Investigation of metal ion binding in phosphonoacetaldehyde hydrolase identifies sequence markers for metal-activated enzymes of the HAD enzyme superfamily.
The 2-haloalkanoic acid dehalogenase (HAD) family, which contains both carbon and phosphoryl transferases, is one of the largest known enzyme superfamilies. HAD members conserve an alpha,beta-core domain that frames the four-loop active-site platform. Each loop contributes one or more catalytic groups, which function in mediating the core chemistry (i.e., group transfer). In this paper, we provide evidence that the number of carboxylate residues on loop 4 and their positions (stations) on the loop are determinants, and therefore reliable sequence markers, for metal ion activation among HAD family members. Using this predictor, we conclude that the vast majority of the HAD members utilize a metal cofactor. Analysis of the minimum requirements for metal cofactor binding was carried out using Mg(II)-activated Bacillus cereus phosphonoacetaldehyde hydrolase (phosphonatase) as an experimental model for metal-activated HAD members. Mg(II) binding occurs via ligation to the loop 1 Asp12 carboxylate and Thr14 backbone carbonyl and to the loop 4 Asp186 carboxylate. The loop 4 Asp190 forms a hydrogen bond to the Mg(II) water ligand. X-ray structure determination of the D12A mutant in the presence of the substrate phosphonoacetaldehyde showed that replacement of the loop 1 Asp, common to all HAD family members, with Ala shifts the position of Mg(II), thereby allowing innersphere coordination to Asp190 and causing a shift in the position of the substrate. Kinetic analysis of the loop 4 mutants showed that Asp186 is essential to cofactor binding while Asp190 simply enhances it. Within the phosphonatase subfamily, Asp186 is stringently conserved, while either position 185 or position 190 is used to position the second loop 4 Asp residue. Retention of a high level of catalytic activity in the G185D/D190G phosphonatase mutant demonstrated the plasticity of the metal binding loop, reflected in the variety of combinations in positioning of two or three Asp residues along the seven-residue motif of the 2700 potential HAD sequences that were examined.


Whale shark economics: a valuation of wildlife tourism in South Ari Atoll, Maldives.
Whale sharks attract large numbers of tourists, divers and snorkelers each year to South Ari Atoll in the Republic of Maldives. Yet without information regarding the use and economic extent of the attraction, it is difficult to prioritize conservation or implement effective management plans. We used empirical recreational data and generalized mixed statistical models to conduct the first economic valuation (with direct spend as the primary proxy) of whale shark tourism in Maldives. We estimated that direct expenditures for whale shark focused tourism in the South Ari Marine Protected Area for 2012 and 2013 accounted for US$7.6 and $9.4 million respectively. These expenditures are based on an estimate of 72,000-78,000 tourists who are involved in whale shark excursions annually. That substantial amount of income to resort owners and operators, and tourism businesses in a relatively small area highlights the need to implement regulations and management that safeguard the sustainability of the industry through ensuring guest satisfaction and whale shark conservation.


Orienting of attention and Parkinson's disease: tactile inhibition of return and response inhibition.
There is growing evidence for cognitive impairments in Parkinson's disease (PD), including in the orienting of attention and inhibition of return (IOR). IOR refers to the slowing of a response to a target stimulus presented in the same location as a previous stimulus. While some researchers have reported normal levels of visual IOR in PD patients using cue-target tasks, others have reported significant reductions in IOR in this patient group. However, the inhibitory effects observed in cue-target tasks may reflect non-ocular response inhibition associated with withholding a response from the cue stimulus, rather than attentional or oculomotor processes. Many researchers working with normal participants have circumvented this confound by using a target-target task, in which a response is made to all peripheral stimuli. Here, we compared IOR measured in cue-target and target-target tasks, using tactile rather than visual stimuli. Both the PD and the control groups exhibited significant inhibitory effects in the cue-target task, but only the control group exhibited significant IOR in the target-target task. Our results demonstrate a reduction, or elimination, of IOR in PD and this change may have been underestimated in previous studies, in which methodologically flawed cue-target tasks were used. This reduction in IOR may reflect impaired inhibitory processes or hyper-reflexive orienting in parkinsonian patients.


Characterization of Fusarium isolates from asparagus fields in southwestern Ontario and influence of soil organic amendments on Fusarium crown and root rot.
Fusarium crown and root rot (FCRR) of asparagus has a complex etiology with several soilborne Fusarium spp. as causal agents. Ninety-three Fusarium isolates, obtained from plant and soil samples collected from commercial asparagus fields in southwestern Ontario with a history of FCRR, were identified as Fusarium oxysporum (65.5%), F. proliferatum (18.3%), F. solani (6.4%), F. acuminatum (6.4%), and F. redolens (3.2%) based on morphological or cultural characteristics and polymerase chain reaction (PCR) analysis with species-specific primers. The intersimple-sequence repeat PCR analysis of the field isolates revealed considerable variability among the isolates belonging to different Fusarium spp. In the in vitro pathogenicity screening tests, 50% of the field isolates were pathogenic to asparagus, and 22% of the isolates caused the most severe symptoms on asparagus. The management of FCRR with soil organic amendments of pelleted poultry manure (PPM), olive residue compost, and fish emulsion was evaluated in a greenhouse using three asparagus cultivars of different susceptibility in soils infested with two of the pathogenic isolates (F. oxysporum Fo-1.5 and F. solani Fs-1.12). Lower FCRR symptom severity and higher plant weights were observed for most treatments on 'Jersey Giant' and 'Grande' but not on 'Mary Washington'. On all three cultivars, 1% PPM consistently reduced FCRR severity by 42 to 96% and increased plant weights by 77 to 152% compared with the Fusarium control treatment. Populations of Fusarium and total bacteria were enumerated after 1, 3, 7, and 14 days of soil amendment. In amended soils, the population of Fusarium spp. gradually decreased while the population of total culturable bacteria increased. These results indicate that soil organic amendments, especially PPM, can decrease disease severity and promote plant growth, possibly by decreasing pathogen population and enhancing bacterial activity in the soil.


Photoaffinity labeling with 2(-)[2-(4-azido-3(-)[125I]- iodophenyl)ethylamino]adenosine and autoradiography with 2(-)[2-(4-amino-3(-)[125I]iodophenyl)ethylamino]adenosine of A2a adenosine receptors in rat brain.
The A2a adenosine receptor agonist 2(-)[2-(4-amino-3- iodophenyl)ethylamino]adenosine is a potent coronary vasodilator. The corresponding radioiodinated ligand, [125I]APE, discriminates between high- and low-affinity conformations of A2a adenosine receptors. In this study, [125I]APE was used for rapid (24-h) autoradiography in rat brain sections. The pattern of [125I]APE binding is consistent with that expected of an A2a-selective radioligand. It is highest in striatum, nucleus accumbens, and olfactory tubercle, with little binding to cortex and septal nuclei. Specific [125I]APE binding to these brain regions is abolished by 1 microM 2-p-(2-carboxyethyl)phenethylamino-5'-N-ethylcarboxamidoadenosine (CGS-21680) but is little affected by 100 nM 8-cyclopentyl-1,3-dipropylxanthine. Conversion of [125I]APE to the corresponding arylazide results in [125I]AzPE. The rank-order potency of compounds to compete for [125I]AzPE binding in the dark is CGS-21680 > D-(R)-N6-phenylisopropyladenosine > N6- cyclopentyladenosine, indicating that it also is an A2a-selective ligand. Specific photoaffinity labeling by [125I]AzPE of a single polypeptide (42 kDa) corresponding to A2a adenosine receptors is reduced 55 +/- 4% by 100 microM guanosine 5'-O-(3-thiotriphosphate) and 91 +/- 1.3% by 100 nM CGS-21680. [125I]APE and [125I]AzPE are valuable new tools for characterizing A2a adenosine receptors and their coupling to GTP-binding proteins by autoradiography and photoaffinity labeling.


Promoting physical activity using an activity monitor and a tailored web-based advice: design of a randomized controlled trial [ISRCTN93896459].
Ageing is associated with a decrease in physical activity. This decrease particularly occurs during specific transitional life stages. Especially during adolescence and young adulthood a steep decrease in physical activity is observed. Inactive people are often not aware of their inactivity. Providing feedback on the actual physical activity level by an activity monitor can increase awareness and may in combination with an individually tailored physical activity advice stimulate a physically active lifestyle. In a randomized controlled trial the effectiveness of providing an activity monitor in combination with a personal physical activity advice through the Internet will be examined. Outcome measures are level of physical activity, determinants of physical activity, quality of life, empowerment, aerobic fitness and body composition. Participants are relatively inactive adolescents and young adults who are measured at baseline, after 3 months intervention and 5 months after the end of the intervention. In addition, facilitating and hindering factors for implementation of the intervention will be investigated. The use of a personal activity monitor in combination with web-based assisted individually tailored health promotion offers a good opportunity to work interactively with large groups of adolescents and young adults and provide them with advice based on their actual activity level. It has great potential to motivate people to change their behaviour and to our knowledge has not been evaluated before.


Low-Scaling Algorithm for Nudged Elastic Band Calculations Using a Surrogate Machine Learning Model.
We present the incorporation of a surrogate Gaussian process regression (GPR) atomistic model to greatly accelerate the rate of convergence of classical nudged elastic band (NEB) calculations. In our surrogate model approach, the cost of converging the elastic band no longer scales with the number of moving images on the path. This provides a far more efficient and robust transition state search. In contrast to a conventional NEB calculation, the algorithm presented here eliminates any need for manipulating the number of images to obtain a converged result. This is achieved by inventing a new convergence criteria that exploits the probabilistic nature of the GPR to use uncertainty estimates of all images in combination with the force in the saddle point in the target model potential. Our method is an order of magnitude faster in terms of function evaluations than the conventional NEB method with no accuracy loss for the converged energy barrier values.


