Eutrophication mediates a common off-flavor compound, 2-methylisoborneol, in a drinking water reservoir.
Off-flavors, such as 2-methylisoborneol (MIB) and geosmin, cause drinking water to have earthy or musty tastes and odors. Humans can detect such compounds at minute concentrations (10 and 30 ng/L for MIB and geosmin, respectively), and, although not a health risk, off-flavors can promote consumer distrust. Removal of these compounds is costly and often unreliable or only suitable under certain conditions. Minimizing off-flavor production at the watershed-scale may be more cost-effective in addition to improving ecosystem health and aesthetics. Cyanobacteria are considered to be the primary drivers of off-flavors in freshwater systems. Due to their ability to produce toxins, cyanobacteria have been under particular scrutiny, and environmental factors promoting cyanobacterial blooms are relatively well-studied. Using this body of literature, we conducted a seven-week, limnocorral experiment where we manipulated nitrogen and nitrogen-to-phosphorus concentrations to influence phytoplankton community structure and off-flavor production. The addition of a single nutrient across broad ranges (nitrogen or phosphorus) had no effect on MIB. However, the addition of both nitrogen and phosphorus promoted high concentrations of MIB relative to treatments that received no nutrients (448% increase) or only nitrogen or phosphorus (722% increase). Interestingly, cyanobacteria waned during the experiment and were replaced by diatoms, which were the dominant taxa by the end of the experiment. Our findings clearly show that eutrophication affects MIB production, but mechanisms leading to the production of this compound may differ from what has been previously predicted.


Heteroduplex molecules cause sexing errors in a standard molecular protocol for avian sexing.
Molecular methods are a necessary tool for sexing monomorphic birds. These molecular approaches are usually reliable, but sexing protocols should be evaluated carefully because biochemical interactions may lead to errors. We optimized laboratory protocols for genetic sexing of a monomorphic shorebird, the upland sandpiper (Bartramia longicauda), using two independent sets of primers, P2/P8 and 2550F/2718R, to amplify regions of the sex-linked CHD-Z and CHD-W genes. We discovered polymorphisms in the region of the CHD-Z intron amplified by the primers P2/P8 which caused four males to be misidentified as females (n = 90 mated pairs). We cloned and sequenced one CHD-W allele (370 bp) and three CHD-Z alleles in our population: Z° (335 bp), Z' (331 bp) and Z″ (330 bp). Normal (Z°Z°) males showed one band in agarose gel analysis and were easily differentiated from females (Z°W), which showed two bands. However, males heterozygous for CHD-Z alleles (Z'Z″) unexpectedly showed two bands in a pattern similar to females. While the Z' and Z″ fragments contained only short deletions, they annealed together during the polymerase chain reaction (PCR) process and formed heteroduplex molecules that were similar in size to the W fragment. Errors previously reported for molecular sex-assignment have usually been due to allelic dropout, causing females to be misidentified as males. Here, we report evidence that events in PCRs can lead to the opposite error, with males misidentified as females. We recommend use of multiple primer sets and large samples of known-sex birds for validation when designing protocols for molecular sex analysis.


Quantitative analysis of the 2002 phocine distemper epidemic in the Netherlands.
Phocine distemper virus (PDV) caused thousands of deaths among harbor seals (Phoca vitulina) from the North Sea in 1988 and 2002. To examine the effects of different factors on the pathology of phocine distemper, we performed necropsies and laboratory analyses on 369 harbor seals that stranded along the Dutch coast during the 2002 PDV epidemic. Diagnostic tests for morbillivirus infection indicated a differential temporal presence of morbillivirus in lung and brain. Seals of 3 years or older were significantly more often IgG positive than younger seals. The most frequent lesions in PDV cases were bronchopneumonia, broncho-interstitial pneumonia, and interstitial emphysema. Extra-thoracic emphysema was rare in <1-year-olds compared with older seals, even though severe pneumonia was more common. PDV cases generally had empty stomachs and less blubber than by-caught seals from before the epidemic. In PDV cases involving older animals, lung, kidney, and adrenal weights were significantly increased. Bordetella bronchiseptica was isolated from lungs in two thirds of the PDV cases examined. Our results indicate that brain should be included among the tissues tested for PDV by RT-PCR; that either phocine distemper has a longer duration in older seals or that there are age-related differences in immunity and organ development; that dehydration could play a role in the course and outcome of phocine distemper; and that bacterial coinfections in lungs are more frequent in PDV cases than gross lesions suggest. These results illustrate how quantitative analysis of pathology data from such epidemics can improve understanding of the causative disease.


Changes in Blood Pressure and Urine Albumin-Creatinine Ratio in a Randomized Clinical Trial Comparing Aflibercept, Bevacizumab, and Ranibizumab for Diabetic Macular Edema.
To compare blood pressure and urine albumin-creatinine ratio over time for participants receiving aflibercept, bevacizumab, or ranibizumab. Preplanned secondary analyses from a randomized trial comparing aflibercept, bevacizumab, and ranibizumab for diabetic macular edema (DME). The Diabetic Retinopathy Clinical Research Network (DRCR.net) enrolled 660 participants with DME and visual acuity 20/32 or worse in at least one eye. Eyes received intravitreous injections of 2.0 mg aflibercept, 1.25 mg bevacizumab, or 0.3 mg ranibizumab based on a structured retreatment protocol over 2 years. Main outcome measures were (1) a change in blood pressure at 2 years, and (2) a change in urine albumin-creatinine ratio (UACR) at 1 year. At baseline, 95 participants (14%) had normal blood pressure, 220 (33%) had borderline blood pressure elevation, 206 (31%) had mild blood pressure elevation, and 139 (21%) had moderate blood pressure elevation. Average change in mean arterial pressure from baseline to 2 years was -1.2 ± 15, -1.8 ± 13.5, -2.6 ± 14.4 mm Hg in the aflibercept, bevacizumab, and ranibizumab groups, respectively (global P = 0.69). At baseline 247 participants (38%) had no albuminuria (<30 mg/g), 195 (30%) had microalbuminuria (30-300 mg/g), and 212 (32%) had macroalbuminuria (>300 mg/g). Changes in UACR category were not different among treatment groups at the 52-week visit (global P = 0.29). There do not appear to be treatment group differences for changes in blood pressure or UACR as a reflection of kidney function in patients with DME treated with aflibercept, bevacizumab, or ranibizumab.


Risk of venous thrombosis in carriers of the prothrombin G20210A variant and factor V Leiden and their interaction with oral contraceptives.
The prothrombin G20210A mutation and factor V Leiden have been found to be associated with an increased risk of venous thrombosis, but the reported prevalences of the prothrombin gene variant both in the normal population and in patients with deep venous thrombosis (DVT) vary greatly in the literature. Moreover, the influence of oral contraceptives (OC) on thrombotic events in patients with the prothrombin G20210A variant has not been well established. In this study we evaluate both circumstances. A case-control study was run on 229 patients with DVT and 246 healthy controls. The patients' history of thrombosis and acquired thrombotic risk factors, especially OC, were recorded. Prothrombin G20210A mutation, factor V Leiden, antithrombin, heparin II cofactor, plasminogen and proteins C and S were evaluated. Seven and a half percent of the patients and 2.9% of the controls were carriers of the prothrombin mutation, while 12.2% of the patients and 1.6% of the controls had factor V Leiden. Among the 229 DVT patients there were 130 patients with clinically suspected thrombophilia (first thrombotic event occurring before the age of 45 years or positive family history of thrombosis or recurrent venous thrombosis). Ten percent of these 130 patients were carriers of the prothrombin G20210A mutation and 18.5% had the factor V Leiden mutation. The odds ratios (OR) for DVT risk were: 2.4 (95% CI, 1.0-6.3) for the total DVT patients and 5.2 (95% CI, 1.4-19.5) for the patients with clinically suspected thrombophilia with the prothrombin mutation. The risk of thrombosis was 6.9 (95% CI, 2.3-20.6) for the DVT patients and 14.3 (95% CI, 3.3-64.6) for the patients with clinically suspected thrombophilia with factor V Leiden. Fifty-five percent of the patients with combined congenital defects (prothrombin mutation G20210A plus another congenital defect) had recurrent thrombosis. In women receiving OC the risk of DVT was 3.5 (95% CI, 1.5-8.2) that of the patients not receiving OC. When women with combined defects were also taking OC, the risk of thrombosis increased significantly. The prevalence of the prothrombin G20210A mutation in the healthy population in our study is similar to that observed in other southern European countries. The prothrombin G20210A mutation does not by itself seem to be a high thrombotic risk factor. However, when it is present together with other thrombotic risk factors, the predicted risk of thrombotic events increases. The use of OC by women with the prothrombin G20210A variant or FV Leiden, either alone or combined with other thrombotic risk factors, was associated with a significant increase in the risk of venous thrombosis.


Epidemiology of Colorectal Cancer in Average Risk Adults 20-39 Years of Age: A Population-Based National Study.
While overall rates of colorectal cancer (CRC) have declined in individuals aged above 50 years of age, this decline has not been seen in younger individuals who do not benefit from current screening guidelines. We sought to describe the prevalence of CRC in adults 20-39 years of age without family history of CRC or inflammatory bowel disease as early-onset CRC (EoCRC), evaluate associated signs and symptoms and medical comorbidities in EoCRC, and compare them with individuals aged 20-39 years without CRC (NoCRC). Our secondary aim was to compare EoCRC with individuals aged 40 years and above with CRC (LoCRC). Utilizing a commercial database (Explorys Inc, Cleveland, OH), we identified a cohort of patients aged 20-39 years with first ever diagnosis of CRC between 2013 and 2018 based on the Systematized Nomenclature of Medicine-Clinical Terms. We calculated the overall prevalence rate of EoCRC, described age, race, and gender-based prevalence rates of EoCRC, and identified associated symptoms and medical comorbidities associated with EoCRC. The overall rate of EoCRC was 18.9/100,000. Compared to NoCRC, EoCRC patients were more likely to be Caucasian and female, with predominant symptoms of hematochezia, anemia, and decreased appetite. EoCRC group had higher prevalence rates of medical comorbidities such as diabetes, smoking, and obesity. Compared to LoCRC, EoCRC group presented more frequently with left-sided CRC and rectal cancers. This is one of the largest studies to date to describe the epidemiology of EoCRC in USA. We found EoCRC to occur predominantly in the Caucasian and female population. EoCRC presented more frequently with left-sided and rectal CRC. We also identified signs/symptoms as well as comorbidities associated with EoCRC. Patients with these features may benefit from earlier screening.


