Gaze bias both reflects and influences preference.
Emotions operate along the dimension of approach and aversion, and it is reasonable to assume that orienting behavior is intrinsically linked to emotionally involved processes such as preference decisions. Here we describe a gaze 'cascade effect' that was present when human observers were shown pairs of human faces and instructed to decide which face was more attractive. Their gaze was initially distributed evenly between the two stimuli, but then gradually shifted toward the face that they eventually chose. Gaze bias was significantly weaker in a face shape discrimination task. In a second series of experiments, manipulation of gaze duration, but not exposure duration alone, biased observers' preference decisions. We thus conclude that gaze is actively involved in preference formation. The gaze cascade effect was also present when participants compared abstract, unfamiliar shapes for attractiveness, suggesting that orienting and preference for objects in general are intrinsically linked in a positive feedback loop leading to the conscious choice.


Stability, accumulation and cytotoxicity of an albumin-cisplatin adduct.
The accumulation and cytotoxicity of a 10 μmol L⁻¹ equimolar human serum albumin-cisplatin adduct (HSA-Pt) was investigated in suspension Ehrlich Ascites Tumor Cells (EATC) and adherent Ehrlich Lettré Ascites Cells (Lettré). HSA-Pt did not induce apoptosis nor was it taken up by the cells to any significant amount within 24 h incubation. The accumulation and cytotoxicity of HSA-Pt was compared to 10 μmol L⁻¹ cisplatin for which a larger accumulation and cytotoxicity were observed in EATC compared to Lettré. The experiment was performed with cell medium exchange every fourth hour as HSA-Pt and cisplatin were not stable in RPMI-1640 with 10% serum. The stability was determined using size exclusion chromatography-inductively coupled plasma-mass spectrometry (SEC-ICP-MS) and after 4 h new platinum peaks were observed. These findings indicate that before conducting cell experiments, the stability of the compound in the cell medium should be investigated especially when long exposure times are applied. Furthermore, HSA-Pt was found to be stable in Hanks Balanced Saline Solution (HBSS) and in Phosphate Buffered Saline (PBS) at pH 5.3, 6.1 and 7.4. Thus, the shift in pH when HSA-cisplatin passes from blood (pH 7.4) to tumor tissue (pH 5-6) is not capable of releasing cisplatin from HSA.


Pharmacokinetics of 2-ethyl-1,3-hexanediol. II. Nonsystemic disposition following single percutaneous or peroral doses in Fischer 344 rats.
The pharmacokinetics of [1,3-14C]-2-ethyl-1,3-hexanediol (EHD) were investigated following single percutaneous doses of 150 mg/kg, applied to male and female Fischer 344 rats, or single peroral doses of 1.5 or 150 mg EHD/kg given by gavage to male Fischer 344 rats. EHD-derived radioactivity was slowly absorbed through skin and relatively rapidly excreted through the urine in a first-order manner over 48 hr postdosing. Skin penetration of 14C was sufficiently slow that the terminal rate constant for the plasma concentration data had to be derived from the absorption phase of this curve, based on the terminal rate constant for a comparable intravenous dose plasma curve [Frantz et al.: Drug Metab. Dispos. 19, 881 (1991)]. Plasma data from perorally doses rats exhibited dose-linearity over a 1.5-150 mg/kg range, with plasma 14C concentration vs. time plots for oral doses of EHD resembling the iv time-course data. This resulted from a very rapid absorption phase (5.5 min t1/2), with plasma 14C levels for both dose levels decreasing in a biexponential manner. The major route of excretion after peroral doses was in urine, making this mode of excretion consistent for both routes of administration evaluated in this study and including the doses given in previous iv work. Kinetic analysis confirmed that this route of excretion was first-order. HPLC analysis of urine from both routes demonstrated that EHD was metabolized and excreted as at least two major, water-soluble urinary metabolites; these metabolites were not identified in this investigation. No unmetabolized EHD was detected in urine, indicating that EHD may be completely metabolized in the rat. Overall, EHD was absorbed, distributed, metabolized, and eliminated from the Fischer rat in a first-order manner following either cutaneous or peroral doses. The results of this study indicate that the kinetic patterns observed experimentally will be dose-proportional for doses administered in the range of 1.5-150 mg/kg.


Change blindness and the primacy of object appearance.
A large body of work suggests that the visual system is particularly sensitive to the appearance of new objects. This is based partly on evidence from visual search studies showing that onsets capture attention whereas many other types of visual event do not. Recently, however, the notion that object onset has a special status in visual attention has been challenged. For instance, an object that looms toward an observer has also been shown to capture attention. In two experiments, we investigated whether onset receives processing priority over looming. Observers performed a change detection task in which one of the display objects either loomed or receded, or a new object appeared. Results showed that looming objects were more resistant to change blindness than receding objects. Crucially, however, the appearance of a new object was less susceptible to change blindness than both looming and receding. We argue that the visual system is particularly sensitive to object onsets.


Secretoneurin gene therapy improves blood flow in an ischemia model in type 1 diabetic mice by enhancing therapeutic neovascularization.
Deficient angiogenesis after ischemia may contribute to worse outcome of peripheral arterial disease in patients with diabetes mellitus. Based on our previous work where we demonstrated that Secretoneurin (SN) is up-regulated under hypoxic conditions and enhances angiogenesis, we analyzed the therapeutic potential of SN gene therapy using a model of severe hind limb ischemia in streptozotocin-induced diabetic mice (STZ-DM). After induction of hind limb ischemia, blood flow was assessed by means of laser Doppler perfusion imaging (LDPI) and increased blood perfusion in the SN-treated animal group was observed. These results were complemented by the clinical observation of reduced necrosis and by an increased number of capillaries and arterioles in the SN-treated animal group. In vitro, we found that SN is capable of promoting proliferation and chemotaxis and reduces apoptosis in HUVECs cultured under hyperglycemic conditions. Additionally, SN activated ERK, eNOS and especially AKT as well as EGF-receptor in hyperglycemic HUVECs. In conclusion, we show that SN gene therapy improves post-ischemic neovascularization in diabetic mice through stimulation of angiogenesis and arteriogenesis indicating a possible therapeutic role of this factor in ischemia-related diseases in diabetic patients.


A polymerase chain reaction method for identification of five major meloidogyne species.
A polymerase chain reaction (PCR) method for discriminating Meloidogyne incognita, M. arenaria, M. javanica, M. hapla, and M. chitwoodi was developed. Single juveniles were ruptured in a drop of water and added directly to a PCR reaction mixture in a microcentrifuge tube. Primer annealing sites were located in the 3' portion of the mitochondrial gene coding for cytochrome oxidase subunit II and in the 16S rRNA gene. Following PCR amplification, fragments of three sizes were detected. The M. incognita and M. javanica reactions produced a 1.7-kb fragment; the M. arenaria reaction, a 1.1-kb fragment; and the M. hapla and M. chitwoodi reactions resulted in a 0.52-kb fragment. Digestion of the amplified product with restriction endonucleases allowed discrimination among species with identically sized amplification products. Dra I digestions of the 0.52-kb amplification product produced a characteristic three-banded pattern in M. chitwoodi, versus a two-banded pattern in M. hapla. Hinf I digestion of the 1.7-kb fragment produced a two-banded pattern in M. javanica, versus a three-banded pattern in M. incognita. Amplification and digestion of DNA from juveniles from single isolates of M. marylandi, M. naasi, and M. nataliei indicated that the diagnostic application of this primer set may extend to less frequently encountered Meloidogyne species.


Treatment of superficial cutaneous vascular lesions: experience with the KTP 532 nm laser.
Whilst most facial telangiectasias respond well to short-pulse-duration pulsed dye laser therapy, studies have shown that for the treatment of larger vessels these short-duration pulses are sub-optimal. Long-pulse frequency-doubled neodymium:YAG lasers have been introduced with pulse durations ranging from 1-50 ms and treatment beam diameters of up to 4 mm. We report the results of KTP/532 nm laser treatment for superficial vascular skin lesions. The aim was to determine the efficacy of the KTP/532 nm laser in the treatment of superficial cutaneous vascular lesions at a regional dermatology centre in a 2 year retrospective analysis. Patients were referred from general dermatology clinics to a purpose-built laser facility. A test dose was performed at the initial consultation and thereafter patients were reviewed and treated at 6 week intervals. Outcome was graded into five classifications by the patient and operator independently based on photographic records: clear, marked improvement, partial response, poor response, and no change or worsening. Over the 2 year period, 204 patients with 246 diagnoses were treated [156 female; median age 41 (range 1-74) years; Fitzpatrick skin types I-III]. Equal numbers of spider angioma (102) and facial telangiectasia (102) were treated. Of those patients who completed treatment and follow up, 57/58 (98%) of spider angiomas and 44/49 (90%) of facial telangiectasia markedly improved or cleared. Satisfactory treatment outcomes, with one clearance and two partial responses, occurred in three of five patients with port-wine stain. Few patients experienced adverse effects: two declined further treatment due to pain, and a small area of minimal superficial scarring developed in one case. Two patients developed mild persistent post-inflammatory hyperpigmentation, and one subject experienced an episode of acute facial erythema, swelling and blistering after one treatment. The KTP/532 nm frequency-doubled neodymium:YAG laser is a safe and effective treatment for common superficial cutaneous vascular lesions in patients with Fitzpatrick skin types I-III.


Atypical adenomatous hyperplasia (adenosis) of the prostate: a case report with review of the literature.
A 62-year-old male presented with symptoms of urinary obstruction and elevated serum prostate-specific antigen level of 3.61 ng/mL. Prostate needle biopsies showed benign prostatic tissue with a focus of crowded glands with minimal cytological atypia, fairly well-circumscribed with infiltrative appearance of glands at the edges. This focus had both small and larger glands with similar histological features. This focus was strongly positive for alpha-methylacyl-coenzyme A-racemase (AMACR), but showed scattered patchy staining with basal cell markers (p63 and CK903/34betaE12). Hence, the final histologic diagnosis was benign prostatic tissue with a focus of florid adenosis. Two subsequent follow-up prostate needle biopsies performed six and 12 months later both showed benign prostatic tissue with atrophic changes. This case highlights the utility of these three immunostains (AMACR, p63 and CK903/34betaE12) in the accurate diagnosis of adenosis of the prostate on needle biopsy, and avoiding its misinterpretation as prostate adenocarcinoma.


