. Environmental water deficiency triggers an osmotic stress signalling cascade, which induces short-term cellular responses to reduce water loss and long-term responses to remodel the transcriptional network and physiological and developmental processes. Several signalling components that have been identified by extensive genetic screens for altered sensitivities to osmotic stress seem to function downstream of the perception of osmotic stress. It is known that hyperosmolality and various other stimuli trigger increases in cytosolic free calcium concentration ([Ca(2+)]i). Considering that in bacteria and animals osmosensing Ca(2+) channels serve as osmosensors, hyperosmolality-induced [Ca(2+)]i increases have been widely speculated to be involved in osmosensing in plants. However, the molecular nature of corresponding Ca(2+) channels remain unclear. Here we describe a hyperosmolality-gated calcium-permeable channel and its function in osmosensing in plants. Using calcium-imaging-based unbiased forward genetic screens we isolated Arabidopsis mutants that exhibit low hyperosmolality-induced [Ca(2+)]i increases. These mutants were rescreened for their cellular, physiological and developmental responses to osmotic stress, and those with clear combined phenotypes were selected for further physical mapping. One of the mutants, reduced hyperosmolality-induced [Ca(2+)]i increase 1 (osca1), displays impaired osmotic Ca(2+) signalling in guard cells and root cells, and attenuated water transpiration regulation and root growth in response to osmotic stress. OSCA1 is identified as a previously unknown plasma membrane protein and forms hyperosmolality-gated calcium-permeable channels, revealing that OSCA1 may be an osmosensor. OSCA1 represents a channel responsible for [Ca(2+)]i increases induced by a stimulus in plants, opening up new avenues for studying Ca(2+) machineries for other stimuli and providing potential molecular genetic targets for engineering drought-resistant crops.


Brain imaging and fluid biomarker analysis in young adults at genetic risk for autosomal dominant Alzheimer's disease in the presenilin 1 E280A kindred: a case-control study.
We have previously characterised functional brain abnormalities in young adults at genetic risk for late-onset Alzheimer's disease. To gain further knowledge on the preclinical phase of Alzheimer's disease, we sought to characterise structural and functional MRI, CSF, and plasma biomarkers in a cohort of young adults carrying a high-penetrance autosomal dominant mutation that causes early-onset Alzheimer's disease. Between January and August, 2010, 18-26-year-old presenilin 1 (PSEN1) E280A mutation carriers and non-carriers from the Colombian Alzheimer's Prevention Initiative Registry in Medellín Antioquia, Colombia, had structural MRI, functional MRI during associative memory encoding and novel viewing and control tasks, and cognitive assessments. Consenting participants also had lumbar punctures and venepunctures. Outcome measures were task-dependent hippocampal or parahippocampal activations and precuneus or posterior cingulate deactivations, regional grey matter reductions, CSF Aβ(1-42), total tau and phospho-tau(181) concentrations, and plasma Aβ(1-42) concentrations and Aβ(1-42):Aβ(1-40) ratios. Structural and functional MRI data were compared using automated brain mapping algorithms and search regions related to Alzheimer's disease. Cognitive and fluid biomarkers were compared using Mann-Whitney tests. 44 participants were included: 20 PSEN1 E280A mutation carriers and 24 non-carriers. The carrier and non-carrier groups did not differ significantly in their dementia ratings, neuropsychological test scores, or proportion of apolipoprotein E (APOE) ɛ4 carriers. Compared with non-carriers, carriers had greater right hippocampal and parahippocampal activation (p=0·001 and p<0·014, respectively, after correction for multiple comparisons), less precuneus and posterior cingulate deactivation (all p<0·010 after correction), and less grey matter in several parietal regions (all p<0·002 uncorrected and corrected p=0·009 in the right parietal search region). In the 20 participants (ten PSEN1 E280A mutation carriers and ten non-carriers) who had lumbar punctures and venepunctures, mutation carriers had higher CSF Aβ(1-42) concentrations (p=0·008) and plasma Aβ(1-42) concentrations (p=0·01) than non-carriers. Young adults at genetic risk for autosomal dominant Alzheimer's disease have functional and structural MRI findings and CSF and plasma biomarker findings consistent with Aβ(1-42) overproduction. Although the extent to which the underlying brain changes are either neurodegenerative or developmental remain to be determined, this study shows the earliest known biomarker changes in cognitively normal people at genetic risk for autosomal dominant Alzheimer's disease. Banner Alzheimer's Foundation, Nomis Foundation, Anonymous Foundation, Forget Me Not Initiative, Boston University Department of Psychology, Colciencias, National Institute on Aging, National Institute of Neurological Disorders and Stroke, and the State of Arizona.