A mutation in Orai1 causes immune deficiency by abrogating CRAC channel function.
Antigen stimulation of immune cells triggers Ca2+ entry through Ca2+ release-activated Ca2+ (CRAC) channels, promoting the immune response to pathogens by activating the transcription factor NFAT. We have previously shown that cells from patients with one form of hereditary severe combined immune deficiency (SCID) syndrome are defective in store-operated Ca2+ entry and CRAC channel function. Here we identify the genetic defect in these patients, using a combination of two unbiased genome-wide approaches: a modified linkage analysis with single-nucleotide polymorphism arrays, and a Drosophila RNA interference screen designed to identify regulators of store-operated Ca2+ entry and NFAT nuclear import. Both approaches converged on a novel protein that we call Orai1, which contains four putative transmembrane segments. The SCID patients are homozygous for a single missense mutation in ORAI1, and expression of wild-type Orai1 in SCID T cells restores store-operated Ca2+ influx and the CRAC current (I(CRAC)). We propose that Orai1 is an essential component or regulator of the CRAC channel complex.


Evaluation of a novel assessment form for observing medical residents: a randomised, controlled trial.
Teaching faculty cannot reliably distinguish between satisfactory and unsatisfactory resident performances and give non-specific feedback. This study aimed to test whether a novel rating form can improve faculty accuracy in detecting unsatisfactory performances, generate more rater observations and improve feedback quality. Participants included two groups of 40 internal medicine residency faculty staff. Both groups received 1-hour training on how to rate trainees in the mini-clinical evaluation exercise (mini-CEX) format. The intervention group was given a new rating form structured with prompts, space for free-text comments, behavioural anchors and fewer scoring levels, whereas the control group used the current American Board of Internal Medicine Mini-CEX form. Participants watched and scored six scripted videotapes of resident performances 2-3 weeks after the training session. Intervention group participants were more accurate in discriminating satisfactory from unsatisfactory performances (85% versus 73% correct; odds ratio [OR] 2.13, 95% confidence interval [CI] 1.16-3.14, P = 0.02) and yielded more correctly identified unsatisfactory performances (96% versus 52% correct; OR 25.35, 95% CI 9.12-70.46), but were less accurate in identifying satisfactory performances (73% versus 95% correct; OR 0.15, 95% CI 0.05-0.39). Intervention group participants averaged one fewer declared intended feedback item (4.7 versus 5.7) and showed no difference in the amount of feedback that was above minimal in quality. Intervention group participants generated more written evaluative observations (10.8 versus 5.7). Inter-rater agreement improved with the new form (Fleiss' kappa, 0.52 versus 0.30). Modifying the currently used direct observations process may produce more recorded observations, increase inter-rater agreement and improve overall rater accuracy, but it may also increase severity error.


Genome-wide association analysis and functional annotation of positional candidate genes for feed conversion efficiency and growth rate in pigs.
Feed conversion efficiency is a measure of how well an animal converts feed into live weight and it is typically expressed as feed conversion ratio (FCR). FCR and related traits like growth rate (e.g. days to 110 kg-D110) are of high interest for animal breeders, farmers and society due to implications on animal performance, feeding costs and environmental sustainability. The objective of this study was to identify genomic regions associated with FCR and D110 in pigs. A total of 952 terminal line boars, showing an individual variation in FCR, were genotyped using 60K SNP-Chips. Markers were tested for associations with estimated breeding values (EBV) for FCR and D110. For FCR, the largest number of associated SNPs was located on chromosomes 4 (30 SNPs), 1 (25 SNPs), X (15 SNPs) and 6 (12 SNPs). The most prominent genomic regions for D110 were identified on chromosomes 15 (10 SNPs), 1 and 4 (both 9 SNPs). The most significantly associated SNPs for FCR and D110 mapped 129.8 Kb from METTL11B (chromosome 4) and 32Kb from MBD5 (chromosome 15), respectively. A list of positional genes, closest to significantly associated SNPs, was used to identify enriched pathways and biological functions related to the QTL for both traits. A number of candidate genes were significantly overrepresented in pathways of immune cell trafficking, lymphoid tissue structure, organ morphology, endocrine system function, lipid metabolism, and energy production. After resequencing the coding region of selected positional and functional candidate genes, six SNPs were genotyped in a subset of boars. SNPs in PRKDC, SELL, NR2E1 and AKRIC3 showed significant associations with EBVs for FCR/D110. The study revealed a number of chromosomal regions and candidate genes affecting FCR/D110 and pointed to corresponding biological pathways related to lipid metabolism, olfactory reception, and also immunological status.


Factors associated with early maternal postpartum discharge from the hospital.
To examine if mothers discharged from the hospital after 1 night's stay or less (early discharge) differ from those discharged from the hospital after 2 or 3 nights' stay (regular discharge) in demographic, economic, behavioral, health status, and health services risk factors associated with maternal and infant well-being. Bivariate and logistic comparisons of women discharged from the hospital early or after a regular hospitalization as reported in the 1988 National Maternal and Infant Health Survey. Postpartum women. Postpartum discharge from the hospital after 1 night's stay or less in the hospital. Early discharge from the hospital occurred for 12.3% of 3,865 366 women. Logistic regression revealed 8 factors that were associated with early discharge from the hospital: less than a high school education, aged older than 35 years, inadequate prenatal care, lack of private insurance or receipt of Medicaid for delivery, giving birth in the western United States, plans to breast-feed, and care by midwives. Two factors were associated with an increased likelihood of a regular discharge from the hospital: report of a hospitalization during pregnancy and giving birth in the Midwest or Northeast. Significant numbers of mothers discharged from the hospital early manifested health and social risk factors associated with poor health outcomes. Economic factors seemed an important motivation for early discharge from the hospital, as did inadequate use of health services. Since brief hospitalization is prevalent, clinicians and administrators must assure that the objectives of medical contact during childbirth are met, if not through hospitalization, then through other mechanisms.


Omega-3 fatty acids status in human subjects estimated using a food frequency questionnaire and plasma phospholipids levels.
Intakes of omega-3 (n-3) fatty acids (FA) are associated with several health benefits. The aim of this study was to verify whether intakes of n-3 FA estimated from a food frequency questionnaire (FFQ) correlate with n-3 FA levels measured in plasma phospholipids (PL). The study sample consisted of 200 French-Canadians men and women aged between 18 to 55 years. Dietary data were collected using a validated FFQ. Fasting blood samples were collected and the plasma PL FA profile was measured by gas chromatography. Low intakes of n-3 long-chain FA together with low percentages of n-3 long-chain FA in plasma PL were found in French-Canadian population. Daily intakes of eicosapentaenoic acid (EPA), docosapentaenoic acid (DPA) and docosahexaenoic acid (DHA) were similar between men and women. Yet, alpha-linolenic acid (ALA) and total n-3 FA intakes were significantly higher in men compared to women (ALA: 2.28 g and 1.69 g, p < 0.0001, total n-3 FA: 2.57 g and 1.99 g, p < 0.0001; respectively). In plasma PL, DPA and DHA percentages were significantly different between men and women (DPA: 1.03% and 0.88%, p < 0.0001, DHA: 3.00% and 3.43%, p = 0.0005; respectively). Moreover, DHA (men: r = 0.52, p < 0.0001; women: r = 0.57, p < 0.0001) and total n-3 FA (men: r = 0.47, p < 0.0001; women: r = 0.52, p < 0.0001) intakes were positively correlated to their respective plasma PL FA levels. In women, EPA (r = 0.44, p < 0.0001) and DPA (r = 0.23, p = 0.02) intakes were also correlated respectively with EPA and DPA plasma PL FA percentages. Estimated n-3 long-chain FA intake among this young and well-educated French-Canadian population is lower than the recommendations. Further, FFQ data is comparable to plasma PL results to estimate DHA and total n-3 FA status in healthy individuals as well as to evaluate the EPA and DPA status in women. Overall, this FFQ could be used as a simple, low-cost tool in future studies to rank n-3 FA status of individuals.


A Penalized Likelihood Framework for High-Dimensional Phylogenetic Comparative Methods and an Application to New-World Monkeys Brain Evolution.
Working with high-dimensional phylogenetic comparative data sets is challenging because likelihood-based multivariate methods suffer from low statistical performances as the number of traits $p $ approaches the number of species $n $ and because some computational complications occur when $p $ exceeds $n$. Alternative phylogenetic comparative methods have recently been proposed to deal with the large $p $ small $n $ scenario but their use and performances are limited. Herein, we develop a penalized likelihood (PL) framework to deal with high-dimensional comparative data sets. We propose various penalizations and methods for selecting the intensity of the penalties. We apply this general framework to the estimation of parameters (the evolutionary trait covariance matrix and parameters of the evolutionary model) and model comparison for the high-dimensional multivariate Brownian motion (BM), Early-burst (EB), Ornstein-Uhlenbeck (OU), and Pagel's lambda models. We show using simulations that our PL approach dramatically improves the estimation of evolutionary trait covariance matrices and model parameters when $p$ approaches $n$, and allows for their accurate estimation when $p$ equals or exceeds $n$. In addition, we show that PL models can be efficiently compared using generalized information criterion (GIC). We implement these methods, as well as the related estimation of ancestral states and the computation of phylogenetic principal component analysis in the R package RPANDA and mvMORPH. Finally, we illustrate the utility of the new proposed framework by evaluating evolutionary models fit, analyzing integration patterns, and reconstructing evolutionary trajectories for a high-dimensional 3D data set of brain shape in the New World monkeys. We find a clear support for an EB model suggesting an early diversification of brain morphology during the ecological radiation of the clade. PL offers an efficient way to deal with high-dimensional multivariate comparative data.


A graphene/single GaAs nanowire Schottky junction photovoltaic device.
A graphene/nanowire Schottky junction is a promising structure for low-cost high-performance optoelectronic devices. Here we demonstrate a graphene/single GaAs nanowire Schottky junction photovoltaic device. The Schottky junction is fabricated by covering a single layer graphene onto an n-doped GaAs nanowire. Under 532 nm laser excitation, the device exhibits a high responsivity of 231 mA W-1 and a short response/recover time of 85/118 μs at zero bias. Under AM 1.5 G solar illumination, the device has an open-circuit voltage of 75.0 mV and a short-circuit current density of 425 mA cm-2, yielding a remarkable conversion efficiency of 8.8%. The excellent photovoltaic performance of the device is attributed to the strong built-in electric field in the Schottky junction as well as the transparent property of graphene. The device is promising for self-powered high-speed photodetectors and low-cost high-efficiency solar cells.


