A study on the protective effect of molecular hydrogen on osteoradionecrosis of the jaw in rats.
The aim of this study was to investigate the protective effect of hydrogen in a rat model of osteoradionecrosis of the jaw (ORNJ). The rats and bone marrow-derived mesenchymal stem cells (BMSCs) were pre-treated with hydrogen before receiving irradiation (7Gy per fraction, five fractions in total once a day for rats, 4Gy for BMSCs). Reactive oxygen species (ROS) and cell differentiation were measured in the BMSCs. Also, the radioprotective effect of hydrogen for ORNJ in Sprague-Dawley rats was examined by gross clinical manifestations, micro-computed tomography, and histology. Hydrogen significantly reduced the production of ROS in BMSCs after irradiation. The cell viability was significantly decreased after irradiation (P= 0.001), but pre-treatment with hydrogen before irradiation increased the cell viability (P= 0.025). Hydrogen considerably increased the cellular differentiation potential of the irradiated cells. Comparing with the rats underwent irradiaton only, those rats treated by hydrogen-rich saline significantly appeared improved occlusion, salivation, alopecia, oral ulcer, and less bone necrosis. Myofibroblasts accumulated overwhelmingly in the fibrosis medulla and around the sequestrum after irradiation, and this was decreased in the group pre-treated with hydrogen. Hydrogen may represent a strategy for the prevention and treatment of ORNJ. Its high efficacy and low toxicity suggest possible therapeutic application.


Inhibition of intraocular neovascularization caused by retinal ischemia in pigs by PKCbeta inhibition with LY333531.
The authors tested the antiangiogenic properties of an orally administered protein kinase-Cbeta inhibitor, LY333531, in a pig model of preretinal neovascularization caused by retinal branch vein occlusion to determine the effectiveness of this therapy in preventing intraocular neovascularization from an ischemic stimulus. In 20 eyes of 10 pigs, branch retinal vein occlusions were created in a standardized manner using photodynamic thrombosis with rose bengal dye and thermal burns from an argon laser with green light. Five animals received 1 mg/kg LY333531 daily in two oral doses, and five animals were untreated. The eyes were followed clinically for 12 weeks with ophthalmoscopy, fundus photography, and fluorescein angiography. A standardized grading system permitted masked assessment of disc proliferations using stereo fundus photographs. After the pigs were killed, all neovascularization was confirmed histopathologically in a masked fashion and a final grade was assigned to each eye. The Mann-Whitney test was used for statistical analysis of the median values of the unpaired data between the two eyes of each animal (data were rounded up). Significant inhibition of neovascularization was observed in eyes from animals treated with the study drug (P = 0.03). Although some of the treated eyes demonstrated no clinically evident new vessels, histopathologic and photographic analysis demonstrated fine new vessels on the optic disc in all eyes (mean grade 1.9). All the untreated eyes developed clinically evident neovascularization (mean grade 3.1). The oral drug was well tolerated, and no side effects were documented. A specific protein kinase-Cbeta inhibitor, LY333531, effectively inhibited preretinal and optic nerve head neovascularization in the pig model of branch retinal vein occlusion. This was consistent with the known pathways of signal transduction by growth factors in activated cells and suggested that inhibition of this key regulatory isozyme is effective in the treatment of ischemia-mediated neovascular diseases.


Preventive or late administration of anti-NGF therapy attenuates tumor-induced nerve sprouting, neuroma formation, and cancer pain.
Early, preemptive blockade of nerve growth factor (NGF)/tropomyosin receptor kinase A (TrkA) attenuates tumor-induced nerve sprouting and bone cancer pain. A critical unanswered question is whether late blockade of NGF/TrkA can attenuate cancer pain once NGF-induced nerve sprouting and neuroma formation has occurred. By means of a mouse model of prostate cancer-induced bone pain, anti-NGF was either administered preemptively at day 14 after tumor injection when nerve sprouting had yet to occur, or late at day 35, when extensive nerve sprouting had occurred. Animals were humanely killed at day 70 when, in vehicle-treated animals, significant nerve sprouting and neuroma formation was present in the tumor-bearing bone. Although preemptive and sustained administration (days 14-70) of anti-NGF more rapidly attenuated bone cancer nociceptive behaviors than late and sustained administration (days 35-70), by day 70 after tumor injection, both preemptive and late administration of anti-NGF significantly reduced nociceptive behaviors, sensory and sympathetic nerve sprouting, and neuroma formation. In this model, as in most cancers, the individual cancer cell colonies have a limited half-life because they are constantly proliferating, metastasizing, and undergoing necrosis as the parent cancer cell colony outgrows its blood supply. Similarly, the sensory and sympathetic nerve fibers that innervate the tumor undergo sprouting at the viable/leading edge of the parent tumor, degenerate as the parent cancer cell colony becomes necrotic, and resprout in the viable, newly formed daughter cell colonies. These results suggest that preemptive or late-stage blockade of NGF/TrkA can attenuate nerve sprouting and cancer pain.


The risk of major cardiac malformations associated with paroxetine use during the first trimester of pregnancy: a systematic review and meta-analysis.
The aim of this study was to perform an up-to-date meta-analysis on the risk of cardiac malformations associated with gestational exposure to paroxetine, taking into account indication, study design and reference category. A systematic review of studies published between 1966 and November 2015 was conducted using embase and MEDLINE. Studies reporting major malformations with first trimester exposure to paroxetine were included. Potentially relevant articles were assessed and relevant data extracted to calculate risk estimates. Outcomes included any major malformations and major cardiac malformations. Pooled odds ratios and 95% confidence intervals were calculated using random-effects models. Twenty-three studies were included. Compared with non-exposure to paroxetine, first trimester use of paroxetine was associated with an increased risk of any major congenital malformations combined (pooled OR 1.23, 95% CI 1.10, 1.38; n = 15 studies), major cardiac malformations (pooled OR 1.28, 95% CI 1.11, 1.47; n = 18 studies), specifically bulbus cordis anomalies and anomalies of cardiac septal closure (pooled OR 1.42, 95% CI 1.07, 1.89; n = 8 studies), atrial septal defects (pooled OR 2.38, 95% CI 1.14, 4.97; n = 4 studies) and right ventricular outflow track defect (pooled OR 2.29, 95% CI 1.06, 4.93; n = 4 studies). Although the estimates varied depending on the comparator group, study design and malformation detection period, a trend towards increased risk was observed. Paroxetine use during the first trimester of pregnancy is associated with an increased risk of any major congenital malformations and cardiac malformations. The increase in risk is not dependent on the study method or population.


Vegetable consumption and blood lead concentrations.
Women resident in an area heavily contaminated by spoil from old lead mining have blood lead concentrations that are about 50% higher (p less than 0.001) than those of women living in a "control" area some distance away. Blood lead concentrations were related to the consumption of home grown produce. Those with the highest consumptions had blood lead concentrations that were 28% higher (p less than 0.001) than those of women who consumed no locally grown vegetables. Nevertheless, in the total population in the area this effect seemed likely to account for only about 5% of the population mean blood lead concentration. The data suggested that an increase in soil lead of 1000 micrograms/g is associated with an increase in blood lead of about 0.20 microM/1 (4.2 micrograms/dl).


Transverse light guides in microstructured optical fibers.
A novel class of microstructured optical fiber coupler is introduced that operates by resonant, rather than proximity, energy transfer by means of transverse light guides built into a fiber cross section. Such a design permits significant spatial separation between interacting fibers, which, in turn, eliminates intercore cross talk owing to proximity coupling. A controllable energy transfer between the cores is then achieved by localized and highly directional transmission through a transverse light guide. The main advantage of this coupling scheme is its inherent scalability, as one can integrate additional fiber cores into the existing fiber cross section simply by placing the cores far enough from the existing optical circuitry to prevent proximity cross talk and then making the necessary intercore connections with transverse light wires, in direct analogy with on-chip electronics integration.


Depletion of the Receptor-Interacting Protein Kinase 3 (RIP3) Decreases Photoreceptor Cell Death During the Early Stages of Ocular Murine Cytomegalovirus Infection.
The purpose of this study was to determine if the receptor-interacting protein kinase 3 (RIP3) plays a significant role in innate immune responses and death of bystander retinal neurons during murine cytomegalovirus (MCMV) retinal infection, by comparing the innate immune response and cell death in RIP3-depleted mice (Rip3-/-) and Rip3+/+ control mice. Rip3-/- and Rip3+/+ mice were immunosuppressed (IS) and inoculated with MCMV via the supraciliary route. Virus-injected and mock-injected control eyes were removed at days 4, 7, and 10 post infection (p.i.) and markers of innate immunity and cell death were analyzed. Compared to Rip3+/+ mice, significantly more MCMV was recovered and more MCMV-infected RPE cells were observed in injected eyes of Rip3-/- mice at days 4 and 7 p.i. In contrast, fewer TUNEL-stained photoreceptors were observed in Rip3-/- eyes than in Rip3+/+ eyes at these times. Electron microscopy showed that significantly more apoptotic photoreceptor cells were present in Rip3+/+ mice than in Rip3-/- mice. Immunohistochemistry showed that the majority of TUNEL-stained photoreceptors died via mitochondrial flavoprotein apoptosis-inducing factor (AIF)-mediated, caspase 3-independent apoptosis. The majority of RIP3-expressing cells in infected eyes were RPE cells, microglia/macrophages, and glia, whereas retinal neurons contained much lower amounts of RIP3. Western blots showed significantly higher levels of activated nuclear factor-κB and caspase 1 were present in Rip3+/+ eyes compared to Rip3-/- eyes. Our results suggest that RIP3 enhances innate immune responses against ocular MCMV infection via activation of the inflammasome and nuclear factor-κB, which also leads to inflammation and death of bystander cells by multiple pathways including apoptosis and necroptosis.


