Global health burden and needs of transgender populations: a review.
Transgender people are a diverse population affected by a range of negative health indicators across high-income, middle-income, and low-income settings. Studies consistently document a high prevalence of adverse health outcomes in this population, including HIV and other sexually transmitted infections, mental health distress, and substance use and abuse. However, many other health areas remain understudied, population-based representative samples and longitudinal studies are few, and routine surveillance efforts for transgender population health are scarce. The absence of survey items with which to identify transgender respondents in general surveys often restricts the availability of data with which to estimate the magnitude of health inequities and characterise the population-level health of transgender people globally. Despite the limitations, there are sufficient data highlighting the unique biological, behavioural, social, and structural contextual factors surrounding health risks and resiliencies for transgender people. To mitigate these risks and foster resilience, a comprehensive approach is needed that includes gender affirmation as a public health framework, improved health systems and access to health care informed by high quality data, and effective partnerships with local transgender communities to ensure responsiveness of and cultural specificity in programming. Consideration of transgender health underscores the need to explicitly consider sex and gender pathways in epidemiological research and public health surveillance more broadly.


In July 2019 the Portuguese Society of Myology was officially born: the first message from the founders.
The amazing behaviour and adaptation capacity of the skeletal muscle system call the attention of several scientists, including us. Thus, we have the pleasure to announce that it was born in the city of Coimbra, Portugal, the Portuguese Society of Myology (SPMyo), a society which the main aim is to spread skeletal muscle knowledge in the areas of muscle physiology, physiopathology and pharmacology in a multidisciplinary organization that reflects the different areas of the skeletal muscle study. The commitment of SPMyo is the progress of skeletal muscle awareness as well as its investigation, encouraging interdisciplinary, national and international collaboration. These goals will be achieved by organization of scientific meetings, formation courses, publications of books and other scientific and didactic materials. It is also a goal of the SPMyo to organize public events allowing general public to understand skeletal muscle system.


Evaluation of formalin-inactivated Clostridium difficile vaccines administered by parenteral and mucosal routes of immunization in hamsters.
Clostridium difficile produces toxins that cause inflammation, necrosis, and fluid in the intestine and is the most important cause of nosocomial antibiotic-associated diarrhea and colitis. We evaluated C. difficile antigens as vaccines to protect against systemic and intestinal disease in a hamster model of clindamycin colitis. Formalin-inactivated culture filtrates from a highly toxigenic strain were administered by mucosal routes (intranasal, intragastric, and rectal) with cholera toxin as a mucosal adjuvant. A preparation of culture filtrate and killed whole cells was also tested rectally. The toxoid was also tested parenterally (subcutaneously and intraperitoneally) and by a combination of three intranasal immunizations followed by a combined intranasal-intraperitoneal boost. Serum antibodies against toxins A and B and whole-cell antigen were measured by enzyme-linked immunosorbent assay, neutralization of cytotoxic activity, and bacterial agglutination. The two rectal immunization regimens induced low antibody responses and protected only 20% of hamsters against death and 0% against diarrhea. The intragastric regimen induced high antibody responses but low protection, 40% against death and 0% against diarrhea. Hamsters immunized by the intranasal, intraperitoneal, and subcutaneous routes were 100% protected against death and partially protected (40, 40, and 20%, respectively) against diarrhea. Among the latter groups, intraperitoneally immunized animals had the highest serum anticytotoxic activity and the highest agglutinating antibody responses. Hamsters immunized intranasally and revaccinated intraperitoneally were 100% protected against both death and diarrhea. Protection against death and diarrhea correlated with antibody responses to all antigens tested. The results indicate that optimal protection against C. difficile disease can be achieved with combined parenteral and mucosal immunization.


Coupling of flagellar gene expression to flagellar assembly in Salmonella enterica serovar typhimurium and Escherichia coli.
How do organisms assess the degree of completion of a large structure, especially an extracellular structure such as a flagellum? Bacteria can do this. Mutants that lack key components needed early in assembly fail to express proteins that would normally be added at later assembly stages. In some cases, the regulatory circuitry is able to sense completion of structures beyond the cell surface, such as completion of the external hook structure. In Salmonella and Escherichia coli, regulation occurs at both transcriptional and posttranscriptional levels. One transcriptional regulatory mechanism involves a regulatory protein, FlgM, that escapes from the cell (and thus can no longer act) through a complete flagellum and is held inside when the structure has not reached a later stage of completion. FlgM prevents late flagellar gene transcription by binding the flagellum-specific transcription factor sigma(28). FlgM is itself regulated in response to the assembly of an incomplete flagellum known as the hook-basal body intermediate structure. Upon completion of the hook-basal body structure, FlgM is exported through this structure out of the cell. Inhibition of sigma(28)-dependent transcription is relieved, and genes required for the later assembly stages are expressed, allowing completion of the flagellar organelle. Distinct posttranscriptional regulatory mechanisms occur in response to assembly of the flagellar type III secretion apparatus and of ring structures in the peptidoglycan and lipopolysaccharide layers. The entire flagellar regulatory pathway is regulated in response to environmental cues. Cell cycle control and flagellar development are codependent. We discuss how all these levels of regulation ensure efficient assembly of the flagellum in response to environmental stimuli.


Human papillomavirus infection of the uterine cervix of women without cytological signs of neoplasia.
One hundred and six patients were studied whose cervical smears showed only non-specific inflammatory changes. Screening for genital pathogens yielded only a few positive cases. Histological examination of biopsy specimens taken by colposcopically directed tissue sampling showed cervical intraepithelial neoplasia in 13 of the women (12.3%). Deoxyribonucleic acid (DNA) hybridisation techniques were used to detect human papillomavirus, which was found in 24 patients (22.6%). In a second group of 104 patients with normal cervical cytology tissue biopsy samples were obtained and examined histologically but in no case was cervical intraepithelial neoplasia found. On DNA hybridisation, however, 12 patients (11.5%) were found to be positive for human papillomavirus. In this group finding human papillomavirus DNA was usually associated with a columnar ectopy. An association between human papillomavirus type 16 DNA and both cervical intraepithelial neoplasia and cervical cancer is well established. In this study it was type 16 which occurred most frequently in both groups.


Dehydroepiandrosterone sulphate reduces chronic hypoxic pulmonary hypertension in rats.
Pathogenesis of pulmonary hypertension includes vascular smooth muscle cell membrane depolarisation and consequent calcium influx. Usually, calcium-gated potassium channels are activated under such conditions and repolarise the membrane. However, in pulmonary hypertension they are downregulated. The authors hypothesised that pharmacological augmentation of these channels would reduce pulmonary hypertension. Dehydroepiandrosterone sulphate (DHEA-S, 0.1 mg x mL(-1)), a recently characterised activator of calcium-gated potassium channels, was given to rats in drinking water. Pulmonary arterial blood pressure, increased by 4 weeks of hypoxia (from 15 +/- 0.2 to 29.4 +/- 2.5 mmHg), was selectively attenuated in rats treated with DHEA-S for the whole duration of the hypoxic exposure (23.9 +/- 0.9 mmHg) and in rats given DHEA-S only after pulmonary hypertension had fully developed (last 2 weeks of hypoxia; 24.4 +/- 1.4 mmHg). Pulmonary vascular remodelling and right ventricular hypertrophy associated with pulmonary hypertension were also reduced by DHEA-S. Cardiac index and systemic arterial blood pressure did not differ among the groups. The authors conclude that treatment with an activator of calcium-gated potassium channels, dehydroepiandrosterone sulphate, known to be well tolerated by humans, reduces hypoxic pulmonary hypertension in rats.


Rethinking the Baseline in Diversity Research: Should We Be Explaining the Effects of Homogeneity?
It is often surprisingly difficult to make definitive scientific statements about the functional value of group diversity. We suggest that one clear pattern in the group diversity literature is the prevailing convention of interpreting outcomes as the effect of diversity alone. Although work in this arena typically compares diverse groups with homogeneous ones, we most often conceive of homogeneous groups as a baseline-a reference point from which we can understand how diversity has changed behavior or what type of response is "normal." In this article, we offer a new perspective through a focus on two propositions. The first proposition is that homogeneity has independent effects of its own-effects that, in some cases, are robust in comparison with the effects of diversity. The second proposition is that even though subjective responses in homogeneous groups are often treated as a neutral indicator of how people would ideally respond in a group setting, evidence suggests that these responses are often less objective or accurate than responses in diverse groups. Overall, we believe that diversity research may unwittingly reveal important insights regarding the effects of homogeneity.