Age and gender differences in the physical activity patterns of urban schoolchildren in Korea and China.
Physical activity in childhood is important as it may establish adult behavior. However, few studies on physical activity in children have been conducted, especially in Asian children. We performed anthropometric measurements of 159 school children in two grades (grade 5: 10-11 years old and grade 8: 13-14 years old) from urban areas of Korea (n=79) and China (n=80). The total daily energy expenditure (TEE) was estimated for 7 consecutive days using an accelerometer. The mean height, weight, and body mass index (BMI) for boys and girls in both countries exceeded the US national reference median (CDC, 2000). Physical activity levels (PALs) were significantly higher in the grade 5 group (10-11 years old) and in girls than in boys for both grades. No significant difference in PALs or daily step-counts (STPs) was observed between 'normal' and 'overweight' subgroups based on BMI, although negative correlations were found between weight, BMI, or %body fat vs. PAL or STP among Korean girls and Chinese boys (r=0.32-0.38, all p<0.05). Daily variation in physical activity was observed in Korean children. In the Koreans (boys and girls, both grades pooled), TEE and STP were significantly lower than the 7-day average on Sundays, whereas for the Chinese population, STP did not clearly differ between the weekends and the week averages. In summary, PALs were higher in the fifth grade boys and girls than in the eighth grade children; interestingly, girls tended to have higher PALs than boys. Daily variation in physical activity was observed in Korea; children were less active on Sundays.


Anti-infectious and anti-inflammatory effects of peptide fragments sequentially derived from the antimicrobial peptide centrocin 1 isolated from the green sea urchin, Strongylocentrotus droebachiensis.
Bacterial resistance against antibiotic treatment has become a major threat to public health. Antimicrobial peptides (AMPs) have emerged as promising alternative agents for treatment of infectious diseases. This study characterizes novel synthetic peptides sequentially derived from the AMP centrocin 1, isolated from the green sea urchin, for their applicability as anti-infective agents.The microbicidal effect of centrocin 1 heavy chain (CEN1 HC-Br), its debrominated analogue (CEN1 HC), the C-terminal truncated variants of both peptides, i.e. CEN1 HC-Br (1-20) and CEN1 HC (1-20), as well as the cysteine to serine substituted equivalent CEN1 HC (Ser) was evaluated using minimal microbicidal concentration assay. The anti-inflammatory properties were assessed by measuring the inhibition of secretion of pro-inflammatory cytokines. All the peptides tested exhibited marked microbicidal and anti-inflammatory properties. No difference in efficacy was seen comparing CEN1 HC-Br and CEN1 HC, while the brominated variant had higher cytotoxicity. C-terminal truncation of both peptides reduced salt-tolerability of the microbicidal effect as well as anti-inflammatory actions. Also, serine substitution of cysteine residue decreased the microbicidal effect. Thus, from the peptide variants tested, CEN1 HC showed the best efficacy and safety profile. Further, CEN1 HC significantly reduced bacterial counts in two different animal models of infected wounds, while Staphylococcus aureus and methicillin-resistant S. aureus (MRSA) failed to develop resistance against this peptide under continued selection pressure. In summary, CEN1 HC appears a promising new antimicrobial agent, and clinical studies are warranted to evaluate the applicability of this AMP for local treatment of infections in man.


Determining the Molecular Pathways Underlying the Protective Effect of Non-Steroidal Anti-Inflammatory Drugs for Alzheimer's Disease: A Bioinformatics Approach.
Alzheimer's disease (AD) represents a substantial unmet need, due to increasing prevalence in an ageing society and the absence of a disease modifying therapy. Epidemiological evidence shows a protective effect of non steroidal anti inflammatory (NSAID) drugs, and genome wide association studies (GWAS) show consistent linkage to inflammatory pathways; both observations suggesting anti-inflammatory compounds might be effective in AD therapy although clinical trials to date have not been positive. In this study, we use pathway enrichment and fuzzy logic to identify pathways (KEGG database) simultaneously affected in both AD and by NSAIDs (Sulindac, Piroxicam, Paracetamol, Naproxen, Nabumetone, Ketoprofen, Diclofenac and Aspirin). Gene expression signatures were derived for disease from both blood (n = 344) and post-mortem brain (n = 690), and for drugs from immortalised human cell lines exposed to drugs of interest as part of the Connectivity Map platform. Using this novel approach to combine datasets we find striking overlap between AD gene expression in blood and NSAID induced changes in KEGG pathways of Ribosome and Oxidative Phosphorylation. No overlap was found in non NSAID comparison drugs. In brain we find little such overlap, although Oxidative Phosphorylation approaches our pre-specified significance level. These findings suggest that NSAIDs might have a mode of action beyond inflammation and moreover that their therapeutic effects might be mediated in particular by alteration of Oxidative Phosphorylation and possibly the Ribosome pathway. Mining of such datasets might prove increasingly productive as they increase in size and richness.


Evaluation of agonist-antagonist properties of nitrogen mustard and cyano derivatives of delta 8-tetrahydrocannabinol.
delta 8-Tetrahydrocannabinol (delta 8-THC) is a naturally occurring cannabinoid with a characteristic pharmacological profile of in vivo effects. Previous studies have shown that modification of the structure of delta 8-THC by inclusion of a nitrogen-containing functional group alters this profile and may alkylate the cannabinoid receptor, similar to the manner in which beta-funaltrexamine (beta-FNA) alkylates the micro-opioid receptor. Two novel analogs of delta 8-THC were synthesized: a nitrogen mustard analog with a dimethylheptyl side chain (NM-delta 8-THC) and a cyano analog with a dimethylpentyl side chain (CY-delta 8-THC). Both analogs showed high affinity for brain cannabinoid receptors and when administered acutely, produced characteristic delta 9-THC-like effects in mice, including locomotor suppression, hypothermia, antinociception and catalepsy. CY-delta 8-THC shared discriminative stimulus effects with CP 55,940; for NM-delta 8-THC, these effects also occurred, but were delayed. Although both compounds attenuated the effects of delta 9-THC in the mouse behavioral tests, evaluation of potential antagonist effects of these compounds was complicated by the fact that two injections of delta 9-THC produced similar results, suggesting that acute tolerance or desensitization might account for the observations. NM-delta 8-THC, but not CY-delta 8-THC, attenuated the discriminative stimulus effects of CP 55,940 in rats several days following injection. Hence, addition of a nitrogen-containing functional group to a traditional cannabinoid structure does not eliminate agonist effects and may produce delayed attenuation of cannabinoid-induced pharmacological effects.


Vedolizumab use is not associated with increased malignancy incidence: GEMINI LTS study results and post-marketing data.
Vedolizumab is a gut-selective antibody to α4 β7 integrin approved to treat moderate-to-severe Crohn's disease and ulcerative colitis in adults. Inflammatory bowel disease (IBD) and immunosuppressant use are associated with increased risk of malignancy. To analyse the incidence of malignancy with vedolizumab treatment in the GEMINI long-term safety (LTS) study and post-marketing (PM) setting. Malignancy data from the LTS study (May 2009 to May 2018), and data from the vedolizumab Global Safety Database (20 May 2014 to 19 May 2018), were identified using Medical Dictionary for Regulatory Activities coding. The number of patients experiencing malignancies in the LTS study (excluding malignancies within 1 year following vedolizumab initiation) was indirectly standardised against the number expected, using age- and sex-specific rates in patients with IBD from Optum's Clinformatics™ Data Mart (CDM) database. Among 1785 patients with ≥1 year of follow-up post-vedolizumab initiation in the LTS study (total 5670 patient-years), observed numbers of malignancies were similar to those expected compared with CDM data (31 vs 29; ratio of observed to expected events = 1.08; P = 0.71; 95% confidence intervals [CI] 0.73, 1.53). The most common malignancies were renal and bladder (6). PM, 293 patients reported 299 malignancies (including malignancies within 1 year following vedolizumab initiation), in approximately 208 050 patient-years of vedolizumab exposure. Lower gastrointestinal malignancies were most common (59). The number of malignancies in the LTS study was similar to that expected from an IBD population with no statistically significant differences, although few confounders could be corrected for. Limitations of PM safety reporting require consideration; however, the number of malignancies with vedolizumab appeared low.


Common fears and their relationship to dental fear and utilization of the dentist.
Common fears were studied by household telephone interviews and mail survey in Seattle, Washington, to determine their relationship to dental fear and to utilization of the dentist. Dental fear was either the first or second most common fear, with a prevalence estimated between 183 and 226 persons per 1000 population. Dental fear was associated with fears of heights, flying, and enclosures. Respondents with multiple common fears other than fear of dentistry were more likely to delay or cancel dental appointments, report a longer period since their last visit to the dentist, and report poorer oral health and less satisfaction with oral appearance. Over 22 percent of the dentally fearful group reported two or more accompanying common fears.


Electrically tunable organic-inorganic hybrid polaritons with monolayer WS2.
Exciton-polaritons are quasiparticles consisting of a linear superposition of photonic and excitonic states, offering potential for nonlinear optical devices. The excitonic component of the polariton provides a finite Coulomb scattering cross section, such that the different types of exciton found in organic materials (Frenkel) and inorganic materials (Wannier-Mott) produce polaritons with different interparticle interaction strength. A hybrid polariton state with distinct excitons provides a potential technological route towards in situ control of nonlinear behaviour. Here we demonstrate a device in which hybrid polaritons are displayed at ambient temperatures, the excitonic component of which is part Frenkel and part Wannier-Mott, and in which the dominant exciton type can be switched with an applied voltage. The device consists of an open microcavity containing both organic dye and a monolayer of the transition metal dichalcogenide WS2. Our findings offer a perspective for electrically controlled nonlinear polariton devices at room temperature.


Cancer Hallmarks, Biomarkers and Breast Cancer Molecular Subtypes.
Breast cancer is a complex disease encompassing multiple tumor entities, each characterized by distinct morphology, behavior and clinical implications. Besides estrogen receptor, progesterone receptor and human epidermal growth factor receptor 2, novel biomarkers have shown their prognostic and predictive values, complicating our understanding towards to the heterogeneity of such cancers. Ten cancer hallmarks have been proposed by Weinberg to characterize cancer and its carcinogenesis. By reviewing biomarkers and breast cancer molecular subtypes, we propose that the divergent outcome observed from patients stratified by hormone status are driven by different cancer hallmarks. 'Sustaining proliferative signaling' further differentiates cancers with positive hormone receptors. 'Activating invasion and metastasis' and 'evading immune destruction' drive the differentiation of triple negative breast cancers. 'Resisting cell death', 'genome instability and mutation' and 'deregulating cellular energetics' refine breast cancer classification with their predictive values. 'Evading growth suppressors', 'enabling replicative immortality', 'inducing angiogenesis' and 'tumor-promoting inflammation' have not been involved in breast cancer classification which need more focus in the future biomarker-related research. This review novels in its global view on breast cancer heterogeneity, which clarifies many confusions in this field and contributes to precision medicine.