Non-clinical interventions for reducing unnecessary caesarean section.
Caesarean section rates are increasing globally. The factors contributing to this increase are complex, and identifying interventions to address them is challenging. Non-clinical interventions are applied independently of a clinical encounter between a health provider and a patient. Such interventions may target women, health professionals or organisations. They address the determinants of caesarean births and could have a role in reducing unnecessary caesarean sections. This review was first published in 2011. This review update will inform a new WHO guideline, and the scope of the update was informed by WHO's Guideline Development Group for this guideline. To evaluate the effectiveness and safety of non-clinical interventions intended to reduce unnecessary caesarean section. We searched CENTRAL, MEDLINE, Embase, CINAHL and two trials registers in March 2018. We also searched websites of relevant organisations and reference lists of related reviews. Randomised trials, non-randomised trials, controlled before-after studies, interrupted time series studies and repeated measures studies were eligible for inclusion. The primary outcome measures were: caesarean section, spontaneous vaginal birth and instrumental birth. We followed standard methodological procedures recommended by Cochrane. We narratively described results of individual studies (drawing summarised evidence from single studies assessing distinct interventions). We included 29 studies in this review (19 randomised trials, 1 controlled before-after study and 9 interrupted time series studies). Most of the studies (20 studies) were conducted in high-income countries and none took place in low-income countries. The studies enrolled a mixed population of pregnant women, including nulliparous women, multiparous women, women with a fear of childbirth, women with high levels of anxiety and women having undergone a previous caesarean section.Overall, we found low-, moderate- or high-certainty evidence that the following interventions have a beneficial effect on at least one primary outcome measure and no moderate- or high-certainty evidence of adverse effects.Interventions targeted at women or familiesChildbirth training workshops for mothers alone may reduce caesarean section (risk ratio (RR) 0.55, 95% confidence interval (CI) 0.33 to 0.89) and may increase spontaneous vaginal birth (RR 2.25, 95% CI 1.16 to 4.36). Childbirth training workshops for couples may reduce caesarean section (RR 0.59, 95% CI 0.37 to 0.94) and may increase spontaneous vaginal birth (RR 2.13, 95% CI 1.09 to 4.16). We judged this one study with 60 participants to have low-certainty evidence for the outcomes above.Nurse-led applied relaxation training programmes (RR 0.22, 95% CI 0.11 to 0.43; 104 participants, low-certainty evidence) and psychosocial couple-based prevention programmes (RR 0.53, 95% CI 0.32 to 0.90; 147 participants, low-certainty evidence) may reduce caesarean section. Psychoeducation may increase spontaneous vaginal birth (RR 1.33, 95% CI 1.11 to 1.61; 371 participants, low-certainty evidence). The control group received routine maternity care in all studies.There were insufficient data on the effect of the four interventions on maternal and neonatal mortality or morbidity.Interventions targeted at healthcare professionalsImplementation of clinical practice guidelines combined with mandatory second opinion for caesarean section indication slightly reduces the risk of overall caesarean section (mean difference in rate change -1.9%, 95% CI -3.8 to -0.1; 149,223 participants). Implementation of clinical practice guidelines combined with audit and feedback also slightly reduces the risk of caesarean section (risk difference (RD) -1.8%, 95% CI -3.8 to -0.2; 105,351 participants). Physician education by local opinion leader (obstetrician-gynaecologist) reduced the risk of elective caesarean section to 53.7% from 66.8% (opinion leader education: 53.7%, 95% CI 46.5 to 61.0%; control: 66.8%, 95% CI 61.7 to 72.0%; 2496 participants). Healthcare professionals in the control groups received routine care in the studies. There was little or no difference in maternal and neonatal mortality or morbidity between study groups. We judged the certainty of evidence to be high.Interventions targeted at healthcare organisations or facilitiesCollaborative midwifery-labourist care (in which the obstetrician provides in-house labour and delivery coverage, 24 hours a day, without competing clinical duties), versus a private practice model of care, may reduce the primary caesarean section rate. In one interrupted time series study, the caesarean section rate decreased by 7% in the year after the intervention, and by 1.7% per year thereafter (1722 participants); the vaginal birth rate after caesarean section increased from 13.3% before to 22.4% after the intervention (684 participants). Maternal and neonatal mortality were not reported. We judged the certainty of evidence to be low.We studied the following interventions, and they either made little or no difference to caesarean section rates or had uncertain effects.Moderate-certainty evidence suggests little or no difference in caesarean section rates between usual care and: antenatal education programmes for physiologic childbirth; antenatal education on natural childbirth preparation with training in breathing and relaxation techniques; computer-based decision aids; individualised prenatal education and support programmes (versus written information in pamphlet).Low-certainty evidence suggests little or no difference in caesarean section rates between usual care and: psychoeducation; pelvic floor muscle training exercises with telephone follow-up (versus pelvic floor muscle training without telephone follow-up); intensive group therapy (cognitive behavioural therapy and childbirth psychotherapy); education of public health nurses on childbirth classes; role play (versus standard education using lectures); interactive decision aids (versus educational brochures); labourist model of obstetric care (versus traditional model of obstetric care).We are very uncertain as to the effect of other interventions identified on caesarean section rates as the certainty of the evidence is very low. We evaluated a wide range of non-clinical interventions to reduce unnecessary caesarean section, mostly in high-income settings. Few interventions with moderate- or high-certainty evidence, mainly targeting healthcare professionals (implementation of guidelines combined with mandatory second opinion, implementation of guidelines combined with audit and feedback, physician education by local opinion leader) have been shown to safely reduce caesarean section rates. There are uncertainties in existing evidence related to very-low or low-certainty evidence, applicability of interventions and lack of studies, particularly around interventions targeted at women or families and healthcare organisations or facilities.


Role of pyruvate dehydrogenase kinase 4 in regulation of blood glucose levels.
In the well-fed state a relatively high activity of the pyruvate dehydrogenase complex (PDC) reduces blood glucose levels by directing the carbon of pyruvate into the citric acid cycle. In the fasted state a relatively low activity of the PDC helps maintain blood glucose levels by conserving pyruvate and other three carbon compounds for gluconeogenesis. The relative activities of the pyruvate dehydrogenase kinases (PDKs) and the opposing pyruvate dehydrogenase phosphatases determine the activity of PDC in the fed and fasted states. Up regulation of PDK4 is largely responsible for inactivation of PDC in the fasted state. PDK4 knockout mice have lower fasting blood glucose levels than wild type mice, proving that up regulation of PDK4 is important for normal glucose homeostasis. In type 2 diabetes, up regulation of PDK4 also inactivates PDC, which promotes gluconeogenesis and thereby contributes to the hyperglycemia characteristic of this disease. When fed a high fat diet, wild type mice develop fasting hyperglycemia but PDK4 knockout mice remain euglycemic, proving that up regulation of PDK4 contributes to hyperglycemia in diabetes. These finding suggest PDK4 inhibitors might prove useful in the treatment of type 2 diabetes.


Deep Sequencing of B Cell Receptor Repertoires From COVID-19 Patients Reveals Strong Convergent Immune Signatures.
Deep sequencing of B cell receptor (BCR) heavy chains from a cohort of 31 COVID-19 patients from the UK reveals a stereotypical naive immune response to SARS-CoV-2 which is consistent across patients. Clonal expansion of the B cell population is also observed and may be the result of memory bystander effects. There was a strong convergent sequence signature across patients, and we identified 1,254 clonotypes convergent between at least four of the COVID-19 patients, but not present in healthy controls or individuals following seasonal influenza vaccination. A subset of the convergent clonotypes were homologous to known SARS and SARS-CoV-2 spike protein neutralizing antibodies. Convergence was also demonstrated across wide geographies by comparison of data sets between patients from UK, USA, and China, further validating the disease association and consistency of the stereotypical immune response even at the sequence level. These convergent clonotypes provide a resource to identify potential therapeutic and prophylactic antibodies and demonstrate the potential of BCR profiling as a tool to help understand patient responses.


[Muscle-invasive bladder cancer].
Muscle invasive bladder cancers represent 20 to 30% of bladder tumors and are metastatic at diagnosis in 10% of cases. The diagnosis is based on histological analysis of transuretral resection pieces. An invasion of the detrusor (stage pT2) signs the infiltrating nature of the tumor. The local and general staging is based on thoraco-abdominopelvic CT. The standard treatment for localized stages is cystoprostatectomy in men and anterior pelvic exenteration in women. For urinary diversion, the two choices are cutaneous non continent ileal conduit (Bricker) and ileal neobladder. Neoadjuvant chemotherapy can be discussed for locally advanced stages. The concomitant chemoradiotherapy is an alternative to surgical treatment in elderly or frail patients. The management of metastatic bladder cancer is based on systemic chemotherapy.


The 'non-falciparum' malarias: the roles of epidemiology, parasite biology, clinical syndromes, complications and diagnostic rigour in guiding therapeutic strategies.
Plasmodium vivax, P. ovale, P. malariae and P. falciparum routinely infect humans. The infections caused by these parasites are loosely referred to as vivax (or benign tertian), ovale, malariae (or quartan) and falciparum (or malignant tertian) malaria, respectively. Recently, P. knowlesi, a parasite of macaque monkeys in South-east Asia, has been identified as the cause of uncomplicated and severe human malaria in Malaysian Borneo. The prescription of appropriate therapies for reliably diagnosed malaria requires a grasp of the epidemiology of the 'non-falciparum' malarias, the biology of the parasites involved, the chemotherapeutic strategies that are available and the problems of emerging drug resistance and changing clinical syndromes. This review is intended to increase clinicians' understanding of how these factors relate to the selection of the antimalarial drugs to be given to a case of 'non-falciparum' malaria, with the aims of improving outcomes and preventing relapses and recrudescences.


Influence of Melt-Pool Stability in 3D Printing of NdFeB Magnets on Density and Magnetic Properties.
The current work presents the results of an investigation focused on the influence of process parameters on the melt-track stability and its consequence to the sample density printed out of NdFeB powder. Commercially available powder of Nd7.5Pr0.7Fe75.4Co2.5B8.8Zr2.6Ti2.5 alloy was investigated at the angle of application in selective laser melting of permanent magnets. Using single track printing the stability of the melt pool was investigated under changing process parameters. The influence of changing laser power, scanning speed, and powder layer thickness on density, porosity structure, microstructure, phase composition, and magnetic properties were investigated. The results showed that energy density coupled with powder layer thickness plays a crucial role in melt-track stability. It was possible to manufacture magnets of both high relative density and high magnetic properties. Magnetization tests showed a significant correlation between the shape of the demagnetization curve and the layer height. While small layer heights are beneficial for sufficient magnetic properties, the remaining main parameters tend to affect the magnetic properties less. A quasi-linear correlation between the layer height and the magnetic properties remanence (Jr), coercivity (HcJ) and maximum energy product ((BH)max) was found.