High resolution grating-assisted surface plasmon resonance fiber optic aptasensor.
A surface plasmon resonance biochemical sensor based on a tilted fiber Bragg grating imprinted in a single mode fiber core is demonstrated. A 30-50 nm thick gold coating on the cladding of the fiber provides the support for surface plasmon waves whose interaction with attached biomolecules is monitored at near infrared wavelengths near 1,550 nm. The transmission spectrum of the sensor provides a fine comb of narrowband resonances that overlap with the broader absorption of the surface plasmon and thus provide a unique tool to measure small shifts of the plasmon with high accuracy. The attachment on the gold surfaces of aptamers with specific affinities for proteins provides the required target-analyte system and is shown to be functional in the framework of our sensing device. The implementation of the sensor either as a stand-alone device or as part of a multi-sensor platform is also described.


Nonviral siRNA delivery to the lung: investigation of PEG-PEI polyplexes and their in vivo performance.
This study describes the physicobiological characterization of PEI- and PEG-PEI polyplexes containing partially 2'-OMe modified 25/27mer dicer substrate siRNAs (DsiRNAs) and their in vivo behavior regarding biodistribution and systemic bioavailability after pulmonary application as well as their ability to knock down gene expression in the lung. Biophysical characterization included circular dichroism of siRNA in polyplexes, condensation efficiency of polymers and in vitro stability. After in vivo application, biodistribution and kinetics of radiolabeled polyplexes were quantified and recorded over time in three-dimensional SPECT images and by end point scintillation counting. The influence on lung tissue and on the humoral and cellular immunosystem was investigated, and finally knockdown of endogenous gene expression in the lung was determined qualitatively. While all of the polymers used in our study were proven to effectively condense siRNA, stability of the complexes depended on the PEG grafting degree. Interestingly, PEI 25 kDa, which showed the least interaction with mucin or surfactant in vitro, performed poorly in vivo. Our nuclear imaging approach enabled us to follow biodistribution of the instilled nanocarriers over time and indicated that PEGylated nanocarriers are more suitable for lung application. While moderate proinflammatory effects were attributed to PEI25k-PEG(2k)(10) nanocarriers, none of the treatments caused histological abnormalities. Our preliminary in vivo knockdown experiment suggests that PEG-PEI/siRNA complexes are promising nanomedicines for pulmonary siRNA delivery. These results encouraged us to further investigate possible adverse effects and to quantify in vivo gene silencing in the lung after intratracheal instillation of PEG-PEI/siRNA complexes.


Efficacy of Intravitreal Injections of Triamcinolone Acetonide in a Rodent Model of Nonarteritic Anterior Ischemic Optic Neuropathy.
To investigate effects of intravitreal injections of triamcinolone acetonide (IVI-TA) at different times in a rodent model of nonarteritic anterior ischemic optic neuropathy (rAION). After inducing ischemic optic neuropathy, the rats received either IVI-TA (0.32 mg/2 μL) at 1 day (1d-TA), 1 week (7d-TA), 2 weeks (14d-TA), or PBS. The density of retinal ganglion cells (RGCs) was calculated using retrograde Fluorogold labeling. Electrophysiological visual function was assessed by flash visual evoked potentials (FVEPs). Apoptosis assays of the retinal sections and immunohistochemistry of ED1 staining of the optic nerves were performed. Four weeks postinfarct, the 1d- and 7d-TA groups had significantly rescued RGCs in the central (2160 ± 250 mm(2), P = 0.004; 2050 ± 660, P = 0.008, respectively) and midperipheral retinas (1240 ± 130; 1250 ± 220, respectively, both P = 0.004) compared with those of the PBS-treated rats. Flash visual evoked potentials demonstrated improvements in P1 amplitude (μV) in the 1d- and 7d-TA groups (both P < 0.05). Assays of TUNEL showed a decrease in the number of apoptotic cells in the RGC layers of 1d- and 7d-TA-treated retinas compared with the PBS-treated group (both P = 0.004). Cells ED1-positive were significantly decreased in the optic nerve (ON) of the 1d- and 7d-TA groups compared with the PBS group (P = 0.004 and 0.02, respectively). Within 1 week postinfarct of rAION, IVI-TA had neuroprotective effects on RGC survival with an increase in the electrophysiological amplitude of VEPs and a decrease in microglial infiltration in the ONs.


Multifocal pupillography in early age-related macular degeneration.
To investigate the potential of multifocal pupillographic objective perimetry to assess changes in retinal function with clinical severity of age-related macular degeneration (AMD). Pupil responses were recorded from 40 subjects with AMD and 23 normal control subjects (mean ± SD age, 71.3 ± 5.1 years). Age-related macular degeneration subjects were classified according to the Age-Related Eye Disease Study (AREDS) classification system and allocated into one of four AMD severity groups. Three multifocal pupillographic objective perimetry stimulus variants that were identical in luminance but varied in spatiotemporal sequence were used. In one of the three protocols, stimuli were presented with a pedestal flicker for 266 milliseconds at 15 Hz. On average, response amplitudes demonstrated a significant change in sensitivity with progression from early-stage (0.32 ± 0.08 dB, t = 3.88) to late-stage (-1.60 ± 0.12 dB, t = -12.7) age-related macular degeneration. Response delays followed a similar trend with the longest delays in AREDS4 (57.2 ± 1.9 milliseconds, t = 29.5). Ring analysis identified the largest mean effect on responses within the central 6 degrees of fixation. The NewStimuli protocol achieved the best diagnostic accuracy across all severity groups with area under the curve values of 0.85 ± 0.066 (AREDS1), 0.908 ± 0.085 (AREDS2), 0.929 ± 0.040 (AREDS3), and 1.0 ± 0.0 (AREDS4). The mean effect of AMD on contraction amplitudes and response delays reflected the severity of disease, and the NewStimuli protocol achieved good diagnostic accuracy across all AMD severity groups. Multifocal pupillographic objective perimetry may potentially be a useful method in monitoring progression of AMD and assessing change in retinal function with novel interventions in early AMD. Longitudinal studies are required to identify biomarkers that predict eyes at risk of progression.


The Genomic Impacts of Drift and Selection for Hybrid Performance in Maize.
Although maize is naturally an outcrossing organism, modern breeding utilizes highly inbred lines in controlled crosses to produce hybrids. The U.S. Department of Agriculture's reciprocal recurrent selection experiment between the Iowa Stiff Stalk Synthetic (BSSS) and the Iowa Corn Borer Synthetic No. 1 (BSCB1) populations represents one of the longest running experiments to understand the response to selection for hybrid performance. To investigate the genomic impact of this selection program, we genotyped the progenitor lines and >600 individuals across multiple cycles of selection using a genome-wide panel of ∼40,000 SNPs. We confirmed previous results showing a steady temporal decrease in genetic diversity within populations and a corresponding increase in differentiation between populations. Thanks to detailed historical information on experimental design, we were able to perform extensive simulations using founder haplotypes to replicate the experiment in the absence of selection. These simulations demonstrate that while most of the observed reduction in genetic diversity can be attributed to genetic drift, heterozygosity in each population has fallen more than expected. We then took advantage of our high-density genotype data to identify extensive regions of haplotype fixation and trace haplotype ancestry to single founder inbred lines. The vast majority of regions showing such evidence of selection differ between the two populations, providing evidence for the dominance model of heterosis. We discuss how this pattern is likely to occur during selection for hybrid performance and how it poses challenges for dissecting the impacts of modern breeding and selection on the maize genome.


Cytoprotective effect of 20S-Rg3 on benzo[a]pyrene-induced DNA damage.
Benzo[a]pyrene (BaP) is a polycyclic aromatic hydrocarbon ubiquitously existing in the environment. Its metabolites have been shown to cause DNA damage and cellular dysfunction in humans. Panax ginseng C.A. Meyer is a Chinese medicinal herb, and ginsenosides are the main active constituent of ginseng. Accumulating evidence had indicated that ginseng extract and ginsenosides possess cytoprotective effects. In this study, the protective effect of ginsenosides on BaP-induced DNA damage in human dermal fibroblasts (HDFs) and HepG2 cells was investigated. The genotoxic effect of BaP was measured by the comet assay. Results showed that tail moment was increased in BaP-treated cells, but cotreatment of ginsenoside 20(S)-Rg3 can significantly decrease BaP-induced DNA damage. A downstream mechanistic study revealed that 20(S)-Rg3 increased the gene expression of an important phase II detoxifying enzyme NAD(P)H:quinine oxidoreductase 1. The effect was also associated with the activation of protein kinase B (Akt) and nuclear translocation of nuclear factor erythroid 2-related factor 2 (Nrf2). These results indicated that 20(S)-Rg3 might protect HDFs from BaP-induced DNA damage through the activation of the phosphatidylinositol 3-kinase/Akt/Nrf2 pathway. Our results also demonstrated that 20(S)-Rg3 is a functional ligand of pregnane X receptor (PXR), a nuclear receptor that mediates the induction of drug clearance pathways. Subsequent knockdown of PXR expression by small interfering RNA confirmed the involvement of PXR on the protective effects of 20(S)-Rg3 against BaP-induced DNA damage. In summary, ginsenoside 20(S)-Rg3 can protect against BaP-induced genotoxicity in human cells, suggesting that ginseng may serve as a natural cytoprotective agent against environmental carcinogens.


Fluorescence recovery assay: a continuous assay for processive DNA polymerases applied specifically to DNA polymerase III holoenzyme.
A continuous assay was developed for processive DNA polymerases. The specific enzyme used to develop the assay was the most processive polymerase known, Escherichia coli DNA polymerase III holoenzyme. The assay was based upon the recovery of the intrinsic fluorescence of single-stranded DNA binding protein (SSB) as it was displaced from the DNA template during DNA synthesis. The intrinsic fluorescence of SSB was quenched by as much as 80% when it bound to single-stranded DNA. As the DNA was replicated, SSB was displaced and recovered its fluorescence. The amount of fluorescence recovered was directly proportional to the amount of DNA synthesized and was used to quantitate the rate of DNA synthesis. However, since 50 to 60 nucleotides must be replicated for every SSB tetramer released, the assay is expected to work best for processive DNA polymerases. The only requirement for using this assay with other DNA polymerases is that they be able to synthesize DNA on a template coated with SSB. The replication SSBs do not pose an obstacle to the assay because they all appear to have intrinsic fluorescence that is sensitive to their ssDNA-bound state.


Effects of a multi-nutrient supplement on exercise performance and hormonal responses to resistance exercise.
The purpose of this study was to determine the influence of a comprehensive multi-component nutritional supplement on performance, hormonal, and metabolic responses to an acute bout of resistance exercise. Nine healthy subjects ingested either Muscle Fuel (MF) or a matched placebo (PL) for 7 days. Subjects then reported to the laboratory, ingested the corresponding supplement, and performed two consecutive days of heavy resistance exercise testing with associated blood draws. MF supplementation improved vertical jump (VJ) power output and the number of repetitions performed at 80% of one repetition maximum (1RM). Additionally, MF supplementation potentiated growth hormone (GH), testosterone, and insulin-like growth factor-1 responses to exercise. Concentrations of circulating myoglobin and creatine kinase (CK) were attenuated immediately following resistance exercise during the MF trial, indicating that MF partially mediated some form of exercise-induced muscle tissue damage. In summary MF enhanced performance and hormonal responses associated with an acute bout of resistance exercise. These responses indicate that MF supplementation augments the quality of an acute bout of resistance exercise thereby increasing the endocrine signaling and recovery following heavy resistance exercise.