Correlation between Kind of Delivery and Posttraumatic Stress Disorder.
Posttraumatic stress disorder (PTSD) is a very common mental condition and a unique anxiety disorder. The present study tried to examine the correlation between kind of delivery and PTSD. This prospective study evaluated 240 Iranian female residents of Tehran, Iran, during the third trimester of their pregnancy and 6-8 weeks after labor. Data were collected using the customized screening form, the Symptom Checklist-90, PTSD Symptom Scale (PSS), and Social Support Questionnaire. The collected data were then analyzed with SPSS software. According to the participants' responses to the subscales of the PSS, reexperiencing, avoidance, and hyperarousal symptoms were present in 100, 25, and 77 participants, respectively. Moreover, 15 individuals manifested all three groups of symptoms. Therefore, the prevalence of PTSD in the studied population was 6.2%. The logistic regression analysis revealed PTSD to be 0.06 times more prevalent in women with poor social support after delivery than in those enjoying a desirable level of support (P < 0.01; odds ratio = 0.06). Based onChisquare test results, there was no significant correlation between the kind of delivery and the incidence of PTSD after delivery (P = 0.48). Since PTSD was more common after cesarean sections (than after vaginal deliveries), health policymakers need to develop efficient strategies to promote vaginal delivery.


Effect of velocity on ground reaction forces in dogs with lameness attributable to tearing of the cranial cruciate ligament.
To ascertain the effectiveness of evaluating ground reaction forces (GRFs) at velocities during walking and trotting in dogs with naturally occurring lameness and determine whether walking would provide sufficient motion to adequately characterize GRFs with respect to trotting. 29 dogs with a naturally occurring tear of the cranial cruciate ligament. Dogs were walked and trotted over a force platform, and GRFs were recorded during the stance phase. Correlation was used to assess the agreement between walking and trotting for GRF The coefficient of variation was calculated to assess the relative variation of outcome variables among the gaits. Group means for walking GRF were compared between dogs that trotted and that failed to trot. GRFs during walking and trotting were highly correlated. The coefficient of variation was smaller for GRFs during walking than during trotting. Dogs that failed to trot had significantly smaller mean values of peak vertical force and vertical impulse during walking, compared with values for dogs that were able to trot. Either velocity is acceptable for GRF evaluation in dogs. Mean GRF during walking was significantly different between dogs that could and could not trot, principally because dogs with the most severe lameness failed to trot. These dogs would be eliminated from a clinical study, and thus, that study would become biased toward dogs that were less lame. In that situation, differences between interventions may be less pronounced, because they would be evaluated on dogs with less lameness.


Sleep homeostasis in the rat is preserved during chronic sleep restriction.
Sleep is homeostatically regulated in all animal species that have been carefully studied so far. The best characterized marker of sleep homeostasis is slow wave activity (SWA), the EEG power between 0.5 and 4 Hz during nonrapid eye movement (NREM) sleep. SWA reflects the accumulation of sleep pressure as a function of duration and/or intensity of prior wake: it increases after spontaneous wake and short-term (3-24 h) sleep deprivation and decreases during sleep. However, recent evidence suggests that during chronic sleep restriction (SR) sleep may be regulated by both allostatic and homeostatic mechanisms. Here, we performed continuous, almost completely artifact-free EEG recordings from frontal, parietal, and occipital cortex in freely moving rats (n = 11) during and after 5 d of SR. During SR, rats were allowed to sleep during the first 4 h of the light period (4S(+)) but not during the following 20 h (20S(-)). During the daily 20S(-) most sleep was prevented, whereas the number of short (<20 s) sleep attempts increased. Low-frequency EEG power (1-6 Hz) in both sleep and wake also increased during 20S(-), most notably in the occipital cortex. In all animals NREM SWA increased above baseline levels during the 4S(+) periods and in post-SR recovery. The SWA increase was more pronounced in frontal cortex, and its magnitude was determined by the efficiency of SR. Analysis of cumulative slow wave energy demonstrated that the loss of SWA during SR was compensated by the end of the second recovery day. Thus, the homeostatic regulation of sleep is preserved under conditions of chronic SR.


An indirect influence of phenylephrine on the release of endothelium-derived vasodilators in rat small mesenteric artery.
1. The possibility that stimulation of smooth muscle alpha(1)-adrenoceptors modulates contraction via the endothelium was examined in rat small mesenteric arteries. 2. N(omega)-nitro-L-arginine methyl ester, (L-NAME, 100 microM to inhibit NO synthase) increased contraction to single concentrations of phenylephrine (1 - 3 microM) by approximately 2 fold (from a control level of 14.2+/-3.0 to 34. 1+/-4.2% of the maximum contraction of the artery, n=20). The action of L-NAME was abolished by disrupting the endothelium. 3. The subsequent addition of apamin (to inhibit small conductance Ca(2+)-activated K(+) channels, 50 nM) further augmented phenylephrine contractions, in an endothelium-dependent manner, to more than 3 fold above control (50.4+/-5.3% of the maximum contraction, n=11). 4.Charybdotoxin (non-selective inhibitor of large conductance Ca(2+)-activated K(+) channels, BK(Ca), 50 nM) plus L-NAME augmented the level of phenylephrine contraction to 4 - 5-fold above control (64.1+/-3.1%, n=5), but this effect was independent of the endothelium. The potentiation of contraction by charybdotoxin could be mimicked with the selective BK(Ca) inhibitor, iberiotoxin,. 5. Apamin together with L-NAME and charybdotoxin further significantly increased the phenylephrine contraction by 5 - 6-fold, to 79.9+/-3.5% of the maximum contraction of the artery (n=13). 6. Phenylephrine failed directly to increase the intracellular Ca(2+) concentration in endothelial cells freshly isolated from the small mesenteric artery. 7. Stimulation of smooth muscle alpha(1)-adrenoceptors in the mesenteric artery induces contraction that is markedly suppressed by the endothelium. The attenuation of contraction appears to reflect both the release of NO from the endothelium and the efflux of K(+) from both endothelial and smooth muscle cells. This suggests that the release of NO and endothelium-derived hyperpolarizing factor can be evoked indirectly by agents which act only on the smooth muscle cells.


Continuity in primary school children's eating problems and the influence of parental feeding strategies.
Eating problems are highly prevalent and seem to show continuity in children. Nevertheless, the effect of different maternal and paternal feeding practices on changes in these problems is not fully understood yet. This study examines short-term continuity in primary school children's overeating, loss of control (over eating), restraint and concerns (about eating, body shape and weight) and the predictive value of parental feeding strategies on change in these eating problems. Children (8-12 years, n = 613, 46.5 % girls) completed the Child Eating Disorder Examination Questionnaire (ChEDE-Q) twice with the second measurement (T2) taking place 6 months after the first (T1). Parents (n = 714, 57.7 % mothers) completed the Child Feeding Questionnaire (CFQ) at T1 to assess the level of controlling feeding strategies. More overeating, restraint and concerns at T1 predicted higher levels of the same eating problems 6 months later, whereas more of loss of control at T1 predicted lower levels of loss of control 6 months later. Additionally, the interaction between maternal pressure and the gender of the child was predictive for increases in overeating, whereas the interaction between paternal monitoring and gender of the child was a marginally significant predictor for decreases in overeating. These findings point to the possible negative and positive effects of parental feeding practices as well as to the possible larger influence of mothers on their daughters' and of fathers on their sons' eating behaviors. The importance of studying mothers and fathers, as well as daughters and sons, and of including different feeding strategies and eating problems is discussed.


Peptide permeases from Streptococcus pneumoniae affect adherence to eucaryotic cells.
To gain access to tissues within the human host, Streptococcus pneumoniae initially colonizes the nasopharynx and then interacts with glycoconjugates on the surfaces of target cells at various sites of infection. Although pneumococcal adhesins are currently unknown, exported proteins on the bacterial surface are potential candidates. To identify bacterial elements involved in this process, mutants of S. pneumoniae with defects in exported proteins were screened for the inability to adhere to cells representative of three in vivo niches: (i) agglutination of bovine erythrocytes, which reflects adherence to cells which reside in the nasopharynx; (ii) human type II pneumocytes (lung cells [LC]), representing the alveolar site of infection; and (iii) human vascular endothelial cells (EC), representing the endovascular site. The capacity of the mutants to adhere during the course of pneumococcal disease was also assessed by using cytokine-activated LC and EC. All of the 30 mutants analyzed produced hemagglutination values comparable with those of the parent strain. Four independent mutants demonstrated a greater than 50% decrease in adherence to both LC and EC. Sequence analysis of the altered alleles from these strains showed that mutations had occurred in two previously identified loci, plpA and ami, which belong to the family of genes encoding protein-dependent peptide permeases. Mutations in the ami locus resulted in an inability to recognize the GalNAc beta 1-4Gal glycoconjugate receptor present on resting LC and EC, whereas mutations in plpA resulted in a failure to recognize a GalNAc beta 1-3Gal glycoconjugate receptor also present on resting cells. Mutations in neither allele affected recognition of GlcNAc receptors present on cytokine-activated LC and EC. These results suggest that peptide permeases modulate pneumococcal adherence to epithelial and endothelial cells either by acting directly as adhesins or by modulating the expression of adhesins on the pneumococcal surface during the initial stages of colonization of the lung or the vascular endothelium.


Self-monitoring of blood pressure in pregnancy: A mixed methods evaluation of a national roll-out in the context of a pandemic.
To evaluate how English maternity units implemented self-monitoring of blood pressure (SMBP) in pregnancy in response to the COVID-19 pandemic. Mixed methods including surveys, anonymised patient data and in-depth interviews with women. Maternity units across England. 45 maternity units completed a survey about the implementation of SMBP (supported by the provision of guidance and blood pressure monitors) during the pandemic, 166 women completed a survey about their experiences of SMBP, and 23 women took part in in-depth interviews. Clinical data from 627 women undertaking SMBP were available from 13 maternity units. SMBP was predominantly used to provide additional BP monitoring for hypertensive or high-risk pregnant women. Overall maternity units and women were positive about its use in terms of reducing the need for additional face-to-face contacts and giving women more control and insight into their own BP. However, there were challenges in setting up SMBP services rapidly and embedding them within existing care pathways, particularly around interpreting readings and managing the provision of monitors. A considerable proportion of maternity units in England commenced a SMBP service for hypertensive or high-risk women from March 2020. There is a need for further research into appropriate care pathways, including guidance around white coat or masked hypertension and the use of SMBP postnatally.