AZIN1 RNA editing confers cancer stemness and enhances oncogenic potential in colorectal cancer.
Adenosine-to-inosine (A-to-I) RNA editing, a process mediated by adenosine deaminases that act on the RNA (ADAR) gene family, is a recently discovered epigenetic modification dysregulated in human cancers. However, the clinical significance and the functional role of RNA editing in colorectal cancer (CRC) remain unclear. We have systematically and comprehensively investigated the significance of the expression status of ADAR1 and of the RNA editing levels of antizyme inhibitor 1 (AZIN1), one of the most frequently edited genes in cancers, in 392 colorectal tissues from multiple independent CRC patient cohorts. Both ADAR1 expression and AZIN1 RNA editing levels were significantly elevated in CRC tissues when compared with corresponding normal mucosa. High levels of AZIN1 RNA editing emerged as a prognostic factor for overall survival and disease-free survival and were an independent risk factor for lymph node and distant metastasis. Furthermore, elevated AZIN1 editing identified high-risk stage II CRC patients. Mechanistically, edited AZIN1 enhances stemness and appears to drive the metastatic processes. We have demonstrated that edited AZIN1 functions as an oncogene and a potential therapeutic target in CRC. Moreover, AZIN1 RNA editing status could be used as a clinically relevant prognostic indicator in CRC patients.


Road traffic noise, blood pressure and heart rate: Pooled analyses of harmonized data from 88,336 participants.
Exposure to road traffic noise may increase blood pressure and heart rate. It is unclear to what extent exposure to air pollution may influence this relationship. We investigated associations between noise, blood pressure and heart rate, with harmonized data from three European cohorts, while taking into account exposure to air pollution. Road traffic noise exposure was assessed using a European noise model based on the Common Noise Assessment Methods in Europe framework (CNOSSOS-EU). Exposure to air pollution was estimated using a European-wide land use regression model. Blood pressure and heart rate were obtained by trained clinical professionals. Pooled cross-sectional analyses of harmonized data were conducted at the individual level and with random-effects meta-analyses. We analyzed data from 88,336 participants, across the three participating cohorts (mean age 47.0 (±13.9) years). Each 10dB(A) increase in noise was associated with a 0.93 (95% CI 0.76;1.11) bpm increase in heart rate, but with a decrease in blood pressure of 0.01 (95% CI -0.24;0.23) mmHg for systolic and 0.38 (95% CI -0.53; -0.24) mmHg for diastolic blood pressure. Adjustments for PM10 or NO2 attenuated the associations, but remained significant for DBP and HR. Results for BP differed by cohort, with negative associations with noise in LifeLines, no significant associations in EPIC-Oxford, and positive associations with noise >60dB(A) in HUNT3. Our study suggests that road traffic noise may be related to increased heart rate. No consistent evidence for a relation between noise and blood pressure was found.


Tuning thermal transport in Si nanowires by isotope engineering.
We study thermal transport in isotopically disordered Si nanowires, discussing the feasibility of phonon engineering for thermoelectric applications within these systems. To this purpose, we carry out atomistic molecular dynamics and nonequilibrium Green's function calculations to characterize the dependence of the thermal conductance as a function of the isotope concentration, isotope radial distribution and temperature. We show that a reduction of the conductivity of up to 20% can be achieved with suitable isotope blends at room temperature and approximately 50% at low temperature. Interestingly, precise control of the isotope composition or radial distribution is not needed. An isotope disordered nanowire roughly behaves like a low-pass filter, as isotope impurities are transparent for long wave-length acoustic phonons, while only mid- and high-frequency optical phonons undergo significant scattering.


An omics perspective to the molecular mechanisms of anticancer metallo-drugs in the computational microscope era.
Metallo-drugs have attracted enormous interest for cancer treatment. The achievements of this drug-type are summarized by the success story of cisplatin. That being said, there have been many drawbacks with its clinical use, which prompted decades worth of research efforts to move towards safer and more effective agents, either containing platinum or different metals. Areas covered: In this review, the authors provide an atomistic picture of the molecular mechanisms involving selected metallo-drugs from structural and molecular simulation studies. They also provide an omics perspective, pointing out many unsettled aspects of the most relevant families of metallo-drugs at an epigenetic level. Expert opinion: Molecular simulations are able to provide detailed information at atomistic and temporal (ps) resolutions that are rarely accessible to experiments. The increasing accuracy of computational methods and the growing performance of computational platforms, allow us to mirror wet lab experiments in silico. Consequently, the molecular mechanisms of drugs action/failure can be directly viewed on a computer screen, like a 'computational microscope', allowing us to harness this knowledge for the design of the next-generation of metallo-drugs.


Copy number gains of FGFR1 and 3q chromosome in squamous cell carcinoma of the lung.
Squamous cell carcinoma of the lung (SQCCL) remains a leading cause of cancer-related death. Unlike non-smoker adenocarcinoma of the lung, where highly efficient tyrosine kinase inhibitors are available for treating mutant EGFR or ALK-rearranged, no targetable biomarkers are available for SQCCL. The frequent and focal amplification of FGFR1 has generated great expectations in offering new therapeutical options in case of 16-22% of SQCCL patients. Broad 3q chromosome amplification is widely recognized as the most common chromosomal aberration found in SQCCL, where PIK3CA, SOX2, ACK1, PRKCI, TP63, PLD1, ECT2, and others genes are located. Although SOX2 has been postulated as a key regulator of basal stem cells transformation and tumor progression, it seems to confer a good prognosis in SQCCL. It is known that each patient might carry a different length of 3q chromosome amplicon. Thus, we suggest that the number and the biological importance of the genes spanned along each patient's 3q amplicon might help to explain inter-individual outcome variations of the disease and its potential predictive value, especially when relevant oncogenes such as those mentioned above are implicated. Currently, there is no clinical predictive data available from clinical trials. In this review, we have focused on the potential role of FGFR1 in SQCCL prognosis. Additionally, we have explored recently available public data on the comprehensive genomic characterization of SQCCL, in relation to the protein-coding genes that have a strong gene copy number - mRNA correlation in 3q chromosome, that were previously described as potential driver oncogenes or its modifiers in SQCCL.


Sustained systemic response paralleled with ovarian metastasis progression by sunitinib in metastatic renal cell carcinoma: Is this an anti-angiogenic potentiation of cancer?
Metastatic renal cell cancer is associated with poor prognosis and survival and is resistant to conventional chemotherapy. Therapeutic targeting of molecular pathways for tumor angiogenesis and other specific activation mechanisms offers improved tumor response and prolonged survival. A 48-year-old, female patient presented with large right renal mass with features suggesting of renal cell cancer without metastasis on contrast enhanced computed tomography (CT). Right radical nephrectomy was done. After 9 months of surgery, she got metastasis in lung, liver and ovary. The patient received sunitinib via an expanded access program. After eight 6-week cycles of sunitinib, a reassessment CT scan confirmed an excellent partial response with the almost complete disappearance (90%) of liver and lung metastasis but the adnexal mass had increased in size (>10 times) and the possibility was thought of second malignancy. Excision of the mass performed. Histopathology of the mass depicted metastatic renal cell cancer. There is possibility of a 'site-specific anti-angiogenic potentiation mechanism' of malignancy in relation to sunitinib based upon the preclinical studies, in reference to the index case. Regression of one site with concurrent progression is possible. The exact mechanism of site-specific response, especially organ specific progression by vascular endothelial growth factor inhibitors in metastatic renal cell cancer warrants further study.


Mental impairment in cytogenetically positive fragile X females.
Prenatal diagnosis is now available to fragile X (fra[X]) syndrome families and has proven reliable when testing male fetuses. It has been reported that approximately one-third of heterozygous fra(X) females demonstrate mental impairment. Based on this, families usually continue pregnancies involving a female fetus. The purpose of this study is to investigate whether a 35% risk for mental impairment is appropriate when counseling heterozygous women carrying fra(X)-positive female fetuses. Forty-three cytogenetically positive (greater than or equal to 2%) daughters of known fra(X) carrier women were ascertained postnatally in an unbiased fashion. Their mother's carrier status was determined on the basis of at least one son with Martin-Bell syndrome. In addition to peripheral blood cytogenetic studies, all daughters had cognitive testing to determine full-scale IQ. In this study, 55.8% (24/43) were mentally impaired (IQ less than 85) compared with the expected 35%. Of these, 42% were mentally retarded (IQ less than 70). Although we do not know the correlation between percent fragility by peripheral blood compared with the percent fragility by amniocentesis or CVS, we assume that they are relatively comparable such that a female who is positive with greater than or equal to 2% fragility would probably be positive by all methods. This report suggests that the penetrance of mental impairment in females with a percent fragility of greater than or equal to 2% may be as high as 55%. Further studies are necessary to clarify this issue so that accurate information can be given in genetic counseling.


Enhancement of MHC class I-restricted peptide-specific T cell induction by a DNA prime/MVA boost vaccination regime.
Human immunodeficiency virus (HIV) vaccine candidates were previously constructed as a string of cytotoxic T lymphocyte (CTL) epitopes delivered and expressed using DNA and modified virus Ankara (MVA; an attenuated vaccinia virus) vectors. These vaccines were shown to induce interferon (IFN)-gamma-producing and cytolytic CD8+ T cells after a single vaccine administration. In the course of this work, immunization protocols were sought which would improve the levels of induced HIV-specific T cells. It was found that previous immunological exposure to MVA reduced the efficiency of subsequent priming and boosting using the same vaccine vehicle. However, a combined regime whereby the animals were first primed with the DNA vaccine and then boosted with MVA was the most potent protocol for the induction of both interferon-gamma-producing and cytolytic T cells against two CTL epitopes simultaneously. The general applicability of this novel vaccination method for induction of major histocompatibility complex class I-restricted T cells is discussed.


Crystals on the move: mechanical effects in dynamic solids.
When exposed to external stimuli such as heat or light, certain single crystals can acquire momentum and undergo motion. On a molecular scale, the motility of such dynamic single crystals is triggered by a phase transition or chemical reaction without gaseous products, and macroscopically manifests as either slow (reversible or irreversible) deformation, or as rapid, almost instantaneous propulsion of the crystals that is oftentimes accompanied by disintegration. While the elastic energy of the slow reconfiguration processes such as bending, twisting and coiling can be utilized for actuation of other objects, the fast disintegrative processes could be exploited to initiate pressure-sensitive applications. This short review intends to summarize recent developments in the growing research on dynamic crystals, especially aspects of the mechanism of rapid motion of thermosalient and photosalient (leaping) crystals. The collective evidence indicates that these solids are organic-based analogues of the inorganic martensitic materials. While qualitative explanation of the molecular processes that lead to the related dynamic phenomena can be provided, quantification of their kinematics, estimation of the useful work that can be extracted, and prediction of their occurrence are yet to be established. Harnessing the potential of these materials to rapidly and efficiently perform the fundamentally important process of transduction of heat or light into kinetic energy appears as a prospective basis for their application in motion gears and devices.