Adherence to bisphosphonates and hormone replacement therapy in a tertiary care setting of patients in the CANDOO database.
Therapies for osteoporosis must be taken for at least 1 year to be effective. The purpose of this study was to determine the difference in adherence to etidronate, alendronate and hormone replacement therapy in a group of patients seen at our tertiary care centres. The Canadian Database of Osteoporosis and Osteopenia (CANDOO), a prospective observational database designed to capture clinical data, was searched for patients who started therapy following entry into CANDOO. There were 1196 initiating etidronate, 477 alendronate and 294 hormone replacement therapy women and men aged (mean, SD) 65.8 (8.7) years in the study. A Cox proportional hazards regression model was used to assess differences between treatment groups in the time to discontinuation of therapy. Several potential covariates such as anthropometry, medications, illnesses, fractures and lifestyle factors were entered into the model. A forward selection technique was used to generate the final model. Hazard ratios and 95% confidence intervals (CI) were calculated. Adjusted results indicated that alendronate-treated patients were more likely to discontinue therapy as compared with etidronate-treated patients (1.404; 95% CI: 1.150, 1.714). After 1 year, 90.3% of patients were still taking etidronate compared with 77.6% for alendronate. No statistically significant differences were found between hormone replacement therapy and etidronate users (0.971; 95% CI: 0.862, 1.093) and hormone replacement therapy and alendronate users (0.824; 95% CI: 0.624, 1.088) after controlling for potential covariates. After 1 year, 80.1% of patients were still taking hormone replacement therapy, which decreased to 44.5% after 6 years. Increasing age and presence of incident non-vertebral fractures were found to be independent predictors of adherence. In conclusion, alendronate users were more likely to discontinue therapy than etidronate users over the follow-up period. Potential barriers to long-term patient adherence to osteoporosis therapies need to be evaluated.


Advances in the Evaluation of Longitudinal Construct Validity of Clinical Outcome Assessments.
Historically, the assessment of longitudinal construct validity in the field of psychosocial measurement involved defining hypotheses and calculating correlation coefficients using scores based on 2 measures at 2 or more time points. In the context of patient-reported outcomes, this evolved into sensitivity to change and responsiveness, including the computation of effect size estimates of change, standardized response means, and indices such as Guyatt's statistic. Cross-sectional analyses or analyses based on 2 time points have been the standard practice. Evolving conceptualizations have incorporated more than 2 time points and have included depictions of individual trajectories of change in multiple measures, structural equation models, and mixed modeling techniques. The focus of this article is on methods to evaluate longitudinal construct validity. We describe a sample of these methods and provide considerations and recommendations for designing a thoughtful longitudinal construct validity evaluation of clinical outcome assessments.


Two distinct DNA ligase activities in mitotic extracts of the yeast Saccharomyces cerevisiae.
Four biochemically distinct DNA ligases have been identified in mammalian cells. One of these enzymes, DNA ligase I, is functionally homologous to the DNA ligase encoded by the Saccharomyces cerevisiae CDC9 gene. Cdc9 DNA ligase has been assumed to be the only species of DNA ligase in this organism. In the present study we have identified a second DNA ligase activity in mitotic extracts of S. cerevisiae with chromatographic properties different from Cdc9 DNA ligase, which is the major DNA joining activity. This minor DNA joining activity, which contributes 5-10% of the total cellular DNA joining activity, forms a 90 kDa enzyme-adenylate intermediate which, unlike the Cdc9 enzyme-adenylate intermediate, reacts with an oligo (pdT)/poly (rA) substrate. The levels of the minor DNA joining activity are not altered by mutation or by overexpression of the CDC9 gene. Furthermore, the 90 kDa polypeptide is not recognized by a Cdc9 antiserum. Since this minor species does not appear to be a modified form of Cdc9 DNA ligase, it has been designated as S. cerevisiae DNA ligase II. Based on the similarities in polynucleotide substrate specificity, this enzyme may be the functional homolog of mammalian DNA ligase III or IV.


Iodide as a fluorescence quencher and promoter--mechanisms and possible implications.
In this work, fluorescence correlation spectroscopy (FCS) was used to investigate the effects of potassium iodide (KI) on the electronic-state population kinetics of a range of organic dyes in the visible wavelength range. Apart from a heavy atom effect promoting intersystem crossing to the triplet states in all dyes, KI was also found to enhance the triplet-state decay rate by a charge-coupled deactivation. This deactivation was only found for dyes with excitation maximum in the blue range, not for those with excitation maxima at wavelengths in the green range or longer. Consequently, under excitation conditions sufficient for triplet state formation, KI can promote the triplet state buildup of one dye and reduce it for another, red-shifted dye. This anticorrelated, spectrally separable response of two different dyes to the presence of one and the same agent may provide a useful readout for biomolecular interaction and microenvironmental monitoring studies. In contrast to the typical notion of KI as a fluorescence quencher, the FCS measurements also revealed that when added in micromolar concentrations KI can act as an antioxidant, promoting the recovery of photo-oxidized fluorophores. However, in millimolar concentrations KI also reduces intact, fluorescently viable fluorophores to a considerable extent. In aqueous solutions, for the dye Rhodamine Green, an optimal concentration of KI of approximately 5 mM can be defined at which the fluorescence signal is maximized. This concentration is not high enough to allow full triplet state quenching. Therefore, as a fluorescence enhancement agent, it is primarily the antioxidative properties of KI that play a role.


Metabolite profiling of Arabidopsis inoculated with Alternaria brassicicola reveals that ascorbate reduces disease severity.
The interaction between the pathogenic ascomycete Alternaria brassicicola and Arabidopsis was investigated by metabolite profiling. The effect of A. brassicicola challenge on metabolite levels was substantial, with nearly 50% of detected compounds undergoing significant changes. Mutations blocking ethylene, jasmonic acid, or ethylene signaling had little effect on metabolite levels. The effects of altering levels of some metabolites were tested by exogenous application during A. brassicicola inoculation. Gamma amino-butyric acid (GABA) or xylitol promoted, while trehalose and ascorbate inhibited, disease severity. GABA promoted, and ascorbate strongly inhibited, fungal growth in culture. Arabidopsis vtc1 and vtc2 mutants, that have low levels of ascorbate, were more susceptible to A. brassicicola. Ascorbate levels declined following A. brassicicola inoculation while levels of dehydroascorbate increased, resulting in a shift of the redox balance between these compounds in the direction of oxidation. These results demonstrate that ascorbate is an important component of resistance to this pathogen.


Environmental Panels as a Proxy for Nursing Facility Patients With Methicillin-Resistant Staphylococcus aureus and Vancomycin-Resistant Enterococcus Colonization.
Most nursing facilities (NFs) lack methicillin-resistant Staphylococcus aureus (MRSA) and vancomycin-resistant Enterococcus (VRE) surveillance programs due to limited resources and high costs. We investigated the utility of environmental screening of high-touch surfaces in patient rooms as a way to circumvent these challenges. We compared MRSA and VRE culture data from high-touch surfaces in patients' rooms (14450 samples from 6 NFs) and ranked each site's performance in predicting patient colonization (7413 samples). The best-performing sites were included in a MRSA- and a VRE-specific panel that functioned as a proxy for patient colonization. Molecular typing was performed to confirm available concordant patient-environment pairs. We identified and validated a MRSA panel that consisted of the bed controls, nurse call button, bed rail, and TV remote control. The VRE panel included the toilet seat, bed controls, bed rail, TV remote control, and top of the side table. Panel colonization data tracked patient colonization. Negative predictive values were 89%-92% for MRSA and 82%-84% for VRE. Molecular typing confirmed a strong clonal type relationship in available concordant patient-environment pairs (98% for MRSA, 91% for VRE), pointing to common epidemiological patterns for environmental and patient isolates. Environmental panels used as a proxy for patient colonization and incorporated into facility surveillance protocols can guide decolonization strategies, improve awareness of MRSA and VRE burden, and inform efforts to reduce transmission. Targeted environmental screening may be a viable surveillance strategy for MRSA and VRE detection in NFs.


Parasite-derived plasma microparticles contribute significantly to malaria infection-induced inflammation through potent macrophage stimulation.
There is considerable debate as to the nature of the primary parasite-derived moieties that activate innate pro-inflammatory responses during malaria infection. Microparticles (MPs), which are produced by numerous cell types following vesiculation of the cellular membrane as a consequence of cell death or immune-activation, exert strong pro-inflammatory activity in other disease states. Here we demonstrate that MPs, derived from the plasma of malaria infected mice, but not naive mice, induce potent activation of macrophages in vitro as measured by CD40 up-regulation and TNF production. In vitro, these MPs induced significantly higher levels of macrophage activation than intact infected red blood cells. Immunofluorescence staining revealed that MPs contained significant amounts of parasite material indicating that they are derived primarily from infected red blood cells rather than platelets or endothelial cells. MP driven macrophage activation was completely abolished in the absence of MyD88 and TLR-4 signalling. Similar levels of immunogenic MPs were produced in WT and in TNF(-/-), IFN-gamma(-/-), IL-12(-/-) and RAG-1(-/-) malaria-infected mice, but were not produced in mice injected with LPS, showing that inflammation is not required for the production of MPs during malaria infection. This study therefore establishes parasitized red blood cell-derived MPs as a major inducer of systemic inflammation during malaria infection, raising important questions about their role in severe disease and in the generation of adaptive immune responses.


Ubiquitylation and isgylation: overlapping enzymatic cascades do the job.
Ubiquitylation-that is, the covalent attachment of ubiquitin polypeptides to target proteins-involves the sequential action of the E1 ubiquitin-activating enzyme, E2 ubiquitin-conjugating enzymes, and E3 ubiquitin-protein ligases. Similarly, the conjugation of ubiquitin-like proteins is thought to occur through parallel, but nonidentical, cascades. This concept of strict separation of these modification pathways is now being challenged by the evidence, showing that there are enzymes that play a role both in ubiquitylation and isgylation.