Kinetic optimisation of the reversed phase liquid chromatographic separation of proanthocyanidins on sub-2 μm and superficially porous phases.
Phenolic compounds, and proanthocyanidins in particular, are important natural molecules which are of significant importance due to their sensory and biological activities. The analysis of proanthocyanidins in natural products is very challenging due to their complex nature. In this study, the kinetic performance of a range of recently developed C18 columns, including sub-2 μm fully porous and 2.6 μm superficially porous particle-packed columns, was evaluated for improved proanthocyanidin analysis. The kinetic plot method was employed to compare the ultimate performance limits of each column in terms of efficiency and speed for different maximum pressures and temperatures using representative proanthocyanidins comprising a range of molecular weights and functionalities as test analytes. By combining plate height data with relevant parameters such as column permeability and mobile phase viscosity, plots of practically attainable efficiencies as a function of analysis time for specific experimental configurations were obtained and performance limits for all investigated supports could accurately be compared. Both fully- and superficially porous particles provided significant speed and/or efficiency gains compared to conventional 5 μm particle packed columns. Analyte properties, particle size and packing quality as well as analysis temperature were all found to have a significant influence on the performance of the presently investigated chromatographic supports. For smaller compounds, higher optimal linear velocities and better performance in the low-efficiency range were observed, while the lower diffusion coefficients of larger procyanidins resulted in lower optimal linear velocities and better performance in the high-efficiency regime. Faster analyses become possible at higher temperatures due to decreased eluent viscosity and faster mass transfer, which was especially beneficial for larger compounds and resulted in dramatic improvement in efficiency. A possible explanation of the abnormal behaviour of oligomeric procyanidins is presented. Our findings indicate that new column formats, when used under optimal conditions, significantly improve the speed and/or efficiency of reversed phase liquid chromatographic analyses of proanthocyanidins.


What's in a name?: popular names are less common on frontiers.
Voluntary settlement on a frontier may promote values of independence. At present, however, researchers know little about the behavioral consequences of this process. In this study, we examined regional variations in baby naming. Because baby naming is an act of considerable personal and familial significance, it reflects prevalent cultural values. In support of the hypothesized link between frontier settlement and independence, we found that babies receive popular names less frequently in western regions of the United States than in its eastern regions (Study 1). The same pattern holds in Canada (Study 2), with popular names being less frequent in western provinces than in eastern provinces. Moreover, popular names are less frequently given to babies in world regions in which Europeans have settled (e.g., Australia, the United States) than in European countries (Study 3). These findings have implications for cross-generational transmission of cultural values.