THE INFLUENCE OF THE SUPRARENAL GLAND ON THE THYMUS : I. REGENERATION OF THE THYMUS FOLLOWING DOUBLE SUPRARENALECTOMY IN THE RAT.
Experimental evidence has been presented showing that the thymus gland of the rat enlarges rapidly in those animals surviving double suprarenalectomy. The enlargement has been observed to follow bilateral ablation with such constancy that it may be said to occur in all rats which survive double suprarenalectomy in good condition for from 3 to 5 weeks. The thymus enlarges even though the animal loses some weight after operation. Our data, though inconclusive, would seem to indicate a sex difference, the secondary hyperplasia of the thymus being more marked in the female than in the male. This difference, we believe, may shed some light on the mechanism of thymus enlargement following double suprarenalectomy. In the rat there is a definite sex difference in the weight of the suprarenals (20), the glands being much larger in the female than in the male. This weight difference becomes greater as the rat grows, and it appears at an early period of life; indeed, it is obvious at about 35 days of age, in spite of the fact that sexual maturity is seldom attained before 60 to 90 days. This difference in the suprarenals is independent of pregnancy. It is highly probable that the suprarenal glands stand in closer relation to the sex organs of the females than in the male, and that removal of these glands disrupts the sex interrelationships between gonads, suprarenals, and thymus, this disturbance being more definitely expressed in the female than in the male. The great majority of the rats were operated on when they were between 6 and 7 months of age; that is, after thymus involution would normally have set in. Thymuses examined when they had reached their largest size resembled both grossly and microscopically the growing thymuses of much younger animals, the only evidence of previous involutionary change being the presence of thickening of the vessels. We assume, therefore, from indirect, but statistical data, that regeneration of the involuted thymus must have taken place. The regeneration following bilateral ablation describes a curve which reaches its height between the 3rd and 5th weeks after operation. After the height of enlargement is attained, there is a gradual decline in the size of the gland, but involutionary changes may be retarded for months. Pregnancy prior to suprarenalectomy does not prevent thymus regeneration following this operation. Thymic enlargement does not ordinarily take place following a unilateral ablation. Severe chronic infections may sometimes bring about such a pronounced pathological involution of the thymus that the hyperplasia may be prevented. There can as yet be only speculation as to the mechanism involved in the regeneration of the thymus which follows suprarenalectomy. This secondary thymic hyperplasia may be one manifestation of the generalized lymphoid hyperplasia that follows sublethal but sufficient suprarenal injury, and which is characterized by the appearance of prominent lymphoid foci in the various organs, particularly in the thyroid, a generalized hyperplasia of the lymph nodes and bone marrow, and the appearance of a lymphocytosis in the blood. The evidence at present available would suggest that both the thymic and general lymphoid hyperplasia may be effected through the gonads and parasex tissues and that the thymus especially regenerates because of a disruption of the interrelations between the interrenal gland, the gonads, and the thymus. The close association between the gonads and the suprarenals (21) is well known. The latter undergo enlargement in pregnancy, ovulation, and castration, and the ovaries hypertrophy after suprarenalectomy. Tumors of the suprarenals are associated frequently with sex perversion and precocity both in the male and female. The interrelation between the gonads and thymus is classical, castration before puberty delaying thymus involution for a long time. A number of diseases present thymus enlargement among the major pathological findings at autopsy. The most outstanding of these are Addison's disease, status lymphaticus, and Graves' disease. It is now generally recognized that the suprarenal glands play an important r61e in the etiology both of Addison's disease and status lymphaticus. Whether the interrenal or the chromophil system is primarily involved or whether disturbances in function of both systems are at the basis of these diseases is still undetermined. The newer evidence, including our own work on lymphoid regeneration following suprarenalectomy supports the view that the lymphoid hyperplasia occurring in both Addison's disease and status lymphaticus is dependent upon insufficiency of the interrenal system (suprarenal cortex). While anatomical studies have not disclosed any constant lesions in the suprarenals in Graves' disease, evidence is being accumulated which would indicate that functionally these glands also play an important if not a primary role in the production of the clinical syndrome of Graves' disease (14, 18, 19). We are of the belief that the persistence of the thymus which occurs in status lymphaticus, and the regeneration which occurs in Addison's and Graves' diseases, are brought about by the same disturbances in glandular interrelations which bring about regeneration of the thymus in the experimental animal after suprarenalectomy.


Development of clinician-educator faculty track at the University of Pennsylvania.
As resources grow scarcer , universities have become reluctant to increase the long-term institutional commitments involved in faculty appointments with tenure. The restrict tenured appointments, universities in recent years have adopted more stringent criteria for approving them and in many cases have set a limit on the number of new appointments and promotions with tenure. The educational and service missions of the medical school, however, necessitate a large cadre of clinical faculty members. The establishment and evolution of the clinician-educator faculty track at the University of Pennsylvania provide a case study of a process taking place in many medical schools to meet the need for nontenured full-time faculty appointments.


Ion-specific effects in foams.
We present a critical review on ion-specific effects in foams in the presence of added salts. We show the theoretical basis developed for understanding experimental data in systems with ionic surfactants, as well as the nascent approaches to modeling the much more difficult systems with non-ionic surfactants, starting with the most recent models of the air-water interface. Even in the case of ionic surfactant systems, we show methods for improving the theoretical understanding and apply them for interpretation of surprising experimental results we have obtained on ion-specific effects in these systems. We report unexpectedly strong ion-specific effects of counter-ions on the stability and the rate of drainage of planar foam films from solutions of 0.5mM sodium dodecyl sulfate (SDS) as a function of concentration of a series of inorganic salts (MCl, M=Li, Na, K). We found that the counter-ions can either stabilize the foam films (up to a critical concentration) or destabilize them beyond it. The ordering for destabilization is in the same order as the Hofmeister series, while for stabilization it is the reverse Therefore, the strongest foam stabilizer (K(+)), becomes the strongest foam destabilizer at and beyond its critical concentration, and vice versa. Though the critical concentration is different for different salts, calculating the critical surfactant adsorption level one could simplify the analysis, with all the critical concentrations occurring at the same surfactant adsorption level. Beyond this level, the foam lifetime decreases and films suddenly start draining faster, which may indicate salt-induced surfactant precipitation. Alternatively, formation of pre-micellar structures may result in slower equilibration and fewer surfactant molecules at the surface, thus leading to unstable foams and films.


Nrf2 is essential for cholesterol crystal-induced inflammasome activation and exacerbation of atherosclerosis.
Oxidative stress and inflammation--two components of the natural host response to injury--constitute important etiologic factors in atherogenesis. The pro-inflammatory cytokine interleukin (IL)-1 significantly enhances atherosclerosis, however, the molecular mechanisms of IL-1 induction within the artery wall remain poorly understood. Here we have identified the oxidative stress-responsive transcription factor NF-E2-related 2 (Nrf2) as an essential positive regulator of inflammasome activation and IL-1-mediated vascular inflammation. We show that cholesterol crystals, which accumulate in atherosclerotic plaques, represent an endogenous danger signal that activates Nrf2 and the NLRP3 inflammasome. The resulting vigorous IL-1 response critically depended on expression of Nrf2, and Nrf2-deficient apolipoprotein E (Apoe)-/- mice were highly protected against diet-induced atherogenesis. Importantly, therapeutic neutralization of IL-1α and IL-1β reduced atherosclerosis in Nrf2+/- Apoe-/- but not in Nrf2-/- Apoe-/- mice, suggesting that the pro-atherogenic effect of Nrf2-signaling was primarily mediated by its permissive role in IL-1 production. Our studies demonstrate a role for Nrf2 in inflammasome activation, and identify cholesterol crystals as disease-relevant triggers of the NLRP3 inflammasome and potent pro-atherogenic cytokine responses. These findings suggest a common pathway through which oxidative stress and metabolic danger signals converge and mutually perpetuate the chronic vascular inflammation that drives atherosclerosis.


Daily inhalation of hydrogen gas has a blood pressure-lowering effect in a rat model of hypertension.
A recent clinical study demonstrated that haemodialysis with a dialysate containing hydrogen (H2) improves blood pressure control in end-stage kidney disease. Herein, we examined whether H2 has a salutary effect on hypertension in animal models. We subjected 5/6 nephrectomised rats to inhalation of either H2 (1.3% H2 + 21% O2 + 77.7% N2) or control (21% O2 + 79% N2) gas mixture for 1 h per day. H2 significantly suppressed increases in blood pressure after 5/6 nephrectomy. The anti-hypertensive effect of H2 was also confirmed in rats in a stable hypertensive state 3 weeks after nephrectomy. To examine the detailed effects of H2 on hypertension, we used an implanted telemetry system to continuously monitor blood pressure. H2 exerted an anti-hypertensive effect not only during daytime rest, but also during night-time activities. Spectral analysis of blood pressure variability revealed that H2 improved autonomic imbalance, namely by suppressing the overly active sympathetic nervous system and augmenting parasympathetic nervous system activity; these effects co-occurred with the blood pressure-lowering effect. In conclusion, 1-h daily exposure to H2 exerts an anti-hypertensive effect in an animal model of hypertension.


Nonreplicating protocells.
Prebiotic soup experiments have shown that the molecular building blocks of life can be built under prebiotically plausible conditions. From this starting point, researchers have launched continued studies of polymerization and explorations of the breadth of RNA function. Recently, effort has intensified to examine experimentally another stage of the origins of life: the assembly of the molecular parts into model protocells intended to represent the first primitive, cell-like systems to emerge on Earth. Although it may not be possible to recreate the precise sequence of events that led to cellular life, laboratory experiments have begun to show what was and was not possible. Prebiotically plausible lipid vesicles form easily and have many properties that are conducive to cellular function. In addition to protecting nascent replicating genetic systems from parasitic sequences, vesicles facilitate evolution. The data thus far suggest that prebiotically plausible vesicles could have grown, divided, and promoted competition between distinct chemical systems. Most protocellular studies to date have probed the role of self-replication, one feature of extant life in the emergence of the first cellular system. Undoubtedly replicating systems were crucial for protocellular evolution, but other features of life must have been important as well. For example, life does not exist in isolation. A living system must cope with and adapt to environmental fluctuations to survive. The protocell must have generated some of these fluctuations because cellular activity necessarily modifies its surroundings by selectively absorbing nutrients and releasing unwanted molecules. It seems likely that life would have faced this challenge early and either emerged in dynamic locales that continuously regenerated conditions conducive to life or exploited mechanisms to physically move to new areas not depleted in resources. Further studies that explore non-replication-based aspects of the origins of life could reveal a more complete picture of the transition from prebiotic chemistry to early life.


Prolactin and breast increase during pregnancy in PCOS: linked to long-term metabolic health?
Objective To explore whether gestational prolactin and breast increase are markers of metabolic health in pregnancy and on long-term, in PCOS. Design Follow-up study. Women with PCOS, according to the Rotterdam criteria (n = 239), former participants of the randomized controlled trial (RCT) PregMet were invited, 131 participated in the current follow-up study, at mean 8 years after pregnancy. Methods Metformin 2000 mg/day or placebo from first trimester to delivery in the original RCT. No intervention in the current study. Prolactin was analyzed in the first trimester and at gestational week 32 and metabolic characteristics which are part of the metabolic syndrome and measures of glucose homeostasis were examined. Metabolic health was also evaluated according to breast increase versus lack of breast increase during pregnancy. Results Prolactin increase in pregnancy was negatively correlated to BMI (P = 0.007) and systolic blood pressure (P ≤ 0.001) in gestational week 32. Prolactin at gestational week 32 was negatively correlated to BMI (P = 0.044) and visceral fat area (P = 0.028) at 8-year follow-up in an unadjusted model. Prolactin at gestational week 32 showed no associations to metabolic health at follow-up when baseline BMI was adjusted for. Women who reported lack of breast increase during pregnancy, had higher BMI (P = 0.034), waist-hip ratio (P = 0.004), visceral fat area (P = 0.050), total cholesterol (P = 0.022), systolic (P = 0.027) and diastolic blood pressure (P = 0.011) at 8-year follow-up. Conclusion High prolactin levels and breast increase in pregnancy were associated with a more favorable long-term metabolic health in women with PCOS. Both prolactin and breast increase may be mediated by gestational BMI.


