Can small institutes address some problems facing biomedical researchers?
At a time of historically low National Institutes of Health funding rates and many problems with the conduct of research (unfunded mandates, disgruntled reviewers, and rampant paranoia), there is a concern that biomedical research as a profession is waning in the United States (see "Rescuing US biomedical research from its systemic flaws" by Alberts and colleagues in the Proceedings of the National Academy of Sciences). However, it is wonderful to discover something new and to tackle tough puzzles. If we could focus more of our effort on discussing scientific problems and doing research, then we could be more productive and perhaps happier. One potential solution is to focus efforts on small thematic institutes in the university structure that can provide a stimulating and supportive environment for innovation and exploration. With an open-lab concept, there are economies of scale that can diminish paperwork and costs, while providing greater access to state-of-the-art equipment. Merging multiple disciplines around a common theme can catalyze innovation, and this enables individuals to develop new concepts without giving up the credit they deserve, because it is usually clear who did the work. Small institutes do not solve larger systemic problems but rather enable collective efforts to address the noisome aspects of the system and foster an innovative community effort to address scientific problems.


Molecular composition and ultrastructure of Jurassic paravian feathers.
Feathers are amongst the most complex epidermal structures known, and they have a well-documented evolutionary trajectory across non-avian dinosaurs and basal birds. Moreover, melanosome-like microbodies preserved in association with fossil plumage have been used to reconstruct original colour, behaviour and physiology. However, these putative ancient melanosomes might alternatively represent microorganismal residues, a conflicting interpretation compounded by a lack of unambiguous chemical data. We therefore used sensitive molecular imaging, supported by multiple independent analytical tests, to demonstrate that the filamentous epidermal appendages in a new specimen of the Jurassic paravian Anchiornis comprise remnant eumelanosomes and fibril-like microstructures, preserved as endogenous eumelanin and authigenic calcium phosphate. These results provide novel insights into the early evolution of feathers at the sub-cellular level, and unequivocally determine that melanosomes can be preserved in fossil feathers.


Gaussian beam propagation in anisotropic turbulence along horizontal links: theory, simulation, and laboratory implementation.
Experimental and theoretical work has shown that atmospheric turbulence can exhibit "non-Kolmogorov" behavior including anisotropy and modifications of the classically accepted spatial power spectral slope, -11/3. In typical horizontal scenarios, atmospheric anisotropy implies that the variations in the refractive index are more spatially correlated in both horizontal directions than in the vertical. In this work, we extend Gaussian beam theory for propagation through Kolmogorov turbulence to the case of anisotropic turbulence along the horizontal direction. We also study the effects of different spatial power spectral slopes on the beam propagation. A description is developed for the average beam intensity profile, and the results for a range of scenarios are demonstrated for the first time with a wave optics simulation and a spatial light modulator-based laboratory benchtop counterpart. The theoretical, simulation, and benchtop intensity profiles show good agreement and illustrate that an elliptically shaped beam profile can develop upon propagation. For stronger turbulent fluctuation regimes and larger anisotropies, the theory predicts a slightly more elliptical form of the beam than is generated by the simulation or benchtop setup. The theory also predicts that without an outer scale limit, the beam width becomes unbounded as the power spectral slope index α approaches a maximum value of 4. This behavior is not seen in the simulation or benchtop results because the numerical phase screens used for these studies do not model the unbounded wavefront tilt component implied in the analytic theory.