Generalized joint hypermobility and perceived harmfulness in healthy adolescents; impact on muscle strength, motor performance and physical activity level.
Objective: The first aim of this study was to determine whether adolescents with asymptomatic Generalized Joint Hypermobility (GJH) have a lower level of physical functioning (physical activity level, muscle strength and performance) compared to non-hypermobile controls. Secondly, to evaluate whether the negative impact of perceived harmfulness on physical functioning was more pronounced in adolescents with asymptomatic GJH.Methods: Cross-sectional study. Sixty-two healthy adolescents (mean age 16.8, range 12-21) participated. Hypermobility (Beighton score), perceived harmfulness (PHODA-youth) and muscle strength (dynamometry), motor performance (Single-Leg-Hop-for-Distance) and physical activity level (PAL) (accelerometry) were measured. Hierarchical regression analyses were used to study differences in physical functioning and perceived harmfulness between asymptomatic GJH and non-hypermobile controls.Results: Asymptomatic GJH was associated with increased knee extensor muscle strength (peak torque/body weight; PT/BW), controlled for age and gender (dominant leg; ß = 0.29; p = .02). No other associations between asymptomatic GJH and muscle strength, motor performance and PAL were found. Perceived harmfulness was not more pronounced in adolescents with asymptomatic GJH.Conclusions: Adolescents with asymptomatic GJH had increased knee extensor muscle strength compared to non-hypermobile controls. No other differences in the level of physical functioning was found and the negative impact of perceived harmfulness was not more pronounced in adolescents with asymptomatic GJH.


Strategies to improve health coverage and narrow the equity gap in child survival, health, and nutrition.
Implementation of innovative strategies to improve coverage of evidence-based interventions, especially in the most marginalised populations, is a key focus of policy makers and planners aiming to improve child survival, health, and nutrition. We present a three-step approach to improvement of the effective coverage of essential interventions. First, we identify four different intervention delivery channels--ie, clinical or curative, outreach, community-based preventive or promotional, and legislative or mass media. Second, we classify which interventions' deliveries can be improved or changed within their channel or by switching to another channel. Finally, we do a meta-review of both published and unpublished reviews to examine the evidence for a range of strategies designed to overcome supply and demand bottlenecks to effective coverage of interventions that improve child survival, health, and nutrition. Although knowledge gaps exist, several strategies show promise for improving coverage of effective interventions-and, in some cases, health outcomes in children-including expanded roles for lay health workers, task shifting, reduction of financial barriers, increases in human-resource availability and geographical access, and use of the private sector. Policy makers and planners should be informed of this evidence as they choose strategies in which to invest their scarce resources.


Regulation of kinase cascade activation and heat shock protein expression by poly(ADP-ribose) polymerase inhibition in doxorubicin-induced heart failure.
Cardiomyopathy is one of the most severe side effects of the chemotherapeutic agent doxorubicin (DOX). The formation of reactive oxygen species plays a critical role in the development of cardiomyopathies, and the pathophysiological cascade activates nuclear enzyme poly(ADP-ribose) polymerase (PARP), and kinase pathways. We characterized the effects of the PARP-inhibitor and kinase-modulator compound L-2286 in DOX-induced cardiac injury models. We studied the effect of the established superoxide dismutase-mimic Tempol and compared the effects of this agent with those of the PARP inhibitor. In the rat H9C2 cardiomyocytes, in which DOX-induced poly(ADP-ribosyl)ation, L-2286 protected them from the DOX-induced injury in a concentration-dependent manner. In the in vivo studies, mice were pretreated (for 1 week) with L-2286 or Tempol before the DOX treatment. Both the agents improved the activation of cytoprotective kinases, Akt, phospho-specific protein kinase C ϵ, ζ/λ and suppressed the activity of cell death promoting kinases glycogen synthase kinase-3β, JNK, and p38 mitogen-activated protein kinase, but the effect of PARP inhibitor was more pronounced and improved the survival as well. L-2286 activated the phosphorylation of proapoptotic transcription factor FKHR1 and promoted the expression of Hsp72 and Hsp90. These data suggest that the mode of the cytoprotective action of the PARP inhibitor may include the modulation of kinase pathways and heat shock protein expression.


Novel lipid transfer property of two mitochondrial proteins that bridge the inner and outer membranes.
This study provides evidence of a novel function for mitochondrial creatine kinase (MtCK) and nucleoside diphosphate kinase (NDPK-D). Both are basic peripheral membrane proteins with symmetrical homo-oligomeric structure, which in the case of MtCK was already shown to allow crossbridging of lipid bilayers. Here, different lipid dilution assays clearly demonstrate that both kinases also facilitate lipid transfer from one bilayer to another. Lipid transfer occurs between liposomes mimicking the lipid composition of mitochondrial contact sites, containing 30 mol % cardiolipin, but transfer does not occur when cardiolipin is replaced by phosphatidylglycerol. Ubiquitous MtCK, but not NDPK-D, shows some specificity in the nature of the lipids transferred and it is not active with phosphatidylcholine alone. MtCK can undergo reversible oligomerization between dimeric and octameric forms, but only the octamer can bridge membranes and promote lipid transfer. Cytochrome c, another basic mitochondrial protein known to bind to anionic membranes but not crosslinking them, is also incapable of promoting lipid transfer. The lipid transfer process does not involve vesicle fusion or loss of the internal contents of the liposomes.


Dynamics of tactical behaviour in association football when manipulating players' space of interaction.
The analysis of positional data in association football allows the spatial distribution of players during matches to be described in order to improve the understanding of tactical-related constraints on the behavioural dynamics of players. The aim of this study was to identify how players' spatial restrictions affected the exploratory tactical behaviour and constrained the perceptual-motor workspace of players in possession of the ball, as well as inter-player passing interactions. Nineteen professional outfield male players were divided into two teams of 10 and 9 players, respectively. The game was played under three spatial constraints: a) players were not allowed to move out of their allocated zones, except for the player in possession of the ball; b) players were allowed to move to an adjacent zone, and; c) non-specific spatial constraints. Positional data was captured using a 5 Hz interpolated GPS tracking system and used to define the configuration states of players for each second in time. The configuration state comprised 37 categories derived from tactical actions, distance from the nearest opponent, distance from the target and movement speed. Notational analysis of players in possession of the ball allowed the mean time of ball possession and the probabilities of passing the ball between players to be calculated. The results revealed that the players' long-term exploratory behaviour decreased and their short-term exploration increased when restricting their space of interaction. Relaxing players' positional constraints seemed to increase the speed of ball flow dynamics. Allowing players to move to an adjacent sub-area increased the probabilities of interaction with the full-back during play build-up. The instability of the coordinative state defined by being free from opponents when players had the ball possession was an invariant feature under all three task constraints. By allowing players to move to adjacent sub-areas, the coordinative state became highly unstable when the distance from the target decreased. Ball location relative to the scoring zone and interpersonal distance constitute key environmental information that constrains the players' coordinative behaviour. Based on our results, dynamic overlap is presented as a good option to capture tactical performance. Moreover, the selected collective (i.e. relational) variables would allow coaches to identify the effects of training drills on teams and players' behaviour. More research is needed considering these type variables to understand how the manipulation of constraints induce a more stable or flexible dynamical structure of tactical behaviour.


Risk factors for explantation due to infection after sacral neuromodulation: a multicenter retrospective case-control study.
Sacral neuromodulation is an effective therapy for overactive bladder, urinary retention, and fecal incontinence. Infection after sacral neurostimulation is costly and burdensome. Determining optimal perioperative management strategies to reduce the risk of infection is important to reduce this burden. We sought to identify risk factors associated with sacral neurostimulator infection requiring explantation, to estimate the incidence of infection requiring explantation, and identify associated microbial pathogens. This is a multicenter retrospective case-control study of sacral neuromodulation procedures completed from Jan. 1, 2004, through Dec. 31, 2014. We identified all sacral neuromodulation implantable pulse generator implants as well as explants due to infection at 8 participating institutions. Cases were patients who required implantable pulse generator explantation for infection during the review period. Cases were included if age ≥18 years old, follow-up data were available ≥30 days after implantable pulse generator implant, and the implant was performed at the institution performing the explant. Two controls were matched to each case. These controls were the patients who had an implantable pulse generator implanted by the same surgeon immediately preceding and immediately following the identified case who met inclusion criteria. Controls were included if age ≥18 years old, no infection after implantable pulse generator implant, follow-up data were available ≥180 days after implant, and no explant for any reason <180 days from implant. Controls may have had an explant for reasons other than infection at >180 days after implant. Fisher exact test (for categorical variables) and Student t test (for continuous variables) were used to test the strength of the association between infection and patient and surgery characteristics. Significant variables were then considered in a multivariable logistic regression model to determine risk factors independently associated with infection. Over a 10-year period at 8 academic institutions, 1930 sacral neuromodulator implants were performed by 17 surgeons. In all, 38 cases requiring device explant for infection and 72 corresponding controls were identified. The incidence of infection requiring explant was 1.97%. Hematoma formation (13% cases, 0% controls; P = .004) and pocket depth of ≥3 cm (21% cases, 0% controls; P = .031) were independently associated with an increased risk of infection requiring explant. On multivariable regression analysis controlling for significant variables, both hematoma formation (P = .006) and pocket depth ≥3 cm (P = .020, odds ratio 3.26; 95% confidence interval, 1.20-8.89) remained significantly associated with infection requiring explant. Of the 38 cases requiring explant, 32 had cultures collected and 24 had positive cultures. All 5 cases with a hematoma had a positive culture (100%). Of the 4 cases with a pocket depth ≥3 cm, 2 had positive cultures, 1 had negative cultures, and 1 had a missing culture result. The most common organism identified was methicillin-resistant Staphylococcus aureus (38%). Infection after sacral neuromodulation requiring device explant is low. The most common infectious pathogen identified was methicillin-resistant S aureus. Demographic and health characteristics did not predict risk of explant due to infection, however, having a postoperative hematoma or a deep pocket ≥3 cm significantly increased the risk of explant due to infection. These findings highlight the importance of meticulous hemostasis as well as ensuring the pocket depth is <3 cm at the time of device implant.