Glucose response during craniotomy: propofol-remifentanil versus isoflurane-remifentanil.
Major surgery is associated with a stress response. The aim of this study was to compare the effects of isoflurane or propofol, both supplemented with remifentanil, on the glucose, cortisol and insulin-based stress responses prospectively. Forty patients undergoing craniotomy randomly received either 1% isoflurane (Group I, N.=20) or propofol 6 mg kg h(-1) (Group P, N.=20) during remifentanil-based (0.125 μg kg min(-1)) anesthesia. Blood glucose was recorded preoperatively, after induction, intubation and pin placement, before and after skin incision, craniotomy, dura incision, 15th, 30th, 60th, 90th, 120th, 150th, 180th min post-dura incision, following dura and skin closure, extubation and at the 1st and 24th postoperative hours. Insulin and cortisol were measured preoperatively, after intubation, dura incision, at the 60th min, extubation and at the 1st and 24th hour postoperatively. The glucose/insulin ratio and glycemic stress index were calculated after all the measurements were obtained. Patient characteristics were comparable in both groups. Blood glucose significantly decreased after induction in comparison to the baseline value in both groups. Blood glucose was significantly higher in Group I than Group P before skin incision, after craniotomy and dura incision and at all measurement time points after the 60th minute following dura incision. There was a significant alteration with time in insulin values in both groups and the insulin values at the 60th min were significantly lower in Group I than in Group P. There was not any difference in the inter-group analysis of cortisol; however, there was a significant change over time in the insulin values in both groups. There was no difference in the intra-group glucose/insulin ratio, however, there was a significant difference between groups at the 60th min and at extubation. The Glycemic Stress Index was comparable between groups (Group I vs. Group P: 2.48±1.15 vs. 2.15±0.86, P=0.465). Isoflurane and propofol, both combined with remifentanil, provided clinically comparable cortisol and insulin responses to surgery in craniotomy operations, whereas propofol attenuated the increase in plasma blood glucose.


Self-reported health, self-reported fitness, and all-cause mortality: prospective cohort study.
Self-reported health has a well established relationship to later mortality, although the reasons are not entirely understood. This study examined the association of a similar self-reported measure of fitness with mortality and compared it to that of self-reported health. The study had a prospective cohort design with multiple sampling points. Participants were 858 men and women from Glasgow and the surrounding area of Scotland, aged 59 when self-reported health and fitness data were first collected in 1991/92. They were re-interviewed at age 64 and 69 and mortality was tracked for 16.5 years in total. Hazard ratios for all cause mortality were estimated for those that reported poor health or poor fitness relative to others their age, taking into account a range of covariates, some of which were also time varying. In both unadjusted and covariate adjusted models, self-reported fitness was at least as good a predictor of mortality as self-reported health. In a mutually adjusted model, both again emerged as significant predictors. Poor subjective health with poor subjective fitness appeared to be a particularly lethal combination. Both self-reported health and self-reported fitness were independent predictors of mortality. Where the objective assessment of aerobic fitness is not feasible, a simple measure of subjective fitness could prove a useful alternative.


Uptake of rosuvastatin by isolated rat hepatocytes: comparison with pravastatin.
1. The liver is the target organ for the lipid-regulating effect of rosuvastatin, a new 3-hydroxy-3-methylglutaryl coenzyme A reductase inhibitor, and liver-selective uptake of this drug is therefore a desirable property. The uptake kinetics of rosuvastatin were investigated and compared with those of pravastatin using isolated rat hepatocytes. 2. Uptake for both drugs involved both active transport and passive diffusion processes. The Michaelis constant (K(m)) of uptake rate for rosuvastatin (9.17 micro M) was approximately half that for pravastatin (16.5 micro M). However, the maximum uptake rate (V(max)) and carrier-mediated uptake clearance (V(max)/K(m)) of rosuvastatin were significantly (p < 0.01) greater than those of pravastatin, and a larger contribution of carrier-mediated uptake clearance to total uptake clearance was shown for rosuvastatin (contribution ratio 0.903 versus pravastatin 0.654). 3. Sodium and chloride ions did not play a significant role in the uptake of rosuvastatin and pravastatin, but the uptake of both drugs was inhibited both by depletion of cellular ATP and by organic anions such as bromosulfophthalein. 4. Rosuvastatin competitively inhibited the uptake of pravastatin, with an inhibition constant (K(i)) (2.75 micro M) relatively similar to its K(m). 5. The results suggest that an organic anion transport protein is the main mediator of the hepatic uptake of rosuvastatin and pravastatin, which occurs in an ATP-dependent manner. Our results indicated that rosuvastatin was taken up by the hepatocytes via the same transport systems as pravastatin, but with a greater affinity and efficiency than pravastatin.


Bubble sorting in pinched microchannels for ultrasound contrast agent enrichment.
Ultrasound contrast agent (UCA) suspensions contain encapsulated microbubbles with a wide size distribution, with radii between 1 and 10 μm. Medical transducers generally operate at a narrow frequency bandwidth, severely limiting the fraction of bubbles that resonates to the driving ultrasound. Thus, the sensitivity of contrast enhanced ultrasound imaging, molecular imaging with targeted bubbles, and drug delivery with microbubbles can be improved by narrowing down the size distribution of the bubble suspension. Here, we use a low-cost lab-on-a-chip method for the sorting of microbubbles by their size without external actuation, based on a microfluidic separation technique known as pinched flow fractionation (PFF). We show by numerical investigation that the inclusion of particle rotation in the pinched segment is essential for an accurate description of the sorting behavior of particles with sizes close to the pinched segment width. Successful enrichment of a polydisperse contrast agent into a bubble suspension with a narrow size distribution (radius 1.56 ± 0.30 μm) was achieved with a PFF-based microdevice. This sorting technique can be readily parallelized, and may thus lead to an easy-to-use and robust device capable of enriching ultrasound contrast agents, leading to an improvement in the sensitivity of contrast ultrasound imaging.


Pharmacokinetics of Escalating Doses of Oral Psilocybin in Healthy Adults.
Psilocybin is a psychedelic tryptamine that has shown promise in recent clinical trials for the treatment of depression and substance use disorders. This open-label study of the pharmacokinetics of psilocybin was performed to describe the pharmacokinetics and safety profile of psilocybin in sequential, escalating oral doses of 0.3, 0.45, and 0.6 mg/kg in 12 healthy adults. Eligible healthy adults received 6-8 h of preparatory counseling in anticipation of the first dose of psilocybin. The escalating oral psilocybin doses were administered at approximately monthly intervals in a controlled setting and subjects were monitored for 24 h. Blood and urine samples were collected over 24 h and assayed by a validated liquid chromatography-tandem mass spectrometry (LC-MS/MS) assay for psilocybin and psilocin, the active metabolite. The pharmacokinetics of psilocin were determined using both compartmental (NONMEM) and noncompartmental (WinNonlin) methods. No psilocybin was found in plasma or urine, and renal clearance of intact psilocin accounted for less than 2% of the total clearance. The pharmacokinetics of psilocin were linear within the twofold range of doses, and the elimination half-life of psilocin was 3 h (standard deviation 1.1). An extended elimination phase in some subjects suggests hydrolysis of the psilocin glucuronide metabolite. Variation in psilocin clearance was not predicted by body weight, and no serious adverse events occurred in the subjects studied. The small amount of psilocin renally excreted suggests that no dose reduction is needed for subjects with mild-moderate renal impairment. Simulation of fixed doses using the pharmacokinetic parameters suggest that an oral dose of 25 mg should approximate the drug exposure of a 0.3 mg/kg oral dose of psilocybin. Although doses of 0.6 mg/kg are in excess of likely therapeutic doses, no serious physical or psychological events occurred during or within 30 days of any dose. NCT02163707.


Spectroscopic correlation analysis of NMR-based metabonomics in exercise science.
Spectroscopic studies of complex clinical fluids have led to the application of a more holistic approach to their chemical analysis becoming more popular and widely employed. The efficient and effective interpretation of multidimensional spectroscopic data relies on many chemometric techniques and one such group of tools is represented by so-called correlation analysis methods. Typical of these techniques are two-dimensional correlation analysis and statistical total correlation spectroscopy (STOCSY). Whilst the former has largely been applied to optical spectroscopic analysis, STOCSY was developed and has been applied almost exclusively to NMR metabonomic studies. Using a (1)H NMR study of human blood plasma, from subjects recovering from exhaustive exercise trials, the basic concepts and applications of these techniques are examined. Typical information from their application to NMR-based metabonomics is presented and their value in aiding interpretation of NMR data obtained from biological systems is illustrated. Major energy metabolites are identified in the NMR spectra and the dynamics of their appearance and removal from plasma during exercise recovery are illustrated and discussed. The complementary nature of two-dimensional correlation analysis and statistical total correlation spectroscopy are highlighted.


2'-O methylation of the viral mRNA cap by West Nile virus evades ifit1-dependent and -independent mechanisms of host restriction in vivo.
Prior studies have shown that 2'-O methyltransferase activity of flaviviruses, coronaviruses, and poxviruses promotes viral evasion of Ifit1, an interferon-stimulated innate immune effector protein. Viruses lacking 2'-O methyltransferase activity exhibited attenuation in primary macrophages that was rescued in cells lacking Ifit1 gene expression. Here, we examined the role of Ifit1 in restricting pathogenesis in vivo of wild type WNV (WNV-WT) and a mutant in the NS5 gene (WNV-E218A) lacking 2'-O methylation of the 5' viral RNA cap. While deletion of Ifit1 had marginal effects on WNV-WT pathogenesis, WNV-E218A showed increased replication in peripheral tissues of Ifit1⁻/⁻ mice after subcutaneous infection, yet this failed to correlate with enhanced infection in the brain or lethality. In comparison, WNV-E218A was virulent after intracranial infection as judged by increased infection in different regions of the central nervous system (CNS) and a greater than 16,000-fold decrease in LD(50) values in Ifit1⁻/⁻ compared to wild type mice. Ex vivo infection experiments revealed cell-type specific differences in the ability of an Ifit1 deficiency to complement the replication defect of WNV-E218A. In particular, WNV-E218A infection was impaired in both wild type and Ifit1⁻/⁻ brain microvascular endothelial cells, which are believed to participate in blood-brain barrier (BBB) regulation of virus entry into the CNS. A deficiency of Ifit1 also was associated with increased neuronal death in vivo, which was both cell-intrinsic and mediated by immunopathogenic CD8⁺ T cells. Our results suggest that virulent strains of WNV have largely evaded the antiviral effects of Ifit1, and viral mutants lacking 2'-O methylation are controlled in vivo by Ifit1-dependent and -independent mechanisms in different cell types.