Antigen receptor proximal signaling in splenic B-2 cell subsets.
Splenic marginal zone (MZ) and follicular mantle (FO) B cells differ in their responses to stimuli in vitro and in vivo. We have previously shown that MZ cells exhibit greater calcium responses after ligation of membrane IgM (mIgM). We have now investigated the molecular mechanism underlying the difference in calcium responses following ligation of mIgM and studied the response to total B cell receptor ligation in these two subsets. We compared key cellular proteins involved in calcium signaling in MZ and FO cells. Tyrosine phosphorylation and activity of phospholipase C-gamma 2 and Syk protein tyrosine kinase were significantly higher in MZ cells than in FO cells after mIgM engagement, providing a likely explanation for our previous findings. Tyrosine phosphorylation of CD22 and expression of Src homology 2-containing inositol phosphatase and Src homology 2-containing protein tyrosine phosphatase-1 were also higher in the MZ cells. Expression and tyrosine phosphorylation of Btk, BLNK, Vav, or phosphatidylinositol 3-kinase were equivalent. In contrast, stimulation with anti-kappa induced equivalent increases in calcium and activation of Syk in the two subsets. These signals were also equivalent in cells from IgM transgenic, J(H) knockout mice, which have equivalent levels of IgM in both subsets. With total spleen B cells, Btk was maximally phosphorylated at a lower concentration of anti-kappa than Syk. Thus, calcium signaling in the subsets of mature B cells reflects the amount of Ig ligated more than the isotype or the subset and this correlates with the relative tyrosine phosphorylation of Syk.


First Report of Lasiodiplodia citricola and Neoscytalidium dimidiatum Causing Death of Graft Union of English Walnut in California.
California produces 99% of the English walnuts (Juglans regia) in the USA. In August 2012 in Tulare County, about 5,000 out of 90,000 trees were killed in a walnut nursery by a distinct black canker that developed around the graft union. The cankers appeared to be initiated at the heading cut on the rootstock, and expanded down to the rootstock and through the budded union up to the scion, resulting in mortality of scion. The walnut nursery was located adjacent to a mature walnut orchard. The fungi isolated from the cankers were identified as Lasiodiplodia citricola and Neoscytalidium dimidiatum based on morphological characteristics and DNA sequence comparisons. L. citricola was isolated from one of the 10 graft unions, while N. dimidiatum from the other nine. L. citricola isolates were characterized by white, aerial mycelium on potato dextrose agar that turned gray after 4 days and produced ellipsoidal to ovoid hyaline one-celled conidia that became 2-celled and brown with thick walls and longitudinal striations in the wall (1). N. dimidiatum isolates were characterized by ellipsoid to ovoid, hyaline conidia with a truncate base and an acutely rounded apex, initially 1-celled, and some becoming brown and 2-celled at maturity; no muriform conidia were produced (3). These identifications were confirmed by analyses of the ITS, BT2, and TEF-1α gene regions. The three gene regions were amplified by using the primers and methods described in (4). For L. citricola (isolates 7E78 to 7E80), a DNA sequence BLASTn at GenBank showed 100% identity with accessions GU945354 (ITS) and GU945340 (TEF-1α) of the ex-type specimen (CBS124707, BT2 sequencing data was not available) (3). For N. dimidiatum (isolates 7E61 to 7E63), a BLASTn search showed a high identify (ITS, 100%; BT2, 99%; TEF-1α, 99%) with reference sequence of N. dimidiatum (ITS, GQ330903; BT2, GU251768; TEF-1α, GU251240). Sequences of the studied DNA regions were deposited to GenBank as KC357298 to KC357303 (ITS); KC357304 to KC357309 (BT2); and, KC357310 to KC357315 (TEF-1α). The pathogenicity of L. citricola in comparison with N. dimidiatum in J. regia cvs. Chandler, Tulare, and Vina was evaluated in an orchard at KARE, by using two isolates each of L. citricola (7E78, 7E80) and N. dimidiatum (7E61, 7E63). Pathogenicity tests were performed by inoculating ten 2-year-old branches per isolate in late September 2012 by the method described in (2). After 3 weeks, the average lesion lengths caused by L. citricola on Chandler, Tulare, and Vina were 152, 156, and 188 mm, respectively, and 32, 38, and 34 mm, respectively, caused by N. dimidiatum. The lesion length averages produced on the three cultivars by the four isolates were all significantly (P < 0.05) longer than their respective controls (average length 10 mm on all cultivars). L. citricola, but not N. dimidiatum, killed branches of Chandler, Tulare, and Vina in 10 days. Both L. citricola and N. dimidiatum were reisolated from the inoculated branches and no fungus was isolated from the control, confirming Koch's postulates. These results confirmed that the walnut graft union canker was caused by either L. citricola or N. dimidiatum. To our knowledge, this is the first report of death of newly grafted walnut trees caused by L. citricola and N. dimidiatum worldwide, and also the first report of L. citricola infecting walnut worldwide. References: (1) J. Abdollahzadeh et al. Persoonia. 25:1, 2010. (2) S. F. Chen et al. Plant Dis. 97:994, 2013. (3) D. Pavlic et al. Mycologia. 106:851, 2008. (4) B. Slippers et al. Mycologia. 96:83, 2004.


Genetic Allee effects and their interaction with ecological Allee effects.
It is now widely accepted that genetic processes such as inbreeding depression and loss of genetic variation can increase the extinction risk of small populations. However, it is generally unclear whether extinction risk from genetic causes gradually increases with decreasing population size or whether there is a sharp transition around a specific threshold population size. In the ecological literature, such threshold phenomena are called 'strong Allee effects' and they can arise for example from mate limitation in small populations. In this study, we aim to (i) develop a meaningful notion of a 'strong genetic Allee effect', (ii) explore whether and under what conditions such an effect can arise from inbreeding depression due to recessive deleterious mutations, and (iii) quantify the interaction of potential genetic Allee effects with the well-known mate-finding Allee effect. We define a strong genetic Allee effect as a genetic process that causes a population's survival probability to be a sigmoid function of its initial size. The inflection point of this function defines the critical population size. To characterize survival-probability curves, we develop and analyse simple stochastic models for the ecology and genetics of small populations. Our results indicate that inbreeding depression can indeed cause a strong genetic Allee effect, but only if individuals carry sufficiently many deleterious mutations (lethal equivalents). Populations suffering from a genetic Allee effect often first grow, then decline as inbreeding depression sets in and then potentially recover as deleterious mutations are purged. Critical population sizes of ecological and genetic Allee effects appear to be often additive, but even superadditive interactions are possible. Many published estimates for the number of lethal equivalents in birds and mammals fall in the parameter range where strong genetic Allee effects are expected. Unfortunately, extinction risk due to genetic Allee effects can easily be underestimated as populations with genetic problems often grow initially, but then crash later. Also interactions between ecological and genetic Allee effects can be strong and should not be neglected when assessing the viability of endangered or introduced populations.


Orientation of glycoprotein galactosyltransferase and sialyltransferase enzymes in vesicles derived from rat liver Golgi apparatus.
UDP-galactose: N-acetylglucosamine galactosyltransferase (GT) and CMP-sialic:desialylated transferrin sialyltransferse (ST) activities of rat liver Golgi apparatus are membrane-bound enzymes that can be released by treatment with Triton X-100. When protein substrates are used to assay these enzymes in freshly prepared Golgi vesicles, both activities are enhanced about eightfold by the addition of Triton X-100. When small molecular weight substrates are used, however, both activities are only enhanced about twofold by the addition of detergent. The enzymes remain inaccessible to large protein substrates even after freezing and storage of the Golgi preparation for 2 mo in liquid nitrogen. Accessibility to small molecular and weight substrates increases significantly after such storage. GT and ST activities in Golgi vesicles are not destroyed by treatment with trypsin, but are destroyed by this treatment if the vesicles are first disrupted with Triton X-100. Treatment of Golgi vesicles with low levels of filipin, a polyene antibiotic known to complex with cholesterol in biological membranes, also results in enhanced trypsin susceptibility of both glycosyltransferases. Maximum destruction of the glycosyltransferase activities by trypsin is obtained at filipin to total cholesterol weight ratios of approximately 1.6 or molar ratios of approximately 1. This level of filipin does not solubilize the enzymes but causes both puckering of Golgi membranes visible by electron microscopy and disruption of the Golgi vesicles as measured by release of serum albumin. When isolated Golgi apparatus is fixed with glutaraldehyde to maintain the three-dimensional orientation of cisternae and secretory vesicles, and then treated with filipin, cisternal membranes on both cis and trans faces of the apparatus as well as secretory granule membranes appear to be affected about equally. These results indicate that liver Golgi vesicles as isolated are largely oriented with GT and ST on the luminal side of the membranes, which corresponds to the cisternal compartment of the Golgi apparatus in the hepatocyte. Cholesterol is an integral part of the membrane of the Golgi apparatus and its distribution throughout the apparatus is similar to that of both transferases.


Directing visual attention with spatially informative and spatially noninformative tactile cues.
We investigated the tactile cuing of visual spatial attention using spatially-informative (75% valid) and spatially-noninformative (25% valid) tactile cues. The participants performed a visual change detection task following the presentation of a tactile spatial cue on their back whose location corresponded to one of the four visual quadrants on a computer monitor. The participants were explicitly instructed to use the spatially-informative tactile cues but to ignore the spatially-noninformative cues. In addition to reaction time data, participants' eye-gaze was monitored as a measure of overt visual attention. The results showed that the spatially-informative tactile cues resulted in initial saccades toward the cued visual quadrants, and significantly reduced the visual change detection latencies. When spatially-noninformative tactile cues were used, the participants were largely successful at ignoring them as indicated by a saccade distribution that was independent of the quadrant that was cued, as well as the lack of a significant change in search time as compared to the baseline measure of no tactile cuing. The eye-gaze data revealed that the participants could not always completely ignore the spatially-noninformative tactile cues. Our results suggest that the tactile cuing of visual attention is natural but not automatic when the tactile cue and visual target are not collocated spatially, and that it takes effort to ignore the cues even when they are known to provide no useful information. In addition, our results confirm previous findings that spatially-informative tactile cues are especially effective at directing overt visual attention to locations that are not typically monitored visually, such as the bottom of a computer screen or the rearview mirror in an automobile.