Tissue Specific Expression of Cre in Rat Tyrosine Hydroxylase and Dopamine Active Transporter-Positive Neurons.
The rat is a preferred model system over the mouse for neurological studies, and cell type-specific Cre expression in the rat enables precise ablation of gene function in neurons of interest, which is especially valuable for neurodegenerative disease modeling and optogenetics. Yet, few such Cre rats are available. Here we report the characterization of two Cre rats, tyrosine hydroxylase (TH)-Cre and dopamine active transporter (DAT or Slc6a3)-Cre, by using a combination of immunohistochemistry (IHC) and mRNA fluorescence in situ hybridization (FISH) as well as a fluorescent reporter for Cre activity. We detected Cre expression in expected neurons in both Cre lines. Interestingly, we also found that in Th-Cre rats, but not DAT-Cre rats, Cre is expressed in female germ cells, allowing germline excision of the floxed allele and hence the generation of whole-body knockout rats. In summary, our data demonstrate that targeted integration of Cre cassette lead to faithful recapitulation of expression pattern of the endogenous promoter, and mRNA FISH, in addition to IHC, is an effective method for the analysis of the spatiotemporal gene expression patterns in the rat brain, alleviating the dependence on high quality antibodies that are often not available against rat proteins. The Th-Cre and the DAT-Cre rat lines express Cre in selective subsets of dopaminergic neurons and should be particularly useful for researches on Parkinson's disease.


Development of Bio-Active Patches Based on Pectin for the Treatment of Ulcers and Wounds Using 3D-Bioprinting Technology.
Biodegradable 3D-printable inks based on pectin have been developed as a system for direct and indirect wound-dressing applications, suitable for 3D printing technologies. The 3D-printable inks formed free-standing transparent films upon drying, with the latter exhibiting fast disintegration upon contact with aqueous media. The antimicrobial and wound-healing activities of the inks have been successfully enhanced by the addition of particles, comprised of chitosan and cyclodextrin inclusion complexes with propolis extract. Response Surface Methodology (RSM) was applied for the optimization of the inks (extrusion-printing pressure, shrinkage minimization over-drying, increased water uptake and minimization of the disintegration of the dry patches upon contact with aqueous media). Particles comprised of chitosan and cyclodextrin/propolis extract inclusion complexes (CCP), bearing antimicrobial properties, were optimized and integrated with the produced inks. The bioprinted patches were assessed for their cytocompatibility, antimicrobial activity and in vitro wound-healing properties. These studies were complemented with ex vivo skin adhesion measurements, a relative surface hydrophobicity and opacity measurement, mechanical properties, visualization, and spectroscopic techniques. The in vitro wound-healing studies revealed that the 3D-bioprinted patches enhanced the in vitro wound-healing process, while the incorporation of CCP further enhanced wound-healing, as well as the antimicrobial activity of the patches.


CD11c/CD18 on neutrophils recognizes a domain at the N terminus of the A alpha chain of fibrinogen.
Fibrinogen and fibrin serve as adhesive substrates for a variety of cells including platelets, endothelial cells, and leukocytes. Previously, we identified the C terminus of the gamma chain of fibrinogen as the region of the fibrinogen molecule that contains a ligand for CD11b/CD18 (complement receptor 3) on phorbol ester-stimulated polymorphonuclear leukocytes. In contrast, we report here that neutrophils stimulated with tumor necrosis factor adhere to fibrinogen-coated surfaces, but not to human serum albumin-coated surfaces, via the integrin CD11c/CD18 (p150/95). Monoclonal antibodies LeuM5 and 3.9, which are directed against the alpha subunit of CD11c/CD18, but not monoclonal antibodies OKM10 and OKM1, which are directed against the alpha subunit of CD11b/CD18, inhibit the adhesion of tumor necrosis factor-stimulated neutrophils to fibrinogen-coated surfaces. To identify the site on fibrinogen recognized by CD11c/CD18, we have examined the adhesion of tumor necrosis factor-stimulated neutrophils to surfaces coated with various fibrinogen fragments. Stimulated neutrophils adhere to surfaces coated with the N-terminal disulfide knot fragment of fibrinogen or fibrinogen fragment E. Moreover, peptides containing the sequence Gly-Pro-Arg (which corresponds to amino acids 17-19 of the N-terminal region of the A alpha chain of fibrinogen), and monoclonal antibody LeuM5, block tumor necrosis factor-stimulated neutrophil adhesion to fibrinogen and to the N-terminal disulfide knot fragment of fibrinogen. Thus, CD11c/CD18 on tumor necrosis factor-stimulated neutrophils functions as a fibrinogen receptor that recognizes the sequence Gly-Pro-Arg in the N-terminal domain of the A alpha chain of fibrinogen.