Longitudinal Change in Peripheral Quantitative Computed Tomography Assessment in Older Adults: The Hertfordshire Cohort Study.
There are few longitudinal data on change in bone structure and muscle mass, strength and function in later life. We report these, and consider bone-muscle interrelationships in older men and women. We studied 188 men and 166 women from the Hertfordshire Cohort Study, who underwent peripheral quantitative computed tomography (pQCT) of the radius and tibia in 2004-2005 and then again in 2011-2012. Grip strength and gait speed were also assessed at both timepoints. Percentage change per year was calculated for grip strength, gait speed, muscle cross-sectional area (mCSA), fat cross-sectional area (fCSA) and diaphyseal bone parameters [total area (Tt.Ar), cortical area (Ct.Ar), cortical density (cBMD) and trabecular density (tBMD)]. The mean (SD) age of men and women at baseline was 68.9 (2.5) and 69.2 (2.6) years, respectively. Rates of muscle area and strength loss did not differ by sex. Tt.Ar increased with age and faster in men [mean (SD) 1.78 (1.64) %/year] than women [mean (SD) 1.03 (1.69) %/year] in the radius (p < 0.001). In both the radius (p = 0.006) and tibia (p < 0.001), Ct.Ar reduced more rapidly in women than men. Change in Ct.Ar was associated with change in muscle area in the corresponding limb (radius; men: regression coefficient 0.36, 95% CI 0.20-0.52, p < 0.001; tibia; men: regression coefficient 0.14, 95% CI 0.00-0.27, p = 0.043, women: regression coefficient 0.16, 95% CI 0.01-0.30, p = 0.032). We have demonstrated that muscle strength and function decrease faster than muscle mass and have provided further evidence that changes in bone structure with age differ by sex.


Summer rains and dry seasons in the upper Blue Nile Basin: the predictability of half a century of past and future spatiotemporal patterns.
During the last 100 years the Ethiopian upper Blue Nile Basin (BNB) has undergone major changes in land use, and is now potentially facing changes in climate. Rainfall over BNB supplies over two-thirds of the water to the Nile and supports a large local population living mainly on subsistence agriculture. Regional food security is sensitive to both the amount and timing of rain and is already an important political challenge that will be further complicated if scenarios of climate change are realized. In this study a simple spatial model of the timing and duration of summer rains (Kiremt) and dry season (Bega), and annual rain over the upper BNB was established from observed data between 1952 and 2004. The model was used to explore potential impacts of climate change on these rains, using a down-scaled ECHAM5/MP1-OM scenario between 2050 and 2100. Over the observed period the amount, onset and duration of Kiremt rains and rain-free Bega days have exhibited a consistent spatial pattern. The spatially averaged annual rainfall was 1490 mm of which 93% was Kiremt rain. The average Kiremt rain and number of rainy days was higher in the southwest (322 days) and decreased towards the north (136 days). Under the 2050-2100 scenario, the annual mean rainfall is predicted to increase by 6% and maintain the same spatial pattern as in the past. A larger change in annual rainfall is expected in the southwest (ca. +130 mm) with a gradually smaller change towards the north (ca. +70 mm). Results highlight the need to account for the characteristic spatiotemporal zonation when planning water management and climate adaptation within the upper BNB. The presented simple spatial resolved models of the presence of Kiremt and annual total rainfall could be used as a baseline for such long-term planning.


Predicting severe pain after root canal therapy in the National Dental PBRN.
Some patients experience severe pain following root canal therapy (RCT) despite advancements in care. We sought to identify factors, which can be measured preoperatively, that predict this negative outcome so that future research may focus on preemptive steps to reduce postoperative pain intensity. Sixty-two practitioners (46 general dentists and 16 endodontists) who are members of the National Dental Practice-Based Research Network enrolled patients receiving RCT for this prospective observational study. Baseline data collected from patients and dentists were obtained before treatment. Severe postoperative pain was defined based on a rating of ≥7 on a scale from 0 (no pain) to 10 (pain as bad as can be) for the worst pain intensity experienced during the preceding week, and this was collected 1 wk after treatment. Multiple logistic regression analyses were used to develop and validate the model. A total of 708 patients were enrolled during a 6-m period. Pain intensity data were collected 1 wk postoperatively from 652 patients (92.1%), with 19.5% (n = 127) reporting severe pain. In multivariable modeling, baseline factors predicting severe postoperative pain included current pain intensity (odds ratio [OR], 1.15; 95% confidence interval [CI], 1.07 to 1.25; P = 0.0003), number of days in the past week that the subject was kept from their usual activities due to pain (OR, 1.32; 95% CI, 1.13 to 1.55; P = 0.0005), pain made worse by stress (OR, 2.55; 95% CI, 1.22 to 5.35; P = 0.0130), and a diagnosis of symptomatic apical periodontitis (OR, 1.63; 95% CI, 1.01 to 2.64; P = 0.0452). Among the factors that did not contribute to predicting severe postoperative pain were the dentist's specialty training, the patient's age and sex, the type of tooth, the presence of swelling, or other pulpal and apical endodontic diagnoses. Factors measured preoperatively were found to predict severe postoperative pain following RCT. Practitioners could use this information to better inform patients about RCT outcomes and possibly use different treatment strategies to manage their patients (Clinicaltrials.gov NCT01201681).


Development of two species-specific fingerprinting probes for broad computer-assisted epidemiological studies of Candida tropicalis.
Candida tropicalis has emerged as the second most frequent colonizing Candida species, and it has been documented in nosocomial infections. To develop an effective fingerprinting system for this species, which is amenable to computer-assisted analyses and which provides information on the genetic relatedness of independent isolates, two DNA fragments, Ct3 (18,000 bp) and Ct14 (20,000 bp), were cloned from a genomic library of Sau3AI partial digestion products. Both probes generate complex Southern blot patterns containing 8 to 20 bands, when hybridized to EcoRI- or EcoRI-HaeIII-digested DNA of independent C. tropicalis isolates. The two probes show no cross-hybridization and are both species specific for C. tropicalis. A comparison of the capacity of the two probes to identify the same strain in different isolates, and differentiate unrelated strains, using computer-assisted computation of similarity coefficients and the genesis of dendrograms, suggests that while Ct14 is more effective in grouping independent isolates, Ct3 is more effective in discriminating small differences in the patterns of highly related isolates and is therefore a more effective probe for determining microevolution within a clonal population and substrain shuffling in recurrent infections. Because of their alternative attributes, it is suggested that both probes be used in fingerprinting studies of C. tropicalis.


Unveiling the carrier transport mechanism in epitaxial graphene for forming wafer-scale, single-domain graphene.
Graphene epitaxy on the Si face of a SiC wafer offers monolayer graphene with unique crystal orientation at the wafer-scale. However, due to carrier scattering near vicinal steps and excess bilayer stripes, the size of electrically uniform domains is limited to the width of the terraces extending up to a few microns. Nevertheless, the origin of carrier scattering at the SiC vicinal steps has not been clarified so far. A layer-resolved graphene transfer (LRGT) technique enables exfoliation of the epitaxial graphene formed on SiC wafers and transfer to flat Si wafers, which prepares crystallographically single-crystalline monolayer graphene. Because the LRGT flattens the deformed graphene at the terrace edges and permits an access to the graphene formed at the side wall of vicinal steps, components that affect the mobility of graphene formed near the vicinal steps of SiC could be individually investigated. Here, we reveal that the graphene formed at the side walls of step edges is pristine, and scattering near the steps is mainly attributed by the deformation of graphene at step edges of vicinalized SiC while partially from stripes of bilayer graphene. This study suggests that the two-step LRGT can prepare electrically single-domain graphene at the wafer-scale by removing the major possible sources of electrical degradation.


High-quality assembly of the reference genome for scarlet sage, Salvia splendens, an economically important ornamental plant.
Salvia splendens Ker-Gawler, scarlet or tropical sage, is a tender herbaceous perennial widely introduced and seen in public gardens all over the world. With few molecular resources, breeding is still restricted to traditional phenotypic selection, and the genetic mechanisms underlying phenotypic variation remain unknown. Hence, a high-quality reference genome will be very valuable for marker-assisted breeding, genome editing, and molecular genetics. We generated 66 Gb and 37 Gb of raw DNA sequences, respectively, from whole-genome sequencing of a largely homozygous scarlet sage inbred line using Pacific Biosciences (PacBio) single-molecule real-time and Illumina HiSeq sequencing platforms. The PacBio de novo assembly yielded a final genome with a scaffold N50 size of 3.12 Mb and a total length of 808 Mb. The repetitive sequences identified accounted for 57.52% of the genome sequence, and 54,008 protein-coding genes were predicted collectively with ab initio and homology-based gene prediction from the masked genome. The divergence time between S. splendens and Salvia miltiorrhiza was estimated at 28.21 million years ago (Mya). Moreover, 3,797 species-specific genes and 1,187 expanded gene families were identified for the scarlet sage genome. We provide the first genome sequence and gene annotation for the scarlet sage. The availability of these resources will be of great importance for further breeding strategies, genome editing, and comparative genomics among related species.


Distribution of bone mineral content is associated with body weight and exercise capacity in patients with chronic obstructive pulmonary disease.
Although low bone mineral density is highly prevalent in patients with chronic obstructive pulmonary disease (COPD), the distribution of the reduced bone mass has not been fully elucidated. To determine regional bone mass loss in patients with COPD and investigate whether the change in distribution may be associated with body weight loss and functional capacity. Body mass index (BMI) was assessed, and height squared indices were derived for the bone mineral content index (BMCI) of the arms, legs and trunk by dual-energy X-ray absorptiometry in 45 male patients with COPD and 12 age- and sex-matched control subjects. Pulmonary function tests were performed, and maximal oxygen uptake (VO2max) was measured. The BMCI was lower in the total bone, legs and trunk of patients with COPD than in control subjects, although the BMCI in the arms was similar between the groups. BMI correlated significantly with the BMCI in all 3 segments. Bone mineral content (BMC) in the trunk, expressed as a percentage of total BMC (BMC trunk/total BMC), correlated significantly with BMI. The BMCI in the trunk was closely related with VO2max but not with airflow limitation. There was a regional difference in BMC reduction, but a predominant reduction of bone mass in the trunk was not associated with the severity of airflow limitation but rather with body weight loss and exercise intolerance. These data suggest that body weight loss and exercise intolerance are important risk factors for vertebral fracture in patients with COPD.