Use of cardioprotective medications in kidney transplant recipients.
Death with function causes half of late kidney transplant failures, and cardiovascular disease (CVD) is the most common cause of death in these patients. We examined the use of potentially cardioprotective medications in a prospective observational study at seven transplant centers in the United States and Canada. Among 935 patients, 87% received antihypertensive medications at both 1 and 6 months after transplantation. Similar antihypertensive regimens were used for patients with and without diabetes and CVD, but with wide variability among centers. In contrast, while 44% of patients were on angiotensin converting enzyme inhibitors (ACEI) or angiotensin receptor blockers (ARB) at the time of transplantation, the proportion taking these agents dropped to 12% at month 1, then increased to 24% at 6 months. Fewer than 30% with CVD or diabetes received ACEI/ARB therapy 6 months posttransplant. Aspirin use was uncommon (<40% of patients). Even among those with diabetes and/or CVD, fewer than 60% received aspirin and only half received a statin at 1 and 6 months. This study demonstrates marked variability in the use of cardioprotective medications in kidney transplant recipients, a finding that may reflect, among several possible explanations, clinical uncertainty due the lack of randomized trials for these medications in this population.


Quantitative comparison of constitutive promoters in human ES cells.
Constitutive promoters that ensure sustained and high level gene expression are basic research tools that have a wide range of applications, including studies of human embryology and drug discovery in human embryonic stem cells (hESCs). Numerous cellular/viral promoters that ensure sustained gene expression in various cell types have been identified but systematic comparison of their activities in hESCs is still lacking. We have quantitatively compared promoter activities of five commonly used constitutive promoters, including the human β-actin promoter (ACTB), cytomegalovirus (CMV), elongation factor-1α, (EF1α), phosphoglycerate kinase (PGK) and ubiquitinC (UbC) in hESCs. Lentiviral gene transfer was used to ensure stable integration of promoter-eGFP constructs into the hESCs genome. Promoter activities were quantitatively compared in long term culture of undifferentiated hESCs and in their differentiated progenies. The ACTB, EF1α and PGK promoters showed stable activities during long term culture of undifferentiated hESCs. The ACTB promoter was superior by maintaining expression in 75-80% of the cells after 50 days in culture. During embryoid body (EB) differentiation, promoter activities of all five promoters decreased. Although the EF1α promoter was downregulated in approximately 50% of the cells, it was the most stable promoter during differentiation. Gene expression analysis of differentiated eGFP+ and eGFP- cells indicate that promoter activities might be restricted to specific cell lineages, suggesting the need to carefully select optimal promoters for constitutive gene expression in differentiated hESCs.


Implementation of a high-alert medication program.
Greater than 500,000 doses of high-alert medications are administered throughout the Kaiser Permanente Northern California (KPNC) Program on an annual basis. High-alert medications (HAM) carry a higher risk of harm than other medications and errors in the administration of HAM can have catastrophic clinical outcomes. The purpose of this project is to ensure safe medication practices and to eliminate medication errors that cause harm to our patients.The Program: KPNC leadership, physicians, nurses, pharmacists, quality leaders, and labor unions worked with regional and local medication safety committees to: 1) standardize high-alert medication-handling practices; 2) enhance education programs related to medication practices, embedding these into annual core competencies of all staff who handle high-alert medications; 3) develop monitoring functions at both the regional and local levels to ensure sustainability and ongoing systems improvements. Begun in December 2005, this program covers the delivery of high-alert medications across the continuum of care and affects all patients receiving HAM. The initial phase of the monitoring process was put in place to measure compliance with implementation. Over the first few months of the program the 90% minimal threshold was surpassed with regional overall compliance of 95%. Following this initial process, the Regional Medication Safety Committee developed monitoring tools. Department managers carry out these concurrent observational audits at the medical centers with oversight by the Assistant Administrators for Quality and Service. These audits are designed to measure whether or not all medications on the HAM list are handled specifically to policy requirements, eg, independent double-checks, HAM stickers, etc. Audit specifications are provided for each audit tool. Medical Center audit results from the third quarter of 2006 through the third quarter of 2007 have shown a regional aggregate of 97.7% compliance. As the high percentages of compliance have held constant over time, more actionable metrics are being put in place for 2008.To determine whether or not the program is reducing HAM errors, data from the regional Quality and Risk database (MIDAS) related to all high-alert medication errors was reviewed. Two interventions were of note: in July of 2005, there was a renewed effort to educate leaders, managers, physicians, and staff on responsible reporting in a "just culture" and the introduction of the new Responsible Reporting Form. An increase in reporting was noted at this time. In December 2005, the HAM program was introduced. There is a statistically significant drop in errors reported for 23 consecutive months following this program. These findings were similar for all phases of the delivery process. A powerful indicator of improvement is the average days between major injury and death. As of November 30, 2007, it has been 232 days since the last significant negative event was reported due to a HAM. This program has been implemented in all of the KPNC Medical Centers and is in the process of being implemented in all KP regions. This spread has been endorsed by the Medical Directors Quality Committee and by the KP Boards of Directors. The Interregional Medication Safety Committee is overseeing the spread process. A toolkit containing all of the required tools plus additional materials and information has been developed and made available throughout KP. The program is the recipient of the 2007 Lawrence Patient Safety Award.


Insect-inspired vision for autonomous vehicles.
Flying insects are being studied these days as if they were agile micro air vehicles fitted with smart sensors, requiring very few brain resources. The findings obtained on these natural fliers have proved to be extremely valuable when it comes to designing compact low-weight artificial optical sensors capable of performing visual processing tasks robustly under various environmental conditions (light, clouds, contrast). Here, we review some outstanding bio-inspired visual sensors, which can be used for either detecting motion in the visible spectrum or controlling celestial navigation in the ultraviolet spectrum and for attitude stabilisation purposes. Biologically inspired visual sensors do not have to comprise a very large number of pixels: they are able to perform both short and long range navigation tasks surprisingly well with just a few pixels and a weak resolution.


Beliefs about prescribed medication among older patients with polypharmacy: a mixed methods study in primary care.
Polypharmacy (≥5 medications) is common in older patients and is associated with adverse outcomes. Patients' beliefs about medication can influence their expectations for medication, adherence, and willingness to deprescribe. Few studies have examined beliefs about prescribed medication among older patients with polypharmacy in primary care. To explore medication-related beliefs in older patients with polypharmacy and factors that might influence beliefs. A mixed methods study utilising data from a randomised controlled trial aiming to decrease potentially inappropriate prescribing in older patients (≥70 years) in Ireland. Beliefs were assessed quantitatively and qualitatively. Participants completed the Beliefs about Medicines Questionnaire by indicating their degree of agreement with individual statements about medicines on a 5-point Likert scale. Semi-structured qualitative interviews were conducted with a purposive sample of participants. Interviews were transcribed verbatim and a thematic analysis conducted. Quantitative and qualitative data were analysed separately and triangulated during the interpretation stage. In total, 196 patients were included (mean age 76.7 years, SD 4.9, 54% male), with a mean of 9.5 (SD 4.1) medications per patient. The majority (96.3%) believed strongly in the necessity of their medication, while 33.9% reported strong concerns. Qualitative data confirmed these coexisting positive and negative attitudes to medications and suggested the importance of patients' trust in GPs in establishing positive beliefs and potential willingness to deprescribe. Participants reported strong beliefs in medications with coexisting positive and negative attitudes. The doctor-patient relationship may have influenced beliefs and attitudes towards medicines, highlighting the importance of strong doctor-patient relationships, which need to be considered in the context of deprescribing.


Visualizing intracellular particles and precise control of drug release using an emissive hydrazone photochrome.
The spatiotemporal control over the structure of nanoparticles while monitoring their localization in tumor cells can improve the precision of controlled drug release, thus enhancing the efficiency of drug delivery. Here, we report on a photochromic nanoparticle system (LSNP), assembled from fluorescent bistable hydrazone photoswitch-modified amphiphilic copolymers. The intrinsic emission of the hydrazone switch allows for the visualization of particle uptake, as well as their intracellular distribution. The Z → E photoswitching of the hydrazone switch within the nanoparticle leads to the expansion of the nanoparticles (i.e., drug release) accompanied by emission quenching, the degree of which can function as an internal indicator for the amount of drug released. The bistability of the switch enables the kinetic trapping of particles of different sizes as a function of irradiation time, and allows for the exhibition of light-dependent cell cytotoxicity in MDA-MB-231 cells using LSNP loaded with doxorubicin.


Numb is required to prevent p53-dependent senescence following skeletal muscle injury.
Regeneration relies on coordinated action of multiple cell types to reconstitute the damaged tissue. Here we inactivate the endocytic adaptor protein Numb in skeletal muscle stem cells prior to chronic or severe muscle injury in mice. We observe two types of senescence in regenerating muscle; a transient senescence in non-myogenic cells of control and Numb mutant mice that partly depends on INK4a/ARF activity, and a persistent senescence in myogenic cells lacking Numb. The senescence levels of Numb-deficient muscle is reduced to wild type levels by an anti-oxidant treatment or p53 ablation, resulting in functional rescue of the regenerative potential in Numb mutants. Ex vivo experiments suggest that Numb-deficient senescent cells recruit macrophages to sustain inflammation and drive fibrosis, two hallmarks of the impaired muscle regeneration in Numb mutants. These findings provide insights into previously reported developmental and oncogenic senescence that are also differentially regulated by p53.