Developmental expression of Musashi-1 and Musashi-2 RNA-binding proteins during spermatogenesis: analysis of the deleterious effects of dysregulated expression.
Spermatogenesis is a complex developmental process whereby diploid spermatogenic stem cells become haploid and undergo a series of morphological changes to produce physically mature spermatozoa. Crucial to this process are a number of RNA-binding proteins, responsible for the posttranscriptional control of essential mRNAs and particularly pertinent to the two periods of inactive transcription that occur in spermatogenesis. One such group of RNA-binding proteins is the Musashi family, specifically Musashi-1 (MSI1) and Musashi-2 (MSI2), which act as key translational regulators in various stem cell populations and have been linked with the induction of tumorigenesis. In the present study, we examined the differential expression of mammalian MSI1 and MSI2 during germ cell development in the mouse testis. MSI1 was found to be predominately localized in mitotic gonocytes and spermatogonia, whereas MSI2 was detected in meiotic spermatocytes and differentiating spermatids. Extensive examination of the function of Musashi in spermatogenesis was achieved through the use of two transgenic mouse models with germ cell-specific overexpression of full-length isoforms of Msi1 or Msi2. These models demonstrated that aberrant expression of either Msi1 or Msi2 has deleterious effects on normal spermatogenesis, with Msi2 overexpression resulting in male sterility. Studies undertaken on human testicular seminoma tumors provide further insights into the relevance of MSI1 and MSI2 overexpression as diagnostic markers to human stem cell cancers. Overall this study provides further evidence for the unique functions that RNA-binding protein isoforms occupy within spermatogenesis, and introduces the potential manipulation of the Musashi family proteins to elucidate the mechanisms of posttranscriptional gene expression during germ cell development.


An acoustic inverse scattering problem for spheres with radially inhomogeneous compressibility.
An inverse acoustic scattering problem the main aim of which is to reconstruct the one-dimensional variation of the acoustical parameters of a spherical object is investigated. The problem is first formulated conventionally through a coupled system of integral equations, and then this system is reduced to one-dimensional form by using the orthogonality properties of spherical harmonics. The inverse problem is solved in an iterative fashion via classical Newton algorithm. Some numerical simulations are carried out to test the feasibility of the method as well as to see the effects of some parameters on the solution. It is shown that the method is very effective for the profiles having smooth variations provided that an appropriate initial guess is chosen. However, some of the classical disadvantages of the Newton type algorithms are also observed in numerical experiments which may limit the applicability of the method to a certain extent.


Influence of bonded enamel margins on dentin bonding stability of one-step self-etching adhesives.
To evaluate the dentin bonding durability of two one-step self-etching adhesive systems with and without the presence of surrounding enamel. Tri-S Bond (Kuraray) and G-Bond (GC), commercially available adhesive systems, were used in this study. The occlusal enamel was removed in order to form a flat, midcoronal dentin surface. The adhesives were applied to both the dentin and the surrounding enamel surfaces according to the manufacturers' instructions. Resin composite crowns (Clearfil AP-X, Kuraray) were incrementally built up with each layer being photocured for 40 s. Half the bonded teeth were sectioned perpendicular to the adhesive interface into 0.7-mm-thick slabs, and then trimmed to an hourglass shape with a 1 mm2 cross-sectional area for microtensile bond strength testing, followed by storage in water for 6 months (direct water exposure group, DE). The other half were left untrimmed and stored in water for 6 months (indirect water exposure group, IE). As control groups, additional bonded teeth were prepared for water storage for 24 h. After the designated periods of water storage, the specimens were subjected to bond strength testing at a crosshead speed of 1.0 mm/min. The microTBS data were statistically analyzed using three-way ANOVA and post-hoc tests (t-test and Tukey's HSD comparison tests) (alpha = 0.05). Indirect water exposure of the resin/dentin interface did not affect the microTBS of either adhesive after 6 months water storage (p < 0.05). However, direct exposure to water significantly reduced the microTBS of both adhesives to dentin (p < 0.05). In addition, there were no significant differences in muTBS between the two adhesive systems for each experimental storage condition (p > 0.05). The durability of resin-dentin bonds was found to be dependent on the method in which the bonded specimens were aged in water. Significantly lower bond strengths were observed for specimens that were sectioned into an hourglass shape before aging compared to those that were sectioned after water storage.