The Diesel Exhaust in Miners Study: IV. Estimating historical exposures to diesel exhaust in underground non-metal mining facilities.
We developed quantitative estimates of historical exposures to respirable elemental carbon (REC) for an epidemiologic study of mortality, including lung cancer, among diesel-exposed miners at eight non-metal mining facilities [the Diesel Exhaust in Miners Study (DEMS)]. Because there were no historical measurements of diesel exhaust (DE), historical REC (a component of DE) levels were estimated based on REC data from monitoring surveys conducted in 1998-2001 as part of the DEMS investigation. These values were adjusted for underground workers by carbon monoxide (CO) concentration trends in the mines derived from models of historical CO (another DE component) measurements and DE determinants such as engine horsepower (HP; 1 HP = 0.746 kW) and mine ventilation. CO was chosen to estimate historical changes because it was the most frequently measured DE component in our study facilities and it was found to correlate with REC exposure. Databases were constructed by facility and year with air sampling data and with information on the total rate of airflow exhausted from the underground operations in cubic feet per minute (CFM) (1 CFM = 0.0283 m³ min⁻¹), HP of the diesel equipment in use (ADJ HP), and other possible determinants. The ADJ HP purchased after 1990 (ADJ HP₁₉₉₀(+)) was also included to account for lower emissions from newer, cleaner engines. Facility-specific CO levels, relative to those in the DEMS survey year for each year back to the start of dieselization (1947-1967 depending on facility), were predicted based on models of observed CO concentrations and log-transformed (Ln) ADJ HP/CFM and Ln(ADJ HP₁₉₉₀(+)). The resulting temporal trends in relative CO levels were then multiplied by facility/department/job-specific REC estimates derived from the DEMS surveys personal measurements to obtain historical facility/department/job/year-specific REC exposure estimates. The facility-specific temporal trends of CO levels (and thus the REC estimates) generated from these models indicated that CO concentrations had been generally greater in the past than during the 1998-2001 DEMS surveys, with the highest levels ranging from 100 to 685% greater (median: 300%). These levels generally occurred between 1970 and the early 1980s. A comparison of the CO facility-specific model predictions with CO air concentration measurements from a 1976-1977 survey external to the modeling showed that our model predictions were slightly lower than those observed (median relative difference of 29%; range across facilities: 49 to -25%). In summary, we successfully modeled past CO concentration levels using selected determinants of DE exposure to derive retrospective estimates of REC exposure. The results suggested large variations in REC exposure levels both between and within the underground operations of the facilities and over time. These REC exposure estimates were in a plausible range and were used in the investigation of exposure-response relationships in epidemiologic analyses.


ATR disruption leads to chromosomal fragmentation and early embryonic lethality.
Although a small decrease in survival and increase in tumor incidence was observed in ATR(+/-) mice, ATR(-/-) embryos die early in development, subsequent to the blastocyst stage and prior to 7.5 days p.c. In culture, ATR(-/-) blastocysts cells continue to cycle into mitosis for 2 days but subsequently fail to expand and die of caspase-dependent apoptosis. Importantly, caspase-independent chromosome breaks are observed in ATR(-/-) cells prior to widespread apoptosis, implying that apoptosis is caused by a loss of genomic integrity. These data show that ATR is essential for early embryonic development and must function in processes other than regulation of p53.


The hypertension of Cushing's syndrome: controversies in the pathophysiology and focus on cardiovascular complications.
Cushing's syndrome is associated with increased mortality, mainly due to cardiovascular complications, which are sustained by the common development of systemic arterial hypertension and metabolic syndrome, which partially persist after the disease remission. Cardiovascular diseases and hypertension associated with endogenous hypercortisolism reveal underexplored peculiarities. The use of exogenous corticosteroids also impacts on hypertension and cardiovascular system, especially after prolonged treatment. The mechanisms involved in the development of hypertension differ, whether glucocorticoid excess is acute or chronic, and the source endogenous or exogenous, introducing inconsistencies among published studies. The pleiotropic effects of glucocorticoids and the overlap of the several regulatory mechanisms controlling blood pressure suggest that a rigorous comparison of in-vivo and in-vitro studies is necessary to draw reliable conclusions. This review, developed during the first 'Altogether to Beat Cushing's syndrome' workshop held in Capri in 2012, evaluates the most important peculiarities of hypertension associated with CS, with a particular focus on its pathophysiology. A critical appraisal of most significant animal and human studies is compared with a systematic review of the few available clinical trials. A special attention is dedicated to the description of the clinical features and cardiovascular damage secondary to glucocorticoid excess. On the basis of the consensus reached during the workshop, a pathophysiology-oriented therapeutic algorithm has been developed and it could serve as a first attempt to rationalize the treatment of hypertension in Cushing's syndrome.


IFNγ Increases M2 Muscarinic Receptor Expression in Cultured Sympathetic Neurons.
M2 muscarinic receptors are expressed on both parasympathetic and sympathetic nerve endings where they function as autoinhibitory receptors to limit release of acetylcholine and norepinephrine, respectively. M2 muscarinic receptor expression on parasympathetic nerves is decreased by viral infection and by gamma-interferon (IFNγ) and increased by dexamethasone; and these effects are of clinical relevance in the etiology and treatment of asthma. Whether IFNγ and dexamethasone similarly modulate M2 receptor expression on sympathetic nerves is not known. To address this question, we examined the effects of IFNγ and dexamethasone on M2 receptor expression at the mRNA and protein level in primary cultures of sympathetic neurons dissociated from the rat superior cervical ganglia (SCG). Semi-quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) indicated that neither IFNγ nor dexamethasone altered M2 receptor transcript levels. However, western blot analyses demonstrated that IFNγ, but not dexamethasone, increases M2 receptor protein expression in sympathetic neurons. Increased expression did not significantly alter subcellular localization of M2 receptors in sympathetic neurons as determined using immunocytochemistry. These findings indicate that M2 receptors are differentially regulated in different types of autonomic neurons, and they suggest a novel mechanism by which IFNγ may contribute to airway hyperreactivity in viral-induced asthma.


An unusual lymphocytic perivascular infiltration in tissues around contemporary metal-on-metal joint replacements.
Metal-on-metal bearing surfaces have been reintroduced for use during total hip replacement. To assess tissue reactions to various types of articulations, we studied the histological appearance of periprosthetic tissues retrieved from around metal-on-metal and metal-on-polyethylene total hip replacements and compared these findings with the appearance of control tissues retrieved at the time of primary arthroplasty. Periprosthetic tissues were obtained at the time of revision of twenty-five cobalt chromium-on-cobalt chromium, nine cobalt chromium-on-polyethylene, and ten titanium-on-polyethylene total hip arthroplasties. Control tissues were obtained from nine osteoarthritic hips at the time of primary total hip arthroplasty. Each tissue sample was processed for routine histological analysis, and sections were stained with hematoxylin and eosin. Quantitative stereological analysis was performed with use of light microscopy. Tissue samples obtained from hips with metal-on-metal implants displayed a pattern of well-demarcated tissue layers. A prominent feature, seen in seventeen of twenty-five tissue samples, was a pattern of perivascular infiltration of lymphocytes. In ten of the tissue samples obtained from hips with metal-on-metal prostheses, there was also an accumulation of plasma cells in association with macrophages that contained metallic wear-debris particles. The surfaces of tissues obtained from hips with metal-on-metal prostheses were more ulcerated than those obtained from hips with other types of implants, particularly in the region immediately superficial to areas of perivascular lymphocytic infiltration. The lymphocytic infiltration was more pronounced in samples obtained at the time of revision because of aseptic failure than in samples retrieved at the time of autopsy or during arthrotomy for reasons other than aseptic failure. Total-joint-replacement and surface-replacement designs of metal-on-metal prostheses were associated with similar results. Tissue samples obtained from hips with metal-on-polyethylene implants showed far less surface ulceration, much less distinction between tissue layers, no pattern of lymphocytic infiltration, and no plasma cells. The inflammation was predominantly histiocytic. Tissues retrieved from hips undergoing primary joint replacement showed dense scar tissue and minimal inflammation. The pattern and type of inflammation seen in periprosthetic tissues obtained from hips with metal-on-metal and metal-on-polyethylene implants are very different. At the present time, we do not know the prevalence or clinical implications of these histologic findings, but we suggest that they may represent a novel mode of failure for some metal-on-metal joint replacements.


An eco-friendly pathway to thermosensitive micellar nanoobjects via photoRAFT PISA: the full guide to poly(N-acryloylpyrrolidin)-block-polystyrene diblock copolymers.
Spherical macromolecular assemblies, so-called latexes, consisting of polystyrene (PS) resemble a relevant class of synthetic polymers used for a plethora of applications ranging from coatings or lubricants to biomedical applications. Their synthesis is usually tailored to the respective application where emulsifiers, radical initiators, or other additives still play a major role in achieving the desired properties. Herein, we demonstrate an alternative based on the photoiniferter reversible addition-fragmentation chain transfer (RAFT) polymerization-induced self-assembly (PISA) of Poly(N-acryloylpyrrolidin)-block-polystyrene (PAPy-b-PS). This approach yields monodisperse nanospheres with tunable sizes based on an aqueous formulation with only two ingredients. These nanospheres are additionally thermosensitive, meaning that they change their hydrodynamic diameter linearly with the temperature in a broad range between 10 °C and 70 °C. Combined with the eco-friendly synthesis in pure water at 40 °C, the herein presented route constitutes an unprecedented pathway to thermosensitive diblock copolymer aggregates in short reaction times without any additives.


Behavioural coping patterns in Parkinson's patients with visual hallucinations.
Visual Hallucinations are considered to affect about 20%-40% of patients with Parkinson's disease. They are generally seen as a side effect of this long-term illness and can severely affect the daily quality of life of patients. The aim of this study was to determine the coping patterns or strategies used by patients and establish whether the phenomenology and behaviours used by patients enabled control of the phenomenon. Demographic and clinical variables were recorded, including motor measures, cognitive status, and depressive symptoms. Patient with hallucinations were at a more advance stage of the disease and displayed more depressive symptoms than their non-hallucinating counterparts. Most patients used more than one constructive coping strategy, the most common were simple behavioural strategies based around motor action or cognitive approaches resulting in visual modification. In addition, humour was a common technique used by the patients to deal with the phenomenon. Emotional responses varied between patients, but it was found that the actual content of the hallucination was not directly associated with whether it caused trouble to the patient, but perceived stress was strongly correlated with the subjective disturbing nature of visual hallucinations (VHs). This study gives insight into the role of cognitive-behavioural approaches when dealing with VHs and opens up avenues for future studies in helping patient to deal with hallucinations.