Identification Problems in Personality Psychology.
This paper discusses and illustrates identification problems in personality psychology. The measures used by psychologists to infer traits are based on behaviors, broadly defined. These behaviors are produced from multiple traits interacting with incentives in situations. In general, measures are determined by these multiple traits and do not identify any particular trait unless incentives and other traits are controlled for. Using two data sets, we show, as an example, that substantial portions of the variance in achievement test scores and grades, which are often used as measures of cognition, are explained by personality variables.


Identification and characterization of a Smad2 homologue from Schistosoma mansoni, a transforming growth factor-beta signal transducer.
Smad proteins are essential intracellular signal transducers of the transforming growth factor-beta (TGF-beta) superfamily. The TGF-beta superfamily signals through phosphorylation and activation of R-Smad proteins, receptor-regulated Smads, by heteromeric complexes of ligand-specific type I and type II serine/threonine kinase receptors. R-Smads receive a signal from the activated receptor complex and transmit it to the nucleus. A cDNA was isolated that encodes a 649-amino acid protein found to be homologous to members of R-Smad subfamily with highest homology scored to clawed African frog and human Smad2. The Schistosoma mansoni homologue (SmSmad2) was overexpressed in bacteria as a Sj26-GST fusion protein and used to raise specific antibodies. The IgG fraction of the immunized rabbit serum identified 70- and 72-kDa protein bands in Western analysis of schistosome extracts. Treatment with alkaline phosphatase removed the 72-kDa band, which indicates that this band represents the phosphorylated form of schistosome Smad2. SmSmad2 was localized in the subtegument, parenchymal cells, and sex organs in both male and female worm cryosections. Similar results were also obtained from the analysis of the Smad2 mRNA distribution pattern revealed by in situ hybridization of adult worm pair paraffin sections. SmSmad2 mRNA levels were determined by reverse transcriptase-polymerase chain reaction in different mammalian host developmental stages and found to be constitutively expressed. SmSmad2 was also found to interact with a previously identified SmTbetaR-I, a serine/threonine type I kinase receptor. Furthermore, SmSmad2 was shown to undergo phosphorylation by constitutively active forms of SmTbetaR-I in vitro. In addition, SmSmad2 localized in the nuclei of mink lung epithelial cells upon treatment with TGF-beta(1). These data indicate that the SmSmad2 responds to the TGF-beta signals by interaction with receptor I, which phosphorylates it, whereupon it translocates into the nucleus presumably to regulate target gene transcription and consequently elicit a specific TGF-beta effect.


Social media and Dentistry: ethical and legal aspects.
In a saturated market with an over-supply of undergraduate and graduate programs, social media have become attractive means of advertising in Dentistry. However, posts frequently contain ethical violations and lead to service commodification, and their contents are often in disagreement with the Code of Consumer Protection. This article, which focuses on ethical and legal developments, contributes to the discussion and elucidation of questions associated with advertising that uses clinical images and photographs of patients in social media for commercial purposes and self-promotion. Social media and networks are valuable tools of dissemination and exchange of information because of their flexibility, democratic character and low cost. However, their abusive or misleading use, as well as the ethical and legal violations associated with the difficulty in controlling their use, may lead to serious damages and unfavorable court decisions.


A leaky voltage sensor domain of cardiac sodium channels causes arrhythmias associated with dilated cardiomyopathy.
Dilated cardiomyopathy (DCM) is a structural heart disease that causes dilatation of cardiac chambers and impairs cardiac contractility. The SCN5A gene encodes Nav1.5, the predominant cardiac sodium channel alpha subunit. SCN5A mutations have been identified in patients with arrhythmic disorders associated with DCM. The characterization of Nav1.5 mutations located in the voltage sensor domain (VSD) and associated with DCM revealed divergent biophysical defects that do not fully explain the pathologies observed in these patients. The purpose of this study was to characterize the pathological consequences of a gating pore in the heart arising from the Nav1.5/R219H mutation in a patient with complex cardiac arrhythmias and DCM. We report its properties using cardiomyocytes derived from patient-specific human induced pluripotent stem cells. We showed that this mutation generates a proton leak (called gating pore current). We also described disrupted ionic homeostasis, altered cellular morphology, electrical properties, and contractile function, most probably linked to the proton leak. We thus propose a novel link between SCN5A mutation and the complex pathogenesis of cardiac arrhythmias and DCM. Furthermore, we suggest that leaky channels would constitute a common pathological mechanism underlying several neuronal, neuromuscular, and cardiac pathologies.


High-magnification vascular imaging to reject false-positive sites in situ during Hexvix® fluorescence cystoscopy.
Fluorescence imaging for detection of non-muscle-invasive bladder cancer is based on the selective production and accumulation of fluorescing porphyrins-mainly, protoporphyrin IX-in cancerous tissues after the instillation of Hexvix®. Although the sensitivity of this procedure is very good, its specificity is somewhat limited due to fluorescence false-positive sites. Consequently, magnification cystoscopy has been investigated in order to discriminate false from true fluorescence positive findings. Both white-light and fluorescence modes are possible with the magnification cystoscope, allowing observation of the bladder wall with magnification ranging between 30× for standard observation and 650×. The optical zooming setup allows adjusting the magnification continuously in situ. In the high-magnification (HM) regime, the smallest diameter of the field of view is 600 microns and the resolution is 2.5 microns when in contact with the bladder wall. With this cystoscope, we characterized the superficial vascularization of the fluorescing sites in order to discriminate cancerous from noncancerous tissues. This procedure allowed us to establish a classification based on observed vascular patterns. Seventy-two patients subject to Hexvix® fluorescence cystoscopy were included in the study. Comparison of HM cystoscopy classification with histopathology results confirmed 32∕33 (97%) cancerous biopsies and rejected 17∕20 (85%) noncancerous lesions.


Hypertension-induced cardiac hypertrophy. Oxygen supply and consumption with pacing.
The purpose of this study was to determine if hypertrophied myocardium was associated with diminished cardiac function, restricted oxygen supply, or oxygen consumption during tachycardia. Myocardial oxygen supply and oxygen consumption were determined during baseline and atrial pacing conditions 30 days after New Zealand White rabbits were prepared as one-kidney, one clip Goldblatt hypertensive or uninephrectomized sham control rabbits. Coronary blood flow and cardiac output, using radioactive microspheres, and small vessel oxygen saturations, using microspectrophotometry, were measured in hypertrophied and nonhypertrophied hearts. After 30 days, baseline blood pressure was significantly higher in the Goldblatt rabbits compared with sham controls, and hypertension was maintained during pacing. The myocardium was hypertrophied in the Goldblatt hypertensive rabbits compared with sham controls. Baseline heart rates were not different between animal groups (242 +/- 32 and 244 +/- 24 beats/min, respectively). Both groups were paced 35% above baseline heart rates; during pacing, cardiac output was similar to baseline values in the sham controls (304 +/- 99 versus 321 +/- 116 ml/min, respectively) but reduced in the hypertensive rabbits (248 +/- 43 versus 325 +/- 62 ml/min). Myocardial oxygen consumption increased twofold in both nonhypertrophied and hypertrophied ventricles during tachycardia. Oxygen extraction was significantly elevated, but coronary blood flow was not altered during pacing in either animal group. Therefore, at the pacing level chosen the diminished function in cardiac hypertrophy was not associated with reduced oxygen consumption. Conversely, reduced efficiency during pacing in the hypertrophied myocardium was suggested.


A comparative study of standard vs. high definition colonoscopy for adenoma and hyperplastic polyp detection with optimized withdrawal technique.
Colonoscopy has a known miss rate for polyps and adenomas. High definition (HD) colonoscopes may allow detection of subtle mucosal change, potentially aiding detection of adenomas and hyperplastic polyps. To compare detection rates between HD and standard definition (SD) colonoscopy. Prospective, cohort study with optimized withdrawal technique (withdrawal time >6 min, antispasmodic, position changes, re-examining flexures and folds). One hundred and thirty patients attending for routine colonoscopy were examined with either SD (n = 72) or HD (n = 58) colonoscopes. Groups were well matched. Sixty per cent of patients had at least one adenoma detected with SD vs. 71% with HD, P = 0.20, relative risk (benefit) 1.32 (95% CI 0.85-2.04). Eighty-eight adenomas (mean +/- standard deviation 1.2 +/- 1.4) were detected using SD vs. 93 (1.6 +/- 1.5) with HD, P = 0.12; however more nonflat, diminutive (<6 mm) adenomas were detected with HD, P = 0.03. Twenty-three proximal hyperplastic polyps (0.32 +/- 0.58) were detected with SD vs. 31 (0.53 +/- 0.86) with HD, P = 0.35. Overall prevalence of proximal large (>9 mm) hyperplastic polyps was 7% (0.09 +/- 0.36). High definition did not lead to a significant increase in adenoma or hyperplastic polyp detection, but may help where comprehensive lesion detection is paramount. High detection rates appear possible with either SD or HD, when using an optimized withdrawal technique.