Operation of an optically coherent frequency comb outside the metrology lab.
We demonstrate a self-referenced fiber frequency comb that can operate outside the well-controlled optical laboratory. The frequency comb has residual optical linewidths of < 1 Hz, sub-radian residual optical phase noise, and residual pulse-to-pulse timing jitter of 2.4 - 5 fs, when locked to an optical reference. This fully phase-locked frequency comb has been successfully operated in a moving vehicle with 0.5 g peak accelerations and on a shaker table with a sustained 0.5 g rms integrated acceleration, while retaining its optical coherence and 5-fs-level timing jitter. This frequency comb should enable metrological measurements outside the laboratory with the precision and accuracy that are the hallmarks of comb-based systems.


Sodium-Glucose Cotransporter 2 Inhibitors and the Risk of Below-Knee Amputation: A Multicenter Observational Study.
Reports of amputations associated with sodium-glucose cotransporter 2 (SGLT2) inhibitors have been inconsistent. We aimed to compare the risk of below-knee amputation with SGLT2 inhibitors versus dipeptidyl peptidase 4 (DPP-4) inhibitors among patients with type 2 diabetes. This multicenter observational study used administrative health care databases from seven Canadian provinces and the U.K. Incident SGLT2 inhibitor users were matched to DPP-4 inhibitor users using a prevalent new-user design and time-conditional propensity scores. Cox proportional hazards models were used to estimate site-specific adjusted hazard ratios (HR) and corresponding 95% CIs of incident below-knee amputation for SGLT2 inhibitor versus DPP-4 inhibitor users. Random effects meta-analyses were used to pool the site-specific results. The study cohort included 207,817 incident SGLT2 inhibitor users matched to 207,817 DPP-4 inhibitor users. During a mean exposed follow-up time of 11 months, the amputation rate was 1.3 per 1,000 person-years among SGLT2 inhibitor users and 1.5 per 1,000 person-years among DPP-4 inhibitor users. The adjusted HR of below-knee amputations associated with SGLT2 inhibitor use compared with DPP-4 inhibitor use was 0.88 (95% CI 0.71-1.09). Similar results were obtained in stratified analyses by specific SGLT2 inhibitor molecule. In this large multicenter observational study, there was no association between SGLT2 inhibitor use and incident below-knee amputations among patients with type 2 diabetes compared with DPP-4 inhibitor use. While these findings provide some reassurance, studies with a longer duration of follow-up are needed to assess potential long-term effects.


Formulation of a mmaA4 gene deletion mutant of Mycobacterium bovis BCG in cationic liposomes significantly enhances protection against tuberculosis.
A new vaccination strategy is urgently needed for improved control of the global tuberculosis (TB) epidemic. Using a mouse aerosol Mycobacterium tuberculosis challenge model, we investigated the protective efficacy of a mmaA4 gene deletion mutant of Mycobacterium bovis BCG (ΔmmaA4BCG) formulated in dimethyl dioctadecyl ammonium bromide (DDA) - D(+) trehalose 6,6 dibenenate (TDB) (DDA/TDB) adjuvant. In previous studies, deletion of the mmaA4 gene was shown to reduce the suppression of IL-12 production often seen after mycobacterial infections. While the non-adjuvanted ΔmmaA4BCG strain did not protect mice substantially better than conventional BCG against a tuberculous challenge in four protection experiments, the protective responses induced by the ΔmmaA4BCG vaccine formulated in DDA/TDB adjuvant was consistently increased relative to nonadjuvanted BCG controls. Furthermore, the ΔmmaA4BCG-DDA/TDB vaccine induced significantly higher frequencies of multifunctional (MFT) CD4 T cells expressing both IFNγ and TNFα (double positive) or IFNγ, TNFα and IL-2 (triple positive) than CD4 T cells derived from mice vaccinated with BCG. These MFT cells were characterized by having higher IFNγ and TNFα median fluorescence intensity (MFI) values than monofunctional CD4 T cells. Interestingly, both BCG/adjuvant and ΔmmaA4BCG/adjuvant formulations induced significantly higher frequencies of CD4 T cells expressing TNFα and IL-2 than nonadjuvanted BCG or ΔmmaA4BCG vaccines indicating that BCG/adjuvant mixtures may be more effective at inducing central memory T cells. Importantly, when either conventional BCG or the mutant were formulated in adjuvant and administered to SCID mice or immunocompromised mice depleted of IFNγ, significantly lower vaccine-derived mycobacterial CFU were detected relative to immunodeficient mice injected with non-adjuvanted BCG. Overall, these data suggest that immunization with the ΔmmaA4BCG/adjuvant formulation may be an effective, safe, and relatively inexpensive alternative to vaccination with conventional BCG.


What Makes Vietnamese (Not) Attend Periodic General Health Examinations? A 2016 Cross-sectional Study.
General health examinations (GHE) have become an increasingly common measure for preventive medicine in Vietnam. However, little is known about the factors among Viet-namese people who attend or miss GHE. Budget or time constraints remain to be evaluated for better-informed policy making. This study investigates factors affecting behaviors in attending periodic GHE. The main objectives are as follows: (1) to explore empirical relationships between influencing factors and periodic GHE frequencies, and (2) to predict the probabilities of attending GHE under associated conditions. The study used a 2,068-observational dataset, obtained from a Vietnamese survey in 2016. The analysis was then performed using the methods of baseline-category logits for establishing relationships between predictor and response variables. Significant relationships were found among the expenditure and time consumption, health priority and sensitivity to health data, insurance status, and frequency of GHE, with most p-values = 0.01. Generally, people attended the GHE when they had the resources and health priorities (72.7% probability). Expenditure and time remain key obstacles to the periodic GHE. Health priority and health data are important in improving rates for GHEs. Health insurance should play a positive role in promoting the GHE.


Occurrence of Transmembrane Protein 119 in the Retina is Not Restricted to the Microglia: An Immunohistochemical Study.
Recently, a new marker protein for microglial cells in the brain was postulated, transmembrane protein 119 (TMEM119), raising the hope for a new opportunity to reliably and unambiguously detect microglial cells in histologic sections. It was of interest whether TMEM119 also was a reliable microglial marker in the retina. Anti-TMEM119 antibodies of two providers were used to label microglia in the murine retina, and labeling properties were compared to those of antibodies against Iba1 and CD11b. As an example of a pathologic situation, labeling for TMEM119 was also performed in eyes treated by an argon laser as an experimental model for choroidal neovascularization. TMEM119 immunoreactivity (IR) was found on microglial cells in the naïve retina. However, specificity and sensitivity of TMEM119 IR varied clearly depending on the source of the antibody, age of the mouse, and location of retinal microglia. After laser treatment, however, microglial cells lost their IR for TMEM119 at the site of the laser spot. Moreover, other cells became positive for TMEM119; for example, Müller cells. TMEM119 is a useful marker for the microglia in the brain. However, retinal microglia shows variable IR for TMEM119, and the microglia is not the only cell showing TMEM IR. Therefore, TMEM119 appears not to be applicable as a general marker for the retinal microglia in pathologic situations. Reliable detection and quantification of microglial cells is of high importance to study disease mechanisms and effects of therapeutic approaches in the retina.


Alpha-synuclein is part of a diverse and highly conserved interaction network that includes PARK9 and manganese toxicity.
Parkinson's disease (PD), dementia with Lewy bodies and multiple system atrophy, collectively referred to as synucleinopathies, are associated with a diverse group of genetic and environmental susceptibilities. The best studied of these is PD. alpha-Synuclein (alpha-syn) has a key role in the pathogenesis of both familial and sporadic PD, but evidence linking it to other predisposition factors is limited. Here we report a strong genetic interaction between alpha-syn and the yeast ortholog of the PD-linked gene ATP13A2 (also known as PARK9). Dopaminergic neuron loss caused by alpha-syn overexpression in animal and neuronal PD models is rescued by coexpression of PARK9. Further, knockdown of the ATP13A2 ortholog in Caenorhabditis elegans enhances alpha-syn misfolding. These data provide a direct functional connection between alpha-syn and another PD susceptibility locus. Manganese exposure is an environmental risk factor linked to PD and PD-like syndromes. We discovered that yeast PARK9 helps to protect cells from manganese toxicity, revealing a connection between PD genetics (alpha-syn and PARK9) and an environmental risk factor (PARK9 and manganese). Finally, we show that additional genes from our yeast screen, with diverse functions, are potent modifiers of alpha-syn-induced neuron loss in animals, establishing a diverse, highly conserved interaction network for alpha-syn.


Use of reflexology in managing secondary lymphoedema for patients affected by treatments for breast cancer: A feasibility study.
The aim of this feasibility study was to examine the use of reflexology lymphatic drainage (RLD) in the treatment of breast-cancer related lymphoedema (BCRL) with a view to further research. An uncontrolled trial was conducted with 26 women who had developed lymphoedema in one arm following treatment for breast cancer. Changes in upper-limb volumes and in participant concerns and wellbeing were measured. Qualitative data were also collected. A significant reduction in the volume of the affected arm was identified at follow-up compared to baseline. This reduction in volume appeared to be maintained for more than six months. Participant concerns were significantly reduced and their wellbeing significantly increased. No serious adverse effects were reported. RLD may be a useful intervention for BCRL although the results could not be attributed to the reflexology intervention because of research design limitations. The main conclusion was, however, that there was sufficient evidence for further research using a randomized controlled trial.


Utility and limitations of measures of health inequities: a theoretical perspective.
This paper examines common approaches for quantifying health inequities and assesses the extent to which they incorporate key theories necessary for explicating the definition of health inequity. The first theoretical analysis examined the distinction between inter-individual and inter-group health inequalities as measures of health inequities. The second analysis considered the notion of fairness in health inequalities from different philosophical perspectives. To understand the extent to which different measures of health inequities incorporate these theoretical explanations, four criteria were used to assess each measure: 1) Does the indicator demonstrate inter-group or inter-individual health inequalities or both; 2) Does it reflect health inequalities in relation to socioeconomic position; 3) Is it sensitive to the absolute transfer of health (outcomes, services, or both) or income/wealth between groups; 4) Could it be used to capture inequalities in relation to other population groupings (other than socioeconomic status)? The measures assessed include: before and after measures within only the disadvantaged population, range, Gini coefficient, Pseudo-Gini coefficient, index of dissimilarity, concentration index, slope and relative indices of inequality, and regression techniques. None of these measures satisfied all the four criteria, except the range. Whereas each measure quantifies a different perspective in health inequities, using a measure within only the disadvantaged population does not measure health inequities in a meaningful way, even using before and after changes. For a more complete assessment of how programs affect health inequities, it may be useful to use more than one measure.