Quantum-dot light-emitting diodes with a perfluorinated ionomer-doped copper-nickel oxide hole transporting layer.
Herein, we report all solution-processed green quantum-dot light-emitting diodes (G-QLEDs) by introducing a perfluorinated ionomer (PFI, Nafion 117) into quantum dots (QDs) to improve hole injection. To reduce the energy level mismatch between the hole transporting layer (HTL) and QDs and exciton quenching on the metal-oxide surface, a PFI-mixed copper-doped nickel oxide (Cu-NiO) HTL was introduced for G-QLEDs. Mixing Cu-NiO with a PFI increases the work function and induces phase separation between Cu-NiO and PFI; thus, energy band bending occurs on the surface such that effective hole injection can be possible. The phase-separated PFI molecules on HTL affect the thickness and compactness of G-QDs and make a smooth interface between G-QDs and HTL. The G-QLED with a PFI and Cu-NiO mixture HTL exhibits the maximum current efficiency (CEmax), power efficiency (PEmax), and external quantum efficiency (EQEmax) of 7.3 cd A-1, 2.1 lm W-1, and 2.14%, respectively, which are about 4 times those of the QLED with a Cu-NiO HTL.


Value of strain-wave sonoelastography as an imaging modality in assessment of benign acute myositis in children
Although sonoelastography is applied in assessment of many organs, studies for evaluation of muscles are very few in number and are mostly limited to adults. With this prospective study, we aimed to evaluate the value of sonoelastography in assessment of influenza related benign acute myositis in children. This study enrolled 25 patients with a clinical diagnosis of benign acute childhood myositis (BACM) and 25 age and sex-matched healthy controls. All patients presented to our emergency department with the complaint of inability to walk and had increased serum creatine kinase (CK) levels. All patients underwent strain elastography of the gastrocnemius muscle, and an elastography score was assigned to each patient by using a previously published 5 point-color scoring system. The findings were compared with those of the control group. No statistically significant difference was detected regarding age, weight, height, or body mass index (BMI) between patient and control groups. A statistically significant difference was found between the final elastography scores of the patient and control groups, mean values being 4.16 ± 0.75 versus 3.08 ±0.40, respectively (p < 0.001). Sonoelastography yielded a sensitivity of 80%, positive predictive value of 87%, specificity 88%, negative predictive value of 81.5%, and an overall accuracy of 84 %. Sonoelastography proves to be a valuable tool for diagnosis of BACM. It is one of the available ultrasound techniques in a radiology department and may particularly evolve to become a useful routine ancillary technique for investigation and follow-up in these cases.


QSAR modeling of the toxicity classification of superparamagnetic iron oxide nanoparticles (SPIONs) in stem-cell monitoring applications: an integrated study from data curation to model development.
The use of in silico approaches for the prediction of biomedical properties of nano-biomaterials (NBMs) can play a significant role in guiding and reducing wetlab experiments. Computational methods, such as data mining and machine learning techniques, can increase the efficiency and reduce the time and cost required for hazard and risk assesment and for designing new safer NBMs. A major obstacle in developing accurate and well-validated in silico models such as Nano Quantitative Structure-Activity Relationships (Nano-QSARs) is that although the volume of data published in the literature is increasing, the data are fragmented in many different publications and are not sufficiently curated for modelling purposes. Moreover, NBMs exhibit high complexity and heterogeneity in their structures, making data collection and curation and QSAR model development more challenging compared to traditional small molecules. The aim of this study was to construct and fully validate a Nano-QSAR model for the prediction of toxicological properties of superparamagnetic iron oxide nanoparticles (SPIONs), focusing on their application as Magnetic Resonance Imaging (MRI) contrast agents for non-invasive stem cell labelling and tracking. To achieve this goal, we first performed an extensive search through the literature for collecting and curating relevant data and we developed a dataset containing both physicochemical and toxicological properties of SPIONs. The data were analysed next, using Automated machine learning (Auto-ML) approaches for optimising the development and validation of nanotoxicity classification QSAR models of SPIONs. Further analysis of relative attribute importances revealed that physicochemical properties such as the size and the magnetic core are the dominant attributes correlated to the toxicity of SPIONs. Our results suggest that as more systematic information from NBM experimental tests becomes available, computational tools could play an important role in supporting the safety-by-design (SbD) concept in regenerative medicine and disease therapeutics.


Hepatocellular carcinoma confirmation, treatment, and survival in surveillance, epidemiology, and end results registries, 1992-2008.
Approaches to the diagnosis and management of hepatocellular carcinoma (HCC) are improving survival. In the Surveillance, Epidemiology, and End Results-13 registries, HCC stage, histological confirmation, and first-course surgery were examined. Among 21,390 HCC cases diagnosed with follow-up of vital status during 1998-2008, there were 4,727 (22%) with reported first-course invasive liver surgery, local tumor destruction, or both. The proportion with reported liver surgery or ablation was 39% among localized stage cases and only 4% among distant/unstaged cases. Though 70% of cases had histologically confirmed diagnoses, the proportion with confirmed diagnoses was higher among cases with reported invasive surgery (99%), compared to cases receiving ablation (81%) or no reported therapy (65%). Incidence rates of histologically unconfirmed HCC increased faster than those of confirmed HCC from 1992 to 2008 (8% versus 3% per year). Two encouraging findings were that incidence rates of localized-stage HCC increased faster than rates of regional- and distant-stage HCC combined (8% versus 4% per year), and that incidence rates of reported first-course surgery or tumor destruction increased faster than incidence rates of HCC without such therapy (11% versus 7%). Between 1975-1977 and 1998-2007, 5-year cause-specific HCC survival increased from just 3% to 18%. Survival was 84% among transplant recipients, 53% among cases receiving radiofrequency ablation at early stage, 47% among cases undergoing resection, and 35% among cases receiving local tumor destruction. Asian or Pacific Islander cases had significantly better 5-year survival (23%) than white (18%), Hispanic (15%), or black cases (12%). HCC survival is improving, because more cases are diagnosed and treated at early stages. Additional progress may be possible with continued use of clinical surveillance to follow individuals at risk for HCC, enabling early intervention.


Combining Biomarkers and Imaging for Short-Term Assessment of Cardiovascular Disease Risk in Apparently Healthy Adults.
Background Current strategies for cardiovascular disease (CVD) risk assessment focus on 10-year or longer timeframes. Shorter-term CVD risk is also clinically relevant, particularly for high-risk occupations, but is under-investigated. Methods and Results We pooled data from participants in the ARIC (Atherosclerosis Risk in Communities study), MESA (Multi-Ethnic Study of Atherosclerosis), and DHS (Dallas Heart Study), free from CVD at baseline (N=16 581). Measurements included N-terminal pro-B-type natriuretic peptide (>100 pg/mL prospectively defined as abnormal); high-sensitivity cardiac troponin T (abnormal >5 ng/L); high-sensitivity C-reactive protein (abnormal >3 mg/L); left ventricular hypertrophy by ECG (abnormal if present); carotid intima-media thickness, and plaque (abnormal >75th percentile for age and sex or presence of plaque); and coronary artery calcium (abnormal >10 Agatston U). Each abnormal test result except left ventricular hypertrophy by ECG was independently associated with increased 3-year risk of global CVD (myocardial infarction, stroke, coronary revascularization, incident heart failure, or atrial fibrillation), even after adjustment for traditional CVD risk factors and the other test results. When a simple integer score counting the number of abnormal tests was used, 3-year multivariable-adjusted global CVD risk was increased among participants with integer scores of 1, 2, 3, and 4, by ≈2-, 3-, 4.5- and 8-fold, respectively, when compared with those with a score of 0. Qualitatively similar results were obtained for atherosclerotic CVD (fatal or non-fatal myocardial infarction or stroke). Conclusions A strategy incorporating multiple biomarkers and atherosclerosis imaging improved assessment of 3-year global and atherosclerotic CVD risk compared with a standard approach using traditional risk factors.


Characterizing fruit ripening in plantain and Cavendish bananas: A proteomics approach.
The fruit physiology of banana cultivars other than Cavendish is poorly understood. To study the ripening process, samples were taken daily from plantain and Cavendish bananas and the ripening stages were determined. We present data from the green to the fully mature stage. By analyzing the protein abundances during ripening we provide some new insights into the ripening process and how plantains fruits are different. Multivariate analysis of the proteins was performed correlated to the starch dynamics. A drop in sucrose synthase and a rise of acid invertase during ripening indicated a change in the balance of the sucrose fate. During ripening, sugars may no longer be available for respiration since they are stored in the vacuoles, making citrate the preferred respiratory substrate. We found significant cultivar specific differences in granule-bound starch synthase, alpha- and beta amylases and cell wall invertase when comparing the protein content at the same ripening stage. This corroborates the difference in starch content/structure between both banana types. Differences in small heat shock proteins and in the cell wall-modifying enzyme xyloglucan endotransglucosylase/hydrolase support respectively the presumed higher carotenoid content and the firmer fruit structure of plantains. SIGNIFICANCE: We follow the proteome during ripening and correlate the proteins to the measured starch content. We discuss the changes in two contrasting genotypes. This gives us for the first time insight into the ripening of plantain and how this is different from the well-known Cavendish banana. This will revive the plantain breeding programs since for the first time we get insight into the plantain ripening.


What will make patients use a patient decision aid? A qualitative study on patients' perspectives on implementation barriers and facilitators.
Few studies focus on patients' views on factors influencing implementation of patient decision aids (PDAs). This study aims to explore patients' views on the factors influencing implementation of an "insulin choice" PDA in a primary care setting. This study used a descriptive qualitative study design. Interviews were conducted using a semistructured interview guide developed based on the theoretical domains framework. Nine in-depth interviews and three focus group discussions were conducted with patients with type 2 diabetes who have been advised to start insulin or were currently using insulin and those who had been seeking diabetes treatment in the clinic for more than 1 year. Interviews were conducted after the participants were familiarized with the PDA. Data were analysed using a thematic approach. Five themes emerged from the data analysis: (a) trust in the physician (patients preferred physicians to other health care providers in delivering the insulin PDA to them as they trusted physicians more when it comes to making decisions such as starting insulin), (b) physician's attitude (patients were more likely to trust a physician who is friendly and sympathetic hence would be more willing to use the insulin PDA), (c) physician's communication style (patients were more willing to use the insulin PDA if the physicians would take time and guide them in the PDA use), (d) conducive environment (patients preferred to read the PDA at home), and (e) cost (patients would not be willing to pay to use the insulin PDA unless they needed it). Patients want physicians to play a major role in the implementation of the insulin PDA; physicians' communication style and commitment may influence implementation outcomes. Health care authorities need to create a conducive environment and provide patients with free access to PDA to promote effective implementation.