Targets and Mechanisms Associated with Protection from Severe Plasmodium falciparum Malaria in Kenyan Children.
Severe malaria (SM) is a life-threatening complication of infection with Plasmodium falciparum Epidemiological observations have long indicated that immunity against SM is acquired relatively rapidly, but prospective studies to investigate its immunological basis are logistically challenging and have rarely been undertaken. We investigated the merozoite targets and antibody-mediated mechanisms associated with protection against SM in Kenyan children aged 0 to 2 years. We designed a unique prospective matched case-control study of well-characterized SM clinical phenotypes nested within a longitudinal birth cohort of children (n= 5,949) monitored over the first 2 years of life. We quantified immunological parameters in sera collected before the SM event in cases and their individually matched controls to evaluate the prospective odds of developing SM in the first 2 years of life. Anti-AMA1 antibodies were associated with a significant reduction in the odds of developing SM (odds ratio [OR] = 0.37; 95% confidence interval [CI] = 0.15 to 0.90; P= 0.029) after adjustment for responses to all other merozoite antigens tested, while those against MSP-2, MSP-3, Plasmodium falciparum Rh2 [PfRh2], MSP-119, and the infected red blood cell surface antigens were not. The combined ability of total IgG to inhibit parasite growth and mediate the release of reactive oxygen species from neutrophils was associated with a marked reduction in the odds of developing SM (OR = 0.07; 95% CI = 0.006 to 0.82;P= 0.03). Assays of these two functional mechanisms were poorly correlated (Spearman rank correlation coefficient [rs] = 0.12;P= 0.07). Our data provide epidemiological evidence that multiple antibody-dependent mechanisms contribute to protective immunity via distinct targets whose identification could accelerate the development of vaccines to protect against SM.


Intracellular calcium dynamics dependent on defined microtopographical features of titanium.
Detailed insights into the complex cellular behavior at the biomaterial interface are crucial for the improvement of implant surfaces with respect to their acceptance and integration. The cells perceive microtopographical features and, in consequence, rearrange their adhesion structures like the actin cytoskeleton and adaptor proteins. But little is known about whether these altered cellular phenotypes have consequences for intracellular calcium signaling and its dynamics. To elucidate if an artificial, geometrical microtopography influences calcium ion (Ca(2+)) mobilization in osteoblasts, human MG-63 cells were stained with the calcium dye Fluo 3-acetoxymethyl ester and set on defined silicon-titanium (Ti) arrays with regular pillar structures (P5, 5 × 5 × 5 μm) and compared with planar Ti. To induce an immediate calcium signal, cells were stimulated with adenosine 5'-triphosphate (ATP). Interestingly, osteoblasts on micropillars expressing a shortened actin cytoskeleton were hampered in their calcium mobilization potential in signal height as well duration. Even the basal level of the intracellular Ca(2+) concentration was reduced, which was accompanied by a disturbed fibronectin synthesis. The expression of the voltage-sensitive calcium channels Cav1.2, Cav1.3 (L-type) and Cav3.1, Cav3.2, Cav3.3 (T-type) as well as the signaling proteins phospho-AKT and phospho-GSK3α/β remained unaffected on pillars. The topography-dependent calcium dynamics observed here provide new insights into how topographical cues alter cell functions - via the intracellular Ca(2+) signaling.