MassSQUIRM: An assay for quantitative measurement of lysine demethylase activity.
In eukaryotes, DNA is wrapped around proteins called histones and is condensed into chromatin. Post-translational modification of histones can result in changes in gene expression. One of the most well-studied histone modifications is the methylation of lysine 4 on histone H3 (H3K4). This residue can be mono-, di- or tri-methylated and these varying methylation states have been associated with different levels of gene expression. Understanding exactly what the purpose of these methylation states is, in terms of gene expression, has been a topic of much research in recent years. Enzymes that can add (methyltransferases) and remove (demethylases) these modifications are of particular interest. The first demethylase discovered, LSD1, is the most well-classified and has been implicated in contributing to human cancers and to DNA damage response pathways. Currently, there are limited methods for accurately studying the activity of demethylases in vitro or in vivo. In this work, we present MassSQUIRM (mass spectrometric quantitation using isotopic reductive methylation), a quantitative method for studying the activity of demethylases capable of removing mono- and di-methyl marks from lysine residues. We focus specifically on LSD1 due to its potential as a prime therapeutic target for human disease. This quantitative approach will enable better characterization of the activity of LSD1 and other chromatin modifying enzymes in vitro, in vivo or in response to inhibitors.


Prescription Birth Control Access Among U.S. Women at Risk of Unintended Pregnancy.
Access to contraception is a vital component of preventing unintended pregnancies. This study was conducted to assess the prevalence of and factors associated with U.S. women's difficulty accessing prescription contraception (pill, patch, or ring). We performed a nationally representative survey of adult women at risk of unintended pregnancy (aged 18-44, not pregnant or seeking pregnancy, sexually active, not sterilized) using a probability-based web panel. In November to December 2011, 2046 women completed the survey. Weighted proportions were calculated and logistic regression was used to identify covariates associated with difficulty obtaining or refilling prescription contraception. A total of 1385 (68%) women had ever tried to get a prescription for hormonal contraception. Among this population, 29% reported ever having problems obtaining a prescription or refills. In multivariable regression, uninsured (vs. privately insured) and Spanish-speaking (vs. English-speaking) women were significantly more likely to report difficulties; women with a high school degree and those with some college (vs. a college degree or higher) were significantly less likely to report difficulty. Difficulties included cost barriers or lack of insurance (14%), challenges obtaining an appointment or getting to a clinic (13%), the clinician requiring a clinic visit, exam, or Pap smear (13%), not having a regular doctor/clinic (10%), difficulty accessing a pharmacy (4%), and other reasons (4%). One-third of adult U.S. women who have ever tried to obtain prescription contraception reported access barriers. While the Affordable Care Act may resolve some of these issues, these data indicate that additional factors may still need to be addressed.


Differential coupling of membrane Ig and CD40 to the extracellularly regulated kinase signaling pathway.
Coupling of membrane Ig (mIg) and CD40 to the extracellularly regulated kinase (ERK) signal transduction pathway was examined in the WEHI-231 B lymphoma and normal mouse B cells. Cross-linking mIg induces ERK activation in both WEHI-231 and normal B cells. In contrast, CD40 cross-linking failed to induce ERK activation in WEHI-231, but signals through CD40 were more effective than mIg as a stimulus for ERK activation in normal B cells. However, several lines of evidence suggest that CD40 and the B cell Ag regulate ERK through distinct pathways that converge at the level of MEK-1, mitogen-activated protein kinase kinase. Abs to mIg or CD40 induced MEK-1 activation with different kinetics. Cross-linking of mIg, but not CD40, induced tyrosine phosphorylation of the SHC adapter molecule that couples receptors to Ras-dependent signaling pathways. Finally, agents that elevate cAMP, causing protein kinase A-mediated inhibition of Raf-1, inhibited activation of ERK in response to mIg cross-linking, but had no affect on ERK activation in response to anti-CD40 or Jun N-terminal kinase activation by signals through either receptor. Thus, CD40 uses an unidentified protein kinase A-insensitive MEK kinase, rather than Raf-1, to regulate ERK activity.


Sudden infant death syndrome and prolongation of the QT interval.
A standard lead II ECG was recorded during either the first or the fourth week of life or at both ages from 30 neonates whose sibling had died of the sudden infant death syndrome (SIDS). Electrocardiographic recordings also were obtained from 75 control neonates and from 52 adults who had had an infant who died of SIDS. The neonatal data revealed that the QT interval, corrected for heart rate (QTC), was longest during NREM (vs rapid eye movement [REM]) sleep. Furthermore, the QTC interval was longer within the fourth week than in the first week of life. However, the QTC interval of siblings of SIDS victims did not differ from that of the control infants, nor did the QT interval of parents of SIDS victims differ from published normal values. One neonate who subsequently died of SIDS did not have an abnormally long QTC interval. These data do not support the hypothesis that genetically determined prolongation of the QT interval plays a major role in SIDS.


The autoimmune risk gene ZMIZ1 is a vitamin D responsive marker of a molecular phenotype of multiple sclerosis.
Multiple Sclerosis (MS) is a neurological condition driven in part by immune cells from the peripheral circulation, the targets for current successful therapies. The autoimmune and MS risk gene ZMIZ1 is underexpressed in blood in people with MS. We show that, from three independent sets of transcriptomic data, expression of ZMIZ1 is tightly correlated with that of hundreds of other genes. Further we show expression is partially heritable (heritability 0.26), relatively stable over time, predominantly in plasmacytoid dendritic cells and non-classical monocytes, and that levels of ZMIZ1 protein expression are reduced in MS. ZMIZ1 gene expression is increased in response to calcipotriol (1,25 Vitamin D3) (p < 0.0003) and associated with Epstein Barr Virus (EBV) EBNA-1 antibody titre (p < 0.004). MS therapies fingolimod and dimethyl fumarate altered blood ZMIZ1 gene expression compared to untreated MS. The phenotype indicates susceptibility to MS, and may correspond with clinical response and represent a novel clinical target.


Vector competence of Aedes aegypti, Culex tarsalis, and Culex quinquefasciatus from California for Zika virus.
Zika virus (ZIKV) has emerged since 2013 as a significant global human health threat following outbreaks in the Pacific Islands and rapid spread throughout South and Central America. Severe congenital and neurological sequelae have been linked to ZIKV infections. Assessing the ability of common mosquito species to transmit ZIKV and characterizing variation in mosquito transmission of different ZIKV strains is important for estimating regional outbreak potential and for prioritizing local mosquito control strategies for Aedes and Culex species. In this study, we evaluated the laboratory vector competence of Aedes aegypti, Culex quinquefasciatus, and Culex tarsalis that originated in areas of California where ZIKV cases in travelers since 2015 were frequent. We compared infection, dissemination, and transmission rates by measuring ZIKV RNA levels in cohorts of mosquitoes that ingested blood meals from type I interferon-deficient mice infected with either a Puerto Rican ZIKV strain from 2015 (PR15), a Brazilian ZIKV strain from 2015 (BR15), or an ancestral Asian-lineage Malaysian ZIKV strain from 1966 (MA66). With PR15, Cx. quinquefasciatus was refractory to infection (0%, N = 42) and Cx. tarsalis was infected at 4% (N = 46). No ZIKV RNA was detected in saliva from either Culex species 14 or 21 days post feeding (dpf). In contrast, Ae. aegypti developed infection rates of 85% (PR15; N = 46), 90% (BR15; N = 20), and 81% (MA66; N = 85) 14 or 15 dpf. Although MA66-infected Ae. aegypti showed higher levels of ZIKV RNA in mosquito bodies and legs, transmission rates were not significantly different across virus strains (P = 0.13, Fisher's exact test). To confirm infectivity and measure the transmitted ZIKV dose, we enumerated infectious ZIKV in Ae. aegypti saliva using Vero cell plaque assays. The expectorated plaque forming units PFU varied by viral strain: MA66-infected expectorated 13±4 PFU (mean±SE, N = 13) compared to 29±6 PFU for PR15-infected (N = 13) and 35±8 PFU for BR15-infected (N = 6; ANOVA, df = 2, F = 3.8, P = 0.035). These laboratory vector competence results support an emerging consensus that Cx. tarsalis and Cx. quinquefasciatus are not vectors of ZIKV. These results also indicate that Ae. aegypti from California are efficient laboratory vectors of ancestral and contemporary Asian lineage ZIKV.


Complementary learning systems within the hippocampus: a neural network modelling approach to reconciling episodic memory with statistical learning.
A growing literature suggests that the hippocampus is critical for the rapid extraction of regularities from the environment. Although this fits with the known role of the hippocampus in rapid learning, it seems at odds with the idea that the hippocampus specializes in memorizing individual episodes. In particular, the Complementary Learning Systems theory argues that there is a computational trade-off between learning the specifics of individual experiences and regularities that hold across those experiences. We asked whether it is possible for the hippocampus to handle both statistical learning and memorization of individual episodes. We exposed a neural network model that instantiates known properties of hippocampal projections and subfields to sequences of items with temporal regularities. We found that the monosynaptic pathway-the pathway connecting entorhinal cortex directly to region CA1-was able to support statistical learning, while the trisynaptic pathway-connecting entorhinal cortex to CA1 through dentate gyrus and CA3-learned individual episodes, with apparent representations of regularities resulting from associative reactivation through recurrence. Thus, in paradigms involving rapid learning, the computational trade-off between learning episodes and regularities may be handled by separate anatomical pathways within the hippocampus itself.This article is part of the themed issue 'New frontiers for statistical learning in the cognitive sciences'.


Locating the two black holes in NGC 6240.
Mergers play an important role in galaxy evolution and are key to understanding the correlation between central-black hole mass and host-galaxy properties. We used the new technology of adaptive optics at the Keck II telescope to observe NGC 6240, a merger between two disk galaxies. Our high-resolution near-infrared images, combined with radio and x-ray positions, revealed the location and environment of two central supermassive black holes. Each is at the center of a rotating stellar disk, surrounded by a cloud of young star clusters. The brightest of these young clusters lie in the plane of each disk, but surprisingly are seen only on the disks' receding side.