Consumption of fructose-sweetened beverages for 10 weeks increases postprandial triacylglycerol and apolipoprotein-B concentrations in overweight and obese women.
Fructose consumption in the USA has increased over the past three decades. During this time, obesity, insulin resistance and the metabolic syndrome have also increased in prevalence. While diets high in fructose have been shown to promote insulin resistance and increase TAG concentrations in animals, there are insufficient data available regarding the long-term metabolic effects of fructose consumption in humans. The objective of the present study was to investigate the metabolic effects of 10-week consumption of fructose-sweetened beverages in human subjects under energy-balanced conditions in a controlled research setting. Following a 4-week weight-maintaining complex carbohydrate diet, seven overweight or obese (BMI 26.8-33.3 kg/m2) postmenopausal women were fed an isoenergetic intervention diet, which included a fructose-sweetened beverage with each meal, for 10 weeks. The intervention diet provided 15 % of energy from protein, 30 % from fat and 55 % from carbohydrate (30 % complex carbohydrate, 25 % fructose). Fasting and postprandial glucose, insulin, TAG and apoB concentrations were measured. Fructose consumption increased fasting glucose concentrations and decreased meal-associated glucose and insulin responses (P = 0.0002, P = 0.007 and P = 0.013, respectively). Moreover, after 10 weeks of fructose consumption, 14 h postprandial TAG profiles were significantly increased, with the area under the curve at 10 weeks being 141 % higher than at baseline (P = 0.04). Fructose also increased fasting apoB concentrations by 19 % (P = 0.043 v. baseline). In summary, consumption of fructose-sweetened beverages increased postprandial TAG and fasting apoB concentrations, and the present results suggest that long-term consumption of diets high in fructose could lead to an increased risk of CVD.


Central vasopressin raises arterial pressure by sympathetic activation and vasopressin release.
Although central administration of arginine vasopressin (AVP) has been reported to increase arterial pressure mediated by activation of the sympathetic system, we found that peripheral blockade of sympathetic transmission did not attenuate this pressor response. To elucidate the mechanism, rats were pretreated with either phentolamine (3 mg/kg), chlorisondamine (2.5 mg/kg), a vasopressin V1 receptor antagonist d(CH2)5Tyr(Me)AVP (AVP-X) (10 micrograms/kg), or the combinations of phentolamine and AVP-X or chlorisondamine and AVP-X. The pressor response to intracerebroventricular injection of AVP in unrestrained conscious rats was reduced but not significantly altered by intravenous injection of phentolamine or AVP-X; however, combined treatment with these agents abolished the response. To determine that the amount of central AVP leaked to the periphery did not contribute to the pressor effect, tritiated AVP and AVP (100 ng total) were injected intracerebroventricularly. Blood samples collected at 0, 3, and 30 minutes after injection showed that radioactivity in plasma was primarily metabolites and that the amount of intact AVP estimated to leak from the brain was too low to produce a pressor effect. Comparative regional hemodynamic studies between intracerebroventricular and intravenous injection of AVP performed in conscious rats instrumented with Doppler flow probes demonstrated a qualitatively similar pattern of increased resistance in the renal, mesenteric, and hindquarters beds. These data suggest that central pressor action of AVP is mediated by both activation of the sympathetic system and release of AVP.


Temperature dependence of anisotropic diffraction in holographic polymer-dispersed liquid crystal memory.
Grating devices using photosensitive organic materials play an important role in the development of optical and optoelectronic systems. High diffraction efficiency and polarization dependence achieved in a holographic polymer-dispersed liquid crystal (HPDLC) grating are expected to provide polarization-controllable optical devices, such as a holographic memory for optically reconfigurable gate arrays (ORGAs). However, the optical property is affected by the thermal modulation around the transition temperature (T(ni)) where the liquid crystal (LC) changes from nematic to isotropic phases. The temperature dependence of the diffraction efficiency in HPDLC grating is investigated using four types of LC composites comprised of LCs and monomers having different physical properties such as T(ni) and anisotropic refractive indices. The holographic memory formed by the LC with low anisotropic refractive index and LC diacrylate monomer implements optical reconfiguration for ORGAs at a high temperature beyond T(ni) of LC.


Using the behaviour change technique taxonomy v1 (BCTTv1) to identify the active ingredients of pharmacist interventions to improve non-hospitalised patient health outcomes.
The aim of this study was to identify which behaviour change techniques (BCTs) were present in intervention and control groups of randomised controlled trials (RCTs) included in a Cochrane systematic review. The RCTs included were conducted in community, primary and/or ambulatory-care settings. The data set was derived from 86 RCTs from an interim update of the Cochrane review of the effectiveness of pharmacist services on non-hospitalised patient outcomes. The primary outcome was the identification of BCTs scheduled for delivery in intervention and control groups of the RCTs. The secondary outcome measure was to identify which BCTs are not being utilised in intervention and control groups of the RCTs. The intervention and control groups included 31 and 12 BCTs, respectively. The number of identifiable BCTs/study ranged from 0 to 12 in the intervention groups (mean 3.01 (SD 2.4)) and 0 to 6 in the control groups (mean 0.38 (SD 0.84)). The most commonly identified BCTs in the intervention groups were: instruction on how to perform the behaviour (55%, n=47) (also the most common BCT in control groups); problem solving (29%, n=25); information about health consequences (24%, n=21); social support (practical) (24%, n=21); and social support (unspecified) (23%, n=20) (the second most common BCT in control groups). Thirteen trials had no identifiable BCTs in either group. The pharmacist interventions presented in this study did not use the full range of available BCTs. Furthermore, the reporting of BCTs was incomplete for both intervention and control groups, thereby limiting the utility and reproducibility of the interventions. Future interventions should be designed and reported using relevant taxonomies and checklists for example, BCT taxonomy and TIDieR (the template for intervention description and replication).


Neighborhood context and the transition to sexual activity among young black women.
Previous studies report that neighborhood characteristics influence pregnancy and childbearing risk among African-American adolescent women. These studies, however, leave unidentified the effects of many neighborhood properties on the proximate determinants of nonmarital fertility. In this study I examine the effects of neighborhood characteristics on the risk of nonmarital first intercourse and on contraceptive use among black female adolescents. The results suggest that neighborhood socioeconomic status, female employment and marital dissolution rates, and peers' departure from mainstream lifecourse trajectories influence young black women's sexual and contraceptive behavior. The effects of female employment and socioeconomic status are greater for teens in urban neighborhoods than for teens living elsewhere.


The Human Gut Microbiome as a Transporter of Antibiotic Resistance Genes between Continents.
Previous studies of antibiotic resistance dissemination by travel have, by targeting only a select number of cultivable bacterial species, omitted most of the human microbiome. Here, we used explorative shotgun metagenomic sequencing to address the abundance of >300 antibiotic resistance genes in fecal specimens from 35 Swedish students taken before and after exchange programs on the Indian peninsula or in Central Africa. All specimens were additionally cultured for extended-spectrum beta-lactamase (ESBL)-producing enterobacteria, and the isolates obtained were genome sequenced. The overall taxonomic diversity and composition of the gut microbiome remained stable before and after travel, but there was an increasing abundance of Proteobacteria in 25/35 students. The relative abundance of antibiotic resistance genes increased, most prominently for genes encoding resistance to sulfonamide (2.6-fold increase), trimethoprim (7.7-fold), and beta-lactams (2.6-fold). Importantly, the increase observed occurred without any antibiotic intake. Of 18 students visiting the Indian peninsula, 12 acquired ESBL-producing Escherichia coli, while none returning from Africa were positive. Despite deep sequencing efforts, the sensitivity of metagenomics was not sufficient to detect acquisition of the low-abundant genes responsible for the observed ESBL phenotype. In conclusion, metagenomic sequencing of the intestinal microbiome of Swedish students returning from exchange programs in Central Africa or the Indian peninsula showed increased abundance of genes encoding resistance to widely used antibiotics.


High-throughput sequencing of a South American Amerindian.
The emergence of next-generation sequencing technologies allowed access to the vast amounts of information that are contained in the human genome. This information has contributed to the understanding of individual and population-based variability and improved the understanding of the evolutionary history of different human groups. However, the genome of a representative of the Amerindian populations had not been previously sequenced. Thus, the genome of an individual from a South American tribe was completely sequenced to further the understanding of the genetic variability of Amerindians. A total of 36.8 giga base pairs (Gbp) were sequenced and aligned with the human genome. These Gbp corresponded to 95.92% of the human genome with an estimated miscall rate of 0.0035 per sequenced bp. The data obtained from the alignment were used for SNP (single-nucleotide) and INDEL (insertion-deletion) calling, which resulted in the identification of 502,017 polymorphisms, of which 32,275 were potentially new high-confidence SNPs and 33,795 new INDELs, specific of South Native American populations. The authenticity of the sample as a member of the South Native American populations was confirmed through the analysis of the uniparental (maternal and paternal) lineages. The autosomal comparison distinguished the investigated sample from others continental populations and revealed a close relation to the Eastern Asian populations and Aboriginal Australian. Although, the findings did not discard the classical model of America settlement; it brought new insides to the understanding of the human population history. The present study indicates a remarkable genetic variability in human populations that must still be identified and contributes to the understanding of the genetic variability of South Native American populations and of the human populations history.


Isopolymolybdate-based inorganic-organic hybrid compounds constructed by multidentate N-donor ligands: syntheses, structures and properties.
Taking advantage of isopolymolybdate [(NH4)6Mo7O24·4H2O] as a POM building block with multidentate N-donor molecules (tris[(2-pyridyl)methyl]amine = TPMA, 1-(tetrazo-5-yl)-3-(triazo-1-yl)benzene = 1,3-ttb) as organic ligands, five different inorganic-organic hybrid compounds were synthesized and characterized by single-crystal X-ray diffraction, infrared spectroscopy (IR) and powder X-ray diffraction (PXRD). Compound 1 is a Mo(vi)-based structure in which the TPMA ligand shows a tridentate coordination mode, which bears an uncoordinated pyridyl arm, and can further react with transition metals. Compound 2 exhibits a Cu(ii)-based zero-dimensional (0D) structure consisting of the TPMA ligand and [β-Mo8O26] polyanion. Compounds 3 and 4 have the same molecular formula with slight structural differences; both have the Co(ii)-based one-dimensional (1D) chain structure. Compared to the β-Mo8O26 polyanion in compound 3, the [Mo8O28]8- units condense via sharing two common terminal O atoms to form infinite [Mo8O26]n4n- chains in compound 4. Compound 5 is a Cu(ii)-based two-dimensional (2D) layer structure consisting of TPMA and 1,3-ttb mixed ligands and [β-Mo8O26]. In addition, the electrochemical properties and photocatalytic activities for the degradation of two types of organic dyes were explored for several compounds.