Feasibility study on equine acellular pericardium matrix (APM): A new tool for breast reconstruction.
The advent of acellular dermal matrix (ADM) for lower pole coverage allows immediate reconstructions with improved aesthetic outcomes and faster recovery. This study describes for the first time, the use of a new acellular pericardium matrix (APM) in implant-based breast reconstruction and characterises its safety profile. Equity is a membrane with a natural cross-linked structure with many of the properties of ADMs, but improved resistance and reduced thickness. A retrospective data collection of all Equity APM reconstructions was conducted at two Italian hospital centres with substantial experience using biomaterials. Between May 2013 and October 2018, a total of 63 APM-assisted breast reconstructions were performed in 55 women. The reconstructed breasts were small to medium and the mean implant weight was 285 g, ranging from 145 g to 685 g. Two patients were previously irradiated while seven received post-operative radiation; five were active smokers and six were hypertensive. Complications included visibility in the upper pole (9.5%), seroma (1.6%), dehiscence, infection and necrosis (3.2% for each). Implant loss occurred in 3.2% of the cases. The patients were highly satisfied, reporting scores above 50 for each section of the Breast-Q questionnaire. With an acceptable complication rate, the use of the equine APM can be considered safe with satisfactory aesthetic results. Although the retrospective nature of this study limits its clinical impact, the use of Equity can be considered a viable alternative to thicker and expensive ADMs.


Improving the Diagnosis and Treatment of Urinary Tract Infection in Young Children in Primary Care: Results from the DUTY Prospective Diagnostic Cohort Study.
Up to 50% of urinary tract infections (UTIs) in young children are missed in primary care. Urine culture is essential for diagnosis, but urine collection is often difficult. Our aim was to derive and internally validate a 2-step clinical rule using (1) symptoms and signs to select children for urine collection; and (2) symptoms, signs, and dipstick testing to guide antibiotic treatment. We recruited acutely unwell children aged under 5 years from 233 primary care sites across England and Wales. Index tests were parent-reported symptoms, clinician-reported signs, urine dipstick results, and clinician opinion of UTI likelihood (clinical diagnosis before dipstick and culture). The reference standard was microbiologically confirmed UTI cultured from a clean-catch urine sample. We calculated sensitivity, specificity, and area under the receiver operator characteristic (AUROC) curve of coefficient-based (graded severity) and points-based (dichotomized) symptom/sign logistic regression models, and we then internally validated the AUROC using bootstrapping. Three thousand thirty-six children provided urine samples, and culture results were available for 2,740 (90%). Of these results, 60 (2.2%) were positive: the clinical diagnosis was 46.6% sensitive, with an AUROC of 0.77. Previous UTI, increasing pain/crying on passing urine, increasingly smelly urine, absence of severe cough, increasing clinician impression of severe illness, abdominal tenderness on examination, and normal findings on ear examination were associated with UTI. The validated coefficient- and points-based model AUROCs were 0.87 and 0.86, respectively, increasing to 0.90 and 0.90, respectively, by adding dipstick nitrites, leukocytes, and blood. A clinical rule based on symptoms and signs is superior to clinician diagnosis and performs well for identifying young children for noninvasive urine sampling. Dipstick results add further diagnostic value for empiric antibiotic treatment.


Flow Diversion for Treatment of Intracranial Aneurysms in Pediatric Patients: Multicenter Case Series.
Though the Pipeline Embolization Device (Medtronic) is approved for use in adults 22 yr and older, the high efficacy and long-term durability of the device is attractive for treatment of intracranial aneurysms in younger patients who often have aneurysms less amenable to traditional endovascular treatments. To report technical, angiographic, and clinical outcomes in patients aged 21 or below undergoing flow-diversion treatment for intracranial aneurysms. Retrospective review across 16 institutions identified 39 patients aged 21 or below undergoing 46 treatment sessions with Pipeline Embolization Device placement between 2012 and 2018. A total of 50 intracranial aneurysms were treated. Details regarding patient demographics, aneurysm characteristics, treatment considerations, clinical outcomes, and aneurysm occlusion were obtained and analyzed in a multicenter database. A total of 70% of patients were male. Nonsaccular morphology was seen in half of identified aneurysms. Six aneurysms were giant, and five patients were treated acutely after ruptured presentation. Eight patients were younger than 10 yr of age. Complete aneurysm occlusion was seen in 74% of treated aneurysms. Three aneurysms (6%) were retreated. A total of 83% of patients had a modified Rankin Scale scores of ≤2 at last clinical follow-up. There were 2 early mortalities (4.3%) in the immediate postprocedure period because of rerupture of a treated ruptured aneurysm. No recanalization of a previously occluded aneurysm was observed. Flow-diversion treatment is a safe and effective treatment for intracranial aneurysms in patients younger than 22 yr. Rates of complete aneurysm occlusion and adverse events are comparable for rates seen in older patients.


Increasing glucose concentrations and prevalence of diabetes mellitus in northern Sweden, 1990-2007.
The prevalence of diabetes in the world is projected to rise from 2.8% in the year 2000 to 4.4% in 2030, an increase suggesting an ongoing global epidemic of diabetes. To examine time trends in fasting and 2-h glucose concentrations, prevalence and 10-year cumulative incidence of diabetes, and the role of education in these trends. Each year the Västerbotten Intervention Programme invites all 40, 50, and 60-year-old individuals to a health survey, which includes a cardiovascular risk factor screening and oral glucose tolerance test. The cross-sectional part of the study is based on health examinations conducted between 1990 and 2007 (n = 102,822). The prospective subset (panel dataset) of the study is based on individuals who have had two health examinations 10 years apart and were not defined as having diabetes at their first health examination (n = 23,546). Between 1990 and 2007, the mean population fasting glucose concentration increased 0.5 mmol/L. Comparing the prevalence in 1990-1995 with 2002-2007 demonstrated a significant 44% increase in men (p < 0.001) and a significant 17% increase in women (p<0.001). Socioeconomic status, here represented by education, clearly influenced both prevalence and incidence of diabetes and glucose concentration. In all time periods and in all age groups, individuals with low education were more likely to have or get diabetes. The 10-year risk of developing diabetes was four to five times higher in the oldest age group (50-60 years) compared with the youngest (30-40 years). A 30% reduction in the 10-year risk of developing diabetes was found in women (p<0.001) between 2000-2003 and 2004-2007. Despite a clear increase in glucose concentrations and diabetes prevalence between 1990 and 2007, especially in men, there was a decline in the 10-year risk of developing diabetes in women between 2000-2003 and 2004-2007.


Kapalabhati--yogic cleansing exercise. II. EEG topography analysis.
Topography of brain electrical activity was studied in 11 advanced yoga practitioners during yogic high-frequency breathing kapalabhati (KB). Alpha activity was increased during the initial five min of KB. Theta activity mostly in the occipital region was increased during later stages of 15 min KB compared to the pre-exercise period. Beta 1 activity increased during the first 10 min of KB in occipital and to a lesser degree in parietal regions. Alpha and beta 1 activity decreased and theta activity was maintained on the level of the initial resting period after KB. The score of General Deactivation factor from Activation Deactivation Adjective Checklist was higher after KB exercise than before the exercise. The results suggest a relative increase of slower EEG frequencies and relaxation on a subjective level as the after effect of KB exercise.


Analysis of mesenchymal stem cell differentiation in vitro using classification association rule mining.
In this paper, data mining is used to analyze the data on the differentiation of mammalian Mesenchymal Stem Cells (MSCs), aiming at discovering known and hidden rules governing MSC differentiation, following the establishment of a web-based public database containing experimental data on the MSC proliferation and differentiation. To this effect, a web-based public interactive database comprising the key parameters which influence the fate and destiny of mammalian MSCs has been constructed and analyzed using Classification Association Rule Mining (CARM) as a data-mining technique. The results show that the proposed approach is technically feasible and performs well with respect to the accuracy of (classification) prediction. Key rules mined from the constructed MSC database are consistent with experimental observations, indicating the validity of the method developed and the first step in the application of data mining to the study of MSCs.


Incoherent policies on universal coverage of health insurance and promotion of international trade in health services in Thailand.
The Thai government has implemented universal coverage of health insurance since October 2001. Universal access to antiretroviral (ARV) drugs has also been included since October 2003. These two policies have greatly increased the demand for health services and human resources for health, particularly among public health care providers. After the 1997 economic crisis, private health care providers, with the support of the government, embarked on new marketing strategies targeted at attracting foreign patients. Consequently, increasing numbers of foreign patients are visiting Thailand to seek medical care. In addition, the economic recovery since 2001 has greatly increased the demand for private health services among the Thai population. The increasing demand and much higher financial incentives from urban private providers have attracted health personnel, particularly medical doctors, from rural public health care facilities. Responding to this increasing demand and internal brain drain, in mid-2004 the Thai government approved the increased production of medical doctors by 10,678 in the following 15 years. Many additional financial incentives have also been applied. However, the immediate shortage of human resources needs to be addressed competently and urgently. Equity in health care access under this situation of competing demands from dual track policies is a challenge to policy makers and analysts. This paper summarizes the situation and trends as well as the responses by the Thai government. Both supply and demand side responses are described, and some solutions to restore equity in health care access are proposed.


Independent association of fibrinogen with carotid intima-media thickness in asymptomatic subjects.
Fibrinogen has been found to be an independent risk factor for cardiovascular disease. Both genetic and environmental factors contribute to its variability in plasma. However, whether the relation between fibrinogen and carotid intima-media thickness (IMT) is independent of those factors has not been established. Therefore, the aim of this study was to investigate the relations of plasma fibrinogens and the -455 G/A Bbeta-fibrinogen polymorphism with the carotid IMT in a series of asymptomatic subjects. Markers of inflammation, C-reactive protein (CRP) and leukocytes, and endothelial perturbation (von Willebrand factor, vWF) were measured in 135 subjects. All individuals underwent a complete clinical examination and lipid measurements (cholesterol and its fractions HDL and LDL and triglycerides). The carotid IMT was measured by B-mode ultrasound in the common carotid artery. Patients in the highest fibrinogen tertile had a significantly higher BMI (p < 0.01), LDL-cholesterol (p < 0.01), leukocyte count, CRP and vWF (p < 0.001). In the univariate model a strong positive relationship was found between plasma fibrinogen and carotid IMT (p < 0.01). Fibrinogen also correlated positively with age, BMI, arterial systolic pressure, cholesterol, cholesterol-LDL, smoking, CRP and vWF (p < 0.01). In the multivariate analysis, the association of fibrinogen with carotid IMT remained significant (p < 0.01) after adjustment for all the parameters analyzed. In a population sample of adults without clinically overt atherosclerotic disease, elevated fibrinogen was related to carotid IMT independent of a wide range of important confounding variables.