[Neuropsychological approach to visual attention].
Visual experience depends critically on visual attention, which selects a particular aspect of a visual display. Recent clinical, neuroimaging, and animal studies revealed that visual attention was divided into active and passive or top-down and bottom-up attention. Although these dichotomies are clear-cut in definition, visual attention could be modulated by many factors. Detailed observation of brain-injured patients provides with evidence for dynamic and fine control of visual attention. We observed patients with dorsal simultanagnosia and that with callosal disconnection syndrome. Patients with dorsal simultanagnosia demonstrated that extent of visual attention was dynamically changed depending on the level of visual processing. Despite the ability to read a kanji character and to describe its components correctly, a patient could not notice a component that he had just written and could not assemble individual components to make up a correct kanji character. He could point to an overlapping area of two figures. But once he started to color the overlapping area, he missed the margin of the area and colored much larger area. Another patient with dorsal simultanagnosia missed borderlines between columns of a newspaper and read letters continuously across columns. In contrast, he could point to lines between figures or meaningless patterns easily. These findings indicated that visual attention was directed automatically to meaningful characters. A patients with callosal disconnection syndrome demonstrated left unilateral spatial neglect only when he used his right hand to draw figures. Right hand movement, controlled by the left hemisphere, elicited visual attention to the right hemispace, resulting in the left unilateral spatial neglect. Thus visual attention is not simply top-down or bottom up, but is implicitly affected by the visual recognition as well as motor component of the task.


Report of case: Partial ureteral obstruction masked by diuretics during intraoperative cystoscopy.
Injury to the lower urinary tract is a potential complication in all major vaginal and urogynecologic surgical procedures. Several authors have recommended the routine use of intraoperative cystoscopy during urogynecologic procedures. To evaluate possible injury to the lower urinary tract during intraoperative cystoscopy, the concomitant use of diuretics with indigo carmine dye has been advocated; efflux of dye is hypothesized to indicate functional patency of the urinary tract. This report describes a case in which a partial ureteral obstruction was present at the time of intraoperative cystoscopy--despite the observation of diuresis caused by furosemide. This case indicates that the efflux of indigo carmine-stained urine from both ureteral orifices is not conclusive evidence of the absence of ureteral insult during intraoperative cystoscopy.


Diversity of antibiotic resistance integrative and conjugative elements among haemophili.
The objective of this study was to investigate the sequence diversity in a single country of a family of integrative and conjugative elements (ICEs) that are vectors of antibiotic resistance in Haemophilus influenzae and Haemophilus parainfluenzae, and test the hypothesis that they emerged from a single lineage. Sixty subjects aged 9 months - 13 years were recruited and oropharyngeal samples cultured. Up to 10 morphologically distinct Pasteurellaceae spp. were purified, and then the species were determined and differentiated by partial sequence analysis of 16S rDNA and mdh, respectively. ICEs were detected by PCR directed at five genes distributed evenly across the ICE. These amplicons were sequenced and aligned by the neighbour-joining algorithm. A total of 339 distinguishable isolates were cultured. ICEs with all 5 genes present were found in 9 of 110 (8 %) H. influenzae and 21 of 211 (10 %) H. parainfluenzae, respectively. ICEs were not detected among the other Pasteurellaceae. A total of 20 of 60 (33 %) children carried at least 1 oropharyngeal isolate with an ICE possessing all 5 genes. One of the five genes, integrase, however, consisted of two lineages, one of which was highly associated with H. influenzae. The topology of neighbour-joining trees of the remaining four ICE genes was compared and showed a lack of congruence; though, the genes form a common pool among H. influenzae and H. parainfluenzae. This family of antibiotic resistance ICEs was prevalent among the children studied, was genetically diverse, formed a large gene pool, transferred between H. influenzae and H. parainfluenzae, lacked population structure and possessed features suggestive of panmixia, all indicating it has not recently emerged from a single source.


Effect of nilvadipine on the development of neurological deficits in stroke-prone spontaneously hypertensive rats.
Several types of antihypertensive drugs have been reported to protect stroke-prone spontaneously hypertensive rats from stroke. However, the clinical relevance remains unclear. This study was performed to investigate the effect of nilvadipine, a calcium channel blocker, on the development of neurological deficits in stroke-prone spontaneously hypertensive rats. In addition, plasma levels of nilvadipine were measured to determine the clinical relevance. Salt-loaded stroke-prone spontaneously hypertensive rats were orally administered nilvadipine mixed with a powder diet (0.01% and 0.03%, wt/wt). Non-salt-loaded rats were maintained on tap water. Chronological changes in neurological deficit scores and systolic blood pressure were recorded. After 6 weeks of medication, measurement of plasma levels of nilvadipine, serum biochemical analysis, and pathological observation of both the brain and the kidney were performed. In the salt-loaded control group, both severe hypertension and neurological deficit developed, and the final survival rate was 30%. Systolic blood pressure decreased significantly in the high-dose nilvadipine-treated group but not in the low-dose nilvadipine-treated group. However, the development of neurological deficit was almost completely inhibited in both nilvadipine-treated groups that had no deaths (P < .01). The mean plasma levels of nilvadipine in the low-dose group and in the high-dose group at the time of death were 0.21 ng/mL and 0.61 ng/mL, respectively. Nilvadipine inhibited the development of neurological deficit in stroke-prone spontaneously hypertensive rats at plasma concentrations lower than that in clinical use. Thus, nilvadipine might prevent cerebral vascular disorders at doses routinely used for essential hypertension.


[Surgical alternatives in complex pancreatic injuries].
Objective of this study is to analyze the treatment outcomes of patients undergoing surgery for complex pancreatic injuries in our center over a 20-year period. Between January 1980 and December 2001, medical records of 11 patients who underwent surgical intervention for complex pancreatic trauma were investigated retrospectively. The study group consisted of nine males and two female patients with a mean age of 36 years. Six patients had stab wounds, three had motor vehicle accidents and two had gunshot wounds. The mean value of Injury Severity Score (ISS) was 32. Among six patients with Grade III injury, five were managed by distal pancreatectomy and one by external drainage. In three patients with Grade IV injury each underwent distal pancreatectomy, subtotal pancreatectomy and pancreaticoduodenectomy. The remaining two patients with Grade V injury were managed by pancreaticoduodenectomy. In the postoperative course, pancreatic fistula was observed in two patients and peripancreatic abscess and pancreatitis in one patient each. Endocrine insufficiency occurred in one patient who underwent subtotal pancreatectomy. Operative mortality was 18.2%. Owing to the retroperitoneal location of the pancreas and its proximity to major vascular structures, associated organ injuries play a significant role in morbidity and mortality.


Clinical pathologists should limit modifier terms used to denote probability of a diagnosis: a survey-based study.
To examine the probability estimates for modifying terms used by clinical pathologists when interpreting cytologic samples and compare these to probability estimates assigned to these terms by clinicians, and to provide restricted, standardizing terms used in cytology reports. 49 clinical pathologists and 466 Veterinary Information Network members responded to 2 similar surveys. Online surveys were distributed to diplomates of the European College of Veterinary Clinical Pathologists and clinician members of the Veterinary Information Network, made available between March 17, 2022, through May 5, 2022. Respondents assigned a range of probabilities to each of 18 modifier terms used by clinical pathologists to denote probability associated with diagnoses; clinicians identified terms that would affect their treatment decisions in cases of canine lymphoma. Respondents then provided thoughts about restricting and standardizing modifying terms and assigning numeric estimates in reports. 49 clinical pathologists and 466 clinicians provided responses. For many terms, probability ranges agreed between the 2 groups. However, differences in estimated probability inferred by a term existed for at least 6 terms. Modifying terms could be restricted to 7 largely nonoverlapping terms that spanned the range of probabilities. Clinicians preferred having numeric estimates of probability, but clinical pathologists resisted providing such estimates in reports. Reducing and standardizing the number of modifying terms to reflect specific probability ranges would reduce disagreement between the clinical pathologist's intended probability range and the clinician's interpretation of a modifying term. This could result in fewer errors in interpretation and better patient care.


Topology optimized mode conversion in a photonic crystal waveguide fabricated in silicon-on-insulator material.
We have designed and for the first time experimentally verified a topology optimized mode converter with a footprint of ~6.3 μm × ~3.6 μm which converts the fundamental even mode to the higher order odd mode of a dispersion engineered photonic crystal waveguide. 2D and 3D topology optimization is utilized and both schemes result in designs theoretically showing an extinction ratio larger than 21 dB. The 3D optimized design has an experimentally estimated insertion loss lower than ~2 dB in an ~43 nm bandwidth. The mode conversion is experimentally confirmed in this wavelength range by recording mode profiles using vertical grating couplers and an infrared camera. The experimentally determined extinction ratio is > 12 dB and is believed to be limited by the spatial resolution of our setup.


Disturbing post-operative symptoms are not reduced by prophylactic antiemetic treatment in patients at high risk of post-operative nausea and vomiting.
To give prophylactics or timely treatment for post-operative nausea and vomiting (PONV) is the question. We compared the intensity and number of disturbing post-operative symptoms (i.e. pain, PONV, headache, fatigue, etc.) after prophylactic antiemetic treatment in a group of patients with >30% risk for post-operative vomiting. Four hundred and ninety-five patients, from three hospitals, planned for gynaecological surgery were randomized double blind. They were given granisetron 3 mg, droperidol 1.25 mg or no prophylactic antiemetic. Post-operative symptoms were followed for 24 h using a questionnaire. Symptoms were analyzed both according to their intensity and in a dichotomous fashion. The intensity of different symptoms differed depending on whether droperidol, granisetron or no antiemetic had been given (P = 0.005) but the overall incidence of moderate to very severe symptoms was similar in all groups. No group fared better in general. The total number of symptoms was higher in the groups given prophylactic treatment (P < 0.05). The relative risk reduction for PONV with granisetron or droperidol prophylaxis was 27%[95% confidence interval (CI) 8-43] and 22% (2-38), respectively. The NNT (number needed to treat) for granisetron (0-24 h) was 7 and for droperidol 8. The NNH (number needed to harm) (0-24 h) for headache and visual disturbances was 6 and 13 (NS) for granisteron and, 50 (NS) and 6 for droperidol. The intensity of symptoms or the total number of disturbing symptoms did not decrease after prophylactic antiemetic treatment in a group of patients, but the profile of disturbing symptoms changed. The relevance of post-operative symptoms in terms of patients' well-being needs to be addressed.