Hemocue, an accurate bedside method of hemoglobin measurement?
Evaluate the accuracy of this bedside method to determine hemoglobin (Hb) concentration in general surgery over a wide range of Hb values and to determine potential sources of error. Accuracy of Hb measurement using HemoCue (AB Leo Diagnostics, Helsinborg, Sweden) was assessed in 140 surgical blood samples using 7 HemoCue devices in comparison with a CO-Oximeter (IL 482, Instrumentation Laboratory, Lexington, MA). To analyze potential sources of error, packed red cells and fresh frozen plasma were reconstituted to randomized Hb levels of 2-18 g/dL. In the surgical blood samples, the Hb concentration determined by the CO-Oximeter (HbCOOX) ranged from 5.1 to 16.7 g/dL and the Hb concentration measured by HemoCue (HbHC) from 4.7 to 16.0 g/dL. Bias (HbCOOX - HbHC) between HbCOOX and HbHC was 0.6+/-0.6 g/dL (mean +/- SD) or 5.4+/-5.0% (p < 0.001). Also in the reconstituted blood, the bias between HbCOOX and HbHC was significant (0.2+/-0.3 g/dL or 2.1+/-3.2%; p < 0.001). The microcuvette explained 68% of the variability between HbCOOX and HbHC. HemoCue thus underestimates the Hb concentration by 2-5% and exhibits a 8-10 times higher variability with only 86.4% of HbHC being within +/- 10% of HbCOOX. CONCLUSION. Although the mean bias between HbCOOX and HbHC was relatively low, Hb measurement by HemoCue exhibited a significant variability. Loading multiple microcuvettes and averaging the results may increase the accuracy of Hb measurement by HemoCue.


A Coarse Grained Model for a Lipid Membrane with Physiological Composition and Leaflet Asymmetry.
The resemblance of lipid membrane models to physiological membranes determines how well molecular dynamics (MD) simulations imitate the dynamic behavior of cell membranes and membrane proteins. Physiological lipid membranes are composed of multiple types of phospholipids, and the leaflet compositions are generally asymmetric. Here we describe an approach for self-assembly of a Coarse-Grained (CG) membrane model with physiological composition and leaflet asymmetry using the MARTINI force field. An initial set-up of two boxes with different types of lipids according to the leaflet asymmetry of mammalian cell membranes stacked with 0.5 nm overlap, reliably resulted in the self-assembly of bilayer membranes with leaflet asymmetry resembling that of physiological mammalian cell membranes. Self-assembly in the presence of a fragment of the plasma membrane protein syntaxin 1A led to spontaneous specific positioning of phosphatidylionositol(4,5)bisphosphate at a positively charged stretch of syntaxin consistent with experimental data. An analogous approach choosing an initial set-up with two concentric shells filled with different lipid types results in successful assembly of a spherical vesicle with asymmetric leaflet composition. Self-assembly of the vesicle in the presence of the synaptic vesicle protein synaptobrevin 2 revealed the correct position of the synaptobrevin transmembrane domain. This is the first CG MD method to form a membrane with physiological lipid composition as well as leaflet asymmetry by self-assembly and will enable unbiased studies of the incorporation and dynamics of membrane proteins in more realistic CG membrane models.


Differential susceptibility of C2C12 myoblasts and myotubes to group II phospholipase A2 myotoxins from crotalid snake venoms.
Group II phospholipase A(2) (PLA(2)) myotoxins isolated from Viperidae/Crotalidae snake venoms induce a rapid cytolytic effect upon diverse cell types in vitro. Previous studies suggested that this effect could be more pronounced on skeletal muscle myotubes than on other cell types, including undifferentiated myoblasts. This study utilized the murine skeletal muscle C2C12 cell line to investigate whether differentiated myotubes are more susceptible than myoblasts, and if this characteristic is specific for the group II myotoxic PLA(2)s. The release of lactic dehydrogenase was quantified as a measure of cytolysis, 3 h after cell exposure to different group II PLA(2)s purified from Bothrops asper, Atropoides nummifer, Cerrophidion godmani, and Bothriechis schlegelii venoms. In addition, susceptibility to lysis induced by synthetic melittin and group III PLA(2) from bee (Apis mellifera) venom, as well as by anionic, cationic, and neutral detergents, was comparatively evaluated on the two cultures. Myotubes were significantly more susceptible to group II PLA(2) myotoxins, but not to the other agents tested, under the same conditions. Moreover, the increased susceptibility of myotubes over myoblasts was also demonstrated with two cytolytic synthetic peptides, derived from the C-terminal region of Lys49 PLA(2) myotoxins, that reproduce the action of their parent proteins. These results indicate that fusion and differentiation of myoblasts into myotubes induce changes that render these cells more susceptible to the toxic mechanism of group II PLA(2) myotoxins, but not to general perturbations of membrane homeostasis. Such changes are likely to involve myotoxin acceptor site(s), which remain(s) to be identified.


Internal oblique radiographs for diagnosis of nondisplaced or minimally displaced lateral condylar fractures of the humerus in children.
Several radiographic criteria based on only standard anteroposterior and lateral radiographs have been suggested for predicting the stability of lateral condylar fractures of the humerus in children. Despite adherence to these guidelines, further displacement often occurs after the initial diagnosis and stability assessment. The purpose of this study was to define the usefulness of an internal oblique radiograph in the assessment of these fractures. We prospectively studied fifty-four lateral humeral condylar fractures in fifty-four children seen between March 2002 and December 2004 to determine the efficacy of internal oblique radiographs in gauging the amount of displacement and the fracture pattern. Plain anteroposterior, lateral, internal oblique, and external oblique radiographs were initially made for all fifty-four patients, and three-dimensional computed tomography scans were performed for seven patients. A large percentage (70%) of the fractures were seen to have different amounts of displacement on the anteroposterior and internal oblique radiographs, and a large percentage (75%) were seen to have different fracture patterns, according to the criteria suggested by Finnbogason et al., on the two views. The internal oblique radiograph was more accurate for demonstrating the fracture gap (thirty cases were seen to have more displacement on that view) and for demonstrating the fracture pattern (twenty cases were seen to have more instability on the internal oblique view). We also confirmed the direction of the fracture line and the location of the fragment more accurately with three-dimensional computed tomography. It is not optimal to evaluate the amount of displacement and the stability of a lateral condylar fracture of the humerus in children on the basis of just anteroposterior and lateral elbow radiographs. Classifications should be based on the greatest displacement seen on at least three radiographic views, especially the internal oblique view.


Correction of chromosomal mutation and random integration in embryonic stem cells with helper-dependent adenoviral vectors.
For gene therapy of inherited diseases, targeted integration/gene repair through homologous recombination (HR) between exogenous and chromosomal DNA would be an ideal strategy to avoid potentially serious problems of random integration such as cellular transformation and gene silencing. Efficient sequence-specific modification of chromosomes by HR would also advance both biological studies and therapeutic applications of a variety of stem cells. Toward these goals, we developed an improved strategy of adenoviral vector (AdV)-mediated HR and examined its ability to correct an insertional mutation in the hypoxanthine phosphoribosyl transferase (Hprt) locus in male mouse ES cells. The efficiency of HR was compared between four types of AdVs that contained various lengths of homologies at the Hprt locus and with various multiplicities of infections. The frequency of HR with helper-dependent AdVs (HD AdVs) with an 18.6-kb homology reached 0.2% per transduced cell at a multiplicity of infection of 10 genomes per cell. Detection of random integration at DNA levels by PCR revealed extremely high efficiency of 5% per cell. We also isolated and characterized chromosomal sites where HD AdVs integrated in a random manner. In contrast to retroviral, lentiviral, and adeno-associated viral vectors, which tend to integrate into genes, the integration sites of AdV was distributed randomly inside and outside genes. These findings suggest that HR mediated by HD AdVs is efficient and relatively safe and might be a new viable option for ex vivo gene therapy as well as a tool for chromosomal manipulation of a variety of stem cells.


The amino-terminal region of the retinoblastoma gene product binds a novel nuclear matrix protein that co-localizes to centers for RNA processing.
The tumor suppressing capacity of the retinoblastoma protein (p110RB) is dependent on interactions made with cellular proteins through its carboxy-terminal domains. How the p110RB amino-terminal region contributes to this activity is unclear, though evidence now indicates it is important for both growth suppression and regulation of the full-length protein. We have used the yeast two-hybrid system to screen for cellular proteins which bind to the first 300 amino acids of p110RB. The only gene isolated from this screen encodes a novel 84-kD nuclear matrix protein that localizes to subnuclear regions associated with RNA processing. This protein, p84, requires a structurally defined domain in the amino terminus of p110RB for binding. Furthermore, both in vivo and in vitro experiments demonstrate that p84 binds preferentially to the functionally active, hypophosphorylated form of p110RB. Thus, the amino terminus of p110RB may function in part to facilitate the binding of growth promoting factors at subnuclear regions actively involved in RNA metabolism.


The spatial and temporal scales of local dengue virus transmission in natural settings: a retrospective analysis.
Dengue is a vector-borne disease caused by the dengue virus (DENV). Despite the crucial role of Aedes mosquitoes in DENV transmission, pure vector indices poorly correlate with human infections. Therefore there is great need for a better understanding of the spatial and temporal scales of DENV transmission between mosquitoes and humans. Here, we have systematically monitored the circulation of DENV in individual Aedes spp. mosquitoes and human patients from Caratinga, a dengue endemic city in the state of Minas Gerais, in Southeast Brazil. From these data, we have developed a novel stochastic point process pattern algorithm to identify the spatial and temporal association between DENV infected mosquitoes and human patients. The algorithm comprises of: (i) parameterization of the variogram for the incidence of each DENV serotype in mosquitoes; (ii) identification of the spatial and temporal ranges and variances of DENV incidence in mosquitoes in the proximity of humans infected with dengue; and (iii) analysis of the association between a set of environmental variables and DENV incidence in mosquitoes in the proximity of humans infected with dengue using a spatio-temporal additive, geostatistical linear model. DENV serotypes 1 and 3 were the most common virus serotypes detected in both mosquitoes and humans. Using the data on each virus serotype separately, our spatio-temporal analyses indicated that infected humans were located in areas with the highest DENV incidence in mosquitoes, when incidence is calculated within 2.5-3 km and 50 days (credible interval 30-70 days) before onset of symptoms in humans. These measurements are in agreement with expected distances covered by mosquitoes and humans and the time for virus incubation. Finally, DENV incidence in mosquitoes found in the vicinity of infected humans correlated well with the low wind speed, higher air temperature and northerly winds that were more likely to favor vector survival and dispersal in Caratinga. We have proposed a new way of modeling bivariate point pattern on the transmission of arthropod-borne pathogens between vector and host when the location of infection in the latter is known. This strategy avoids some of the strong and unrealistic assumptions made by other point-process models. Regarding virus transmission in Caratinga, our model showed a strong and significant association between high DENV incidence in mosquitoes and the onset of symptoms in humans at specific spatial and temporal windows. Together, our results indicate that vector surveillance must be a priority for dengue control. Nevertheless, localized vector control at distances lower than 2.5 km around premises with infected vectors in densely populated areas are not likely to be effective.