Docosahexaenoic acid supplementation and cognitive decline in Alzheimer disease: a randomized trial.
Docosahexaenoic acid (DHA) is the most abundant long-chain polyunsaturated fatty acid in the brain. Epidemiological studies suggest that consumption of DHA is associated with a reduced incidence of Alzheimer disease. Animal studies demonstrate that oral intake of DHA reduces Alzheimer-like brain pathology. To determine if supplementation with DHA slows cognitive and functional decline in individuals with Alzheimer disease. A randomized, double-blind, placebo-controlled trial of DHA supplementation in individuals with mild to moderate Alzheimer disease (Mini-Mental State Examination scores, 14-26) was conducted between November 2007 and May 2009 at 51 US clinical research sites of the Alzheimer's Disease Cooperative Study. Participants were randomly assigned to algal DHA at a dose of 2 g/d or to identical placebo (60% were assigned to DHA and 40% were assigned to placebo). Duration of treatment was 18 months. Change in the cognitive subscale of the Alzheimer's Disease Assessment Scale (ADAS-cog) and change in the Clinical Dementia Rating (CDR) sum of boxes. Rate of brain atrophy was also determined by volumetric magnetic resonance imaging in a subsample of participants (n = 102). A total of 402 individuals were randomized and a total of 295 participants completed the trial while taking study medication (DHA: 171; placebo: 124). Supplementation with DHA had no beneficial effect on rate of change on ADAS-cog score, which increased by a mean of 7.98 points (95% confidence interval [CI], 6.51-9.45 points) for the DHA group during 18 months vs 8.27 points (95% CI, 6.72-9.82 points) for the placebo group (linear mixed-effects model: P = .41). The CDR sum of boxes score increased by 2.87 points (95% CI, 2.44-3.30 points) for the DHA group during 18 months compared with 2.93 points (95% CI, 2.44-3.42 points) for the placebo group (linear mixed-effects model: P = .68). In the subpopulation of participants (DHA: 53; placebo: 49), the rate of brain atrophy was not affected by treatment with DHA. Individuals in the DHA group had a mean decline in total brain volume of 24.7 cm(3) (95% CI, 21.4-28.0 cm(3)) during 18 months and a 1.32% (95% CI, 1.14%-1.50%) volume decline per year compared with 24.0 cm(3) (95% CI, 20-28 cm(3)) for the placebo group during 18 months and a 1.29% (95% CI, 1.07%-1.51%) volume decline per year (P = .79). Supplementation with DHA compared with placebo did not slow the rate of cognitive and functional decline in patients with mild to moderate Alzheimer disease. clinicaltrials.gov Identifier: NCT00440050.


Cellular and synaptic mechanisms of anti-NMDA receptor encephalitis.
We recently described a severe, potentially lethal, but treatment-responsive encephalitis that associates with autoantibodies to the NMDA receptor (NMDAR) and results in behavioral symptoms similar to those obtained with models of genetic or pharmacologic attenuation of NMDAR function. Here, we demonstrate that patients' NMDAR antibodies cause a selective and reversible decrease in NMDAR surface density and synaptic localization that correlates with patients' antibody titers. The mechanism of this decrease is selective antibody-mediated capping and internalization of surface NMDARs, as Fab fragments prepared from patients' antibodies did not decrease surface receptor density, but subsequent cross-linking with anti-Fab antibodies recapitulated the decrease caused by intact patient NMDAR antibodies. Moreover, whole-cell patch-clamp recordings of miniature EPSCs in cultured rat hippocampal neurons showed that patients' antibodies specifically decreased synaptic NMDAR-mediated currents, without affecting AMPA receptor-mediated currents. In contrast to these profound effects on NMDARs, patients' antibodies did not alter the localization or expression of other glutamate receptors or synaptic proteins, number of synapses, dendritic spines, dendritic complexity, or cell survival. In addition, NMDAR density was dramatically reduced in the hippocampus of female Lewis rats infused with patients' antibodies, similar to the decrease observed in the hippocampus of autopsied patients. These studies establish the cellular mechanisms through which antibodies of patients with anti-NMDAR encephalitis cause a specific, titer-dependent, and reversible loss of NMDARs. The loss of this subtype of glutamate receptors eliminates NMDAR-mediated synaptic function, resulting in the learning, memory, and other behavioral deficits observed in patients with anti-NMDAR encephalitis.


Characterizing the encapsulation and release of lentivectors and adeno-associated vectors from degradable alginate hydrogels.
Gene therapy using viral vectors has been licensed for clinical use both in the European Union and the United States. Lentivectors (LV) and adeno-associated vectors (AAV) are two promising and FDA approved gene-therapy viral vectors. Many future applications of these vectors will benefit from targeting specific regions of interest within the body. Therefore, building on the early success of these vectors may depend on finding effective delivery systems to localize therapeutic administration. Degradable alginate hydrogels have been tested as appealing delivery vehicles for the controlled delivery of vector payloads. In this study, we compare the ability of two different degradable alginate hydrogel formulations to efficiently deliver LV and AAV. We propose that release rates of viral vectors are dependent on the physical properties of both the hydrogels and vectors. Here, we demonstrate that the initial strength and degradation rate of alginate hydrogels provides levers of control for tuning LV release but do not provide control in the release of AAV. While both alginate formulations used showed sustained release of both LV and AAV, LV release was shown to be dependent on alginate hydrogel degradation, while AAV release was largely governed by diffusive mechanisms. Altogether, this study demonstrates alginate's use as a possible delivery platform for LV and, for the first time, AAV - highlighting the potential of injectable degradable alginate hydrogels to be used as a versatile delivery tool in gene therapy applications.


Anatomical and functional results after arthroscopic Hill-Sachs remplissage.
Large osseous defects of the posterosuperior aspect of the humeral head can engage the glenoid rim and cause recurrent instability after arthroscopic Bankart repair for glenohumeral dislocation. Filling of the humeral head defect with the posterior aspect of the capsule and the infraspinatus tendon (i.e., Hill-Sachs remplissage) has recently been proposed as an additional arthroscopic procedure. Our hypothesis is that the capsulotenodesis heals in the humeral bone defect without a severe adverse effect on shoulder mobility, allowing return to preinjury sports activity. Of 459 patients operated on for recurrent traumatic anterior shoulder instability, forty-seven (10.2%) underwent arthroscopic Bankart repair combined with Hill-Sachs remplissage with use of suture anchors. All had a large Hill-Sachs lesion (Calandra grade III), engaging over the glenoid rim, without substantial glenoid bone loss. Nine patients had had prior unsuccessful surgery to address glenohumeral instability (three Bankart and six Bristow-Latarjet procedures). The average age at the time of surgery (and standard deviation) was 29 ± 5.4 years. Postoperatively, comparative shoulder motion was precisely measured with use of digital photographic images. Capsulotenodesis healing was assessed on a computed tomography (CT) arthrogram (n = 38) or magnetic resonance image (MRI) (n = 4). The mean duration of follow-up was twenty-four months. Healing of the posterior aspect of the capsule and the infraspinatus tendon into the humeral defect was observed in all forty-two patients who underwent postoperative imaging, and thirty-one (74%) had a remplissage of ≥75%. Compared with the normal (contralateral) side, the mean deficit in external rotation was 8° ± 7° with the arm at the side of the trunk and 9° ± 7° in abduction at the time of the last follow-up. Of forty-one patients involved in sports, thirty-seven (90%) were able to return postoperatively and twenty-eight (68%) returned to the same level of sports, including those involving overhead activities. Ninety-eight percent (forty-six) of the forty-seven patients had a stable shoulder at the time of the last follow-up. Arthroscopic Hill-Sachs remplissage, performed in combination with a Bankart repair, is a potential solution for patients with a large engaging humeral head bone defect but no substantial glenoid bone loss. The posterior capsulotenodesis heals predictably in the humeral defect. The slight restriction in external rotation (approximately 10°) does not significantly affect return to sports, including those involving overhead activities. The procedure, which may also be useful for revision of previous failed glenohumeral instability surgery, is not indicated for patients with glenoid bone deficiency.


Highly efficient upconversion in Er³⁺ doped BaY₂F₈ single crystals: dependence of quantum yield on excitation wavelength and thickness.
This manuscript presents a study of the upconversion (UC) in barium yttrium fluoride (BaY2F8) single crystal doped with trivalent erbium ions (Er3+) under excitation of the 4I(13/2) level at three different wavelengths: 1493 nm, 1524 nm and 1556 nm. The resulting UC emission at around 980 nm has been investigated and it has been found that a thickness optimization is required to reach high quantum yield values, otherwise limited by self-absorption losses. The highest external photoluminescence quantum yield (ePLQY) measured in this study was 12.1±1.2 % for a BaY2F8:30at%Er3+ sample of thickness 1.75±0.01 mm, while the highest internal photoluminescence quantum yield (iPLQY) of 14.6±1.5 % was measured in a BaY2F8:20at%Er3+ sample with a thickness of 0.49±0.01 mm. Both values were obtained under excitation at 1493 nm and an irradiance of 7.0±0.7 Wcm(-2). The reported iPLQY and ePLQY values are among the highest achieved for monochromatic excitation. Finally, the losses due to self-absorption were estimated in order to evaluate the maximum iPLQY achievable by the upconverter material. The estimated iPLQY limit values were ∼19%, ∼25% and ∼30%, for 10%, 20% and 30% Er3+ doping level, respectively.


A2E, a pigment of the lipofuscin of retinal pigment epithelial cells, is an endogenous ligand for retinoic acid receptor.
Lipofuscin contains fluorophores, which represent a biomarker for cellular aging. Although it remains unsubstantiated clinically, experimental results support that the accumulation of lipofuscin is related to an increased risk of choroidal neovascularization due to age-related macular degeneration, a leading cause of legal blindness. Here, we report that a major lipofuscin component, A2E, activates the retinoic acid receptor (RAR). In vitro experiments using luciferase reporter assay, competitional binding assay, analysis of target genes, and chromatin immunoprecipitation (ChIP) assay strongly suggest that A2E is a bona fide ligand for RAR and induces sustained activation of RAR target genes. A2E-induced vascular endothelial growth factor (VEGF) expression in a human retinal pigment epithelial cell line (ARPE-19) and RAR antagonist blocked the up-regulation of VEGF. The conditioned medium of A2E-treated ARPE-19 cells induced tube formation in human umbilical vascular endothelial cells, which was blocked by the RAR antagonist and anti-VEGF antibody. These results suggest that A2E accumulation results in the phenotypic alteration of retinal pigment epithelial cells, predisposing the environment to choroidal neovascularization development. This is mediated through the agonistic function of A2E, at least in part. The results of this study provide a novel potential therapeutic target for this incurable condition.