Nonoperative Treatment of Rotator Cuff Disease With Platelet-Rich Plasma: A Systematic Review of Randomized Controlled Trials.
To perform a systematic review of randomized controlled trials on the use of platelet-rich plasma (PRP) for nonoperative treatment of rotator cuff disease. Using Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines, 2 reviewers independently screened the MEDLINE, Embase, and Cochrane Library databases for prospective, randomized controlled trials comparing PRP with a control in the nonoperative treatment of chronic rotator cuff disease for inclusion. Clinical data were extracted and evaluated. The quality of evidence was assessed using The Cochrane Collaboration risk-of-bias tool. Five randomized controlled trials met the inclusion criteria, with 108 patients treated with PRP and 106 treated with a control. The mean age was 53.7 years, and 61.6% of patients were female patients. All of the studies found that the groups receiving PRP injections experienced improved clinical outcomes at final follow-up compared with baseline. Two studies found that PRP resulted in improved outcomes, mostly pain scores, compared with a control. One study compared PRP with formal exercise versus a saline solution injection with formal exercise therapy. It showed no difference in clinical outcomes between PRP and a saline solution injection when formal exercise therapy was used. Two other studies reported that PRP alone resulted in inferior outcomes to control groups receiving formal exercise therapy. The currently limited available evidence on PRP for nonoperative treatment of chronic rotator cuff disease suggests that in the short term, PRP injections may not be beneficial. When directly compared with exercise therapy, PRP does not result in superior functional outcomes, pain scores, or range of motion. However, interpretation of this literature is confounded by the lack of reporting of the cytology and characteristics of PRP. Level II, systematic review of Level I and II evidence.


pH-Dependent conformational changes in proteins and their effect on experimental pK(a)s: the case of Nitrophorin 4.
The acid-base behavior of amino acids is an important subject of study due to their prominent role in enzyme catalysis, substrate binding and protein structure. Due to interactions with the protein environment, their pK(a)s can be shifted from their solution values and, if a protein has two stable conformations, it is possible for a residue to have different "microscopic", conformation-dependent pK(a) values. In those cases, interpretation of experimental measurements of the pK(a) is complicated by the coupling between pH, protonation state and protein conformation. We explored these issues using Nitrophorin 4 (NP4), a protein that releases NO in a pH sensitive manner. At pH 5.5 NP4 is in a closed conformation where NO is tightly bound, while at pH 7.5 Asp30 becomes deprotonated, causing the conformation to change to an open state from which NO can easily escape. Using constant pH molecular dynamics we found two distinct microscopic Asp30 pK(a)s: 8.5 in the closed structure and 4.3 in the open structure. Using a four-state model, we then related the obtained microscopic values to the experimentally observed "apparent" pK(a), obtaining a value of 6.5, in excellent agreement with experimental data. This value must be interpreted as the pH at which the closed to open population transition takes place. More generally, our results show that it is possible to relate microscopic structure dependent pKa values to experimentally observed ensemble dependent apparent pK(a)s and that the insight gained in the relatively simple case of NP4 can be useful in several more complex cases involving a pH dependent transition, of great biochemical interest.


Kava blocks 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone-induced lung tumorigenesis in association with reducing O6-methylguanine DNA adduct in A/J mice.
We previously reported the chemopreventive potential of kava against 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK)- and benzo(a)pyrene (BaP)-induced lung tumorigenesis in A/J mice during the initiation and postinitiation stages. In this study, we investigated the tumorigenesis-stage specificity of kava, the potential active compounds, and the underlying mechanisms in NNK-induced lung tumorigenesis in A/J mice. In the first experiment, NNK-treated mice were given diets containing kava at a dose of 5 mg/g of diet during different periods. Kava treatments covering the initiation stage reduced the multiplicity of lung adenomas by approximately 99%. A minimum effective dose is yet to be defined because kava at two lower dosages (2.5 and 1.25 mg/g of diet) were equally effective as 5 mg/g of diet in completely inhibiting lung adenoma formation. Daily gavage of kava (one before, during, and after NNK treatment) completely blocked lung adenoma formation as well. Kavalactone-enriched fraction B fully recapitulated kava's chemopreventive efficacy, whereas kavalactone-free fractions A and C were much less effective. Mechanistically, kava and fraction B reduced NNK-induced DNA damage in lung tissues with a unique and preferential reduction in O(6)-methylguanine (O(6)-mG), the highly tumorigenic DNA damage by NNK, correlating and predictive of efficacy on blocking lung adenoma formation. Taken together, these results demonstrate the outstanding efficacy of kava in preventing NNK-induced lung tumorigenesis in A/J mice with high selectivity for the initiation stage in association with the reduction of O(6)-mG adduct in DNA. They also establish the knowledge basis for the identification of the active compound(s) in kava.


[Senescence of mesothelial cells].
The mesothelium is a specific group of cells having characteristics of both mesenchymal and epithelial cells. One of the most unique properties of these cells is a low proliferative capacity and a small number of achievable division. The purpose of this paper was to present the current state of knowledge on the causes of premature senescence of peritoneal mesothelial cells and to discuss the molecular events involved in this process. Particular attention was paid to the role of telomeres, the activity of senescence effectors at the level of the cell cycle, and the action of oxidative stress and transforming growth factor beta1. Moreover, the relationship between senescence of mesothelial cells and the aging of the organism as a whole, as well as the participation of senescent cells in the development of the intraperitoneal cancer metastasis was addressed.


Interaction of PARP-2 with DNA structures mimicking DNA repair intermediates and consequences on activity of base excision repair proteins.
Poly(ADP-ribosyl)ation is a posttranslational protein modification significant for genomic stability and cell survival in response to DNA damage. Poly(ADP-ribosyl)ation is catalyzed by poly(ADP-ribose)polymerases (PARPs). Among the 17 members of the PARP family, PARP-1 and PARP-2 are described as enzymes whose catalytic activity is stimulated by some types of DNA damages. Whereas the role of PARP-1 in response to DNA damage has been widely illustrated, the contribution of another DNA-dependent PARP, PARP-2, is less documented. To find out specific DNA targets of PARP-2 we evaluated by EMSA Kd values of PARP-2-DNA complexes for several DNA structures mimicking intermediates of different DNA metabolizing processes. In addition, we tested these DNA as activators of PARP-1 and PARP-2 in poly(ADP-ribose) synthesis. Like PARP-1, PARP-2 doesn't show correlation between activation efficiency and Kd values for DNA. PARP-2 displayed the highest affinity for flap-containing DNA, but was more efficiently activated by 5'-overhang DNA. Evaluating the influence of PARP-1 and PARP-2 on DNA repair synthesis catalyzed by DNA polymerase β revealed that both PARPs inhibit DNA polymerase β activity. However, unlike PARP-1, poly(ADP-ribosyl)ation of PARP-2 does not result in restoration of DNA synthesis efficiency. Similarly, both PARPs proteins inhibited FEN1 activity, but only activation of PARP-1, not PARP-2, could restore FEN1 activity, and only when PARP-2 was not present. Taken together, our data show that PARP-2 can directly regulate BER proteins but also can modulate the influence of PARP-1 on these BER proteins, by decreasing its poly(ADP-ribosyl)ation activity.


Single-strand conformational polymorphism and denaturing gradient gel electrophoresis in screening for variegate porphyria: identification of two new mutations.
Single-strand conformational polymorphism and denaturing gel electrophoresis were used to screen for mutations in the protoporphyrinogen oxidase gene (PPOX) of three patients with clinically and biochemically proven variegate porphyria in order to select genomic regions for specific DNA sequence analysis. Two previously undescribed mutations were identified: PPOX1423-1426-delATCT and PPOX2272insG. Denaturing gel electrophoresis was able to discern the point mutation in exon 5 (PPOX2272insG) of the PPOX gene. Once an index individual has been identified, single-strand conformational polymorphism and denaturing gel electrophoresis techniques are useful to identify family members who may be unaffected carriers. Such identification can help potential cases to avoid medications and other triggers that could precipitate acute porphyric attacks.


Timp3 deficiency in insulin receptor-haploinsufficient mice promotes diabetes and vascular inflammation via increased TNF-alpha.
Activation of inflammatory pathways may contribute to the beginning and the progression of both atherosclerosis and type 2 diabetes. Here we report a novel interaction between insulin action and control of inflammation, resulting in glucose intolerance and vascular inflammation and amenable to therapeutic modulation. In insulin receptor heterozygous (Insr+/-) mice, we identified the deficiency of tissue inhibitor of metalloproteinase 3 (Timp3, an inhibitor of both TNF-alpha-converting enzyme [TACE] and MMPs) as a common bond between glucose intolerance and vascular inflammation. Among Insr+/- mice, those that develop diabetes have reduced Timp3 and increased TACE activity. Unchecked TACE activity causes an increase in levels of soluble TNF-alpha, which subsequently promotes diabetes and vascular inflammation. Double heterozygous Insr+/-Timp3+/- mice develop mild hyperglycemia and hyperinsulinemia at 3 months and overt glucose intolerance and hyperinsulinemia at 6 months. A therapeutic role for Timp3/TACE modulation is supported by the observation that pharmacological inhibition of TACE led to marked reduction of hyperglycemia and vascular inflammation in Insr+/- diabetic mice, as well as by the observation of increased insulin sensitivity in Tace+/- mice compared with WT mice. Our results suggest that an interplay between reduced insulin action and unchecked TACE activity promotes diabetes and vascular inflammation.


Information-based fitness and the emergence of criticality in living systems.
Empirical evidence suggesting that living systems might operate in the vicinity of critical points, at the borderline between order and disorder, has proliferated in recent years, with examples ranging from spontaneous brain activity to flock dynamics. However, a well-founded theory for understanding how and why interacting living systems could dynamically tune themselves to be poised in the vicinity of a critical point is lacking. Here we use tools from statistical mechanics and information theory to show that complex adaptive or evolutionary systems can be much more efficient in coping with diverse heterogeneous environmental conditions when operating at criticality. Analytical as well as computational evolutionary and adaptive models vividly illustrate that a community of such systems dynamically self-tunes close to a critical state as the complexity of the environment increases while they remain noncritical for simple and predictable environments. A more robust convergence to criticality emerges in coevolutionary and coadaptive setups in which individuals aim to represent other agents in the community with fidelity, thereby creating a collective critical ensemble and providing the best possible tradeoff between accuracy and flexibility. Our approach provides a parsimonious and general mechanism for the emergence of critical-like behavior in living systems needing to cope with complex environments or trying to efficiently coordinate themselves as an ensemble.


Mid-infrared point sensor for in situ monitoring of CO2 emissions from large-scale engines.
This paper describes an optical fiber based system that has been developed for the monitoring of carbon dioxide emissions in situ within engines above 500 kW. Conventional sensors, reviewed here, fail to meet monitoring requirements, such as lifespan, accuracy, and robustness. This paper describes a sensor designed as a single point reflective probe configuration using low cost, compact mid-infrared optical components, making it suitable for insertion in large-scale engines including automotive tailpipes. The response of the sensor to carbon dioxide supplied from a cylinder in the laboratory environment is presented, as well as a number of experimental results taken in situ in an exhaust of an automotive diesel engine (smaller than 500 kW). The sensor is shown to have a long term stable operation over a wide range of concentrations (2%-15% CO(2)) with a lower detection limit smaller than the lowest value encountered in modern day engines.