Cytoplasmic polyadenylation and cytoplasmic polyadenylation element-dependent mRNA regulation are involved in Xenopus retinal axon development.
Translation in axons is required for growth cone chemotropic responses to many guidance cues. Although locally synthesized proteins are beginning to be identified, how specific mRNAs are selected for translation remains unclear. Control of poly(A) tail length by cytoplasmic polyadenylation element (CPE) binding protein 1 (CPEB1) is a conserved mechanism for mRNA-specific translational regulation that could be involved in regulating translation in axons. We show that cytoplasmic polyadenylation is required in Xenopus retinal ganglion cell (RGC) growth cones for translation-dependent, but not translation-independent, chemotropic responses in vitro, and that inhibition of CPE binding through dominant-negative interference severely reduces axon outgrowth in vivo. CPEB1 mRNA transcripts are present at low levels in RGCs but, surprisingly, CPEB1 protein was not detected in eye or brain tissue, and CPEB1 loss-of-function does not affect chemotropic responses or pathfinding in vivo. UV cross-linking experiments suggest that CPE-binding proteins other than CPEB1 in the retina regulate retinal axon development. These results indicate that cytoplasmic polyadenylation and CPE-mediated translational regulation are involved in retinal axon development, but that CPEB1 may not be the key regulator of polyadenylation in the developing retina.


Saturn's Ring and the Satellites of Jupiter: Interpretations of Infrared Spectra.
Infrared spectra of Saturn's ring correspond closely with the reflection spectrum of water ice. This result supports Kuiper's suggestion for the composition of the ring. The absence of the strong absorption at 8873 A in spectra of Io, Ganymede, and Callisto leads to a new upper limit for the abundance of methane in possible atmospheres of these satellites.


Training-Monitoring Engagement: An Evidence-Based Approach in Elite Sport.
Poor athlete buy-in and adherence to training-monitoring systems (TMS) can be problematic in elite sport. This is a significant issue, as failure to record, interpret, and respond appropriately to negative changes in athlete well-being and training status may result in undesirable consequences such as maladaptation and/or underperformance. This study examined the perceptions of elite athletes to their TMS and their primary reasons for noncompletion. Nine national-team sprint athletes participated in semistructured interviews on their perceptions of their TMS. Interview data were analyzed qualitatively, based on grounded theory, and TMS adherence information was collected. Thematic analysis showed that athletes reported their main reason for poor buy-in to TMS was a lack of feedback on their monitoring data from key staff. Furthermore, training modifications made in response to meaningful changes in monitoring data were sometimes perceived to be disproportionate, resulting in dishonest reporting practices. Perceptions of opaque or unfair decision making on training-program modifications and insufficient feedback were the primary causes for poor athlete TMS adherence. Supporting TMS implementation with a behavioral-change model that targets problem areas could improve buy-in and enable limited resources to be appropriately directed.


Linking brain structure and activation in temporoparietal junction to explain the neurobiology of human altruism.
Human altruism shaped our evolutionary history and pervades social and political life. There are, however, enormous individual differences in altruism. Some people are almost completely selfish, while others display strong altruism, and the factors behind this heterogeneity are only poorly understood. We examine the neuroanatomical basis of these differences with voxel-based morphometry and show that gray matter (GM) volume in the right temporoparietal junction (TPJ) is strongly associated with both individuals' altruism and the individual-specific conditions under which this brain region is recruited during altruistic decision making. Thus, individual differences in GM volume in TPJ not only translate into individual differences in the general propensity to behave altruistically, but they also create a link between brain structure and brain function by indicating the conditions under which individuals are likely to recruit this region when they face a conflict between altruistic and selfish acts.


Severe polyposis in Apc(1322T) mice is associated with submaximal Wnt signalling and increased expression of the stem cell marker Lgr5.
Adenomatous polyposis coli (APC) is a tumour suppressor gene mutated in the germline of patients with familial adenomatous polyposis (FAP) and somatically in most colorectal cancers. APC mutations impair β-catenin degradation, resulting in increased Wnt signalling. The most frequent APC mutation is a codon 1309 truncation that is associated with severe FAP. A previous study compared two mouse models of intestinal tumorigenesis, Apc(R850X) (Min) and Apc(1322T) (1322T), the latter a model of human codon 1309 changes. 1322T mice had more severe polyposis but, surprisingly, these tumours had lower levels of nuclear β-catenin than Min tumours. The consequences of these different β-catenin levels were investigated. Enterocytes were isolated from 1322T and Min tumours by microdissection and gene expression profiling was performed. Differentially expressed Wnt targets and other stem cell markers were validated using quantitative PCR, in situ hybridisation and immunohistochemistry. As expected, lower nuclear β-catenin levels in 1322T lesions were associated with generally lower levels of Wnt target expression. However, expression of the Wnt target and stem cell marker Lgr5 was significantly higher in 1322T tumours than in Min tumours. Other stem cell markers (Musashi1, Bmi1 and the Wnt target Cd44) were also at higher levels in 1322T tumours. In addition, expression of the Bmp antagonist Gremlin1 was higher in 1322T tumours, together with lower Bmp2 and Bmp4 expression. The severe phenotype caused by truncation of Apc at codon 1322 is associated with an increased number of stem cells. Thus, a submaximal level of Wnt signalling favours the stem cell phenotype and this may promote tumorigenesis. A level of Wnt signalling exists that is too high for optimal tumour growth.


HLA-DRB1 polymorphisms and autoimmune responses to desmogleins in Japanese patients with pemphigus.
Pemphigus vulgaris (PV) and pemphigus foliaceus (PF) are caused by autoantibodies against keratinocyte adhesion molecules desmoglein 3 (Dsg3) and desmoglein 1 (Dsg1), respectively. To determine possible major histocompatibility complex (MHC) class II associations with autoantibody responses to desmogleins, haplotype and allele distributions, along with molecular polymorphisms of HLA-DR and -DQ genes were analyzed based on the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) results in 85 Japanese patients with pemphigus. Each of 55 PV patients carried at least one allele of HLA-DRB1*04 and DRB1*14 subtypes, with significant increases of HLA-DRB1*0406/DQA1*0301/DQB1*0302, DRB1*14/DQA1*0104/DQB1*05 and DRB1*1406/DQA1*0503/ DQB1*0301 haplotypes compared to normal controls. The HLA-DRB1*04 and DRB*14 alleles carried by PV patients shared hydrophobic amino acid residues Phe26, Leu67 and Val86, as well as hydrophilic amino acid residues at positions 70 and 71 on the DRB1 beta chain. HLA-DR/DQ distributions did not differ among PV patients according to the presence or absence of anti-Dsg1 co-existing with anti-Dsg3. Thirty PF patients, all producing autoantibodies only to Dsg1, showed more diverse HLA-DR/DQ distributions, sharing hydrophobic amino acid residues at positions 26 and 67, as well as hydrophilic amino acid residues at positions 70 and 71, of the DRB1 chain. These findings suggest that autoantibody responses to desmogleins might be regulated by amino acid residues at positions 26, 67, 70, 71 and 86 at peptide binding sites of HLA-DRB1 molecules, and that autoimmune responses to Dsg3 might be more strictly regulated by specific amino acid residues at these positions on the HLA-DRB1 chain than responses to Dsg1.


Graphical Discrimination of New Zealand Honey from International Honey Using Elemental Analysis.
This study investigated the potential of utilising the elemental fingerprinting of honey to differentiate New Zealand (NZ) honey from that of international origin. Twenty elements were analysed by ICP-MS in 352 honeys from 34 various countries. Of these, 323 honeys (245 New Zealand honeys, 78 international) and two subsets of data (NZ and European origin, n = 306, and, NZ and Denmark/Germany, n = 280) were visualised using principal component analysis (PCA). For the NZ/Europe subset, 42.2% of data was explained in the first two principal components. Statistical classification rules were also derived using linear discriminant analysis (LDA) and decision tree analysis. Various combinations of elements were explored for classification, considering the effect of soil-derived elements and those from anthropogenic sources. A high degree of accuracy (at least 90%) for the characterisation of New Zealand honey was observed for all statistical models, showing the robustness of these analyses. When using decision tree analysis to distinguish New Zealand samples from international samples, a tree with five terminal nodes (using Cs, Ba and Rb) was created with 92.4% accuracy. This work has demonstrated that elemental fingerprints of honey are a promising tool for categorising New Zealand honey from other geographical locations.


Mesenchymal stem cell-derived molecules directly modulate hepatocellular death and regeneration in vitro and in vivo.
Orthotopic liver transplantation is the only proven effective treatment for fulminant hepatic failure (FHF), but its use is limited because of organ donor shortage, associated high costs, and the requirement for lifelong immunosuppression. FHF is usually accompanied by massive hepatocellular death with compensatory liver regeneration that fails to meet the cellular losses. Therefore, therapy aimed at inhibiting cell death and stimulating endogenous repair pathways could offer major benefits in the treatment of FHF. Recent studies have demonstrated that mesenchymal stem cell (MSC) therapy can prevent parenchymal cell loss and promote tissue repair in models of myocardial infarction, acute kidney failure, and stroke through the action of trophic secreted molecules. In this study, we investigated whether MSC therapy can protect the acutely injured liver and stimulate regeneration. In a D-galactosamine-induced rat model of acute liver injury, we show that systemic infusion of MSC-conditioned medium (MSC-CM) provides a significant survival benefit and prevents the release of liver injury biomarkers. Furthermore, MSC-CM therapy resulted in a 90% reduction of apoptotic hepatocellular death and a three-fold increment in the number of proliferating hepatocytes. This was accompanied by a dramatic increase in the expression levels of 10 genes known to be up-regulated during hepatocyte replication. Direct antiapoptotic and promitotic effects of MSC-CM on hepatocytes were demonstrated using in vitro assays. These data provide the first clear evidence that MSC-CM therapy provides trophic support to the injured liver by inhibiting hepatocellular death and stimulating regeneration, potentially creating new avenues for the treatment of FHF.