[Unstable blood pressure during anesthesia in diabetic patients with autonomic neuropathy].
Diabetic autonomic neuropathy (AN) as a risk factor of surgical procedures has so far been described in case reports, but no controlled clinical studies are known. Therefore, we intended to study systematically the perioperative relevance of the autonomic dysfunction. 32 ophthalmosurgical patients (20 diabetics, 12 non-diabetics) were investigated. Cardiovascular autonomic function was examined preoperatively by a combination of tests (heart rate variations during deep breathing, Valsalva ratio, 30:15 ratio, postural hypotension, sustained hand grip). A scoring system based on these tests provided the staging of the autonomic involvement of the patients (AN score). The anaesthesiological management (thiopentone, N2O, halothane) and the operative procedure (vitrectomy) were uniform. A significant correlation was found between the AN score and the coefficient of variation of the systolic (r = 0.70, p less than 0.001) resp. the diastolic (r = 0.52, p less than 0.01) blood pressure during anaesthesia. No correlation was found between the intraoperative variability of the arterial blood pressure (coefficient of variation) and the age of the patients resp. the preoperative value of the blood pressure. In diabetics with severe autonomic neuropathy (AN score greater than 5) hypotensive reactions were seen very often during the operation. The results demonstrate that the haemodynamic stability in the perioperative period depends on the severity of the autonomic dysfunction. Diabetics with severe autonomic neuropathy have a high risk of blood pressure instability. Non-invasive diagnostic methods allow to identify these patients preoperatively.


New treatments for bipolar disorder: the role of atypical neuroleptic agents.
Atypical neuroleptic agents are an excellent, safer, and more effective alternative to the widespread practice of maintenance adjunctive treatment with traditional neuroleptic agents in patients with bipolar disorder. Currently, a number of prospective studies are available with clozapine, risperidone, olanzapine, and quetiapine in the treatment of bipolar disorder. Most are short-term studies, although longer-term data are becoming available. Four double-blind studies of acute mania have been conducted with risperidone and olanzapine, leading to recent Food and Drug Administration approval for olanzapine in the indication of acute mania. Given the limited longer-term data, and the evidence for mostly adjunctive benefits with these agents, it seems unlikely that these agents will prove to be primary mood stabilizers in their own right. Nonetheless, they serve an important role as adjunctive treatments along with standard mood stabilizers in the rational polypharmacy of bipolar disorder. To date, differences in efficacy have not been established. However, differences in the side effect of weight gain may be even more relevant in bipolar disorder than in schizophrenia due to the need to use standard mood stabilizers that often potentiate such weight gain.


Revisiting gene delivery to the brain: silencing and editing.
Neurodegenerative disorders, ischemic brain diseases, and brain tumors are debilitating diseases that severely impact a person's life and could possibly lead to their demise if left untreated. Many of these diseases do not respond to small molecule therapeutics and have no effective long-term therapy. Gene therapy offers the promise of treatment or even a cure for both genetic and acquired brain diseases, mediated by either silencing or editing disease-specific genes. Indeed, in the last 5 years, significant progress has been made in the delivery of non-coding RNAs as well as gene-editing formulations to the brain. Unfortunately, the delivery is a major limiting factor for the success of gene therapies. Both viral and non-viral vectors have been used to deliver genetic information into a target cell, but they have limitations. Viral vectors provide excellent transduction efficiency but are associated with toxic effects and have limited packaging capacity; however, non-viral vectors are less toxic and show a high packaging capacity at the price of low transfection efficiency. Herein, we review the progress made in the field of brain gene therapy, particularly in the design of non-toxic and trackable non-viral vectors, capable of controlled release of genes in response to internal/external triggers, and in the delivery of formulations for gene editing. The application of these systems in the context of various brain diseases in pre-clinical and clinical tests will be discussed. Such promising approaches could potentially pave the way for clinical realization of brain gene therapies.


MDCT imaging of the aorta and peripheral vessels.
Since its clinical introduction in 1991, volumetric CT scanning using spiral or helical scanners has resulted in a revolution for diagnostic imaging. Helical CT has improved over the past 8 years with faster gantry rotation, more powerful X-ray tubes, and improved interpolation algorithms, but the greatest advance has been the recent introduction of multi detector-row CT (MDCT) scanners [J. Comput. Assist. Tomogr. 23 (1999) S83]. Currently capable of acquiring four channels of helical data simultaneously, MDCT scanners have achieved the greatest incremental gain in scan speed since the development of helical CT and have profound implications for clinical CT scanning. Fundamental advantages of MDCT include substantially shorter acquisition times, retrospective creation of thinner or thicker sections from the same raw data, and improved three-dimensional (3-D) rendering with diminished helical artifacts. While these features will likely be important to many applications of CT scanning, including the characterization of focal lung and liver lesions through the creation of thin sections retrospectively, the greatest impact has been on CT angiography. The implication for CT angiography is that scans can be performed approximately three-times faster than is possible with the fastest single-detector CT scanner. For example a 1.25 mm nominal thick section (1.6 mm effective section thickness) can be acquired with a table speed of 9.4 mm/s, and a 2.5 mm nominal thick section (3.2 mm effective section thickness) can be acquired with an 18.8 mm/s table speed. The advantages of MDCT for imaging the vascular system can be broken down into three fundamental improvements over single detector-row CT scanners speed (faster), distance (longer), and section thickness (better). The focus of this article will be how multidetector-row CT technology has substantially improved aortoiliac and lower extremity arterial imaging.


Transport of CaM kinase along processes elicited by neuronal contact evokes an inhibition of arborization and outgrowth in D. melanogaster cultured neurons.
Transgenic Drosophila strains expressing an inhibitory peptide of Ca2+/calmodulin dependent protein kinase II (CaM kinase), or a constitutively activated CaM kinase, show altered neuronal process morphology compared to wild type in scanning electron microscopy (SEM) of cultured mature neurons from embryonic neuroblasts. We observed significantly enhanced process growth in cells with inhibited enzyme, and reduced process growth in cells with activated enzyme, suggesting that active CaM kinase is involved in the inhibition of neurite growth during development. The subcellular distribution of CaM kinase in wild type neuronal cultures was determined using a gold particle labeling procedure which allowed the mapping of the enzyme directly in the scanning electron microscope (SEM). Before neuronal contact there was little labeling of processes, but after connections had been made the processes were heavily labeled. Our results suggest that the major transport of CaM kinase to the terminals does not occur until after or during the formation of neuronal connections when a functional synapse might be formed. Taken together, these results suggest a target-dependent transport of the enzyme along processes and an inhibitory role for CaM kinase on neurite branching.


Candidiasis (vulvovaginal).
Vulvovaginal candidiasis is estimated to be the second most common cause of vaginitis after bacterial vaginosis. Candida albicans accounts for 85% to 90% of cases. We conducted a systematic review and aimed to answer the following clinical questions: What are the effects of drug treatments for acute vulvovaginal candidiasis in non-pregnant symptomatic women? What are the effects of alternative or complementary treatments for acute vulvovaginal candidiasis in non-pregnant symptomatic women? What are the effects of treating a male sexual partner to resolve symptoms and prevent recurrence in non-pregnant women with symptomatic acute vulvovaginal candidiasis? What are the effects of alternative or complementary treatments for symptomatic recurrent vulvovaginal candidiasis in non-pregnant women? What are the effects of treating a male sexual partner in non-pregnant women with symptomatic recurrent vulvovaginal candidiasis? What are the effects of treating asymptomatic non-pregnant women with a positive swab for candidiasis? We searched: Medline, Embase, The Cochrane Library, and other important databases up to March 2009 (Clinical Evidence reviews are updated periodically; please check our website for the most up-to-date version of this review). We included harms alerts from relevant organisations such as the US Food and Drug Administration (FDA) and the UK Medicines and Healthcare products Regulatory Agency (MHRA). We found 61 systematic reviews, RCTs, or observational studies that met our inclusion criteria. We performed a GRADE evaluation of the quality of evidence for interventions. In this systematic review, we present information relating to the effectiveness and safety of the following interventions: alternative or complementary treatments; douching; drug treatments; garlic; intravaginal preparations (boric acid, nystatin, imidazoles, tea tree oil); oral fluconazole; oral itraconazole; treating a male sexual partner; and yoghurt containing Lactobacillus acidophilus (oral or vaginal).


The validity and reliability study of the University of California, Los Angeles Scleroderma Clinical Trial Consortium Gastrointestinal Tract (UCLA SCTC GIT) 2.0 questionnaire for the Turkish society.
To translate the University of California, Los Angeles Scleroderma Clinical Trial Consortium Gastrointestinal Tract (UCLA SCTC GIT) 2.0 questionnaire from English to Turkish and to validate it. UCLA SCTC GIT 2.0 was translated into Turkish using the translation-retranslation method. The available Turkish GIT 2.0 and the Short Form 36 (SF-36) were administered to 97 Turkish-speaking patients with systemic sclerosis (Ssc). Internal consistency reliability and structural validity were assessed by analyzing the correlations between the UCLA SCTC GIT 2.0 and the SF-36 scales. Internal consistency was determined by calculating Cronbach's alpha. For evaluation of reliability, the questionnaire scale was repeatedly applied to a subgroup of patients with a 2-week interval, and the intraclass correlation coefficient (ICC) was calculated. The Spearman's correlation coefficients between the GIT and the SF-36 scores were calculated. A group of 97 patients with Ssc with a mean age of 55.37±11.35 years and a female predominance (87.6%) were included in the study. The Cronbach's alpha value for the UCLA SCTC GIT 2.0 scale was 0.894. ICC was 0.821 (p=0.000). The scale showed acceptable reliability, with the exception of the diarrhea subscale (alpha=0.356). There was a moderate correlation between the total GIT score and the Short Form 36 (SF-36) subscales. All of the items in the scale were included in the validity analysis owing to their reliability. The Turkish GIT 2.0 scale showed good internal consistency, high reliability, and an acceptable validity.