Genetic engineering activates biosynthesis of aromatic fumaric acid amides in the human pathogen Aspergillus fumigatus.
The Aspergillus fumigatus nonribosomal peptide synthetase FtpA is among the few of this species whose natural product has remained unknown. Both FtpA adenylation domains were characterized in vitro. Fumaric acid was identified as preferred substrate of the first and both l-tyrosine and l-phenylalanine as preferred substrates of the second adenylation domain. Genetically engineered A. fumigatus strains expressed either ftpA or the regulator gene ftpR, encoded in the same cluster of genes, under the control of the doxycycline-inducible tetracycline-induced transcriptional activation (tet-on) cassette. These strains produced fumaryl-l-tyrosine and fumaryl-l-phenylalanine which were identified by liquid chromatography and high-resolution mass spectrometry. Modeling of the first adenylation domain in silico provided insight into the structural requirements to bind fumaric acid as peptide synthetase substrate. This work adds aromatic fumaric acid amides to the secondary metabolome of the important human pathogen A. fumigatus which was previously not known as a producer of these compounds.


STrategically Acquired Gradient Echo (STAGE) imaging, part III: Technical advances and clinical applications of a rapid multi-contrast multi-parametric brain imaging method.
One major thrust in radiology today is image standardization with a focus on rapidly acquired quantitative multi-contrast information. This is critical for multi-center trials, for the collection of big data and for the use of artificial intelligence in evaluating the data. Strategically acquired gradient echo (STAGE) imaging is one such method that can provide 8 qualitative and 7 quantitative pieces of information in 5 min or less at 3 T. STAGE provides qualitative images in the form of proton density weighted images, T1 weighted images, T2* weighted images and simulated double inversion recovery (DIR) images. STAGE also provides quantitative data in the form of proton spin density, T1, T2* and susceptibility maps as well as segmentation of white matter, gray matter and cerebrospinal fluid. STAGE uses vendors' product gradient echo sequences. It can be applied from 0.35 T to 7 T across all manufacturers producing similar results in contrast and quantification of the data. In this paper, we discuss the strengths and weaknesses of STAGE, demonstrate its contrast-to-noise (CNR) behavior relative to a large clinical data set and introduce a few new image contrasts derived from STAGE, including DIR images and a new concept referred to as true susceptibility weighted imaging (tSWI) linked to fluid attenuated inversion recovery (FLAIR) or tSWI-FLAIR for the evaluation of multiple sclerosis lesions. The robustness of STAGE T1 mapping was tested using the NIST/NIH phantom, while the reproducibility was tested by scanning a given individual ten times in one session and the same subject scanned once a week over a 12-week period. Assessment of the CNR for the enhanced T1W image (T1WE) showed a significantly better contrast between gray matter and white matter than conventional T1W images in both patients with Parkinson's disease and healthy controls. We also present some clinical cases using STAGE imaging in patients with stroke, metastasis, multiple sclerosis and a fetus with ventriculomegaly. Overall, STAGE is a comprehensive protocol that provides the clinician with numerous qualitative and quantitative images.


Finding the pitch of the missing fundamental in infants.
Pitch perception is critical for the perception of speech and music, for object identification, and for auditory scene analysis, whereby representations are derived for each sounding object in the environment from the complex sound wave that reaches the ears. The perceived pitch of a complex sound corresponds to its fundamental frequency. However, removal of energy at the fundamental does not alter the pitch because adults use the harmonics to derive the pitch (Bendor and Wang, 2005; Trainor, 2008). Although sound frequency is represented subcortically, the integration of harmonics into a representation of pitch does not occur until auditory cortex (Bendor and Wang, 2005). Given that auditory cortex is immature in young infants, we examined the development of cortical representations for pitch by measuring electrophysiological (EEG) responses to pitch changes that required processing the pitch of the missing fundamental. Adults and infants 4 months and older showed a mismatch negativity response to these pitch changes, but 3-month-old infants did not. Thus, cortical representations of the pitch of the missing fundamental emerge between 3 and 4 months of age, indicating that there is a profound change in auditory perception for pitch in early infancy.


Impacts of CD44 knockdown in cancer cells on tumor and host metabolic systems revealed by quantitative imaging mass spectrometry.
CD44 expressed in cancer cells was shown to stabilize cystine transporter (xCT) that uptakes cystine and excretes glutamate to supply cysteine as a substrate for reduced glutathione (GSH) for survival. While targeting CD44 serves as a potentially therapeutic stratagem to attack cancer growth and chemoresistance, the impact of CD44 targeting in cancer cells on metabolic systems of tumors and host tissues in vivo remains to be fully determined. This study aimed to reveal effects of CD44 silencing on alterations in energy metabolism and sulfur-containing metabolites in vitro and in vivo using capillary electrophoresis-mass spectrometry and quantitative imaging mass spectrometry (Q-IMS), respectively. In an experimental model of xenograft transplantation of human colon cancer HCT116 cells in superimmunodeficient NOG mice, snap-frozen liver tissues containing metastatic tumors were examined by Q-IMS. As reported previously, short hairpin CD44 RNA interference (shCD44) in cancer cells caused significant regression of tumor growth in the host liver. Under these circumstances, the CD44 knockdown suppressed polyamines, GSH and energy charges not only in metastatic tumors but also in the host liver. In culture, HCT116 cells treated with shCD44 decreased total amounts of methionine-pool metabolites including spermidine and spermine, and reactive cysteine persulfides, suggesting roles of these metabolites for cancer growth. Collectively, these results suggest that CD44 expressed in cancer accounts for a key regulator of metabolic interplay between tumor and the host tissue.


Quantification of syntrophic fatty acid-beta-oxidizing bacteria in a mesophilic biogas reactor by oligonucleotide probe hybridization.
Small-subunit rRNA sequences were obtained for two saturated fatty acid-beta-oxidizing syntrophic bacteria, Syntrophomonas sapovorans and Syntrophomonas wolfei LYB, and sequence analysis confirmed their classification as members of the family Syntrophomonadaceae. S. wolfei LYB was closely related to S. wolfei subsp. wolfei, but S. sapovorans did not cluster with the other members of the genus Syntrophomonas. Five oligonucleotide probes targeting the small-subunit rRNA of different groups within the family Syntrophomonadaceae, which contains all currently known saturated fatty acid-beta-oxidizing syntrophic bacteria, were developed and characterized. The probes were designed to be specific at the family, genus, and species levels and were characterized by temperature-of-dissociation and specificity studies. To demonstrate the usefulness of the probes for the detection and quantification of saturated fatty acid-beta-oxidizing syntrophic bacteria in methanogenic environments, the microbial community structure of a sample from a full-scale biogas plant was determined. Hybridization results with probes for syntrophic bacteria and methanogens were compared to specific methanogenic activities and microbial numbers determined with most-probable-number estimates. Most of the methanogenic rRNA was comprised of Methanomicrobiales rRNA, suggesting that members of this order served as the main hydrogen-utilizing microorganisms. Between 0.2 and 1% of the rRNA was attributed to the Syntrophomonadaceae, of which the majority was accounted for by the genus Syntrophomonas.


Genetic and environmental factors in head and neck cancer genesis.
Head and neck squamous cell carcinoma (HNSCC) include squamous cell carcinomas of the oral cavity, pharynx, and larynx. Epidemiologic data suggest that the etiology and pathogenesis of HNSCC are influenced by environmental and lifestyle-related factors, such as tobacco use, ethanol consumption, papilloma virus infection, dietary factors and exposure to toxic substances. DNA repair systems and carcinogen-metabolizing enzymes can increase the risk for HNSCC but no definite causal mechanism has been demonstrated. There are several well-characterized entities that are associated with risk and prognosis of head and neck cancer, including Lynch-II syndrome, Bloom syndrome, Fanconi's anemia, xeroderma pigmentosum, ataxia telangiectasia, and Li-Fraumeni syndrome. This review aims to present the current status in our understanding of HNSCC and highlight controversies relating to the role of several factors in the genesis of the cancer.


Fas-activated serine/threonine kinase (FAST) phosphorylates TIA-1 during Fas-mediated apoptosis.
We have identified a serine/threonine kinase that is rapidly activated during Fas-mediated apoptosis. Fas-activated serine/threonine kinase (FAST) is phosphorylated on serine and threonine residues in Jurkat cells. In response to Fas ligation, it is rapidly dephosphorylated and concomitantly activated to phosphorylate TIA-1, a nuclear RNA-binding protein that has been implicated as an effector of apoptosis. Phosphorylation of TIA-1 precedes the onset of DNA fragmentation, suggesting a role in signaling downstream events in the apoptotic program. Our results introduce Fast and TIA-1 as components of a molecular cascade involved in signaling Fas-mediated apoptosis.


Spanish menstrual literacy and experiences of menstruation.
There is growing recognition of the important role menstrual health plays in achieving health, education, and gender equity. Yet, stigmatisation and taboo remain present and negative emotions like fear and shame dominate the narrative when speaking about periods. This paper analyses how formal and informal menstrual education is received in Spain, to understand the role of menstrual health literacy in the way menstruation is experienced, and to identify what information would be useful to integrate into formal menstrual education. An online survey with more than 4000 participants (aged between 14 and 80, both people who will/do/have previously menstruate/d and those who do not menstruate) was conducted. Data was gathered using the digital platform Typeform, descriptive and inferential statistical analyses were performed with SPSS software and qualitative data was thematically analysed using Nvivo. Many participants declared not having received sufficient information on menstruation prior to menarche, particularly about how to physically manage it. Furthermore, negative emotions like shame, worry, and fear were recurrently reported to describe menarche; this has not changed between generations. Interestingly, we saw an increase in stress and sadness with an increase in perceived knowledge of the reproductive role of menstruation. We did observe a reduction in negative emotions when people who menstruate perceived they had sufficient information on how to manage their first bleeding. It is recommended that menstrual education beyond reproductive biology, particularly including how to physically manage periods, is integrated into school curricula. Menstrual education of everyone - including those who do not menstruate-can improve how periods are experienced in Spain.