Receptor tyrosine phosphatases regulate axon guidance across the midline of the Drosophila embryo.
Neural receptor-linked protein tyrosine phosphatases (RPTPs) are required for guidance of motoneuron and photoreceptor growth cones in Drosophila. These phosphatases have not been implicated in growth cone responses to specific guidance cues, however, so it is unknown which aspects of axonal pathfinding are controlled by their activities. Three RPTPs, known as DLAR, DPTP69D, and DPTP99A, have been genetically characterized thus far. Here we report the isolation of mutations in the fourth neural RPTP, DPTP10D. The analysis of double mutant phenotypes shows that DPTP10D and DPTP69D are necessary for repulsion of growth cones from the midline of the embryonic central nervous system. Repulsion is thought to be triggered by binding of the secreted protein Slit, which is expressed by midline glia, to Roundabout (Robo) receptors on growth cones. Robo repulsion is downregulated by the Commissureless (Comm) protein, allowing axons to cross the midline. Here we show that the Rptp mutations genetically interact with robo, slit and comm. The nature of these interactions suggests that DPTP10D and DPTP69D are positive regulators of Slit/Roundabout repulsive signaling. We also show that elimination of all four neural RPTPs converts most noncrossing longitudinal pathways into commissures that cross the midline, indicating that tyrosine phosphorylation controls the manner in which growth cones respond to midline signals.


Self-monitoring of blood glucose in diabetes.
A key aim of management in people with diabetes mellitus is to control blood glucose concentrations, in order to avoid the complications of persistently raised levels (with their attendant healthcare costs) and improve quality of life. However, such control is associated with an increased likelihood of hypoglycaemia, particularly among patients on insulin therapy. In pursuit of good glycaemic control while avoiding hypoglycaemia, self-monitoring of blood glucose is widely acknowledged as a routine part of management in patients with type 1 diabetes. However, the cost of monitoring products has now become a major issue for healthcare providers. This has led to some restrictions being placed on self-monitoring, especially for patients with type 2 diabetes, where the evidence for a benefit is less clear-cut. With this in mind, we review the data relating to the benefits of self-monitoring in adults with diabetes, particularly those with type 2 disease.


Crystal structures of alpha-lytic protease complexes with irreversibly bound phosphonate esters.
The structures of the complexes with alpha-lytic protease of both phosphorus stereoisomers of N-[(2S)-2-[[[(1R)-1-[N-[(tert-butyloxycarbonyl)-L-alanyl-L-alanyl- L-prolyl]amino]-2-methylpropyl]-phenoxyphosphinyl]oxy]propanoyl]- L-alanine methyl ester, an analogue of the peptide Boc-Ala-Ala-Pro-Val-Ala-Ala where Val is replaced with an analogous phosphonate phenyl ester and the subsequent Ala is replaced with lactate, have been determined to high resolution (1.9 A) by X-ray crystallography. Both stereoisomers inactivate the enzyme but differ by a factor of 2 in the second-order rate constant for inactivation [Sampson, N. S., & Bartlett, P. A. (1991) Biochemistry (preceding paper in this issue)]. One isomer (B) forms a tetrahedral adduct in which the phosphonate phenyl ester is displaced by the active site serine (S195) and interacts with the enzyme across seven substrate recognition sites that span both sides of the scissile bond. Seven hydrogen bonds are formed with the enzyme, and 510 A2 of hydrophobic surface area is buried when the inhibitor interacts with the enzyme. Although two hydrogen bonds are gained by incorporation of two residues on the C-terminal side of the scissile bond into the inhibitor, there is very little adjustment in the structure of the enzyme in this region. Surprisingly, the active site histidine (H57) does not interact with the phosphonate, apparently because the phosphonate lacks negative charge in or near the oxyanion hole, and instead, the side chain rotates out of the active site cleft and hydrogen bonds with solvent. The other isomer (A) forms a mixture of two different tetrahedral adducts in the active site, both covalently bonded to Ser 195. One adduct, at approximately 58% occupancy, is exactly the same in structure as the complex formed with isomer B, and the other adduct, at 42% occupancy, has lost the two residues C-terminal to the scissile bond by hydrolysis. In the lower occupancy structure, His 57 does not rotate out of the active site and forms a hydrogen bond with the phosphonate oxygen instead. The structures of both complexes were insensitive to pH. As very little change in structure accompanies the histidine rotation, the complex with isomer B provides an excellent mimic for the structure of the transition state (or high-energy reaction intermediate) that spans both sides of the scissile bond.


Maximizing antimalarial efficacy and the importance of dosing strategies.
Artemisinin-based combination therapies (ACTs) are the cornerstone for the treatment of malaria. However, confirmed resistance to artemisinins in South-East Asia, and reports of reduced efficacy of ACTs raise major concerns for malaria treatment and control. Without new drugs to replace artemisinins, it is essential to define dosing strategies that maximize therapeutic efficacy, limit the spread of resistance, and preserve the clinical value of ACTs. It is important to determine the extent to which reduced efficacy of ACTs reflects true resistance versus sub-optimal dosing, and quantify other factors that determine treatment failure. Pooled analyses of individual patient data from multiple clinical trials, by investigators in the Worldwide Antimalarial Resistance Network, have shown high overall efficacy for three widely used ACTs, artemether-lumefantrine, artesunate-amodiaquine, and dihydroartemisinin-piperaquine. Analyses also highlight that suboptimal dosing leads to increased risk of treatment failure, especially among children. In the most recent study, an analysis of clinical trials of artesunate-amodiaquine, widely used among children in Africa, revealed a superior efficacy for fixed-dose combination tablets compared to loose non-fixed dose combinations. This highlights the benefits of fixed-dose combinations as a practical strategy for ensuring optimal antimalarial dosing and maximizing efficacy. Please see related article: http://www.biomedcentral.com/1741-7015/13/66.


Efficacy and tolerability of antibiotics in patients undergoing H. pylori eradication.
Helicobacter pylori (H. pylori) infection is one of the most common gastrointestinal diseases. An increasing number of people undergo different treatment options. Unfortunately, H. pylori therapy may be troublesome for drug side effects and inefficacious for resistance to antibiotics. One hundred and ninety-three (193) H. pylori-positive patients were randomly assigned to one of the following 7-day treatments: Group A (N = 64): amoxicillin, clarithromycin and rabeprazole; Group B (N = 64): tinidazole, clarithromycin and ranitidine bismuth citrate; Group C (N = 65): tinidazole, clarithromycin and rabeprazole Eradication was assessed by 13C-Urea Breath Test 6-8 weeks after the end of the therapy. Not-eradicated patients underwent a second cycle with tinidazole, tetracycline, bismuth and rabeprazole. All patients were asked to complete a validated questionnaire regarding presence and intensity of drug side effects. One hundred and eighty-eight out of the 193 H. pylori-positive patients (96%) completed therapy. No significant difference in eradication rates was observed among the three groups both in intention to treat analysis and in per protocol analysis. No significant difference in incidence of side effects occurred among groups after the first-line regimens: 48% in group A, 44% in group B and 46% in group C. Twenty-two out of the 193 enrolled subjects (11%) were not eradicated after the first-line therapy. Among them, 86% were successfully eradicated by the tinidazole, tetracycline, bismuth and rabeprazole therapy. Moreover, during quadruple therapy, a higher prevalence and intensity of side effects than in each one of the groups submitted to the first-line triple therapy was observed. This study shows that triple rabeprazole-based eradicating regimens are effective and safe. Incidence of side effects seems low and similar in different three-drug regimens used. Quadruple therapy, which appear highly efficacious as a second line therapy, is associated with a significantly higher incidence of side effects when compared to first-line treatment.


Overview of randomized trials of postoperative adjuvant radiotherapy in breast cancer.
An overview of the mortality results of the mature trials in which radiotherapy was a randomized option after simple or radical mastectomy is presented. The principal aim of the overview was to study long-term survival, and recent trials, often including chemotherapy options, have not been included. A total of 7941 patients were entered into these trials and 4148 deaths occurred. No differences were found in survival in the first 10 years of follow-up, in trials employing either radical mastectomy or simple mastectomy. Follow-up information after 10 years came mostly from trials employing radical mastectomy with or without irradiation, and in these trials a significant excess of deaths was observed among patients given radiotherapy (P less than 0.001, all trials; P = 0.002, radical mastectomy trials only). Further data collection will be necessary to determine which causes of death are elevated.


An investigation into the involvement of defense signaling pathways in components of the nonhost resistance of Arabidopsis thaliana to rust fungi also reveals a model system for studying rust fungal compatibility.
Seventeen accessions of Arabidopsis thaliana inoculated with the cowpea rust fungus Uromyces vignae exhibited a variety of expressions of nonhost resistance, although infection hypha growth typically ceased before the formation of the first haustorium, except in Ws-0. Compared with wild-type plants, there was no increased fungal growth in ndr1 or eds1 mutants defective in two of the signal cascades regulated by the major class of Arabidopsis host resistance genes. However, in the Col-0 background, infection hyphae of U. vignae and two other rust fungi were longer in sid2 mutants defective in an enzyme that synthesizes salicylic acid (SA), in npr1 mutants deficient in a regulator of the expression of SA-dependent pathogenesis related (PR) genes, and in NahG plants containing a bacterial salicylate hydroxylase. Infection hyphae of U. vignae and U. appendiculatus but not of Puccinia helianthi were also longer in jar1 mutants, which are defective in the jasmonic acid defense signaling pathway. Nevertheless, haustorium formation increased only for the Uromyces spp. and only in sid2 mutants or NahG plants. Rather than the hypersensitive cell death that usually accompanies haustorium formation in nonhost plants, Arabidopsis typically encased haustoria in calloselike material. Growing fungal colonies of both Uromyces spp., indicative of a successful biotrophic relationship between plant and fungus, formed in NahG plants, but only U. vignae formed growing colonies in the sid2 mutants and cycloheximide-treated wild-type plants. Growing colonies did not develop in NahG tobacco or tomato plants. These data suggest that nonhost resistance of Arabidopsis to rust fungi primarily involves the restriction of infection hypha growth as a result of defense gene expression. However, there is a subsequent involvement of SA but not SA-dependent PR genes in preventing the Uromyces spp. from forming the first haustorium and establishing a sufficient biotrophic relationship to support further fungal growth. The U. vignae-Arabidopsis combination could allow the application of the powerful genetic capabilities of this model plant to the study of compatibility as well as nonhost resistance to rust fungi.