Is Wolff-Parkinson-White syndrome a genetic disease?
Family studies, and more recent molecular genetic investigations, indicate that the Wolff-Parkinson-White (WPW) syndrome and associated preexcitation disorders can have a substantial genetic component. Because preexcitation disorders are sometimes inherited as single gene disorders, key mechanistic insights can be gained that are expected to be relevant also to the more common multifactorial forms of these traits. Potentially, such insights will inform the future management of these conditions. Where WPW is inherited as a familial trait, with or without associated cardiac defects or a systemic syndrome, there are clinical genetic ramifications that are already of practical importance.


The chaperone-mediated autophagy receptor organizes in dynamic protein complexes at the lysosomal membrane.
Chaperone-mediated autophagy (CMA) is a selective type of autophagy by which specific cytosolic proteins are sent to lysosomes for degradation. Substrate proteins bind to the lysosomal membrane through the lysosome-associated membrane protein type 2A (LAMP-2A), one of the three splice variants of the lamp2 gene, and this binding is limiting for their degradation via CMA. However, the mechanisms of substrate binding and uptake remain unknown. We report here that LAMP-2A organizes at the lysosomal membrane into protein complexes of different sizes. The assembly and disassembly of these complexes are a very dynamic process directly related to CMA activity. Substrate proteins only bind to monomeric LAMP-2A, while the efficient translocation of substrates requires the formation of a particular high-molecular-weight LAMP-2A complex. The two major chaperones related to CMA, hsc70 and hsp90, play critical roles in the functional dynamics of the LAMP-2A complexes at the lysosomal membrane. Thus, we have identified a novel function for hsc70 in the disassembly of LAMP-2A from these complexes, whereas the presence of lysosome-associated hsp90 is essential to preserve the stability of LAMP-2A at the lysosomal membrane.


Short-term carbon cycling responses of a mature eucalypt woodland to gradual stepwise enrichment of atmospheric CO2 concentration.
Projections of future climate are highly sensitive to uncertainties regarding carbon (C) uptake and storage by terrestrial ecosystems. The Eucalyptus Free-Air CO2 Enrichment (EucFACE) experiment was established to study the effects of elevated atmospheric CO2 concentrations (eCO2 ) on a native mature eucalypt woodland with low fertility soils in southeast Australia. In contrast to other FACE experiments, the concentration of CO2 at EucFACE was increased gradually in steps above ambient (+0, 30, 60, 90, 120, and 150 ppm CO2 above ambient of ~400 ppm), with each step lasting approximately 5 weeks. This provided a unique opportunity to study the short-term (weeks to months) response of C cycle flux components to eCO2 across a range of CO2 concentrations in an intact ecosystem. Soil CO2 efflux (i.e., soil respiration or Rsoil ) increased in response to initial enrichment (e.g., +30 and +60 ppm CO2 ) but did not continue to increase as the CO2 enrichment was stepped up to higher concentrations. Light-saturated photosynthesis of canopy leaves (Asat ) also showed similar stimulation by elevated CO2 at +60 ppm as at +150 ppm CO2 . The lack of significant effects of eCO2 on soil moisture, microbial biomass, or activity suggests that the increase in Rsoil likely reflected increased root and rhizosphere respiration rather than increased microbial decomposition of soil organic matter. This rapid increase in Rsoil suggests that under eCO2, additional photosynthate was produced, transported belowground, and respired. The consequences of this increased belowground activity and whether it is sustained through time in mature ecosystems under eCO2 are a priority for future research.


Multiple cross mapping (MCM) markedly improves the localization of a QTL for ethanol-induced activation.
This study examines the use of multiple cross mapping (MCM) to reduce the interval for an ethanol response QTL on mouse chromosome 1. The phenotype is the acute locomotor response to a 1.5-g/kg i.p. dose of ethanol. The MCM panel consisted of the six unique intercrosses that can be obtained from the C57BL/6J (B6), DBA/2J (D2), BALB/cJ (C) and LP/J (LP) inbred mouse strains (N > or = 600/cross). Ethanol response QTL were detected only with the B6xD2 and B6xC intercrosses. For both crosses, the D2 and C alleles were dominant and decreased ethanol response. The QTL information was used to develop an algorithm for sorting and editing the chromosome 1 Mit microsatellite marker set (http://www.jax.org). This process yielded a cluster of markers between 82 and 85cM (MGI). Evidence that the QTL was localized in or near this interval was obtained by the analysis of a sample (n = 550) of advanced cross heterogenous stock animals. In addition, it was observed that one of the BXD recombinant inbred strains (BXD-32) had a recombination in the interval of interest which produced the expected change in behavior. Overall, the data obtained suggest that the information available within existing genetic maps coupled with MCM data can be used to reduce the QTL interval. In addition, the MCM data set can be used to interrogate gene expression data to estimate which polymorphisms within the interval of interest are relevant to the QTL.


Chest physiotherapy does not exacerbate gastroesophageal reflux in patients with chronic bronchitis and bronchiectasis.
Chest physiotherapy is beneficial to patients with mucus hypersecretion. However, it is not risk-free. Chest physiotherapy may affect gastrointestinal motility and the competence of the gastroesophageal sphincter during the procedure. Our study was aimed to investigate whether gastroesophageal reflux is exacerbated or induced by chest physiotherapy. Thirty-two adult patients with chronic bronchitis or bronchiectasis received chest physiotherapy. An esophageal pH meter was placed 5 cm above the gastroesophageal junction and monitored the reflux episodes from 8 AM on the first day to 12 noon on the second day. Physiotherapy was performed with the patient in three different positions on the second day for one hour. Thirteen patients were found to have gastroesophageal reflux disease. For the study group as a whole, neither the frequency nor the duration of having a pH < 4 during the physiotherapy was significantly different from the mean hourly frequency and mean hourly duration recorded on the first day or from those values recorded at a corresponding time on the first day. As for the patients with gastroesophageal reflux disease, the frequency and total duration of gastroesophageal reflux during physiotherapy were not significantly different from the mean hourly frequency and the mean hourly duration or from the values recorded at the corresponding time on the first day. There was no significant difference in the gastroesophageal reflux frequency or the duration in different body positions. Chest physiotherapy including postural drainage, percussion and forced expiration techniques in different positions did not induce or increase the incidence of gastroesophageal reflux in patients with chronic bronchitis and bronchiectasis.


Efficacy and safety of programmed cell death-1/programmed cell death ligand-1 inhibitors in advanced urothelial malignancy: A systematic review and meta-analysis.
Programmed cell death-1/programmed cell death ligand-1 (PD-1/PDL-1) inhibitors are the newest class of approved drugs for advanced urothelial cancer (AdUC). This review aims to collate the evidence for their efficacy and safety in various treatment settings. Extensive search of databases was performed (updated May 2018) and the protocol was registered on PROSPERO (CRD42017081568). The review was conducted according to the Preferred Reporting Items for Systematic reviews and Meta-Analysis statement. STATA (v 12) and Revman 5.3.5 were used for data analysis. Ten nonrandomized, open-label clinical trials were included in this review. PD-1/PD-L1 inhibitors were used as second-line, stand-alone in eight trials and as first-line in cisplatin-ineligible in two trials. Heterogeneity was observed for study design, PDL-1 testing methods, cutoff criterias used and translational markers evaluated. The pooled objective response rate (ORR) was 18.2% (95% confidence interval [CI] 15.1-21.2, n = 1785) with PD-1/PDL-1 inhibitors in second-line settings as compared to 12.6% (95% CI 10.3-14.9, n = 736) with second-line chemotherapy and 23.7% (95% CI 19.9-27.4, n = 489) with PD-1/PDL-1 inhibitors as first-line therapy in cisplatin-ineligible patients. The median progression-free survival and overall survival was similar with PD-1/PD-L1 inhibitors in both second- and first-line treatment settings (1.5-2.9 vs. 2.0-2.7 months and 7.9-18.2 vs. 15.9 months) and second-line chemotherapy (3.3-4.0 months and 7.4-8 months). Odds of achieving ORR was 0.10 (95% CI 0.03-0.31, n = 229) in the second-line, stand-alone setting with a combined positive score (CPS) cutoff of 25% and was 0.34 (95% CI 0.19-0.62, n = 265) with a CPS cut-off of 10% in first-line setting in the cisplatin-ineligible. PD-1/PDL-1 inhibitors appear to be promising in the treatment of AdUC and CPS may be a potentially reliable biomarker for predicting response but needs validation. Caution needs to be exercised until more data are available on imAEs and further studies are required to prove their worth as the standard of care.


Characterization of mitophagy in the 6-hydoxydopamine Parkinson's disease model.
In the present study, the activation of autophagy and its interaction with the mitochondrial fission machinery was investigated in an experimental model of Parkinson's disease. The addition of 50µM 6-hydroxydopamine (6-OHDA) to the dopaminergic cell line SH-SY5Y profoundly stimulated formation of autophagosomes within 12h. Under these conditions, mitochondrial fission was also activated in a sustained manner, but this occurred at earlier time points (after 3h). Upon 6-OHDA treatment, dynamin-related protein 1 (Drp1) transiently translocated to mitochondria, with increased levels of mitochondrial Drp1 being observed after 3 and 9h. Pharmacological inhibition of Drp1, through treatment with the mitochondrial-division inhibitor-1 (mdivi-1), resulted in the abrogation of mitochondrial fission and in a decrease of the number of autophagic cells. In addition, 6-OHDA failed to induce the expression of the proapoptotic protein Bax in total cellular extracts although it did induce its migration to mitochondria. In our model, Bax migrated later than Drp1. However, Drp1 inhibition did not block Bax migration. These results show that reactive oxygen species but not quinone derivates act as mediators of autophagy at an early stage of the process. 6-OHDA induces hydrogen peroxide production, which was placed upstream of mitochondrial fission, given that mdivi-1 did not abrogate this increase. Furthermore, the 6-OHDA-induced activation of autophagy was also suppressed by addition of the free radical scavengers TEMPOL and MnTBAP. This effect could be reproduced by the addition of hydrogen peroxide, but not with aged 6-OHDA. To our knowledge, this is the first detailed study highlighting the various mediators that are implicated in mitochondrial alterations and autophagy of cells in response to 6-OHDA.