Collinear holography.
A novel reading and writing technology for a holographic storage system called collinear holography is developed. With this method, two-dimensional page data can be recorded as volumetric holograms generated by a reference beam and a signal beam that are bundled on the same axis and irradiated on the recording medium through a single objective lens. The multiplex recording and reconstructing process is demonstrated, and it is presented that optical configuration and the dichroic media disk structure are suitable for a compact system. This method enables us to construct a small volumetric optical disk storage system with CD and DVD upper compatibility.


Mutagenesis in PMS2- and MSH2-deficient mice indicates differential protection from transversions and frameshifts.
DNA mismatch repair (MMR) deficiency leads to an increased mutation frequency and a predisposition to neoplasia. 'Knockout' mice deficient in the MMR proteins Msh2 and Pms2 crossed with mutation detection reporter (supF, lacI and cII) transgenic mice have been used to facilitate a comparison of the changes in mutation frequency and spectra. We find that the mutation frequency was consistently higher in Msh2-deficient mice than Pms2-deficient mice. The lacI target gene, which is highly sensitive to point mutations, demonstrated that both Msh2- and Pms2-deficient mice accumulate transition mutations as the predominant mutation. However, when compared with Msh2(-/-) mice, lacI and cII mutants from Pms2-deficient mice revealed an increased proportion of +/-1 bp frameshift mutations and a corresponding decrease in transversion mutations. The supF target gene, which is sensitive to frameshift mutations, and the cII target gene revealed a strong tendency for -1 bp deletions over +1 bp insertions in Msh2(-/-) compared with Pms2(-/-) mice. These data indicate that Msh2 and Pms2 deficiency have subtle but differing effects on mutation avoidance which may contribute to the differences in tumor spectra observed in the two 'knockout' mouse models. These variances in mutation accumulation may also play a role, in part, in the differences seen in prevalence of MSH2 and PMS2 germline mutations in hereditary non-polyposis colorectal cancer patients.


Triacylglyceride physiology in the short-finned eel, Anguilla australis--the effects of androgen.
The importance of androgens (especially 11-ketotestosterone) during previtellogenesis in eels is well established. In wild pubertal migrants, circulating 11-ketotestosterone levels correlate with a number of morphological and molecular changes. Here, we test the prediction that this correlation represents a causal relationship by artificially raising the levels of circulating 11-ketotestosterone in prepubertal nonmigratory female and pubertal, migratory male short-finned eels (Anguilla australis) using sustained-release hormone implants. In females, increases in hepatosomatic index and transcript copy numbers of hepatic apolipoprotein B and microsomal triacylglyceride transfer protein indicated increased repackaging of endogenously sourced triacylglycerides. These changes in liver measures were reflected in increased concentrations of serum triacylglycerides. However, despite a small increase in gonadosomatic index, ovarian lipoprotein receptor transcript abundances were not affected by 11-ketotestosterone. Interestingly, no such changes in hepatic gene expression were detected in a dose-response experiment using males. We propose that the androgens are inducing the observed changes in previtellogenic females, although it remains unclear to what extent these effects are direct or indirect.


The 25th anniversary of the retrograde suction decompression technique (Dallas technique) for the surgical management of paraclinoid aneurysms: historical background, systematic review, and pooled analysis of the literature.
Paraclinoid internal carotid artery (ICA) aneurysms frequently require temporary occlusion to facilitate safe clipping. Brisk retrograde flow through the ophthalmic artery and cavernous ICA branches make simple trapping inadequate to soften the aneurysm. The retrograde suction decompression (RSD), or Dallas RSD, technique was described in 1990 in an attempt to overcome some of those treatment limitations. A frequent criticism of the RSD technique is an allegedly high risk of cervical ICA dissection. An endovascular modification was introduced in 1991 (endovascular RSD) but no studies have compared the 2 RSD variations. The authors performed a systematic review of MEDLINE/PubMed and Web of Science and identified all studies from 1990-2016 in which either Dallas RSD or endovascular RSD was used for treatment of paraclinoid aneurysms. A pooled analysis of the data was completed to identify important demographic and treatment-specific variables. The primary outcome measure was defined as successful aneurysm obliteration. Secondary outcome variables were divided into overall and RSD-specific morbidity and mortality rates. Twenty-six RSD studies met the inclusion criteria (525 patients, 78.9% female). The mean patient age was 53.5 years. Most aneurysms were unruptured (56.6%) and giant (49%). The most common presentations were subarachnoid hemorrhage (43.6%) and vision changes (25.3%). The aneurysm obliteration rate was 95%. The mean temporary occlusion time was 12.7 minutes. Transient or permanent morbidity was seen in 19.9% of the patients. The RSD-specific complication rate was low (1.3%). The overall mortality rate was 4.2%, with 2 deaths (0.4%) attributable to the RSD technique itself. Good or fair outcome were reported in 90.7% of the patients.Aneurysm obliteration rates were similar in the 2 subgroups (Dallas RSD 94.3%, endovascular RSD 96.3%, p = 0.33). Despite a higher frequency of complex (giant or ruptured) aneurysms, Dallas RSD was associated with lower RSD-related morbidity (0.6% vs 2.9%, p = 0.03), compared with the endovascular RSD subgroup. There was a trend toward higher mortality in the endovascular RSD subgroup (6.4% vs 3.1%, p = 0.08). The proportion of patients with poor neurological outcome at last follow-up was significantly higher in the endovascular RSD group (15.4% vs 7.2%, p < 0.01). The treatment of paraclinoid ICA aneurysms using the RSD technique is associated with high aneurysm obliteration rates, good long-term neurological outcome, and low RSD-related morbidity and mortality. Review of the RSD literature showed no evidence of a higher complication rate associated with the Dallas technique compared with similar endovascular methods. On a subgroup analysis of Dallas RSD and endovascular RSD, both groups achieved similar obliteration rates, but a lower RSD-related morbidity was seen in the Dallas technique subgroup. Twenty-five years after its initial publication, RSD remains a useful neurosurgical technique for the management of large and giant paraclinoid aneurysms.


Plasmodium falciparum-infected erythrocytes induce granzyme B by NK cells through expression of host-Hsp70.
In the early immune response to Plasmodium falciparum-infected erythrocytes (iRBC), Natural Killer (NK) cells are activated, which suggests an important role in innate anti-parasitic immunity. However, it is not well understood whether NK cells directly recognize iRBC or whether stimulation of NK cells depends mainly on activating signals from accessory cells through cell-to-cell contact or soluble factors. In the present study, we investigated the influence of membrane-bound host Heat shock protein (Hsp) 70 in triggering cytotoxicity of NK cells from malaria-naïve donors or the cell line NK92 against iRBC. Hsp70 and HLA-E membrane expression on iRBC and potential activatory NK cell receptors (NKG2C, CD94) were assessed by flow cytometry and immunoblot. Upon contact with iRBC, Granzyme B (GzmB) production and release was initiated by unstimulated and Hsp70-peptide (TKD) pre-stimulated NK cells, as determined by Western blot, RT-PCR and ELISPOT analysis. Eryptosis of iRBC was determined by Annexin V-staining. Our results suggest that presence of Hsp70 and absence of HLA-E on the membrane of iRBC prompt the infected host cells to become targets for NK cell-mediated cytotoxicity, as evidenced by impaired parasite development. Contact of iRBC with NK cells induced release of GzmB. We propose that following GzmB uptake, iRBC undergo eryptosis via a perforin-independent, GzmB-mediated mechanism. Since NK activity toward iRBC could be specifically enhanced by TKD peptide and abrogated to baseline levels by blocking Hsp70 exposure, we propose TKD as an innovative immunostimulatory agent to be tested as an adjunct to anti-parasitic treatments in vivo.


Networks' Characteristics Matter for Systems Biology.
A fundamental goal of systems biology is to create models that describe relationships between biological components. Networks are an increasingly popular approach to this problem. However, a scientist interested in modeling biological (e.g., gene expression) data as a network is quickly confounded by the fundamental problem: how to construct the network? It is fairly easy to construct a network, but is it the network for the problem being considered? This is an important problem with three fundamental issues: How to weight edges in the network in order to capture actual biological interactions? What is the effect of the type of biological experiment used to collect the data from which the network is constructed? How to prune the weighted edges (or what cut-off to apply)? Differences in the construction of networks could lead to different biological interpretations. Indeed, we find that there are statistically significant dissimilarities in the functional content and topology between gene co-expression networks constructed using different edge weighting methods, data types, and edge cut-offs. We show that different types of known interactions, such as those found through Affinity Capture-Luminescence or Synthetic Lethality experiments, appear in significantly varying amounts in networks constructed in different ways. Hence, we demonstrate that different biological questions may be answered by the different networks. Consequently, we posit that the approach taken to build a network can be matched to biological questions to get targeted answers. More study is required to understand the implications of different network inference approaches and to draw reliable conclusions from networks used in the field of systems biology.


Periostin expression in neoplastic and non-neoplastic diseases of bone and joint.
Periostin is a matricellular protein that is expressed in bone and joint tissues. To determine the expression of periostin in primary bone tumours and to assess whether it plays a role in tumour progression, we carried out immunohistochemistry and ELISA for periostin in a range of neoplastic and non-neoplastic bone and joint lesions. 140 formalin-fixed paraffin-embedded sections of bone tumours and tumour-like lesions were stained by an indirect immunoperoxidase technique with a polyclonal anti-periostin antibody. Periostin expression was also assessed in rheumatoid arthritis (RA) and non-inflammatory osteoarthritis (OA) synovium and synovial fluid immunohistochemistry and ELISA respectively. Periostin was most strongly expressed in osteoid/woven bone of neoplastic and non-neoplastic bone-forming lesions, including osteoblastoma, osteosarcoma, fibrous dysplasia, osteofibrous dysplasia, fracture callus and myositis ossificans, and mineralised chondroid matrix/woven bone in chondroblastoma and clear cell chondrosarcoma. Reactive host bone at the edge of growing tumours, particularly in areas of increased vascularity and fibrosis, also stained strongly for periostin. Vascular elements in RA synovium strongly expressed periostin, and synovial fluid levels of periostin were higher in RA than OA. In keeping with its known role in modulating the synthesis of collagen and other extracellular matrix proteins in bone, strong periostin expression was noted in benign and malignant lesions forming an osteoid or osteoid-like matrix. Periostin was also noted in other bone tumours and was found in areas of reactive bone and increased vascularity at the edge of growing tumours, consistent with its involvement in tissue remodelling and angiogenesis associated with tumour progression.