Cost effectiveness of patient education for the prevention of falls in hospital: economic evaluation from a randomized controlled trial.
Falls are one of the most frequently occurring adverse events that impact upon the recovery of older hospital inpatients. Falls can threaten both immediate and longer-term health and independence. There is need to identify cost-effective means for preventing falls in hospitals. Hospital-based falls prevention interventions tested in randomized trials have not yet been subjected to economic evaluation. Incremental cost-effectiveness analysis was undertaken from the health service provider perspective, over the period of hospitalization (time horizon) using the Australian Dollar (A$) at 2008 values. Analyses were based on data from a randomized trial among n = 1,206 acute and rehabilitation inpatients. Decision tree modeling with three-way sensitivity analyses were conducted using burden of disease estimates developed from trial data and previous research. The intervention was a multimedia patient education program provided with trained health professional follow-up shown to reduce falls among cognitively intact hospital patients. The short-term cost to a health service of one cognitively intact patient being a faller could be as high as A$14,591 (2008). The education program cost A$526 (2008) to prevent one cognitively intact patient becoming a faller and A$294 (2008) to prevent one fall based on primary trial data. These estimates were unstable due to high variability in the hospital costs accrued by individual patients involved in the trial. There was a 52% probability the complete program was both more effective and less costly (from the health service perspective) than providing usual care alone. Decision tree modeling sensitivity analyses identified that when provided in real life contexts, the program would be both more effective in preventing falls among cognitively intact inpatients and cost saving where the proportion of these patients who would otherwise fall under usual care conditions is at least 4.0%. This economic evaluation was designed to assist health care providers decide in what circumstances this intervention should be provided. If the proportion of cognitively intact patients falling on a ward under usual care conditions is 4% or greater, then provision of the complete program in addition to usual care will likely both prevent falls and reduce costs for a health service. Australia and New Zealand Clinical Trials Register: ACTRN12608000015347.


Occurrence of Flavobacterium psychrophilum in fish-farming environments.
Occurrence of Flavobacterium psychrophilum in fish farms and fish-farming environments was studied using agar plate cultivation, the immunoflourescence antibody technique (IFAT) and nested PCR. Characteristics of 64 F. psychrophilum isolates from rainbow trout Oncorhynchus mykiss, fish farm rearing water, ovarian fluid and wild fish were serotyped, ribotyped and compared biochemically. Virulence of F. psychrophilum isolates from different sources was compared by injection into rainbow trout. Additionally, the number of F. psychrophilum cells shed by naturally infected rainbow trout was determined. F. psychrophilum was detected and isolated from skin mucus, skin lesions and internal organs of diseased rainbow trout and from fish without clinical disease. The pathogen was also present in wild perch Perca fluviatilis, roach Rutilus rutilus, and ovarian fluids of farmed rainbow trout brood fish. Isolates were biochemically homogenous, excluding the capability to degrade elastin. Five different agglutination patterns with different antisera against F. psychrophilum were found among the isolates studied. Although several different ribopatterns were found (ClaI: 12 ribopatterns and HaeIII: 9 ribopatterns), ribotype A was the most dominant. Farmed rainbow trout brood fish carried a broad-spectrum of serologically and genetically different F. psychrophilum in ovarian fluids. Virulence of the tested isolates in rainbow trout varied and naturally infected rainbow trout shed 10(4) to 10(8) cells fish(-1) h(-1) of F. psychrophilum into the surrounding water.


Expectancies and hypnotic responsiveness: an experimental-design flaw revealed.
Recent research suggests that expectancies about being hypnotized have a determinant role in the hypnotic experience. The authors analyzed the relationship between expectancies and the phenomenology of hypnosis using the Phenomenology of Consciousness Inventory and Hypnotic Assessment Procedure. Participants (115) were assigned either to the imagination (hypnosis labeled as imagination) or the hypnosis conditions. Results revealed only a minor influence of expectancies and none on the label "hypnosis" across all variables. These findings indicate that the methodology commonly used to study the influence of expectancies on hypnotic responsiveness and phenomenology might represent a flaw in favor of a causal relationship between expectancies and hypnotic experience.


Synaptic inputs to retrogradely labeled ganglion cells in the retina of the cane toad, Bufo marinus.
The entire population of ganglion cells in the retina of the toad Bufo marinus was labeled by retrograde transport of a lysine-fixable biotinylated dextran amine of 3000 molecular weight. Synaptic connections between bipolar, amacrine, and ganglion cells in the inner plexiform layer were quantitatively analyzed, with emphasis on synaptic inputs to labeled ganglion cell dendrites. Synapses onto ganglion cell dendrites comprised 47% of a total of 1234 identified synapses in the inner plexiform layer. Approximately half of the bipolar or amacrine cell synapses were directed onto ganglion cell dendrites, while the rest were made mainly onto amacrine cell dendrites. Most of the synaptic inputs to ganglion cell dendrites derived from amacrine cell dendrites (84%), with the rest from bipolar cell terminals. Synaptic inputs to ganglion cell dendrites were distributed relatively uniformly throughout all sublaminae of the inner plexiform layer. The present study provides unambiguous identification of ganglion cell dendrites including very fine processes, enabling a detailed analysis of the types and distribution of synaptic inputs from the bipolar and amacrine cell to the ganglion cells. The retrograde tracing technique used in the present study will prove to be a useful tool for identifying synaptic inputs to ganglion cell dendrites from neurochemically identified bipolar and amacrine cell types in the retina.


Systematic review and meta-analysis of interventions tested in animal models of lacunar stroke.
A total of 25% of strokes are lacunar, and these are pathophysiologically different from large artery strokes. Despite emerging evidence of a substantial impact on physical disability and dementia, little attention has been paid to the development of specific treatments. The optimal use of the animal models of lacunar stroke used to test candidate interventions is not known. We conducted a systematic review and meta-analysis of studies testing candidate interventions in animal models of lacunar stroke. We used random-effects meta-analysis to assess the impact of study characteristics and trim and fill to seek evidence of publication bias. The efficacy of 43 distinct interventions was described in 57 publications. The median number of quality checklist items scored was 3 of 8 (interquartile range, 2-4). Many models reflected mechanisms of limited relevance to lacunar stroke. Meta-analysis of results from 27 studies showed that on average, infarct size and neurobehavioral outcome were improved by 34.2% (24.1-44.2) and 0.82 standardized mean difference (0.51-1.14), respectively. Four interventions improved both infarct size and neurobehavioral outcome but there were insufficient data for this finding to be considered robust. For infarct size, efficacy was lower in studies reporting blinding and higher in studies reporting randomization. For neurobehavior, efficacy was lower in randomized studies. For infarct size there was evidence of publication bias. No intervention has yet been tested in sufficient range and depth to support translation to clinical trial. There is limited reporting of measures to reduce the risk of bias and evidence for a substantial publications bias.


Bayesian hierarchical models suggest oldest known plant-visiting bat was omnivorous.
The earliest record of plant visiting in bats dates to the Middle Miocene of La Venta, the world's most diverse tropical palaeocommunity. Palynephyllum antimaster is known from molars that indicate nectarivory. Skull length, an important indicator of key traits such as body size, bite force and trophic specialization, remains unknown. We developed Bayesian models to infer skull length based on dental measurements. These models account for variation within and between species, variation between clades, and phylogenetic error structure. Models relating skull length to trophic level for nectarivorous bats were then used to infer the diet of the fossil. The skull length estimate for Palynephyllum places it among the larger lonchophylline bats. The inferred diet suggests Palynephyllum fed on nectar and insects, similar to its living relatives. Omnivory has persisted since the mid-Miocene. This is the first study to corroborate with fossil data that highly specialized nectarivory in bats requires an omnivorous transition.


Nemo-like kinase suppresses Notch signalling by interfering with formation of the Notch active transcriptional complex.
The Notch signalling pathway has a crucial function in determining cell fates in multiple tissues within metazoan organisms. On binding to ligands, the Notch receptor is cleaved proteolytically and releases its intracellular domain (NotchICD). The NotchICD enters the nucleus and acts cooperatively with other factors to stimulate the transcription of target genes. High levels of Notch-mediated transcriptional activation require the formation of a ternary complex consisting of NotchICD, CSL (CBF-1, suppressor of hairless, LAG-1) and a Mastermind family member. However, it is still not clear how the formation of the ternary complex is regulated. Here we show that Nemo-like kinase (NLK) negatively regulates Notch-dependent transcriptional activation by decreasing the formation of this ternary complex. Using a biochemical screen, we identified Notch as a new substrate of NLK. NLK-phosphorylated Notch1ICD is impaired in its ability to form a transcriptionally active ternary complex. Furthermore, knockdown of NLK leads to hyperactivation of Notch signalling and consequently decreases neurogenesis in zebrafish. Our results both define a new function for NLK and reveal a previously unidentified mode of regulation in the Notch signalling pathway.


Racial/ethnic variation in clinical presentation, treatment, and survival among breast cancer patients under age 35.
The age specific breast cancer incidence rate for African-American women under age 35 is more than twice the rate for white women of similar age, and the mortality rate is more than three times higher. To determine factors that may explain racial/ethnic variation in outcomes among young women diagnosed with breast cancer, the authors examined the clinical presentation, treatment, and survival of African-American, Hispanic, and white women under age 35 years. Surveillance, Epidemiology, and End Results (SEER) Program data for 1990-1998 and SEER Patterns of Care data for 1990, 1991, and 1995 were used for this analysis. Multivariate logistic regression analyses were performed to examine factors associated with the receipt of selected breast cancer treatments. Kaplan-Meier survival analyses and Cox proportional hazards regression analyses were used to examine 5-year overall survival and disease-specific survival. The authors found racial/ethnic variation in clinical presentation, treatment, and survival. Both African-American and Hispanic women presented with higher disease stage and a higher prevalence of adverse prognostic indicators compared to white women. African-American and Hispanic women received cancer-directed surgery and radiation less frequently after undergoing breast-conserving surgery. Racial/ethnic differences in clinical presentation and treatment were associated with poorer overall survival in unadjusted analyses. African-American and Hispanic women also had poorer overall survival after controlling for clinical and demographic characteristics and type of treatment. Future research studies should further examine the factors that influence racial/ethnic differences in incidence, clinical presentation, and treatment differentials among young women diagnosed with breast cancer. A better understanding of these factors will facilitate the development of strategies to help eliminate this health disparity.