An electrophysiological approach to the diagnosis of neurogenic dysphagia: implications for botulinum toxin treatment.
Botulinum toxin (BTX) injection into the cricopharyngeal (CP) muscle has been proposed for the treatment of neurogenic dysphagia due to CP hyperactivity. The aim was to determine whether an electrophysiological method exploring oropharyngeal swallowing could guide treatment and discriminate responders from non-responders, based on the association of CP dysfunction with other electrophysiological abnormalities of swallowing. Patients with different neurological disorders were examined: Parkinson disease, progressive supranuclear palsy, multiple system atrophy-Parkinson variant, multiple system atrophy cerebellar variant, stroke, multiple sclerosis and ataxia telangiectasia. All patients presented with clinical dysphagia, and with complete absence of CP muscle inhibition during the hypopharyngeal phase of swallowing. Each patient underwent clinical and electrophysiological investigations before and after treatment with BTX into the CP muscle of one side (15 units of Botox). Clinical and electrophysiological procedures were performed in a blind manner by two different investigators. The following electrophysiological measures were analysed: (1) duration of EMG activity of suprahyoid/submental muscles (SHEMG-D); (2) duration of laryngopharyngeal mechanogram (LPM-D); (3) duration of the inhibition of the CP muscle EMG activity (CPEMG-ID); and (4) interval between onset of EMG activity of suprahyoid/submental muscles and onset of laryngopharyngeal mechanogram (I-SHEMG-LPM). Two months after treatment, 50% of patients showed a significant improvement. Patients with prolonged or reduced SHEMG-D values and prolonged I-SHEMG-LPM values did not respond to BTX. Therefore, values for which BTX had no effect (warning values) were identified. This electrophysiological method can recognise swallowing abnormalities which may affect the outcome of the therapeutic approach to dysphagia with BTX treatment.


Markedly Reduced Risk of Internal Malignancies in Patients With Vitiligo: A Nationwide Population-Based Cohort Study.
Recent studies indicated that the autoimmunity of vitiligo exerts effects on cells other than melanocytes, which confer reduced risks of both melanoma and nonmelanoma skin cancers in patients with vitiligo. However, the risk of internal malignancy in patients with vitiligo has not been elucidated. We conducted a population-based retrospective cohort study using data from the Korean National Health Insurance claims database obtained from January 2007 to December 2016. All patients age 20 years or older with vitiligo who had at least two contacts with a physician from 2009 to 2016, during which a principal diagnosis was made, were identified (vitiligo group). Controls were randomly selected (two per patient with vitiligo) after frequency matching with the vitiligo group for age and sex during the same period (control group). A total of 101,078 patients with vitiligo and 202,156 controls without vitiligo were included. The incidence rates of internal malignancies were 612.9 and 708.9 per 100,000 person-years in the vitiligo and control groups, respectively. Patients with vitiligo showed a significantly reduced risk of overall internal malignancies (hazard ratio [HR], 0.86; 95% CI, 0.82 to 0.89; P < .001) compared with controls without vitiligo after adjustments for age, sex, and comorbidities. With regard to organ-specific malignancies, patients with vitiligo showed a remarkably decreased risk of cancer in the colon and rectum (HR, 0.62; 95% CI, 0.55 to 0.69; P < .001), ovary (HR, 0.62; 95% CI, 0.46 to 0.83; P < .001), and lung (HR, 0.75; 95% CI, 0.65 to 0.86; P < .001). Vitiligo was associated with a reduced risk of overall internal malignancies. These findings suggest that autoimmune diseases, including vitiligo, may provide immune surveillance for the development of cancer beyond the targeted organ.


A review on schizophrenia and relapse--a quest for user-friendly psychopharmacotherapy.
Schizophrenia in general is notoriously associated with relapses rendering the illness progressive to worse outcomes, a concept of which is compatible with neurotoxicity. Therefore, relapse prevention is of utmost clinical relevance. In this review, we aim to put relapse into clinical context in the realm of natural history of, or heterogeneity in, schizophrenia and summarize risk factors of relapse. We discuss how to effectively 'define' relapse in schizophrenia and recent meta-analytic studies on this topic to highlight the importance of continuous antipsychotic treatment. The following issues emerged: 'How low maintenance antipsychotic dosage could be?', 'How extended dosing could be?', 'Who could be successfully withdrawn from antipsychotics?' and 'How relapse could be defined in the first place?' The question in particular is how better to deliver antipsychotics at the lowest possible, whereby dose and dosing interval are relevant. While ongoing antipsychotic treatment is the rule, recent works are pointing to a possibility of lower dosage in the maintenance phase of the illness. Bearing in mind that suboptimal adherence and withdrawal from antipsychotics are an established and unequivocal risk factor for relapse, further investigations are certainly needed to explore user-friendly manner of psychopharmacotherapy to prevent relapse in schizophrenia.


Molecular analysis of human metapneumovirus detected in patients with lower respiratory tract infection in upper egypt.
Introduction. Since 2001, when Human metapneumovirus (HMPV) was isolated in the Netherlands, the virus has been detected in several continents. Although reports have confirmed the prevalence of HMPV worldwide, data from Egypt remain limited. HMPV plays an important role in respiratory tract infections in individuals of all ages particularly in children. This study was aimed at estimating the prevalence of HMPV in patients with community-acquired lower respiratory infection in Upper Egypt and characterizing the circulating Egyptian HMPV strains for the first time. Materials and Methods. From 2005 to 2008, respiratory samples from 520 patients were analyzed for the presence of HMPV by real-time RT-PCR. Molecular and phylogenetic analyses were performed on partial fusion gene sequences of HMPV-positive patients. Results. HMPV-positive patients were detected in 2007-2008. The overall infection rate was 4%, while 57% of the patients were children. Sequence analysis demonstrated circulation of subgroup B viruses with predominance of lineage B2. Nucleotide sequence identity within lineage B1 was 98.8%-99.7% and higher than that in lineage B2 (94.3%-100%). Three new amino acid substitutions (T223N, R229K, and D280N) of lineage B2 were observed. Conclusion. HMPV is a major viral pathogen in the Egyptian population especially in children. During 2007-2008, predominantly HMPV B2 circulated in Upper Egypt.


Health workers' participation in voluntary counselling and testing in three districts of Mashonaland East Province, Zimbabwe.
To determine the proportion of health workers who had undergone VCT for HIV in three districts of Mashonaland East Province and to further explore reasons for non-participation in those who had not been tested. The study was conducted in the rural Districts of Murewa, Mutoko and Mudzi in Mashonaland East province. The study was a descriptive cross sectional survey. All categories of health workers in Murewa, Mutoko and Mudzi. Participation in VCT, and reason for non-participation. Out of 200 questionnaires sent out 183 (91.5%) were completed and returned. The majority of the respondents were nurses 142 (77.6%), female 113 (61.7%), married 135 (73.8%). The median age was 33 (Q1; 28: Q3; 38) and they had attained a high school education 137 (74.9%). Of the respondents 160 (87.4%) had not gone for VCT; 141 (77%) did not want to have an HIV test. The reasons for not being willing to have an HIV test included not being ready to go for VCT 154 (84.2%); could not cope with the results, 143 (78.1%), do not have the courage to go 133 (72.7%); no need for testing as there is no cure for HIV/AIDS 106 (57.9%). One hundred and twenty six (69%) indicated that they needed counselling for them to be able to go for VCT. HIV/AIDS programmes have been directed at the community at large neglecting the health worker. This study, therefore, recommends programmes specifically tailor-made for health workers. These programmes should have a strong counselling component and should focus on self-efficacy so that health workers can finally be ready, be able to cope with HIV results and have the courage to participate in VCT.


Robust estimates of environmental effects on population vital rates: an integrated capture-recapture model of seasonal brook trout growth, survival and movement in a stream network.
Modelling the effects of environmental change on populations is a key challenge for ecologists, particularly as the pace of change increases. Currently, modelling efforts are limited by difficulties in establishing robust relationships between environmental drivers and population responses. We developed an integrated capture-recapture state-space model to estimate the effects of two key environmental drivers (stream flow and temperature) on demographic rates (body growth, movement and survival) using a long-term (11 years), high-resolution (individually tagged, sampled seasonally) data set of brook trout (Salvelinus fontinalis) from four sites in a stream network. Our integrated model provides an effective context within which to estimate environmental driver effects because it takes full advantage of data by estimating (latent) state values for missing observations, because it propagates uncertainty among model components and because it accounts for the major demographic rates and interactions that contribute to annual survival. We found that stream flow and temperature had strong effects on brook trout demography. Some effects, such as reduction in survival associated with low stream flow and high temperature during the summer season, were consistent across sites and age classes, suggesting that they may serve as robust indicators of vulnerability to environmental change. Other survival effects varied across ages, sites and seasons, indicating that flow and temperature may not be the primary drivers of survival in those cases. Flow and temperature also affected body growth rates; these responses were consistent across sites but differed dramatically between age classes and seasons. Finally, we found that tributary and mainstem sites responded differently to variation in flow and temperature. Annual survival (combination of survival and body growth across seasons) was insensitive to body growth and was most sensitive to flow (positive) and temperature (negative) in the summer and fall. These observations, combined with our ability to estimate the occurrence, magnitude and direction of fish movement between these habitat types, indicated that heterogeneity in response may provide a mechanism providing potential resilience to environmental change. Given that the challenges we faced in our study are likely to be common to many intensive data sets, the integrated modelling approach could be generally applicable and useful.