BAX channel activity mediates lysosomal disruption linked to Parkinson disease.
Lysosomal disruption is increasingly regarded as a major pathogenic event in Parkinson disease (PD). A reduced number of intraneuronal lysosomes, decreased levels of lysosomal-associated proteins and accumulation of undegraded autophagosomes (AP) are observed in PD-derived samples, including fibroblasts, induced pluripotent stem cell-derived dopaminergic neurons, and post-mortem brain tissue. Mechanistic studies in toxic and genetic rodent PD models attribute PD-related lysosomal breakdown to abnormal lysosomal membrane permeabilization (LMP). However, the molecular mechanisms underlying PD-linked LMP and subsequent lysosomal defects remain virtually unknown, thereby precluding their potential therapeutic targeting. Here we show that the pro-apoptotic protein BAX (BCL2-associated X protein), which permeabilizes mitochondrial membranes in PD models and is activated in PD patients, translocates and internalizes into lysosomal membranes early following treatment with the parkinsonian neurotoxin MPTP, both in vitro and in vivo, within a time-frame correlating with LMP, lysosomal disruption, and autophagosome accumulation and preceding mitochondrial permeabilization and dopaminergic neurodegeneration. Supporting a direct permeabilizing effect of BAX on lysosomal membranes, recombinant BAX is able to induce LMP in purified mouse brain lysosomes and the latter can be prevented by pharmacological blockade of BAX channel activity. Furthermore, pharmacological BAX channel inhibition is able to prevent LMP, restore lysosomal levels, reverse AP accumulation, and attenuate mitochondrial permeabilization and overall nigrostriatal degeneration caused by MPTP, both in vitro and in vivo. Overall, our results reveal that PD-linked lysosomal impairment relies on BAX-induced LMP, and point to small molecules able to block BAX channel activity as potentially beneficial to attenuate both lysosomal defects and neurodegeneration occurring in PD.


Polysialylated N-glycans identified in human serum through combined developments in sample preparation, separations, and electrospray ionization-mass spectrometry.
The N-glycan diversity of human serum glycoproteins, i.e., the human blood serum N-glycome, is both complex and constrained by the range of glycan structures potentially synthesizable by human glycosylation enzymes. The known glycome, however, has been further limited by methods of sample preparation, available analytical platforms, e.g., based upon electrospray ionization-mass spectrometry (ESI-MS), and software tools for data analysis. In this report several improvements have been implemented in sample preparation and analysis to extend ESI-MS glycan characterization and to include polysialylated N-glycans. Sample preparation improvements included acidified, microwave-accelerated, PNGase F N-glycan release to promote lactonization, and sodium borohydride reduction, that were both optimized to improve quantitative yields and conserve the number of glycoforms detected. Two-stage desalting (during solid phase extraction and on the analytical column) increased sensitivity by reducing analyte signal division between multiple reducing-end-forms or cation adducts. Online separations were improved by using extended length graphitized carbon columns and adding TFA as an acid modifier to a formic acid/reversed phase gradient, providing additional resolving power and significantly improved desorption of both large and heavily sialylated glycans. To improve MS sensitivity and provide gentler ionization conditions at the source-MS interface, subambient pressure ionization with nanoelectrospray (SPIN) was utilized. When these improved methods are combined together with the Glycomics Quintavariate Informed Quantification (GlyQ-IQ) recently described (Kronewitter et al. Anal. Chem. 2014, 86, 6268-6276), we are able to significantly extend glycan detection sensitivity and provide expanded glycan coverage. We demonstrated the application of these advances in the context of the human serum glycome, and for which our initial observations included the detection of a new class of heavily sialylated N-glycans, including polysialylated N-glycans.


Effects of dietary carbohydrate restriction versus low-fat diet on flow-mediated dilation.
We previously reported that a carbohydrate-restricted diet (CRD) ameliorated many of the traditional markers associated with metabolic syndrome and cardiovascular risk compared with a low-fat diet (LFD). There remains concern how CRD affects vascular function because acute meals high in fat have been shown to impair endothelial function. Here, we extend our work and address these concerns by measuring fasting and postprandial vascular function in 40 overweight men and women with moderate hypertriacylglycerolemia who were randomly assigned to consume hypocaloric diets (approximately 1500 kcal) restricted in carbohydrate (percentage of carbohydrate-fat-protein = 12:59:28) or LFD (56:24:20). Flow-mediated dilation of the brachial artery was assessed before and after ingestion of a high-fat meal (908 kcal, 84% fat) at baseline and after 12 weeks. Compared with the LFD, the CRD resulted in a greater decrease in postprandial triacylglycerol (-47% vs -15%, P = .007), insulin (-51% vs -6%, P = .009), and lymphocyte (-12% vs -1%, P = .050) responses. Postprandial fatty acids were significantly increased by the CRD compared with the LFD (P = .033). Serum interleukin-6 increased significantly over the postprandial period; and the response was augmented in the CRD (46%) compared with the LFD (-13%) group (P = .038). After 12 weeks, peak flow-mediated dilation at 3 hours increased from 5.1% to 6.5% in the CRD group and decreased from 7.9% to 5.2% in the LFD group (P = .004). These findings show that a 12-week low-carbohydrate diet improves postprandial vascular function more than a LFD in individuals with atherogenic dyslipidemia.


Positive and negative thymocyte selection induced by different concentrations of a single peptide.
T lymphocyte maturation is dependent on interactions between the T cell receptor (TCR) expressed on the developing thymocyte and intrathymic major histocompatibility complex (MHC)-peptide ligands. The relation between the peptide-MHC complex that results in negative or positive selection has not been identified. Here, the requirements for the maturation of thymocytes expressing a defined transgenic TCR specific for a viral peptide are studied in fetal thymic organ culture. Low concentrations of the viral peptide antigen recognized by this transgenic TCR can mediate positive selection, whereas high concentrations result in thymocyte tolerance. These findings support the affinity-avidity model of thymocyte selection.


Quality of compounded hydrocortisone capsules used in the treatment of children.
Due to the lack of paediatric-licensed formulations, children are often treated with individualized pharmacy-compounded adult medication. An international web-based survey about the types of medication in children with adrenal insufficiency (AI) revealed that the majority of paediatric physicians are using pharmacy-compounded medication to treat children with AI. Observations of loss of therapy control in children with congenital adrenal hyperplasia with compounded hydrocortisone capsules and regained control after prescribing a new hydrocortisone batch led to this 'real world' evaluation of pharmacy-compounded paediatric hydrocortisone capsules. Capsule samples were collected randomly from volunteering parents of treated children suffering from congenital adrenal hyperplasia from all over Germany. Analysis of net mass and hydrocortisone content by high-performance liquid chromatography with ultraviolet (HPLC-UV) detection method was performed based on the European Pharmacopeia. In a total of 61 batches that were sent, 5 batches could not be analysed because of missing dose information, insufficient number of capsules or were not possible to be evaluated. Fifty-six batches containing 1125 capsules were evaluated. 21.4% of the batches revealed insufficiency in uniformity of net mass or drug content and additional 3.6% failed because they did not contain the labelled drug. Compounded medication is a possible cause of variation of steroid doses in children with adrenal insufficiency or congenital adrenal hyperplasia, putting these vulnerable patients at risk of poor disease control and adrenal crisis. These data may apply to other individualized compounded oral medication as well, emphasizing the need for development of licensed paediatric formulations approved by regulatory authorities.


A role for oxidative stress in suppressing serum immunoglobulin levels in lead-exposed Fisher 344 rats.
Evidence implicating oxidative stress in toxicity during lead intoxication in vivo has opened new avenues for investigation of the mechanisms of lead-induced immunosuppression. The current study explores the possibility that lead-induced oxidative stress contributes to the immunosuppression observed during lead poisoning. Fisher 344 rats were exposed to 2,000 ppm lead acetate in their drinking water for 5 weeks. One week following removal of lead from the drinking water, significant reductions in serum levels of IgA, IgM, and IgG were detected. Significant increases in oxidative damage, based on malondialdehyde (MDA) content, were observed in peripheral blood mononuclear cells (PMCs) collected during the same experiments. In addition, MDA content increased in livers from lead-exposed rats. Following 5 weeks of lead exposure, administration of either 5.5 mmol/kg N-acetylcysteine (NAC) or 1 mmol/kg meso-2,3-dimercaptosuccinic acid (DMSA) in the drinking water for 1 week significantly reversed the inhibitory effects of lead on serum immunoglobulin (Ig) levels. Also, all parameters indicative of oxidative stress returned to control levels. These results suggest that oxidative stress contributes to suppressed serum Ig levels during lead intoxication in vivo, and that intervention with either a thiol antioxidant (NAC) or a metal chelator (DMSA) will alleviate this lead-induced suppression by correcting the prooxidant/antioxidant imbalance caused by lead exposure.


Multichannel photoinduced intramolecular electron-transfer excitations in a bis-naphthalimide spermine conjugate by time-dependent density functional theory.
Density functional theory was applied to the investigation of photoinduced electron transfer (ET) and the absorption spectrum for a bis-naphthalimide spermine conjugate. The multichannel feature of ET excitation in this system was focused on because four groups may act as electron donors and acceptors. The segment in this conjugate, N-(N-methylpropyl)-1,8-naphthalimide, which contains one donor and acceptor pair, was studied at first. Through theoretical calculation, the absorption band at 340 nm was assigned to the pi-->pi* transition. For the whole system involving four chromophores, this work suggested three types of ET. From the theoretical investigation, the naphthalimide radical anion turned out to be formed via intramolecular ET between the two terminal naphthalimide groups, rather than via the electron transfer between the dialkylamine moiety and the naphthalimide one. Furthermore, the electronic coupling matrix elements according to the generalized Mulliken-Hush theory were estimated and the detailed analyses showed that the strongest absorption was due to the local excitation of the naphthalimide chromophore.