The effectiveness of parents in promoting the development of road crossing skills in young children.
Young children show poor judgment when asked to select a safe place to cross the road, frequently considering dangerous sites to be safe. Correspondingly, child pedestrian accidents are over-represented at such locations. Increasing the child's ability to recognise such dangers is a central challenge for road safety education. Practical training methods have proved effective in improving such judgments but are labour-intensive, time-consuming and therefore difficult to implement on a realistic scale. The study examined the possibility that volunteers from the local community might be capable of using such methods to promote children's pedestrian competence. Sixty children from the Primary 1 (Reception) classes of three Glasgow schools took part. Volunteers were ordinary parents from the same areas. None had 'formal' experience of working with children other than through being parents. Volunteers received experience of training children at courses organised in each school. Children learned in small groups, receiving two sessions of roadside training followed by four on a table-top model. Pre- and post-tests allowed the effectiveness of training to be assessed. Significant improvements relative to controls were found in all children following training. Improvements proved robust and no deterioration was observed two months after the programme ended. Comparison with a previous study in which training was undertaken by highly qualified staff showed that the volunteers were as effective as 'expert' trainers. Parent volunteers can significantly increase the pedestrian competence of children as young as five years. They constitute a most valuable 'resource' in road safety education. The opportunities afforded by involving the local community in educational interventions should be further explored.


Cross-species systems analysis of evolutionary toolkits of neurogenomic response to social challenge.
Social challenges like territorial intrusions evoke behavioral responses in widely diverging species. Recent work has showed that evolutionary "toolkits"-genes and modules with lineage-specific variations but deep conservation of function-participate in the behavioral response to social challenge. Here, we develop a multispecies computational-experimental approach to characterize such a toolkit at a systems level. Brain transcriptomic responses to social challenge was probed via RNA-seq profiling in three diverged species-honey bees, mice and three-spined stickleback fish-following a common methodology, allowing fair comparisons across species. Data were collected from multiple brain regions and multiple time points after social challenge exposure, achieving anatomical and temporal resolution substantially greater than previous work. We developed statistically rigorous analyses equipped to find homologous functional groups among these species at the levels of individual genes, functional and coexpressed gene modules, and transcription factor subnetworks. We identified six orthogroups involved in response to social challenge, including groups represented by mouse genes Npas4 and Nr4a1, as well as common modulation of systems such as transcriptional regulators, ion channels, G-protein-coupled receptors and synaptic proteins. We also identified conserved coexpression modules enriched for mitochondrial fatty acid metabolism and heat shock that constitute the shared neurogenomic response. Our analysis suggests a toolkit wherein nuclear receptors, interacting with chaperones, induce transcriptional changes in mitochondrial activity, neural cytoarchitecture and synaptic transmission after social challenge. It shows systems-level mechanisms that have been repeatedly co-opted during evolution of analogous behaviors, thus advancing the genetic toolkit concept beyond individual genes.


Biglycan potentially regulates angiogenesis during fracture repair by altering expression and function of endostatin.
The small proteoglycan biglycan (Bgn) is highly expressed in the organic matrix of bone and plays a role in bone formation. Previous work implicated Bgn in vessel growth during bone healing [1]. By infusing barium sulfate (BaSO4) into WT and Bgn-deficient mice we discovered the positive effect of Bgn in modulating angiogenesis during fracture healing. Using micro-computed tomography angiography we found significant differences in the vessel size and volume among other parameters. To further understand the mechanistic basis for this, we explored the relationship between Bgn and the anti-angiogenic protein endostatin. Immunohistochemistry (IHC) showed co-localization of Bgn and endostatin in regions of bone formation, with increased endostatin staining in Bgn-KO compared to WT at 14days post-fracture. To further elucidate the relationship between Bgn and endostatin, an endothelial cell tube formation assay was used. This study showed that endothelial cells treated with endostatin had significantly decreased vessel length and vessel branches compared to untreated cells, while cells treated with endostatin and Bgn at a 1:1M ratio had vessel length and vessel branches comparable to untreated cells. This indicated that Bgn was able to mitigate the inhibitory effect of endostatin on endothelial cell growth. In summary, these results suggest that Bgn is needed for proper blood vessel formation during fracture healing, and one mechanism by which Bgn impacts angiogenesis is through inhibition of endostatin.


Large-scale mutation in the evolution of a gene complex for cryptic coloration.
The types of mutations affecting adaptation in the wild are only beginning to be understood. In particular, whether structural changes shape adaptation by suppressing recombination or by creating new mutations is unresolved. Here, we show that multiple linked but recombining loci underlie cryptic color morphs of Timema chumash stick insects. In a related species, these loci are found in a region of suppressed recombination, forming a supergene. However, in seven species of Timema, we found that a megabase-size "supermutation" has deleted color loci in green morphs. Moreover, we found that balancing selection likely contributes more to maintaining this mutation than does introgression. Our results show how suppressed recombination and large-scale mutation can help to package gene complexes into discrete units of diversity such as morphs, ecotypes, or species.


Simultaneous imaging of oxygen tension and blood flow in animals using a digital micromirror device.
In this study we present a novel imaging method that combines high resolution cerebral blood flow imaging with a highly flexible map of absolute pO(2). In vivo measurements of pO(2) in animals using phosphorescence quenching is a well established method, and is preferable over electrical probes which are inherently invasive and are limited to single point measurements. However, spatially resolved pO(2) measurements using phosphorescence lifetime quenching typically require expensive cameras to obtain images of pO(2) and often suffer from poor signal to noise. Our approach enables us to retain the high temporal resolution and sensitivity of single point detection of phosphorescence by using a digital micromirror device (DMD) to selectively illuminate arbitrarily shaped regions of tissue. In addition, by simultaneously using Laser Speckle Contrast Imaging (LSCI) to measure relative blood flow, we can better examine the relationship between blood flow and absolute pO(2). We successfully used this instrument to study changes that occur during ischemic conditions in the brain with enough spatial resolution to clearly distinguish different regions. This novel instrument will provide researchers with an inexpensive and improved technique to examine multiple hemodynamic parameters simultaneously in the brain as well as other tissues.


Non-invasive ventilation for cystic fibrosis.
Non-invasive ventilation (NIV) may be a means to temporarily reverse or slow the progression of respiratory failure in cystic fibrosis (CF). To compare the effect of NIV versus no NIV in people with CF. We searched the Cochrane Cystic Fibrosis and Genetic Disorders Group Trials Register comprising references identified from comprehensive electronic database searches, handsearching relevant journals and abstract books of conference proceedings. We searched the reference lists of each trial for additional publications possibly containing other trials.Most recent search: June 2008. Randomised controlled trials comparing a form of pressure preset or volume preset NIV to no NIV in people with acute or chronic respiratory failure in CF. Three reviewers independently assessed trials for inclusion criteria and methodological quality, and extracted data. Fifteen trials were identified; seven trials met the inclusion criteria with a total of 106 participants. Six trials evaluated single treatment sessions and one evaluated a six-week intervention.Four trials (79 participants) evaluated NIV for airway clearance compared with an alternative chest physiotherapy method and showed that airway clearance may be easier with NIV and people with CF may prefer it. We were unable to find any evidence that NIV increases sputum expectoration, but it did improve some lung function parameters.Three trials (27 participants) evaluated NIV for overnight ventilatory support, measuring lung function, validated quality of life scores and nocturnal transcutaneous carbon dioxide. Due to the small numbers of participants and statistical issues, there were discrepancies in the results between the RevMan and the original trial analyses. No clear differences were found between NIV compared with oxygen or room air except for exercise performance, which significantly improved with NIV compared to room air over six weeks. Non-invasive ventilation may be a useful adjunct to other airway clearance techniques, particularly in people with CF who have difficulty expectorating sputum. Non-invasive ventilation, used in addition to oxygen, may improve gas exchange during sleep to a greater extent than oxygen therapy alone in moderate to severe disease. These benefits of NIV have largely been demonstrated in single treatment sessions with small numbers of participants. The impact of this therapy on pulmonary exacerbations and disease progression remain unclear. There is a need for long-term randomised controlled trials which are adequately powered to determine the clinical effects of non-invasive ventilation in CF airway clearance and exercise.


Distribution and physiology of aerobic bacteria containing bacteriochlorophyll a on the East and west coasts of australia.
Aerobic heterotrophic bacteria containing bacteriochlorophyll were isolated from specimens from a wide variety of marine environments on the west (Shark Bay, Lake Clifton, Lake Heyward, and Perth) and east (near Townsville and Brisbane) coasts of Australia. The bacteria were found in a high proportion (10 to 30%) of the total heterotrophic bacterial strains isolated from marine algae, seagrasses, stromatolites, the epiphytes on stromatolites, seawater, and sands; in some cases they constituted up to 49% of the total. This is much higher than the previous report of 6% from Japan. A high percentage, 13%, was also found in the seawater of Hamelin Pool, at Shark Bay, where the salinity was 66%. The number of these bacteria was generally low in seawater and sands, with a few exceptions. There were no aerobic bacteriochlorophyll-containing bacteria on sponges or corals. The isolated strains were orange or pink, and most had absorption maxima around 800 and 850 to 870 nm, the latter range being the absorption of bacteriochlorophyll a in vivo. The maximum bacteriochlorophyll content was 1 nmol/mg (dry weight) of bacterial cells. Most of the bacteria did not grow phototrophically under anaerobic conditions in a broth medium containing succinate. Cells and cell extracts grown under aerobic conditions had photochemical activities such as reversible photooxidations of the reaction center and cytochrome(s). Some strains showed denitrifying activity. The optimal salinity for bacterial growth varied between strains.


SUR1 regulates PKA-independent cAMP-induced granule priming in mouse pancreatic B-cells.
Measurements of membrane capacitance were applied to dissect the cellular mechanisms underlying PKA-dependent and -independent stimulation of insulin secretion by cyclic AMP. Whereas the PKA-independent (Rp-cAMPS-insensitive) component correlated with a rapid increase in membrane capacitance of approximately 80 fF that plateaued within approximately 200 ms, the PKA-dependent component became prominent during depolarizations >450 ms. The PKA-dependent and -independent components of cAMP-stimulated exocytosis differed with regard to cAMP concentration dependence; the K(d) values were 6 and 29 micro M for the PKA-dependent and -independent mechanisms, respectively. The ability of cAMP to elicit exocytosis independently of PKA activation was mimicked by the selective cAMP-GEFII agonist 8CPT-2Me-cAMP. Moreover, treatment of B-cells with antisense oligodeoxynucleotides against cAMP-GEFII resulted in partial (50%) suppression of PKA-independent exocytosis. Surprisingly, B-cells in islets isolated from SUR1-deficient mice (SUR1(-/-) mice) lacked the PKA-independent component of exocytosis. Measurements of insulin release in response to GLP-1 stimulation in isolated islets from SUR1(-/-) mice confirmed the complete loss of the PKA-independent component. This was not attributable to a reduced capacity of GLP-1 to elevate intracellular cAMP but instead associated with the inability of cAMP to stimulate influx of Cl(-) into the granules, a step important for granule priming. We conclude that the role of SUR1 in the B cell extends beyond being a subunit of the plasma membrane K(ATP)-channel and that it also plays an unexpected but important role in the cAMP-dependent regulation of Ca(2+)-induced exocytosis.