Circulating half-life of PEGylated recombinant methioninase holoenzyme is highly dose dependent on cofactor pyridoxal-5'-phosphate.
Recombinant methioninase (rMETase) has been shown to target the elevated methionine (MET) dependence of tumor cells and arrest their growth as well as make tumors more sensitive to standard chemotherapy agents. Polyethylene glycol (PEG)-modified rMETase (PEG-rMETase) has reduced antigenicity compared with unmodified rMETase. However, PEG-rMETase has a limited active circulating half-life due to rapid in vivo dissociation of its cofactor pyridoxal-5'-phosphate (PLP), a surprising finding, because PLP is tightly bound to PEG-rMETase in buffer. The question asked in the current study was on the effect of increasing doses of PLP to extend the circulating half-life of active PEG-rMETase holoenzyme in vivo. rMETase was conjugated with methoxypolyethylene glycol succinimidyl glutarate 5000 (MEGC-PEG). Miniosmotic pumps containing various concentrations of PLP were implanted in BALB-C mice. PLP-infused mice were then injected with a single dose of 4000 or 8000 units/kg PEG-rMETase. Mice infused with 5, 50, 100, 200, and 500 mg/ml PLP-containing miniosmotic pumps increased plasma PLP to 7, 24, 34, 60, and 95 microm, respectively, from the PLP baseline of 0.3 microm. PLP increased the half-life of MEGC-PEG-rMETase holoenzyme in a dose-dependent manner. Pumps containing 500 mg/ml PLP increased the half-life of MEGC-PEG-rMETase holoenzyme 4.5-fold from 1.5 to 7 h. Infused PLP did not extend the half-life of MEGC-PEG-rMETase apoenzyme. With a dose of 4000 units/kg MEGC-PEG-rMETase in the mice infused with 5, 50, 200, and 500 mg/ml PLP, plasma MET was depleted from 50 microm to < or = 5 microm for 8, 24, 72, and 72 h, respectively. Thus, PLP infusion could extend the period of MET depletion by MEGC-PEG-rMETase by approximately 10-fold in a dose-dependent manner. The mice given 8000 units/kg MEGC-PEG-rMETase showed a similar plasma MET depletion time course, indicating that the limiting factor for MEGC-PEG-rMETase-mediated MET depletion in vivo was PLP. The extended time of MET depletion by MEGC-PEG-rMETase was due to the maintenance of active MEGC-PEG-rMETase holoenzyme by infused PLP. The infused PLP either bound to apo-MEGC-PEG-rMETase and/or inhibited dissociation of PLP from holo-PEG-rMETase, thereby maintaining the holoenzyme form of MEGC-PEG-rMETase in vivo. The combination of MEGC-PEG-rMETase treatment with PLP infusion suggests an effective clinical strategy for long-term MET depletion to arrest cancer growth.


Treatment of chromoblastomycosis with itraconazole, cryosurgery, and a combination of both.
Chromoblastomycosis is a subcutaneous mycosis, seen frequently in tropical areas, and caused by dematiaceous fungi. It produces nodulo-verrucous lesions in the arms and legs. There is no treatment of choice for this disease and sometimes a combination of chemotherapy and physical therapy is necessary. The study included 12 patients diagnosed with chromoblastomycosis by means of fungal and histopathologic tests. The patients were assigned to three treatment groups: patients with small lesions, not greater than 15 cm2 in area, were assigned to Group 1, in which the treatment consisted of itraconazole 300 mg/day, or to Group 2, in which the treatment consisted of one or more sessions of open-spray cryosurgery. Patients with large lesions were assigned to Group 3 and started treatment with itraconazole 300 mg/day, until a maximal reduction of lesions occurred, and then underwent one or several cryosurgery sessions. Clinical, fungal, and laboratory tests were performed in each group before, during, and at the completion of treatment. Positive cultures of Fonsecaea pedrosoi were obtained in 11 out of 12 patients. Two out of four patients in Groups 1 and 3 had a clinical and fungal cure and the remaining patients experienced significant improvement. All four patients included in Group 2 achieved a cure. No important side-effects were seen among the patients included in any of the two itraconazole groups, and only two out of eight patients reported gastric discomfort. The cryosurgery group reported only normal complications of the process, such as edema and pain; two out of eight patients had a superimposed infection. The results of itraconazole and cryosurgery were good in cases with small lesions; antifungal therapy being more appropriate for flexion areas. The combination of itraconazole, to reduce the size of the lesions, with subsequent treatment of the remaining lesions with cryosurgery, represents a new alternative in the treatment of patients with large lesions. Both types of therapy are considered safe, with few side-effects.


Anti-inflammatory activity of extracts from leaves of Phyllanthus emblica.
Leaves and fruits of Phyllanthus emblica L. have been used for the anti-inflammatory and antipyretic treatment of rural populations in its growing areas in subtropical and tropical parts of China, India, Indonesia, and the Malay Peninsula. In the present study, leaves of Ph. emblica were extracted with ten different solvents (n-hexane, diethyl ether, methanol, tetrahydrofuran, acetic acid, dichloromethane, 1,4-dioxane, toluene, chloroform, and water). The inhibitory activity of the extracts against human polymorphonuclear leukocyte (PMN) and platelet functions was studied. Methanol, tetrahydrofuran, and 1,4-dioxane extracts (50 micrograms/ml) inhibited leukotriene B4-induced migration of human PMNs by 90% and N-formyl-L-methionyl-L-leucyl-L-phenylalanine (FMLP)-induced degranulation by 25-35%. The inhibitory activity on receptor-mediated migration and degranulation of human PMNs was associated with a high proportion of polar compounds in the extracts as assessed by normal phase thin layer chromatography. Diethyl ether extract (50 micrograms/ml) inhibited calcium ionophore A23187-induced leukotriene B4 release from human PMNs by 40%, thromboxane B2 production in platelets during blood clotting by 40% and adrenaline-induced platelet aggregation by 36%. Ellagic acid, gallic acid and rutin, all compounds isolated earlier from Ph. emblica, could not explain these inhibitory activities on PMNs or platelets by Ph. emblica extracts. These results show that the leaves of Ph. emblica have inhibitory activity on PMNs and platelets, which confirm the anti-inflammatory and antipyretic properties of this plant as suggested by its use in traditional medicine. The data suggest that the plant leaves contain as yet unidentified polar compound(s) with potent inhibitory activity on PMNs and chemically different apolar molecule(s) which inhibit both prostanoid and leukotriene synthesis.


Does drinking water influence hospital-admitted sialolithiasis on an epidemiological level in Denmark?
Sialolithiasis, or salivary stones, is not a rare disease of the major salivary glands. However, the aetiology and incidence remain largely unknown. Since sialoliths are comprised mainly of calcium phosphate salts, we hypothesise that drinking water calcium levels and other elements in drinking water could play a role in sialolithiasis. Owing to substantial intermunicipality differences in drinking water composition, Denmark constitutes a unique environment for testing such relations. An epidemiological study based on patient data extracted from the National Patient Registry and drinking water data from the Geological Survey of Denmark and Greenland retrieved as weighted data on all major drinking water constituents for each of the 3364 waterworks in Denmark. All patient cases with International Statistical Classification of Diseases 10th Revision (ICD-10) codes for sialolithiasis registered between the years 2000 and 2010 were included in the study (n=3014) and related to the drinking water composition on a municipality level (n=98). Multiple regression analysis using iterative search and testing among all demographic and drinking water variables with sialolithiasis incidence as the outcome in search of possible relations among the variables tested. The nationwide incidence of hospital-admitted sialolithiasis was 5.5 cases per 100,000 citizens per year in Denmark. Strong relations were found between the incidence of sialolithiasis and the drinking water concentration of calcium, magnesium and hydrogen carbonate, however, in separate models (p<0.001). Analyses also confirmed correlations between drinking water calcium and magnesium and their concentration in saliva whereas this was not the case for hydrogen carbonate. Differences in drinking water calcium and magnesium may play a role in the incidence of sialolithiasis. These findings are of interest because many countries have started large-scale desalination programmes of drinking water.


Differential induction of c-Fos and phosphorylated ERK by a noxious stimulus after peripheral nerve injury.
In this study, we compared induction of c-Fos and phosphorylated extracellular signal-regulated kinase (p-ERK) in the spinal dorsal horn after peripheral nerve injury. We examined the spinal dorsal horn for noxious heat-induced c-Fos and p-ERK protein-like immunoreactive (c-Fos- and p-ERK-IR) neuron profiles after tibial nerve injury. The effect of administration of a MEK 1/2 inhibitor (PD98059) on noxious heat-induced c-Fos expression was also examined after tibial nerve injury. A large number of c-Fos- and p-ERK-IR neuron profiles were induced by noxious heat stimulation to the hindpaw in sham-operated animals. A marked reduction in the number of c-Fos- and p-ERK-IR neuron profiles was observed in the medial 1/3 (tibial territory) of the dorsal horn at 3 and 7 days after nerve injury. Although c-Fos-IR neuron profiles had reappeared by 14 days after injury, the number of p-ERK-IR neuron profiles remained decreased in the tibial territory of the superficial dorsal horn. Double immunofluorescence labeling for c-Fos and p-ERK induced by noxious heat stimulation to the hindpaw at different time points revealed that a large number of c-Fos-IR, but not p-ERK-IR, neuron profiles were distributed in the tibial territory after injury. Although administration of a MEK 1/2 inhibitor to the spinal cord suppressed noxious heat-induced c-Fos expression in the peroneal territory, this treatment did not alter c-Fos induction in the tibial territory after nerve injury. ERK phosphorylation may be involved in c-Fos induction in normal nociceptive responses, but not in exaggerated c-Fos induction after nerve injury.


Optimal Antiplatelet Therapy in Moderate to Severe Asymptomatic and Symptomatic Carotid Stenosis: A Comprehensive Review of the Literature.
Carotid stenosis patients are at risk of vascular events despite antiplatelet therapy. Data on prescribed antiplatelet regimens have not been comprehensively collated from trials to guide optimal therapy in this population. This review was conducted in line with the current Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) statement. Medline, Ovid, Embase, Web of Science, and Google Scholar from 1988 to 2018 were searched using the search terms "carotid stenosis", "asymptomatic", "symptomatic", "antiplatelet", and "anti-platelet" to identify randomised trials in patients with asymptomatic or symptomatic extracranial moderate-severe carotid stenosis on any form of antiplatelet therapy in which vascular events and pre specified composite outcome events were reported. Twenty-five studies were judged eligible for inclusion. Data from one randomised controlled trial showed no significant difference in benefit with aspirin versus placebo in asymptomatic carotid stenosis, but it is still reasonable to recommend aspirin (81-325 mg daily) for prevention of vascular events in these patients. Low to medium dose aspirin (81-325 mg daily) is superior to higher doses (>650 mg daily) at preventing recurrent vascular events in patients undergoing endarterectomy. Data from endovascular treatment (EVT) trials support peri-procedural treatment of asymptomatic and symptomatic patients with 81-325 mg of aspirin daily. The use of peri-procedural aspirin-clopidogrel in patients undergoing EVT is based on one pilot trial, but appears safe. Short-term aspirin-dipyridamole or aspirin-clopidogrel treatments are equally effective at reducing micro-embolic signals on transcranial Doppler ultrasound in patients with ≥50% symptomatic carotid stenosis. There is insufficient evidence to recommend routine aspirin-clopidogrel combination therapy to reduce the risk of recurrent clinical ischaemic events in patients with symptomatic moderate-severe carotid stenosis. This comprehensive review outlines an evidence based approach to antiplatelet therapy in carotid stenosis patients. Future trials should randomise such patients to receive different antiplatelet regimens to assess their efficacy and safety and to optimise peri-procedural and long-term preventive treatment in this patient cohort.