Neuropeptide Y inhibits cholangiocarcinoma cell growth and invasion.
No information exists on the role of neuropeptide Y (NPY) in cholangiocarcinoma growth. Therefore, we evaluated the expression and secretion of NPY and its subsequent effects on cholangiocarcinoma growth and invasion. Cholangiocarcinoma cell lines and nonmalignant cholangiocytes were used to assess NPY mRNA expression and protein secretion. NPY expression was assessed by immunohistochemistry in human liver biopsies. Cell proliferation and migration were evaluated in vitro by MTS assays and matrigel invasion chambers, respectively, after treatment with NPY or a neutralizing NPY antibody. The effect of NPY or NPY depletion on tumor growth was assessed in vivo after treatment with NPY or the neutralizing NPY antibody in a xenograft model of cholangiocarcinoma. NPY secretion was upregulated in cholangiocarcinoma compared with normal cholangiocytes. Administration of exogenous NPY decreased proliferation and cell invasion in all cholangiocarcinoma cell lines studied and reduced tumor cell growth in vivo. In vitro, the effects of NPY on proliferation were blocked by specific inhibitors for NPY receptor Y2, but not Y1 or Y5, and were associated with an increase in intracellular d-myo-inositol 1,4,5-trisphosphate and PKCα activation. Blocking of NPY activity using a neutralizing antibody promoted cholangiocarcinoma growth in vitro and in vivo and increased the invasiveness of cholangiocarcinoma in vitro. Increased NPY immunoreactivity in human tumor tissue occurred predominantly in the center of the tumor, with less expression toward the invasion front of the tumor. We demonstrated that NPY expression is upregulated in cholangiocarcinoma, which exerts local control on tumor cell proliferation and invasion. Modulation of NPY secretion may be important for the management of cholangiocarcinoma.


Satisfaction with Care in Late Stage Parkinson's Disease.
In late stage Parkinson's disease (PD) (i.e., Hoehn and Yahr (HY) stages IV-V), both motor and nonmotor symptoms (NMS) are pronounced, and the patients become increasingly dependent on help in their daily life. Consequently, there is an increasing demand on health-care and social care resources for these patients and support for their informal caregivers. The aim of this study was to assess satisfaction with care in late stage PD patients and to identify factors associated with satisfaction with care. Moreover, to assess their informal caregivers' satisfaction with support and to identify factors associated with caregivers' satisfaction with support. Factors potentially associated with satisfaction with care/support were assessed in 107 late stage PD patients and their informal caregivers (n=76) and entered into multivariable logistic regression analyses. Fifty-eight (59%) of the patients and 45 (59%) of the informal caregivers reported satisfaction with their overall care/support. Patients satisfied with their care reported higher independence in activities of daily living (ADL) (Katz ADL index; P=0.044), less depressive symptoms (Geriatric Depression Scale, GDS-30; P=0.005), and higher individual quality of life (QoL) (Schedule for the Evaluation of Individual Quality of Life Questionnaire, SEIQoL-Q; P=0.036). Multivariable logistic regression analyses identified depressive symptoms (P=0.015) and independence in ADL (P=0.025) as independently associated with satisfaction with care. For informal caregivers, the analyses identified patients' HY stage (P=0.005) and caregivers' QoL (Alzheimer's Carers Quality of Life Inventory, ACQLI; P=0.012) as independently associated with satisfaction with caregiver support. The results indicate that an effective both pharmacological and nonpharmacological PD therapy is important, to adequately treat motor and NMS (e.g., depressive symptoms) in order to improve depressive symptoms and patient independence in ADL. This may benefit not only the patients, but also their informal caregivers.


Active site closure stabilizes the backtracked state of RNA polymerase.
All cellular RNA polymerases (RNAP) occasionally backtrack along the template DNA as part of transcriptional proofreading and regulation. Here, we studied the mechanism of RNAP backtracking by one nucleotide using two complementary approaches that allowed us to precisely measure the occupancy and lifetime of the backtracked state. Our data show that the stability of the backtracked state is critically dependent on the closure of the RNAP active site by a mobile domain, the trigger loop (TL). The lifetime and occupancy of the backtracked state measurably decreased by substitutions of the TL residues that interact with the nucleoside triphosphate (NTP) substrate, whereas amino acid substitutions that stabilized the closed active site increased the lifetime and occupancy. These results suggest that the same conformer of the TL closes the active site during catalysis of nucleotide incorporation into the nascent RNA and backtracking by one nucleotide. In support of this hypothesis, we construct a model of the 1-nt backtracked complex with the closed active site and the backtracked nucleotide in the entry pore area known as the E-site. We further propose that 1-nt backtracking mimics the reversal of the NTP substrate loading into the RNAP active site during on-pathway elongation.


First Report of Web Blight on Oregano (Origanum vulgare L.) Caused by Rhizoctonia solani AG-1-IB in Italy.
Origanum vulgare L., common name oregano, family Labiatae, is grown for its aromatic and medicinal properties and as ornamental. In the fall of 2012, a blight was observed in a farm located near Albenga (northern Italy) on 6% of 30,000 50-day-old plants, grown in trays in a peat/perlite mix. Semicircular, water soaked lesions appeared on leaves and stems, starting from the basal ones. As the disease progressed, blighted leaves turned brown, withered, clung to the shoots, and matted on the surrounding foliage. Eventually, infected plants died. Leaf and stem fragments taken from the margin of the diseased tissues belonging to 10 plants were disinfected for 10 s in 1% NaOCl, rinsed with sterile water, and plated on potato dextrose agar (PDA). A fungus with the morphological characters of Rhizoctonia solani was consistently recovered. Three isolates of R. solani obtained from affected plants were successfully anastomosed with R. solani isolate AG 1 (ATCC 58946). Three pairings were made for each tester strain. The hyphal diameter at the point of anastomosis was reduced, the anastomosis point was obvious, and death of adjacent cells was observed. Results were consistent with other reports on anastomosis reactions (2). Isolates from oregano were paired with R. solani isolates AG 2, 3, 4, 6, 7, or 11 and examined microscopically. Anastomosis was not observed in any of the pairings. Tests were conducted twice. Mycelium of 10-day-old isolates from oregano appeared reddish brown, coarse, and radiate. Numerous dark brown sclerotia, 0.3 to 1.0 mm diameter (average 0.7) developed within 10 days after transfer of mycelia to PDA in 90 mm diameter petri dishes at 21 to 24°C. The descriptions of mycelium and sclerotia were typical for subgroup IB Type 1 (4). The internal transcribed spacer (ITS) region of rDNA was amplified using the primers ITS1/ITS4 and sequenced. BLASTn analysis (1) of the 538 bp showed a 99% homology with the sequence of R. solani FJ746937, confirming the morphological identification of the species. The nucleotide sequence has been assigned the GenBank Accession KC493638. For pathogenicity tests, one of the isolates assigned to the anastomosis group AG-1-IB was tested by placing 9 mm diameter mycelial disks removed from PDA 10-day-old cultures of the fungus on leaves of 90-day-old oregano plants (n = 35). Thirty-five plants inoculated with non-inoculated PDA disks served as controls. Plants were covered with plastic bags and maintained in a growth chamber at 25 ± 1°C with 12 h light/dark. The first symptoms, similar to those observed in the farm, developed 3 days after inoculation. Nine days after the artificial inoculation, 50% of plants were dead. About 10 colonies of R. solani were reisolated from infected leaves of inoculated plants. Control plants remained healthy. The pathogenicity test was carried out twice with similar results. Symptoms caused by R. solani have been recently observed on O. vulgare in Greece (3). This is, to our knowledge, the first report of blight of O. vulgare caused by R. solani in Italy. References: (1) S. F. Altschul et al. Nucleic Acids Res., 25:3389, 1997. (2) D. E. Carling. Grouping in Rhizoctonia solani by hyphal anastomosis reactions. In: Rhizoctonia Species: Taxonomy, Molecular Biology, Ecology, Pathology and Disease control. Kluwer Academic Publishers, The Netherlands, pp. 37-47, 1996. (3) C. D. Holevas et al. Benaki Phytopathol. Inst., Kiphissia, Athens, 19:1-96, 2000. (4) R. T. Sherwood. Phytopathology 59:1924, 1969.


Targeted in-gel MRM: a hypothesis driven approach for colorectal cancer biomarker discovery in human feces.
Colorectal cancer (CRC) is the second most common cause of cancer-related deaths in both men and women. The fecal occult blood test is currently the first line method for CRC screening but has an unacceptably low sensitivity and specificity. Improved screening tests are therefore urgently required for early stage CRC screening. We have described a hypothesis-driven approach for a rapid biomarker discovery process whereby selected proteins previously implicated as colorectal cancer-associated proteins (CCAP), which can potentially be shed into the feces from a colorectal tumor, are targeted for excision from 1D-SDS-PAGE based on their predicted molecular weight followed by directed identification and relative quantification using multiple reaction monitoring (MRM). This approach can significantly reduce the time for clinical assay development with the added advantage that many proteins will have been validated by previous in vitro and/or in vivo studies. Sixty potential CCAPs were selected from the literature and appropriate MRM conditions were established for measurement of proteotypic peptides. Nineteen of these proteins were detected in the feces from a patient with colorectal cancer. Relative quantitation of these 19 CCAP across 5 CRC patients and 5 healthy volunteers were carried out, revealing hemoglobin, myeloperoxidase, S100A9, filamin A and l-plastin to be present only in the feces of CRC patients.


Factors leading to obstructive granulation tissue formation after ultraflex stenting in benign tracheal narrowing.
This retrospective study aimed to determine the factors leading to obstructive granulation tissue formation after the placement of a self-expandable metallic stent (SEMS) in patients with benign tracheal disease. From 2001 to 2007, a total of 67 patients (age: 62.1 +/- 15.4 years; range: 23-87 years) with benign tracheal disease received 75 ultraflex SEMS in our institution. There were 35 SEMSs complicated by obstructive granulation tissue formation out of the 75 stents placed in patients with tracheal disease, giving an incidence of 47.8 % (32/67 patients). The median time until developing granulation tissue was 106 days (IQR, 46-396). Structural airway obstruction prior to SEMS implantation independently predicted obstructive granulation tissue formation after SEMS implantation (odds ratio: 3.84; 95 % CI: 1.01-8.7; P = 0.04). Time to granulation tissue detection was shorter in patients with structural airway obstruction before SEMS implantation (structural airway obstruction vs. dynamic collapse airway: median [IQR] 95 [38-224, n = 26] vs. 396 days [73-994, n = 9]; P = 0.02). Obstructive granulation tissue formation is not uncommon after SEMS implantation and structural airway obstruction prior to SEMS implantation is an independent predictor. Although SEMS implantation should be restricted to a select population, it may be placed in patients not suitable for surgical intervention or rigid bronchoscopy with anesthesia because of poor pulmonary function.