Untargeted Liquid Chromatography-Mass Spectrometry-Based Metabolomics Analysis of Wheat Grain.
Understanding the interactions between genes, the environment and management in agricultural practice could allow more accurate prediction and management of product yield and quality. Metabolomics data provides a read-out of these interactions at a given moment in time and is informative of an organism's biochemical status. Further, individual metabolites or panels of metabolites can be used as precise biomarkers for yield and quality prediction and management. The plant metabolome is predicted to contain thousands of small molecules with varied physicochemical properties that provide an opportunity for a biochemical insight into physiological traits and biomarker discovery. To exploit this, a key aim for metabolomics researchers is to capture as much of the physicochemical diversity as possible within a single analysis. Here we present a liquid chromatography-mass spectrometry-based untargeted metabolomics method for the analysis of field-grown wheat grain. The method uses the liquid chromatograph quaternary solvent manager to introduce a third mobile phase and combines a traditional reversed-phase gradient with a lipid-amenable gradient. Grain preparation, metabolite extraction, instrumental analysis and data processing workflows are described in detail. Good mass accuracy and signal reproducibility were observed, and the method yielded approximately 500 biologically relevant features per ionization mode. Further, significantly different metabolite and lipid feature signals between wheat varieties were determined.


Implication of the recent positive endovascular intervention trials for organizing acute stroke care: European perspective.
Timely recanalization leads to improved patient outcomes in acute ischemic stroke. Recent trial results demonstrated a strong benefit for endovascular therapies over standard medical care in patients with acute ischemic stroke and a major intracranial artery occlusion≤6 hours or even beyond from symptom onset and independent of patients' age. Previous studies have shown the benefit of intravenous thrombolysis that had gradually, albeit slowly, reshaped acute stroke care worldwide. Now, given the superior benefits of endovascular intervention, the whole structure of acute stroke care needs to be reorganized to meet patient needs and to deliver evidence-based treatments effectively. However, a blueprint for success with novel stroke treatments should be composed of numerous elements and requires efforts from various parties. Regarding the endovascular therapies, the strengths of Europe include highly organized democratic society structures, high rate of urbanization, well-developed revenue-based healthcare systems, and high income levels, whereas the obstacles include the east-west disparity in wealth, the ongoing economic crisis hindering spread of fairly costly new treatments, and the quickly aging population putting more demands on health care in general. Regional and national plans for covering whole population with 24/7 adequate acute stroke care are necessary in close cooperation of professionals and decision-makers. Europe-wide new training programs for expert physicians in stroke care should be initiated shortly. European Stroke Organisation has a unique role in providing expertise, consultation, guidelines, and versatile training in meeting new demands in stroke care. This article discusses the current situation, prospects, and challenges in Europe offering personal views on potential solutions.


Continuous protein purification using functionalized magnetic nanoparticles in aqueous micellar two-phase systems.
A novel technique for technical-scale continuous purification of proteins is presented. It is based on the combined use of functionalized magnetic nano-particles and an Aqueous Micellar Two-Phase System featuring the non-ionic surfactant, Eumulgin ES, which undergoes temperature induced phase separation at ∼25°C. In the first step, conducted below the transition temperature (i.e. 15°C), the magnetic sorbent particles are added into the single dispersed phase and bind the protein of interest. Next, on raising the temperature to 30°C the protein-laden magnetic particles partition strongly into the micelle-rich top phase of the micellar two-phase system that's formed. The magnetically susceptible top phase is then continuously separated from the micelle-poor phase in a flowthrough tailor-made magnetic extractor featuring a permanent magnet providing an upwardly acting magnetic force. This separation device was shown to be effective for continuous separation of a wide range of differently sized magnetic particle sorbents (i.e. from 2μm diameter to as small as 25nm) from a 10% (w/w) Eumulgin ES system; high separation efficiencies were recorded for the phase-forming surfactant (87 to >98%), and all magnetic sorbent particles tested (95-99.9%). Finally, protein purification by continuous magnetic extraction was demonstrated at 15L scale for the recovery of an antibody fragment, A33 Fab', from a crude extract of Escherichia coli periplasm. Nearly 70% of the A33 Fab' initially present in the extract at 15.6% of the total protein content was recovered in a 2-fold concentrated and highly purified (>98%) state. Further, the amounts of magnetic sorbent and phase-forming surfactant lost in the process were very small; thus recycling of both components into subsequent rounds of continuous magnetic extraction is highly feasible.


Generation of a high-affinity Fcgamma receptor by Ig-domain swapping between human CD64A and CD16A.
A recombinant soluble version of the human high-affinity receptor for IgG, rh-FcgammaRIA or CD64A, was expressed in mammalian cells and purified from their conditioned media. As assessed by circular dichroism, size exclusion chromatography and dynamic light scattering, incubation of rh-FcgammaRIA at 37 degrees C resulted in time-dependent formation of soluble aggregates caused by protein unfolding and loss of native structure. Aggregate formation was irreversible, temperature-dependent and was independent of rh-FcgammaRIA concentration. Aggregated rh-FcgammaRIA lost its ability to inhibit immune complex precipitation and failed to bind to IgG-Sepharose. Addition of human IgG1 to rh-FcgammaRIA prior to incubation at 37 degrees C blocked the formation of rh-FcgammaRIA aggregates. Production of soluble monomeric rh-FcgammaRIA was limited by aggregate formation during cell culture. Substitution of the membrane distal D1 Ig domain of FcgammaRIA with the D1 Ig domain of FcgammaRIIIA or CD16A resulted in a chimeric receptor, FcgammaR3A1A, with enhanced temperature stability. Relative to native rh-FcgammaRIA, FcgammaR3A1A exhibited less aggregation in Chinese hamster ovary cell-conditioned media or when purified receptor was incubated for up to 24 h at 37 degrees C. Both receptors bound to immobilized human IgG1 with high affinity and were equipotent at blockade of immune complex-mediated cytokine production from cultured mast cells. Equivalent dose-dependent reductions in edema and neutrophil infiltration in the cutaneous Arthus reaction in mice were noted for rh-FcgammaRIA and FcgammaR3A1A. These data demonstrate that the D1 Ig domains of FcgammaRIA and FcgammaRIIIA are functionally interchangeable and further suggest that the chimeric receptor FcgammaR3A1A is an effective inhibitor of type III hypersensitivity in mice.


Venous thromboembolic events during warm autoimmune hemolytic anemia.
Thrombotic manifestations are a hallmark of many auto-immune diseases (AID), specially of warm autoimmune hemolytic anemia (wAIHA), as 15 to 33% of adults with wAIHA experience venous thromboembolic events (VTE). However, beyond the presence of positive antiphospholipid antibodies and splenectomy, risk factors for developing a VTE during wAIHA have not been clearly identified. The aim of this retrospective study was to characterize VTEs during wAIHA and to identify risk factors for VTE. Forty-eight patients with wAIHA were included, among whom 26 (54%) had secondary wAIHA. Eleven (23%) patients presented at least one VTE, that occurred during an active phase of the disease for 10/11 patients (90%). The frequency of VTE was not different between primary and secondary AIHA (23.7 vs. 19.2%; p = 0.5). The Padua prediction score based on traditional risk factors was not different between patients with and without VTE. On multivariate analysis, total bilirubin ≥ 40 μmol/L [odds ratio (OR) = 7.4; p = 0.02] and leucocyte count above 7x10(9)/L (OR = 15.7; p = 0.02) were significantly associated with a higher risk of thrombosis. Antiphospholipid antibodies were screened in 9 out the 11 patients who presented a VTE and were negative. Thus, the frequency of VTE is high (23%) during wAIHA and VTE preferentially occur within the first weeks of diagnosis. As no clinically relevant predictive factors of VTE could be identified, the systematic use of a prophylactic anticoagulation should be recommended in case of active hemolysis and its maintenance after hospital discharge should be considered. The benefit of a systematic screening for VTE and its procedure remain to be determined.


Effects of leg massage on recovery from high intensity cycling exercise.
The effect of massage on recovery from high intensity exercise is debatable. Many studies on massage suffer from methodological flaws such as poor standardisation of previous exercise, lack of dietary control, and inappropriate massage duration. To examine the effects of leg massage compared with passive recovery on lactate clearance, muscular power output, and fatigue characteristics after repeated high intensity cycling exercise, with the conditions before the intervention controlled and standardised. Nine male games players participated. They attended the laboratory on two occasions one week apart and at the same time of day. Dietary intake and activity were replicated for the two preceding days on each occasion. After baseline measurement of heart rate and blood lactate concentration, subjects performed a standardised warm up on the cycle ergometer. This was followed by six standardised 30 second high intensity exercise bouts, interspersed with 30 seconds of active recovery. After five minutes of active recovery and either 20 minutes of leg massage or supine passive rest, subjects performed a second standardised warm up and a 30 second Wingate test. Capillary blood samples were drawn at intervals, and heart rate, peak power, mean power, and fatigue index were recorded. There were no significant differences in mean power during the initial high intensity exercise bouts (p = 0.92). No main effect of massage was observed on blood lactate concentration between trials (p = 0.82) or heart rate (p = 0.81). There was no difference in the maximum power (p = 0.75) or mean power (p = 0.66) in the subsequent Wingate test, but a significantly lower fatigue index was observed in the massage trial (p = 0.04; mean (SD) fatigue index 30.2 (4.1)% v 34.2 (3.3)%). No measurable physiological effects of leg massage compared with passive recovery were observed on recovery from high intensity exercise, but the subsequent effect on fatigue index warrants further investigation.