Examining Key Stakeholder and Community Residents' Understanding of Environmental Influences to Inform Place-Based Interventions to Reduce Obesity in Rural Communities, Kentucky 2015.
Rural residents report high rates of obesity, physical inactivity, and poor eating habits. The objectives of this study were to (1) use the collective impact model to guide efforts to elicit community members' perceptions of county-specific factors influencing high obesity rates; (2) determine the association between utilization of food retail venues and concern about obesity and healthy eating; and (3) determine community members' utilization of physical activity infrastructure and concern about physical inactivity. The study was conducted in 6 rural counties in Kentucky with adult obesity prevalence rates >40%. Community stakeholders met to assess counties' needs and assets in implementing interventions to reduce obesity in their communities. A random-digit dial survey (n = 756) also was conducted to examine awareness and availability of community resources for healthy eating and physical activity. Stakeholders identified lack of access to fruits and vegetables and poor physical activity infrastructure as contributors to obesity. Reporting moderate and serious concern about obesity and healthy eating was associated with higher odds of shopping at a supercenter compared with those expressing little concern. Reported access to information about physical activity opportunities was associated with higher odds of reporting the availability of safe places for physical activity, sidewalks, and trails compared with those who reported that information was difficult to obtain. This study elicits community-identified barriers to healthy behaviors and provides foundational data to inform future place-based obesity reduction interventions.


Cardiac ferroportin regulates cellular iron homeostasis and is important for cardiac function.
Iron is essential to the cell. Both iron deficiency and overload impinge negatively on cardiac health. Thus, effective iron homeostasis is important for cardiac function. Ferroportin (FPN), the only known mammalian iron-exporting protein, plays an essential role in iron homeostasis at the systemic level. It increases systemic iron availability by releasing iron from the cells of the duodenum, spleen, and liver, the sites of iron absorption, recycling, and storage respectively. However, FPN is also found in tissues with no known role in systemic iron handling, such as the heart, where its function remains unknown. To explore this function, we generated mice with a cardiomyocyte-specific deletion of Fpn. We show that these animals have severely impaired cardiac function, with a median survival of 22 wk, despite otherwise unaltered systemic iron status. We then compared their phenotype with that of ubiquitous hepcidin knockouts, a recognized model of the iron-loading disease hemochromatosis. The phenotype of the hepcidin knockouts was far milder, with normal survival up to 12 mo, despite far greater iron loading in the hearts. Histological examination demonstrated that, although cardiac iron accumulates within the cardiomyocytes of Fpn knockouts, it accumulates predominantly in other cell types in the hepcidin knockouts. We conclude, first, that cardiomyocyte FPN is essential for intracellular iron homeostasis and, second, that the site of deposition of iron within the heart determines the severity with which it affects cardiac function. Both findings have significant implications for the assessment and treatment of cardiac complications of iron dysregulation.


Comparative effects of teriparatide and strontium ranelate in the periosteum of iliac crest biopsies in postmenopausal women with osteoporosis.
The periosteum contains osteogenic cells that regulate the outer shape of bone and contribute to determine its cortical thickness, size and position. We assessed the effects of subcutaneous injections of teriparatide (TPTD, 20μg/day) or oral strontium ranelate (SrR, 2g/day) in postmenopausal women with osteoporosis on new bone formation activity at the periosteal and endosteal bone surfaces using dynamic histomorphometric measurements. Evaluable tetracycline-labeled transiliac crest bone biopsies were analyzed from 27 patients in the TPTD group, and 22 in the SrR group after six months of treatment. Measurements were conducted on the thicker and thinner cortices separately, and comparisons between the thicker, thinner and combined cortices were carried out. At the combined periosteal cortex, the mineralization surface as a percent of bone surface (MS/BS%) was greater for TPTD (mean±SE: 8.08±1.22%) than SrR (3.22±1.05%) (p<0.005). The difference in mineral apposition rate (MAR) between TPTD (0.35±0.06μm/day) and SrR (0.14±0.06μm/day) was also significant (p<0.05), while that of bone formation rate per bone surface (BFR/BS) between TPTD (0.014±0.004 mm(3)/mm(2)/year) and SrR (0.004±0.003 mm(3)/mm(2)/year) was not (p=0.057). Statistically significant differences between the two treatments were also observed for MS/BS%, BFR/BS, MAR and the double-labeled perimeter in the periosteum of the thicker, but not thinner, iliac crest cortices. The comparison between the thicker and thinner cortices of both periosteal and endosteal surfaces showed statistically significant differences for MAR and the double-labeled perimeter for TPTD treated women. There were no statistically significant differences in any bone formation dynamic measurements between the two cortices in the SrR group. In conclusion, most of the bone formation and mineralization variables were significantly higher for TPTD- than SrR-treated women at both the periosteal and endosteal combined cortices. The response to TPTD for dynamic bone formation measurements in the periosteal surface was greater for the thicker than thinner cortex, but this difference was not significant in SrR treated patients. This may reflect a greater ability of TPTD to enhance responsiveness of bone to the mechanical loading environment. These effects on bone formation may underlie the improvement in bone quality in patients with osteoporosis treated with TPTD.


Oncology nursing in the Global South during COVID-19.
In mid-2020, a call was made to oncology nurses in the Global South to share their experiences managing patient care during the coronavirus disease 2019 (COVID-19) pandemic. Eighteen submissions were received from 16 countries across Latin America, Africa, Europe and Asia. Three were research-based and 15 were personal narratives on the psychosocial impact of COVID-19 on the nurses, colleagues, patients and families. Three narratives were from oncology nurses working with cancer-related non-governmental organisations locally or, in one case, internationally. A simultaneous literature search for publications (including grey literature) was performed to identify themes of COVID-19's impact in these 16 countries and specifically on oncology nurses and patients/families. Four themes were identified: a) interruptions to care; b) support/resource shortages; c) psychosocial impact on nurses and patients and d) staffing and nursing role impacts. The three research-based studies describe oncology nursing in-depth efforts to explore the impact of COVID-19. Findings in the 15 narratives are briefly presented according to the four themes identified in the literature. Due to the severe shortage of physician adult and paediatric oncology specialists, oncology nurses in the Global South often shoulder much of the care for patients with cancer and even more so during COVID-19 with attendant oncology nursing shortages due to reassignment to COVID-19 units. It is important to hear from these critical members of the oncology nursing workforce who often lack the time, resources or training to publish in peer-reviewed journals in English, particularly in the middle of a pandemic. Giving voice to these nurses documents the reality of their work and ability to continue to provide care despite the chaos and rapidly changing guidelines and government action. Lessons learned by these nurses to improve mental health and psychosocial support of the nurses as well as their patients/families will be essential for the next global pandemic.


Fixation mechanisms of nanoparticles on substrates by electron beam irradiation.
For applications such as the fabrication of plasmonic waveguides we developed a patterning technique to fabricate an array of nanoparticles on a substrate using focused electron beams (Noriki, T.; Abe, S.;.Kajikawa, K.; Shimojo, M. Beilstein J. Nanotechnol.2015,6, 1010-1015). This technique consists of three steps: Firstly, nanoparticles are placed over the entire surface of a substrate. Secondly, the nanoparticles are fixed on the substrate by focused electron beam irradiation. The electron beam decomposes the organic molecules located around the particle into amorphous carbon. The amorphous carbon immobilizes the particle on the substrate. Finally, the unfixed nanoparticles are removed. However, in this original technique, the area in which the nanoparticles were fixed was wider than the electron-probe size of a few nanometers. To understand this widening mechanisms, the effects of accelerating voltage, particle size and substrate material are investigated by means of both experiments and simulation. It is demonstrated that the fixing area is greatly affected by the electrons back-scattered by the substrate. The back-scattering leads to an increase in line width and thus reduces the resolution of this patterning technique.


Diversity and host range of foliar fungal endophytes: are tropical leaves biodiversity hotspots?
Fungal endophytes are found in asymptomatic photosynthetic tissues of all major lineages of land plants. The ubiquity of these cryptic symbionts is clear, but the scale of their diversity, host range, and geographic distributions are unknown. To explore the putative hyperdiversity of tropical leaf endophytes, we compared endophyte communities along a broad latitudinal gradient from the Canadian arctic to the lowland tropical forest of central Panama. Here, we use molecular sequence data from 1403 endophyte strains to show that endophytes increase in incidence, diversity, and host breadth from arctic to tropical sites. Endophyte communities from higher latitudes are characterized by relatively few species from many different classes of Ascomycota, whereas tropical endophyte assemblages are dominated by a small number of classes with a very large number of endophytic species. The most easily cultivated endophytes from tropical plants have wide host ranges, but communities are dominated by a large number of rare species whose host range is unclear. Even when only the most easily cultured species are considered, leaves of tropical trees represent hotspots of fungal species diversity, containing numerous species not yet recovered from other biomes. The challenge remains to recover and identify those elusive and rarely cultured taxa with narrower host ranges, and to elucidate the ecological roles of these little-known symbionts in tropical forests.


Environmental stratifications for soybean cultivar recommendation and its consistency over time.
Stratification of environments is a strategy to capitalize genotype x environment (GxE) interaction, which can optimize the process of assessment and cultivar recommendation, increasing productivity in a target environmental population. The objective of this study was to assess environmental stratification methods based on the analysis of GxE interaction, to identify consistent agronomic zones across time for soybean. Grain yield data of inbred lines from three maturity groups (early, medium, and late) were used. Lines and cultivars were tested in regional variety trials during three growing seasons at eighteen locations in the tropics of Central Brazil. Three methods were applied to stratify the environments. The first was based on joint analyses of variance for all the pairs of locations within each growing year. The second was based on a distance measure between each pair of locations, which was related to the GxE interaction estimated via additive main effects and multiplicative interaction analysis. The third was based on the approach of winning genotypes. The stratification results from the first two methods were not consistent across the growing seasons. However, the winning genotype approach provided consistent environmental stratification across years. From locations used in the genotypic assessment, three environmental clusters were identified for the early and medium maturity groups of soybean, and four clusters for the late maturity group. The use of different genotypic sets across years reinforces the predictive value of the environmental stratification established.