Effect of prenatal exposure to diclofenac sodium on the male rat arteries: a stereological and histopathological study.
In this study, we investigated the morphometric and histological alterations of the aorta, brachial, and femoral arteries in 4- and 20-week-old rats that were prenatally exposed to diclofenac sodium (DS). For this purpose, pregnant rats were divided into three groups: control, saline injected, and drug treated. Beginning from day 5 after mating through day 15 of pregnancy, saline or DS (1 mg/kg daily) was intraperitoneally injected into groups 2 and 3. No injection was given to the rats in the control group. After spontaneous delivery, male offspring were obtained. At the end of weeks 4 and 20, vessel samples were removed. After dissection and routine histological preparation, histopathological and stereological investigations were made. Our results indicate that both saline and DS application lead to a decrease in the mean volume fraction of tunica media in all vessel walls, but result in an increase of the same fraction of lumen to the whole vessel wall, especially in 4-week-old rats. Elastic fibers of the vessel wall were affected by DS treatment, because a decrease of the elastic fiber was observed in this group. Finally, in light of our findings, we suggest that DS or saline may lead to vascular changes (i.e., vasodilatation or vasoconstriction) in rats that are prenatally subjected to increased volume of maternal blood resulting from injection.


The epidemiology of nosocomial epidemic Pseudomonas cepacia infections.
Pseudomonas cepacia has occasionally been identified as an epidemic and endemic nosocomial pathogen. In outbreaks, usually one clinical site predominates but many may be involved. Detailed investigations have usually implicated a contaminated liquid reservoir or moist environmental surface as the source. Liquid sources have included a number of different classes of antiseptics and disinfectants such as quaternary ammonium chlorides, biguanides, hexachlorophene, and iodophors. Environmental and patient isolates have had multiply resistant antimicrobial susceptibility patterns. The clinical distinction between colonization and infection may be difficult and may challenge the skills of the clinician. Expenditure of resources needed to solve epidemics is justified in view of the potential virulence of this organism and the high likelihood that an unrecognized but easily eliminated liquid environmental reservoir may be the source.


Multicentre study of the in vitro activity of cefepime, a broad-spectrum cephalosporin, compared to other broad-spectrum agents.
The in vitro activity of cefepime was compared to that of a range of other broad-spectrum agents, using a gradient diffusion MIC method, against 995 recent clinical isolates of Enterobacteriaceae, Pseudomonas aeruginosa, other nonfermentative gram-negative bacilli, staphylococci (except oxacillin-resistant Staphylococcus aureus), streptococci, enterococci, and aerobic gram-positive bacilli. Cefepime had excellent activity against Enterobacteriaceae, including eight presumptive extended-spectrum beta-lactamase producers and 33 stably derepressed mutants of natural cephalosporinases. Activity against Pseudomonas aeruginosa was similar to ceftazidime and superior to cefpirome. Its activity against gram-positive cocci was also good, being more active against staphylococci and only slightly less active against streptococci than ceftriaxone. Cefepime maintained activity against bacteria resistant to aminoglycosides and ciprofloxacin. Enterococci, Bacillus species, Burkholderia cepacia and Stenotrophomonas maltophilia were predicably resistant. Cefepime has a spectrum of activity almost as broad as that of the carbapenems.


Sex dependent risk factors for mortality after myocardial infarction: individual patient data meta-analysis.
Although a number of risk factors are known to predict mortality within the first years after myocardial infarction, little is known about interactions between risk factors, whereas these could contribute to accurate differentiation of patients with higher and lower risk for mortality. This study explored the effect of interactions of risk factors on all-cause mortality in patients with myocardial infarction based on individual patient data meta-analysis. Prospective data for 10,512 patients hospitalized for myocardial infarction were derived from 16 observational studies (MINDMAPS). Baseline measures included a broad set of risk factors for mortality such as age, sex, heart failure, diabetes, depression, and smoking. All two-way and three-way interactions of these risk factors were included in Lasso regression analyses to predict time-to-event related all-cause mortality. The effect of selected interactions was investigated with multilevel Cox regression models. Lasso regression selected five two-way interactions, of which four included sex. The addition of these interactions to multilevel Cox models suggested differential risk patterns for males and females. Younger women (age<50) had a higher risk for all-cause mortality than men in the same age group (HR 0.7 vs. 0.4), while men had a higher risk than women if they had depression (HR 1.4 vs. 1.1) or a low left ventricular ejection fraction (HR 1.7 vs. 1.3). Predictive accuracy of the Cox model was better for men than for women (area under the curves: 0.770 vs. 0.754). Interactions of well-known risk factors for all-cause mortality after myocardial infarction suggested important sex differences. This study gives rise to a further exploration of prediction models to improve risk assessment for men and women after myocardial infarction.


Self-splicing of Tetrahymena rRNA can proceed with phosphorothioate substitution at the splice sites.
The self-excision of a 413-base intervening sequence of the 26S rRNA of Tetrahymena thermophila has been investigated using phosphorothioate-substituted RNA. Transcripts containing this intron were prepared by T7 RNA polymerase-catalyzed polymerisation using a M13 mICE10 vector in the presence of various nucleoside alpha-thiotriphosphate analogues. Wild-type transcripts incorporating phosphorothioates 5' to adenosine or uridine were inactive, whereas incorporation 5' to cytidine or guanosine allowed splicing. The first two substitutions place phosphorothioates inter alia at the 5' and 3' splice sites respectively. Mutagenesis at either site allowed phosphorothioate substitution 5' to guanosine at each splice site. This did not block splicing, suggesting that substitution at internal sites within the intron has more effect.


Inhaled hydrogen gas therapy for prevention of lung transplant-induced ischemia/reperfusion injury in rats.
Successful abrogation of ischemia/reperfusion (I/R) injury of lung grafts could significantly improve short- and long-term outcomes for lung transplant (LTx) recipients. Hydrogen gas has potent antioxidant and antiapoptotic properties and has been recently used in number of experimental and clinical studies. The purpose of this research was to investigate whether inhaled hydrogen gas could reduce graft I/R injury during lung transplantation. Orthotopic left LTxs were performed in syngenic Lewis rats. Grafts were perfused with and stored in low potassium dextran solution at 4°C for 6 hr. The recipients received 100% O2 or 98% O2 with 2% N2, 2% He, or 2% H2 during surgery and 1 hr after reperfusion. The effects of hydrogen were assessed by functional, pathologic, and molecular analysis. Gas exchange was markedly impaired in animals exposed to 100% O2, 2% N2, or 2% He. Hydrogen inhalation attenuated graft injury as indicated by significantly improved gas exchange 2 hr after reperfusion. Graft lipid peroxidation was significantly reduced in the presence of hydrogen, demonstrating antioxidant effects of hydrogen in the transplanted lungs. Lung cold I/R injury causes the rapid production and release of several proinflammatory mediators and epithelial apoptosis. Exposure to 2% H2 significantly blocked the production of several proinflammatory mediators and reduced apoptosis with induction of the antiapoptotic molecules B-cell lymphoma-2 and B-cell lymphoma-extra large. Treatment of LTx recipients with inhaled hydrogen can prevent lung I/R injury and significantly improve the function of lung grafts after extended cold preservation, transplant, and reperfusion.


Adoptive cellular gene therapy of autoimmune disease.
Autoimmune disorders represent inappropriate immune responses directed at self-tissue. Because CD4+ T cells are important mediators in the pathogenesis of autoimmune disease, they are ideal candidates for cell-based gene therapy. Using retrovirally-transduced cells and luciferase bioluminescence, we have demonstrated that primary T cells and hybridomas, rapidly and preferentially home to the sites of inflammation in organ-specific autoimmune disease. These cells, transduced with retroviral vectors to drive expression of various 'regulatory proteins', such as IL-4, IL-10 and IL-12p40, deliver these immunoregulatory proteins to the inflamed lesions, providing therapy for experimental models of autoimmune disease such as EAE, CIA and NOD mice. This technique was originally developed in our lab in the murine model of multiple sclerosis, EAE, where T cell hybridomas reactive with myelin basic protein (MBP) were transduced to express and used to deliver the modulatory cytokine, IL-4. Recently we have observed that the cytokine receptor antagonist, IL-12p40 transduced anti-myelin basic protein (MBP) TCR-transgenic T cells (but not CII-reactive T cells) were effective in preventing EAE whereas the CII-reactive, but not MBP-reactive T cells, transduced to express IL-12p40, would treat CIA.


Arabidopsis ATPase family gene 1-like protein 1 is a calmodulin-binding AAA+-ATPase with a dual localization in chloroplasts and mitochondria.
Members of the AAA(+)-ATPase superfamily (ATPases associated with various cellular activities) are found in all kingdoms of life and they are involved in very diverse cellular processes, including protein degradation, membrane fusion or cell division. The Arabidopsis genome encodes approximately 140 different proteins that are putative members of this superfamily, although the exact function of most of these proteins remains unknown. Using affinity chromatography on calmodulin-agarose with chloroplast proteins, we purified a 50 kDa protein encoded by AT4G30490 with similarity to the ATPase family gene 1 protein from yeast. Structural analysis showed that the protein possesses a single AAA-domain characteristic for members of the AAA(+)-ATPase superfamily and that this contains all features specific to proteins of the ATPase family gene 1-like subfamily. In vitro pull-down as well as cross-linking assays corroborate calcium-dependent binding of the protein to calmodulin. The calmodulin binding domain could be located to a region of 20 amino acids within the AAA-domain in close proximity to the Walker A motif. Our analysis further showed that the protein is localized in both mitochondria and chloroplasts, further supporting the incorporation of both endosymbiotic organelles into the calcium-signaling network of the cell. Localization of the same calmodulin-binding protein into mitochondria and chloroplasts could be a means to provide a coordinated regulation of processes in both organelles by calcium signals.


Modulation of heterochromatin protein 1 dynamics in primary Mammalian cells.
Heterochromatin protein 1 (HP1beta), a key component of condensed DNA, is strongly implicated in gene silencing and centromeric cohesion. Heterochromatin has been considered a static structure, stabilizing crucial aspects of nuclear organization and prohibiting access to transcription factors. We demonstrate here, by fluorescence recovery after photobleaching, that a green fluorescent protein-HP1beta fusion protein is highly mobile within both the euchromatin and heterochromatin of ex vivo resting murine T cells. Moreover, T cell activation greatly increased this mobility, indicating that such a process may facilitate (hetero)chromatin remodeling and permit access of epigenetic modifiers and transcription factors to the many genes that are consequently derepressed.