SVAS3: Strain Vector Aided Sensorization of Soft Structures.
Soft material structures exhibit high deformability and conformability which can be useful for many engineering applications such as robots adapting to unstructured and dynamic environments. However, the fact that they have almost infinite degrees of freedom challenges conventional sensory systems and sensorization approaches due to the difficulties in adapting to soft structure deformations. In this paper, we address this challenge by proposing a novel method which designs flexible sensor morphologies to sense soft material deformations by using a functional material called conductive thermoplastic elastomer (CTPE). This model-based design method, called Strain Vector Aided Sensorization of Soft Structures (SVAS3), provides a simulation platform which analyzes soft body deformations and automatically finds suitable locations for CTPE-based strain gauge sensors to gather strain information which best characterizes the deformation. Our chosen sensor material CTPE exhibits a set of unique behaviors in terms of strain length electrical conductivity, elasticity, and shape adaptability, allowing us to flexibly design sensor morphology that can best capture strain distributions in a given soft structure. We evaluate the performance of our approach by both simulated and real-world experiments and discuss the potential and limitations.


Investigating the genetic variation underlying episodicity in major depressive disorder: suggestive evidence for a bipolar contribution.
Highly recurrent major depressive disorder (MDD) has reportedly increased risk of shifting to bipolar disorder; high recurrence frequency has, therefore, featured as evidence of 'soft bipolarity'. We aimed to investigate the genetic underpinnings of total depressive episode count in recurrent MDD. Our primary sample included 1966 MDD cases with negative family history of bipolar disorder from the RADIANT studies. Total episode count was adjusted for gender, age, MDD duration, study and center before being tested for association with genotype in two separate genome-wide analyses (GWAS), in the full set and in a subset of 1364 cases with positive family history of MDD (FH+). We also calculated polygenic scores from the Psychiatric Genomics Consortium MDD and bipolar disorder studies. Episodicity (especially intermediate episode counts) was an independent index of MDD familial aggregation, replicating previous reports. The GWAS produced no genome-wide significant findings. The strongest signals were detected in the full set at MAGI1 (p=5.1×10(-7)), previously associated with bipolar disorder, and in the FH+ subset at STIM1 (p=3.9×10(-6) after imputation), a calcium channel signaling gene. However, these findings failed to replicate in an independent Munich cohort. In the full set polygenic profile analyses, MDD polygenes predicted episodicity better than bipolar polygenes; however, in the FH+ subset, both polygenic scores performed similarly. Episode count was self-reported and, therefore, subject to recall bias. Our findings lend preliminary support to the hypothesis that highly recurrent MDD with FH+ is part of a 'soft bipolar spectrum' but await replication in larger cohorts.


Delay in onset of insulitis in NOD mice following a single injection of CD 4 and CD 8 antibodies.
Non-obese diabetic (NOD) mice injected with 500 micrograms of both CD 4 and CD 8 antibodies at 5 weeks of age did not develop insulitis until 18 weeks of age, 12 weeks later than the onset of insulitis in parallel uninjected controls. Injection with both antibodies at 2 weeks or 4 weeks of age protected from insulitis for 10 and 14 weeks respectively. Insulitis was not delayed in onset in animals injected at any age with one antibody only, or who were injected at birth. Injection after the onset of insulitis achieved partial but incomplete clearing of islet infiltrates. Salivary gland infiltrates (sialitis) were also delayed in animals injected with both CD 4 and CD 8 antibodies though the degree of protection was less pronounced than that seen for insulitis.


Common complication of crush injury, but a rare compartment syndrome.
Compartment syndrome (CS) is a common complication of crush injury but it is rare to find bilateral gluteal compartment syndrome (BGCS). Only six cases of BGCS have been reported in the literature. This syndrome has been reported after crush injury, drug overdose, surgical positioning, and vascular surgery. Apart from CS, crush injury is associated with multi-system adverse effects and these patients are at high risk for renal failure and sepsis. CS patients may present with dehydration; coagulation disorders; elevated creatine phosphokinase and myoglobin levels; hyperkalemia and hypocalcaemia, which may cause life-threatening arrhythmias and therefore need urgent and aggressive therapy. The early goal in these patients is prevention of acute renal failure with aggressive fluid therapy, alkalinization of urine, and forced diuresis. Early treatment of hyperkalemia, antibiotic therapy, immunoprophylaxis, and wound care will minimize the risk of arrhythmias and sepsis. CS must be considered when any patient is diagnosed with crush injury syndrome. CS is defined as elevation of interstitial/ intracompartmental pressure, leading to microvascular and myoneural dysfunction and secondary hypoxia; it may cause functional loss or even death if not detected early and treated properly. The increase in pressure in one or all compartments of the gluteal region causes CS with devastating effects on muscle and neurovascular bundles. CS is traditionally diagnosed on the basis of five 'p's: pain, pallor, paraesthesia, pulselessness and paralysis. Diagnosis of gluteal CS is difficult as the peripheral pulses are preserved and the condition is usually only diagnosed when neurological abnormality is noticed. Diagnosis of CS can be made by direct measurement of the compartment pressure and magnetic resonance imaging or computerized tomography. Gluteal CS is managed by fasciotomy and debridement of necrosed tissue, with secondary closure of fascia. A high index of suspicion is necessary for the early diagnosis of gluteal CS, and this will reduce the disability and complications as a consequence of this syndrome. The acute-care physician, the intensivist, and the trauma surgeon must be aware of this rare syndrome, as it can result in multiorgan dysfunction and death. Here we report a case of bilateral gluteal CS that was successfully treated in our trauma intensive care unit.


Rapid diagnostic method of tobacco products in saliva by fourier transform infrared spectroscopy (FTIR).
The present study was designed to explore the easy and fast method diagnosis of tobacco products in saliva of tobacco users (TU) by FTIR. Sixty four male tobacco users (TU) with mean age range 15.3 to 30.7 years were randomly selected for collection of saliva samples before and after tobacco use (smoking, chewing and dipping tobacco). Twenty were the smoking tobacco users (STU), 24 were chewing tobacco users (CTU) and 20 were dipping tobacco users (DTU). CTU were the users of Mainpuri (n=10) and users of PEN, FIT, 2100 (n=14). Forty eight saliva samples of age and gender matched healthy individuals with negative personal or family history of any addiction were also collected for comparison which served as controls. All were analyzed for their salivary flow rate, salivary pH and salivary diagnostic bands by FTIR. Significantly increased SFR (p<0.05) and salivary pH were found in after chewing tobacco as compared to before its chewing. The comparison between after tobacco use and controls we found decreased SFR and salivary pH for STU. Significant decreased SFR and increased salivary pH were found before or after use of dipping tobacco as compared to controls. Sharp bands at 735-745 cm-1 were found and may be used as salivary diagnostic bands for STU, 945-949 cm-1 for DTU and 900-915 cm-1 for CTU as well as DTU. In conclusion, the salivary diagnostic bands were found at 735-745 cm-1, 900-915 cm-1 and 945-949 cm-1 for TU by easy and fast method using FTIR.


IMMUNITY FACTORS IN PNEUMOCOCCUS INFECTION IN THE DOG.
Intravenous inoculations of from 1 to 3 cc. per kilo of body weight of a bouillon culture of virulent pneumococci produce septicemia and meningitis in dogs. The injected pneumococci leave the circulation rapidly, but begin to reinvade the blood from 24 to 48 hours later. The septicemia reaches its climax between the 4th and 5th days and then abruptly declines, the blood becoming sterile within from 1 to 3 days after the height of the septicemia is reached. The initial disappearance of the pneumococci from the circulation has been found to be due to agglutination of the diplococci in the blood stream and accumulation of the clumps in the lungs, liver, spleen, etc. If the dogs are reinoculated during the ascension of the septicemia, the injected diplococci leave the circulation as rapidly as in normal dogs. Cultures isolated in this stage of the infection, both before and from 3 to 4 hours after the reinoculation, are resistant to the agglutinins and opsonins of immune sera that agglutinate and opsonize the cultures with which the dogs were originally infected. Thus it follows that the pneumococci are able to reinvade the circulation because they have acquired a fastness to the existing antibodies and not because the antibodies have been bound or exhausted. By reinoculating dogs at the time of the crisis in the septicemia it has been shown that the agglutination of the pneumococci is more rapid and complete and that the diplococci leave the circulation much more rapidly than in normal dogs. Hence acquired antibodies are operative within the animals at this time although they cannot be demonstrated in vitro until from 24 to 48 hours later. Pneumococci isolated as the infection is subsiding are more susceptible to the action of immune sera than the original cultures injected. It is probable that all the dogs would have survived the infection if a meningitis had not developed. In the acutely fatal cases of meningitis few pneumococci are phagocyted, while in the milder and convalescent cases much phagocytosis occurs. It is suggested that the incubation period of infectious diseases is due to the fact that the infecting agents must become adapted to the adverse conditions encountered in the newly infected host before they can multiply sufficiently to produce the symptoms of disease. It is further suggested that epidemics may arise because the infectious agent is passed from person to person in the ascending stage of the disease and thus enters new hosts in a state of maximum resistance to the natural antibodies of such individuals. When early contacts are avoided, epidemics tend to subside because the infectious agent is weakened by the action of acquired antibodies during the period of convalescence.


The Use of a 3D Printer in Pre-operative Planning for a Patient Requiring Acetabular Reconstructive Surgery.
Surgical management of acetabular fractures is often highly complex, and a successful outcome depends upon an appreciation of the fracture pattern and the most appropriate approach to reduce and hold it. Currently, computed tomography (CT) images are used in conjunction with plain x-rays to identify the main fracture components and their spatial relationship to one another, and as such surgeons still have to make decisions based upon their ability to visualise the fracture from the images available. 3D printers have now become widely available and inexpensive, and can be used to rapidly produce life-size models based on CT scans of an individual patient. The availability of patient specific, accurate and detailed models of complex acetabular fractures can aid planning of surgical management on a patient specific basis. This report describes the use of a 3D printer to create a life-size model reconstruction of the pelvis of a 48 year old male patient who sustained a left sided associated both column acetabular fracture following a motorbike accident in the Sahara Desert. The model allowed visualisation of the multiple fracture fragments and their relative displacements. The tactile feedback allowed assessment of the different fracture fragments. The relative displacement of the quadrilateral plate and posterior column fragments could be assessed and the surgeon felt that these would be amenable to reduction from an ilioinguinal approach. An anatomic reduction was achieved and was held with the application of a pelvic brim plate with 2 screws lagging the posterior column/quadrilateral plate fragment. There are previous examples of 3D models being used in orthopaedic surgery through the use of rapid prototyping, however this method is usually expensive and time consuming. Advances in 3D printer technology offer surgeons a number of advantages when treating these complex fractures. With the ever-increasing economy, ease of use and speed of additive processing, the possible applications of this technology within orthopaedic surgery are numerous. Given the possible applications of this technology, and its ever increasing availability, we feel that its use can only improve patient outcomes and so should be explored further for use in orthopaedic surgery.


Regulation of global acetylation in mitosis through loss of histone acetyltransferases and deacetylases from chromatin.
Histone acetylation, a reversible modification of the core histones, is widely accepted to be involved in remodeling chromatin organization for genetic reprogramming. Histone acetylation is a dynamic process that is regulated by two classes of enzymes, the histone acetyltransferases (HATs) and histone deacetylases (HDACs). Although promoter-specific acetylation and deacetylation has received most of the recent attention, it is superimposed upon a broader acting and dynamic acetylation that profoundly affects many nuclear processes. In this study, we monitored this broader histone acetylation as cells enter and exit mitosis. In contrast to the hypothesis that HATs and HDACs remain bound to mitotic chromosomes to provide an epigenetic imprint for postmitotic reactivation of the genome, we observed that HATs and HDACs are spatially reorganized and displaced from condensing chromosomes as cells progress through mitosis. During mitosis, HATs and HDACs are unable to acetylate or deacetylate chromatin in situ despite remaining fully catalytically active when isolated from mitotic cells and assayed in vitro. Our results demonstrate that HATs and HDACs do not stably bind to the genome to function as an epigenetic mechanism of selective postmitotic gene activation. Our results, however, do support a role for spatial organization of these enzymes within the cell nucleus and their relationship to euchromatin and heterochromatin postmitotically in the reactivation of the genome.


Bacterial lipopolysaccharide suppresses the production of catalytically active lysosomal acid hydrolases in human macrophages.
Sub-microgram quantities of bacterial lipopolysaccharide (LPS) have been found to substantially reduce the intracellular catalytic activities of three representative lysosomal enzymes (namely, acid phosphatase, hexosaminidase, and beta-glucuronidase) in human monocyte-derived macrophages. This response was not associated with a concurrent increase in enzyme catalytic activity in the culture supernatant, and hence, could not be explained by mobilization of preformed material. By conducting experiments in the presence and absence of indomethacin, a cyclooxygenase inhibitor, the reduction in lysosomal enzyme catalytic activities was shown not to be dependent on the ability of LPS to induce prostaglandin E2 production. The response was not found to be the result of a more generalized LPS-dependent reduction in the ability of the cells to synthesize protein, since the presence of LPS in macrophage cultures did not appreciably affect the amount of [35S]methionine incorporated into total cellular proteins. A kinetic analysis of the effect of LPS on the down-regulation of enzyme catalytic activities indicated that this was an early response of the cells to LPS exposure. An investigation of the effects of blockade of enzyme catabolism (using the lysosomotropic weak-base, methylamine) indicated that the reduction of catalytic enzyme activities in response to LPS was probably due to a decreased rate of production of active product, rather than an enhanced rate of enzyme catabolism. This suggestion was confirmed by experiments in which the synthesis of pro-hexosaminidase (measured by biosynthetic labeling with [35S]methionine and specific immunoprecipitation of labeled pro-hexosaminidase) was found to be reduced by 42% after a 24-h exposure to LPS (although the synthesis of complement component C3 was stimulated by a factor of 4.5). It is suggested that the ability of LPS to regulate the functional expression of protein products contributes to changes in the overall functional status of these cells in response to this bacterial product.


High risk of silent stroke in older bipolar patients with typical-onset age.
Older patients with bipolar disorder (BD) are at a high risk of stroke. Silent stroke could be neglected in BD patients after middle age, particularly in those with illness onset at the young age. Therefore, we investigated the morbidity and related factors for stroke in older BD patients with typical-onset age. Older patients with bipolar I disorder (age > 50 years) and their illness onset prior to the age of 40 years were recruited. After whole-brain magnetic resonance imaging was conducted, the patients were divided into stroke and non-stroke groups. Clinical data were obtained by reviewing all available medical records and directly interviewing the patients along with their reliable family members. We recruited 62 patients with a mean age of 60.4 years and illness onset at the mean age of 28.4 years. Cerebral infarction or old stroke was observed in 24 (38.7%) patients, including 22 without any reported clinical history of stroke. That is, silent stroke (n = 22) was detected in 36.7% of 60 patients without clinical history of stroke. The stroke group had significantly higher mean numbers of lifetime mood episodes (P = 0.006) than the non-stroke one. Logistic regression analysis showed that 10 or more prior mood episodes (odds ratio = 3.43, 95% confidence interval = 1.12-10.47, P < 0·04) was significantly associated with the occurrence of stroke. The two study groups did not exhibit any other differences in demographic and clinical variables, such as age, laboratory or physical measurements during the last acute psychiatric hospitalisation, body mass index, and substance use problems and concurrent medical diseases. High morbidity of stroke, particularly silent stroke, could be found in older bipolar patients with typical-onset age. In addition to traditional risk factors, the number of recurrent mood episodes in a lifetime may increase the risk of stroke in older BD patients.


Structure-Guided Redesign Improves NFL HIV Env Trimer Integrity and Identifies an Inter-Protomer Disulfide Permitting Post-Expression Cleavage.
Soluble HIV-1 envelope glycoprotein (Env) trimers are under active investigation as vaccine candidates in relevant pre-clinical models. Like SOSIPs, the cleavage-independent native flexibly linked (NFL) trimers are faithful mimics of the Env spike. Here, we analyzed multiple new designs to explore alternative modifications, informing tertiary interactions, while maintaining NFL trimer homogeneity and integrity. Accordingly, we performed a proline (P) substitution screen in the gp41 heptad repeat 1 region, identifying other trimer-enhancing Ps, including L555P. This P improved trimer integrity compared to I559P in selected properties. Next, we screened 15 structure-guided potential cysteine pairs in gp140 and found that A501C-L663C ("CC2") forms an inter-protomer disulfide bond that demonstrably increased NFL trimer thermostability. We combined these two approaches with trimer-derived substitutions, coupled with glycine substitutions at helix-to-coil transitions, developed by our group. To increase the exposure of the fusion peptide (FP) N-terminus, we engineered an enterokinase (EK) cleavage site upstream of the FP for controlled post-expression cleavage. In combination, the redesigns resulted in highly stable and homogeneous NFL mimics derived from different clades. Following recombinant EK cleavage, the NFL trimers retained covalent linkage, maintaining a native-like structure while displaying enhanced stability and favorable antigenic features. These trimers also displayed increased exposure of neutralizing epitopes in the FP and gp120/gp41 interface, while retaining other neutralizing epitopes and occluding non-neutralizing elements. This array of Env-structure-guided designs reveals additional interactive regions in the prefusion state of the HIV Env spike, affording the development of novel antigens and immunogens.


Acoustofluidic particle trapping, manipulation, and release using dynamic-mode cantilever sensors.
We show here that dynamic-mode cantilever sensors enable acoustofluidic fluid mixing and trapping of suspended particles as well as the rapid manipulation and release of trapped micro-particles via mode switching in liquid. Resonant modes of piezoelectric cantilever sensors over the 0 to 8 MHz frequency range are investigated. Sensor impedance response, flow visualization studies using dye and micro-particle tracers (100 μm diameter), and finite element simulations of cantilever modal mechanics and acoustic streaming show fluid mixing and particle trapping configurations depend on the resonant mode shape. We found trapped particles could be: (1) rapidly manipulated on millimeter length scales, and (2) released from the cantilever surface after trapping by switching between low- and high-order resonant modes (less than 250 kHz and greater than 1 MHz, respectively). Such results suggest a potentially promising future for dynamic-mode cantilevers in separations, pumping and mixing applications as well as acoustofluidic-enhanced sensing applications.


Sexual effects of puncturevine (Tribulus terrestris) extract (protodioscin): an evaluation using a rat model.
Apart from its claims for improvement of sexual functions in men, the puncturevine plant (Tribulus terrestris: TT) has long been considered as an energizer and vitalizer in the indigenous system of medicine. Sexual behavior and intracavernous pressure (ICP) measurements were taken in rats to scientifically validate the claim of TT [containing protodioscin (PTN)] as an aphrodisiac. Forty sexually mature male Sprague-Dawley rats were randomly divided into four groups of 10 each. Group I served as a control group and groups II, III, and IV were treated with three different doses of TT extract (2.5, 5 and 10 mg/kg body weight, respectively), orally, once daily for 8 weeks. Weight was recorded and the rats from all four groups were subjected to sexual behavior studies with primed females and various parameters namely mount and intromission frequencies (MF and IF, respectively), mount, intromission and ejaculation latencies (ML, IL, and EL, respectively) as well as postejaculatory interval (PEI) were recorded. In addition, blood pressure and ICP were recorded for all rats at the end of study. Increases in body weight (by 9, 23, and 18% for groups II, III & IV) and ICP (by 43% and 26% for groups III and IV) were statistically significant compared to the control group. Increases in MF (by 27% and 24%) and IF (by 19% and 22%) for the groups III and IV were statistically significant. Decreases in ML (by 16%, 23%, and 22% for groups II, III, and IV) and PEI (by 20% for group III) were statistically significant compared to the control. The weight gain and improvement in sexual behavior parameters observed in rats could be secondary to the androgen increasing property of TT (PTN) that was observed in our earlier study on primates. The increase in ICP which confirms the proerectile aphrodisiac property of TT could possibly be the result of an increase in androgen and subsequent release of nitric oxide from the nerve endings innervating the corpus cavernosum.


A role for SUMOylation in the Formation and Cellular Localization of TDP-43 Aggregates in Amyotrophic Lateral Sclerosis.
In amyotrophic lateral sclerosis, motor neurons undergoing degeneration are characterized by the presence of cytoplasmic aggregates containing TDP-43 protein. SUMOylation, a posttranslational modification of proteins, has been previously implicated in the formation of aggregates positives for SOD1, another protein enriched in a subset of ALS patients. We show in this study that TDP-43 is also a target of SUMOylation. The inhibition of the first step of the SUMOylation process by anacardic acid significantly reduces the presence of TDP-43 aggregates and improves neuritogenesis and cell viability in vitro. Interestingly, the mutation of the unique SUMOylation site on TDP-43, using site-directed mutagenesis, modifies the intracellular localization of TDP-43 aggregates. Instead of being cytoplasmic where they are associated with toxic effects, they are located inside the nucleus. This change of localization results in improvement in cell viability and in global cellular functions. Our results implicate the SUMOylation site of TDP-43 in the formation of cytoplasmic TDP-43 aggregates, a hallmark of ALS, and thus identifies this region as a new target for novel therapeutic strategies.


Biomarkers of mercury toxicity: Past, present, and future trends.
Mercury (Hg) toxicity continues to represent a global health concern. Given that human populations are mostly exposed to low chronic levels of mercurial compounds (methylmercury through fish, mercury vapor from dental amalgams, and ethylmercury from vaccines), the need for more sensitive and refined tools to assess the effects and/or susceptibility to adverse metal-mediated health risks remains. Traditional biomarkers, such as hair or blood Hg levels, are practical and provide a reliable measure of exposure, but given intra-population variability, it is difficult to establish accurate cause-effect relationships. It is therefore important to identify and validate biomarkers that are predictive of early adverse effects prior to adverse health outcomes becoming irreversible. This review describes the predominant biomarkers used by toxicologists and epidemiologists to evaluate exposure, effect and susceptibility to Hg compounds, weighing on their advantages and disadvantages. Most importantly, and in light of recent findings on the molecular mechanisms underlying Hg-mediated toxicity, potential novel biomarkers that might be predictive of toxic effect are presented, and the applicability of these parameters in risk assessment is examined.


Happiness and Social Behavior.
It is often assumed that there is a robust positive symmetrical relationship between happiness and social behavior: Social relationships are viewed as essential to happiness, and happiness is thought to foster social relationships. However, empirical support for this widely held view is surprisingly mixed, and this view does little to clarify which social partner a person will be motivated to interact with when happy. To address these issues, we monitored the happiness and social interactions of more than 30,000 people for a month. We found that patterns of social interaction followed the hedonic-flexibility principle, whereby people tend to engage in happiness-enhancing social relationships when they feel bad and sustain happiness-decreasing periods of solitude and less pleasant types of social relationships that might promise long-term payoff when they feel good. These findings demonstrate that links between happiness and social behavior are more complex than often assumed in the positive-emotion literature.


Fertility treatments and multiple births in the United States.
The advent of fertility treatments has led to an increase in the rate of multiple births in the United States. However, the trends in and magnitude of the contribution of fertility treatments to the increase are uncertain. We derived the rates of multiple births after natural conception from data on distributions of all births from 1962 through 1966 (before fertility treatments were available). Publicly available data on births from 1971 through 2011 were used to determine national multiple birth rates, and data on in vitro fertilization (IVF) from 1997 through 2011 were used to estimate the annual proportion of multiple births that were attributable to IVF and to non-IVF fertility treatments, after adjustment for maternal age. Trends in multiple births were examined starting from 1998, the year when clinical practice guidelines for IVF were developed with an aim toward reducing the incidence of multiple births. We estimated that by 2011, a total of 36% of twin births and 77% of triplet and higher-order births resulted from conception assisted by fertility treatments. The observed incidence of twin births increased by a factor of 1.9 from 1971 to 2009. The incidence of triplet and higher-order births increased by a factor of 6.7 from 1971 to 1998 and decreased by 29% from 1998 to 2011. This decrease coincided with a 70% reduction in the transfer of three or more embryos during IVF (P<0.001) and a 33% decrease in the proportion of triplet and higher-order births attributable to IVF (P<0.001). Over the past four decades, the increased use of fertility treatments in the United States has been associated with a substantial rise in the rate of multiple births. The rate of triplet and higher-order births has declined over the past decade in the context of a reduction in the transfer of three or more embryos during IVF. (Funded by the Centers for Disease Control and Prevention.).


Magnesium in drinking water and ischemic heart disease.
The associations found in the general populations of a number of different countries are suggestive and warrant an integrated program of laboratory and epidemiologic research to reject or confirm the magnesium-IHD hypothesis. Singling out this particular risk factor has two justifications. First, as would be the case with any epidemiologic risk factor for IHD whose attributable risk was large enough to be detectable through epidemiology, applying that attributable risk to the vast annual morbidity and mortality from IHD would translate into tens of thousands of lives benefited and millions of dollars in hospital costs avoided per year. Second, this particular risk factor could conceivably be eliminated by an inexpensive supplementation program. For example, a low-sodium, higher-magnesium and -potassium table salt has been recommended and used in Finland for many years, during a period when the prevalence of hypertension in population surveys was said to decrease (117). Interventions which do not require behavioral change have always been the most cost-effective in public health. We therefore urge funding agencies to give priority to studies determining whether there are unforeseen adverse effects of magnesium for some population subgroups and whether the apparent benefit derived from low doses of magnesium in the development of IHD or IHD death is real. Furthermore, researchers should determine which chemical form of magnesium is best absorbed and most effective. We need to better understand the interrelation of various water and food constituents, as well as individual risk factors, in the pathogenesis of IHD. Susceptible individuals who are at higher risk of being depleted of magnesium need to be identified, and potential untoward effects of magnesium should be studied. Future research must provide better answers about low level waterborne magnesium before recommendations to the public can be made.


Androgen deprivation increases p300 expression in prostate cancer cells.
Standard therapy for nonorgan confined prostate cancer aims to block the production or action of androgens. Although initially successful, antiandrogen therapy eventually fails and androgen depletion independent (ADI) disease emerges. Remarkably, ADI prostate cancers still rely on a functional androgen receptor (AR). Aberrant expression of coregulatory proteins required for the formation of productive AR transcriptional complexes is critical for ADI AR activation. Previously, we have shown that the transcriptional coactivator p300 is required for ADI activation of the AR and is up-regulated in prostate cancer, in which its expression is associated with cell proliferation and predicts aggressive tumor features. The mechanism responsible for the deregulated expression of p300, however, remains elusive. Here, we show that p300 expression in prostate cancer cells is subject to androgen regulation. In several prostate cancer model systems, addition of synthetic and natural androgens led to decreased expression of p300 in a time-dependent and dose-dependent manner. Experiments using AR antagonists or small interfering RNA targeting the AR revealed that down-regulation of p300 depends entirely on the presence of a functional AR. It is noteworthy that androgens down-regulated p300 protein expression while leaving messenger levels unaltered. Conversely, both short-term and long-term androgen deprivation resulted in marked up-regulation of p300 expression. The androgen deprivation-induced increase in p300 expression was not affected by the addition of cytokines or growth factors or by cotreatment with antiandrogens. Moreover, increased p300 expression upon androgen starvation is crucial for prostate cancer cell proliferation, as loss of p300 expression severely reduces expression of cyclins governing G(1)-S and G(2)-M cell cycle transition and decreases 5-bromo-2'-deoxyuridine incorporation.


P-glycoprotein regulates trafficking of CD8(+) T cells to the brain parenchyma.
The trafficking of cytotoxic CD8(+) T lymphocytes across the lining of the cerebral vasculature is key to the onset of the chronic neuro-inflammatory disorder multiple sclerosis. However, the mechanisms controlling their final transmigration across the brain endothelium remain unknown. Here, we describe that CD8(+) T lymphocyte trafficking into the brain is dependent on the activity of the brain endothelial adenosine triphosphate-binding cassette transporter P-glycoprotein. Silencing P-glycoprotein activity selectively reduced the trafficking of CD8(+) T cells across the brain endothelium in vitro as well as in vivo. In response to formation of the T cell-endothelial synapse, P-glycoprotein was found to regulate secretion of endothelial (C-C motif) ligand 2 (CCL2), a chemokine that mediates CD8(+) T cell migration in vitro. Notably, CCL2 levels were significantly enhanced in microvessels isolated from human multiple sclerosis lesions in comparison with non-neurological controls. Endothelial cell-specific elimination of CCL2 in mice subjected to experimental autoimmune encephalomyelitis also significantly diminished the accumulation of CD8(+) T cells compared to wild-type animals. Collectively, these results highlight a novel (patho)physiological role for P-glycoprotein in CD8(+) T cell trafficking into the central nervous system during neuro-inflammation and illustrate CCL2 secretion as a potential link in this mechanism.


Demographic, Mental Health, Behavioral, and Psychosocial Factors Associated with Cigarette Smoking Status Among Young Men Who Have Sex with Men: The P18 Cohort Study.
Young sexual minority men smoke at higher rates relative to heterosexual peers. The purpose of this study was to examine correlates of smoking in a sample of young gay, bisexual, and other men who have sex with men (MSM) who might differ from more general and age-diverse samples of sexual minority individuals and, thus, inform tailored approaches to addressing tobacco use within this population. Data on smoking status were examined in relation to demographics, mental health, substance use behavior, and psychosocial factors. Using multinomial logistic regression, factors were identified that differentiate current and former smokers from never smokers. In bivariate analysis, smoking status was related to demographic, mental health, substance use, and psychosocial factors. Most significantly, smoking status was associated with school enrollment status, current alcohol and marijuana use, and symptoms of depression. Multivariate modeling revealed that, compared to being a never smoker, the odds of current or former smoking were highest among those currently using either alcohol or marijuana. The odds of both current and former smoking were also higher among those reporting greater levels of gay community affinity. Finally, the odds of being a former smoker were higher for those reporting internalized antihomosexual prejudice. This study identifies several factors related to smoking status in a diverse sample of young sexual minority males. These findings should encourage investigations of smoking disparities among younger MSM to look beyond common smoking risk factors in an attempt to understand etiologies that may be unique to this group. Such findings may indicate multiple points of potential intervention aimed at decreasing cigarette smoking within this vulnerable population.


Structural and functional evidence for the interaction of insulin-like growth factors (IGFs) and IGF binding proteins with vitronectin.
Previous studies demonstrated that IGF-II binds directly to vitronectin (VN), whereas IGF-I binds poorly. However, binding of VN to integrins has been demonstrated to be essential for a range of IGF-I-stimulated biological effects, including IGF binding protein (IGFBP)-5 production, IGF type-1 receptor autophosphorylation, and cell migration. Thus, we hypothesized that a link between IGF-I and VN must occur and may be mediated through IGFBPs. This was tested using competitive binding assays with VN and (125)iodine-labeled IGFs in the absence and presence of IGFBPs. IGFBP-4, IGFBP-5, and nonglycosylated IGFBP-3 were shown to significantly enhance binding of IGF-I to VN, whereas IGFBP-2 and glycosylated IGFBP-3 had a smaller effect. Furthermore, binding studies with analogs indicate that glycosylation status and the heparin-binding domain of IGFBP-3 are important in this interaction. To examine the functional significance of IGFs binding to VN, cell migration in MCF7 cells was measured and found to be enhanced when VN was prebound to IGF-I in the presence of IGFBP-5. The effect required IGF:IGFBP:VN complex formation; this was demonstrated by use of a non-IGFBP-binding IGF-I analog. Together, these data indicate the importance of IGFBPs in modulating IGF-I binding to VN and that this binding has functional consequences in cells.


Effects of the herbicide imazapyr on juvenile Oregon spotted frogs.
Conflict between native amphibians and aquatic weed management in the Pacific Northwest is rarely recognized because most native stillwater-breeding amphibian species move upland during summer, when herbicide application to control weeds in aquatic habitats typically occurs. However, aquatic weed management may pose a risk for aquatic species present in wetlands through the summer, such as the Oregon spotted frog (OSF, Rana pretiosa), a state endangered species in Washington. Acute toxicity of herbicides used to control aquatic weeds tends to be low, but the direct effects of herbicide tank mixes on OSFs have remained unexamined. We exposed juvenile OSFs to tank mixes of the herbicide imazapyr, a surfactant, and a marker dye in a 96-h static-renewal test. The tank mix was chosen because of its low toxicity to fish and its effectiveness in aquatic weed control. Concentrations were those associated with low-volume (3.5 L/ha) and high-volume (7.0 L/ha) applications of imazapyr and a clean-water control. Following exposure, frogs were reared for two months in clean water to identify potential latent effects on growth. Endpoints evaluated included feeding behavior, growth, and body and liver condition indices. We recorded no mortalities and found no significant differences for any end point between the herbicide-exposed and clean-water control frogs. The results suggest that imazapyr use in wetland restoration poses a low risk of direct toxic effects on juvenile OSFs.


Human immunodeficiency virus, AIDS, and drug consumption in South America and the Caribbean: epidemiological evidence and initiatives to curb the epidemic.
The paper reviews data on drug use in relation to the spread of human immunodeficiency virus and AIDS in South America and the Caribbean. Information was gathered by thoroughly reviewing major bibliographic databanks, web sites of international institutions and regional networks working with substance misuse or human immunodeficiency virus and AIDS, and abstracts from conferences and meetings. Although some gaps remain, a growing body of evidence documents the significant role of injected cocaine in the Brazilian and Southern Cone epidemics. The Caribbean and the Andean areas have thus far been spared in large part from the spread of injection drug use and its consequences, but the situation has been changing in Southern Cone countries towards a higher prevalence of harmful injection habits. Additional challenges have been posed by the increasing availability of heroin in the Andean Area and the abuse of crack cocaine and its impact on the sexual transmission of human immunodeficiency virus in many cities. Harm reduction strategies have been established in most areas of Brazil and are gaining momentum in Argentina. Other countries in the Region still face serious limitations due to restrictive legislation and lack of broader support. Greater participation of Latin American and Caribbean countries in research protocols and continued debate on both successful and failed experiences should be encouraged in order to minimize existing barriers to the full adoption of effective measures to curb the human immunodeficiency virus and AIDS epidemic in this Region.


Wide occurrence of SSU rDNA intragenomic polymorphism in foraminifera and its implications for molecular species identification.
Ribosomal DNA is commonly used as a marker for protist phylogeny and taxonomy because of its ubiquity and its expected species specificity thanks to the mechanism of concerted evolution. However, numerous studies reported the occurrence of intragenomic (intra-individual) polymorphism in various protists and particularly in Foraminifera. To infer to what extent the SSU rDNA intragenomic variability occurs in Foraminifera, we studied 16 foraminiferal species belonging to single-chambered monothalamids and multi-chambered Globothalamea, with one to six individuals per species. We performed single-cell DNA extractions and PCRs of a 600bp fragment of SSU rDNA, and sequenced 9 to 23 clones per individual for a total of 818 sequences. We found intragenomic variability in almost all species, even after excluding singleton mutations. Intra-individual sequence divergence ranged from 0 to 5.15% and was higher than 1% in 11 species. Variability was usually located at the end of stem-loop structures and included compensatory single nucleotide polymorphisms and expansion segments polymorphisms. However, the polymorphisms did not change the secondary structure of the rRNA. Our results suggest a non-concerted evolution of rRNA genes in Foraminifera. The origin of this variability and its implications for species identification in environmental DNA studies are discussed.


Evidence for renal control of urinary excretion of bile acids and bile acid sulphates in the cholestatic syndrome.
1. The bile acids and bile acid sulphates in the urine, serum and bile of eight cholestatic patients were studied quantitatively by gasliquid chromatography and gas-liquid chromoatography/mass spectrometry. 2. The primary bile acids (cholic acid and chenodeoxycholic acid) comprised on average 94% of the total bile acids in bile, 70% in the serum and 64% in urine. 3. The percentage composition of bile acids in bile was relatively constant and was not influenced by the degree of cholestasis. In contrast, in the serum only the primary bile acids were increased, the concentrations of the secondary bile acids (deoxycholic acid and lithocholic acid) and the minor bile acids remaining constant. 4. The data do not support the hypothesis that monohydroxy bile acids accumulate in cholestasis and are related to the pathogenesis of this syndrome. 5. The pattern of bile acid urinary excretion was similar to that in the serum. But in one patient, 3alpha, 7beta, 12alpha-trihydroxy-5beta-cholan-24-oic acid was a principal urinary bile acid, although very low concentrations of the compound were found in that patient's serum, suggesting that some of the minor bile acids in urine may originate by epimerization in the kidney. 6. In bile only a small proportion of the bile acids was sulphated (range 2.1-4.6%) and in serum the degree of sulphation was very variable (9-50%). However, in urine, sulphate esters accounted for a large proportion of the total bile acids (33-72%). 7. The output of bile acid sulphate in the urine was related to the urine total bile acid output but the serum concentration of bile acid sulphate remained relatively constant. Consequently, in contrast to the non-sulphated bile acids, whose renal clearance was relatively constant, the renal clearance of sulphated bile acids was directly related to the urine total bile acid output. This finding is inconsistent with the earlier hypothesis that their predominance in urine was due to a high renal clearance. It may indicate renal synthesis of some of the bile acid sulphates in the urine and/or inhibition of active renal tubular reabsorption of sulphated bile acids by non-sulphated bile acids.


Echocardiographic findings in former professional cyclists after long-term deconditioning of more than 30 years.
In professional cyclists, typical changes include reversible dilatation of atria and left ventricle (LV), LV hypertrophy but normal diastolic function. Data on long-term outcome are limited. Of all 134 former Swiss professional cyclists (PC) participating >or=1x in the professional bicycle race Tour de Suisse from 1955 to 1975, 62 (42%) were recruited for a prospective case control study. The PC and a control group of 62 golfers (matched for age, gender, hypertension, present physical activity) were screened [clinical examination, history, echocardiography, measurement of proBNP (normal <227 pg/mL)]. The interval since the last bicycle race as PC was 38 (15-49) years. Average age at exam was equal in controls and PC (66+/-6 vs 66+/-7 years; P = 0.73). Percentage of participants undergoing >4 h of endurance training per week was identical (P = 0.72). Total kilometers (km) on the bicycle were higher in PCs with 311,000 (60,000-975,000) than in controls (2500 [0-120,000]; P < 0.0001). PC had larger atrial volume indices (P = 0.002) and tended to have higher LV muscle mass indices (P = 0.07). Multiple regression analysis identified the total number of bicycle km as an independent factor for LV muscle mass. For left atrial size, heart rate at rest, age, years since the last bicycle race and the current hours of endurance training were identified as independent predictors. Long axis function of both ventricles (systolic velocities of mitral and tricuspid annulus) was decreased in PC (P <or= 0.04). There were signs of diastolic dysfunction with lower annular E' and A' velocities. ProBNP levels were comparable in both groups (P = 0.21). Among former PC, there seems to be incomplete cardiac remodelling with differences in systolic and diastolic function between former PCs and controls in the long time follow-up. Former high level endurance training may have a persisting impact on cardiac size and function.


Harris lines revisited: prevalence, comorbidities, and possible etiologies.
The occurrence of transverse radiopaque lines in long bones-Harris lines (HLs)-is correlated with episodes of temporary arrest of longitudinal growth and has been used as an indicator of health and nutritional status of modern and historical populations. However, the interpretation of HLs as a stress indicator remains debatable. The aim of this article is to evaluate the perspectives and the limitations of HLs analyses and to examine their reliability as a stress indicator. The study was conducted on 241 tibiae from a medieval Swiss skeletal material and was carried out using a standardized, semiautomated HL detection and analysis tool developed by the authors. We compared four different age-at-formation estimation methods and analyzed the correlation of HL occurrence to life expectancy, mean-age-at-death, stature, tibia length, and metabolic disorders as expressed by linear enamel hypoplasia and hypothyroidism. The evaluation of the age-at-formation estimation methods showed statistical significant differences. Therefore, a mathematical framework for the conversion between the methods has been developed. Remodeling had eliminated about half of the HLs formed during adolescence, and a further half of the remaining ones during early adulthood, whereas no association between the aforementioned conditions and HL prevalence could be determined. The peaks of high HL frequency among various populations were found to parallel normal growth spurts and growth hormone secretion. We suggest a reconsideration of HLs as more of a result of normal growth and growth spurts, rather than a pure outcome of nutritional or pathologic stress.