The Chemical Synthesis, Stability, and Activity of MAIT Cell Prodrug Agonists That Access MR1 in Recycling Endosomes.
Mucosal-associated invariant T (MAIT) cells are antibacterial effector T cells that react to pyrimidines derived from bacterial riboflavin synthesis presented by the monomorphic molecule MR1. A major challenge in MAIT cell research is that the commonly used MAIT agonist precursor, 5-amino-6-d-ribitylaminouracil (5-A-RU), is labile to autoxidation, resulting in a loss of biological activity. Here, we characterize two independent autoxidation processes by LCMS. To overcome the marked instability, we report the synthesis of a 5-A-RU prodrug generated by modification of the 5-amino group with a cleavable valine-citrulline-p-aminobenzyl carbamate. The compound is stable in prodrug form, with the parent amine (i.e., 5-A-RU) released only after enzymatic cleavage. Analysis of the prodrug in vitro and in vivo showed an enhanced MAIT cell activation profile compared to 5-A-RU, which was associated with preferential loading within recycling endosomes, a route used by some natural agonists. This prodrug design therefore overcomes the difficulties associated with 5-A-RU in biological studies and provides an opportunity to explore different presentation pathways.


What are the long-term symptoms and complications of COVID-19: a protocol for a living systematic review.
Although the majority of patients with COVID-19 will experience mild to moderate symptoms and will recover fully, there is now increasing evidence that a significant proportion will experience persistent symptoms for weeks or months after the acute phase of the illness. These symptoms include, among others, fatigue, problems in breathing, lack of smell and taste, headaches, and also depression and anxiety. It has also become clear that the virus has lasting effects not only on the respiratory system but also on other parts of the body, including the heart, liver, and the nervous system. In this paper we present a protocol for a living systematic review that aims to synthesize the evidence on the prevalence and duration of symptoms and clinical features of post-acute COVID-19 and its long-term complications. The living systematic review will be updated regularly, initially monthly with update cycles under continuous review as the pace of new evidence generated develops through the pandemic. We will include studies that follow up with COVID-19 patients who have experienced persistent mild, moderate or severe symptoms, with no restrictions regarding country, setting, or language. We will use descriptive statistics to analyse the data and our findings will be presented as infographics to facilitate transcription to lay audiences. Ultimately, we aim to support the work of policy makers, practitioners, and patients when planning rehabilitation for those recovering from COVID-19. The protocol has been registered with PROSPERO ( CRD42020211131, 25/09/2020).


Pulmonary nocardiosis developed in a hematopoietic stem cell transplant recipient with bronchiolitis obliterans.
The chronic graft-versus-host disease often requires unceasing immunosuppressive therapy (IST), which increases a risk of infectious complications in hematopoietic stem cell transplantation (HSCT) recipients. We report an adult T-cell leukemia/lymphoma case who developed pulmonary nocardiosis, a rare pulmonary complication, after allogeneic HSCT despite administration of the prophylactic trimethoprim-sulfamethoxazole (TMP/STX). The inhaled corticosteroid in addition to systemic IST had been started for bronchiolitis obliterance 4 months prior to nocardiosis development. The patient was successfully treated with an increased dose of TMP/STX combined with meropenem. Transplantation physicians should keep this rare pulmonary complication in mind during sustained IST.


Stability of diagnosis of obsessive-compulsive disorder in the Epidemiologic Catchment Area study.
This study examined the 1-year temporal stability of a National Institute of Mental Health Diagnostic Interview Schedule (DIS) lifetime diagnosis of obsessive-compulsive disorder in the Epidemiologic Catchment Area (ECA) study. In that study, 20,862 individuals, aged 18 years and over, at five sites were evaluated by lay interviewers using the DIS (wave 1). All of those who were available 12 months later were reinterviewed (wave 2). In the present study, the temporal stability of wave 1 obsessive-compulsive disorder diagnoses at wave 2 was examined, as well as relationships with comorbid diagnoses. The consistency of reports of "new-onset" illness was also examined. Factors contributing to these measures were evaluated. The temporal stability of the diagnosis of obsessive-compulsive disorder was very low. Subjects with a stable diagnosis of obsessive-compulsive disorder had a higher rate of both obsessions and compulsions, an earlier age at onset, and more comorbid anxiety, affective, and alcohol abuse/dependence disorders at initial assessment. The originally reported 1-year incidence estimates for obsessive-compulsive disorder primarily reflect data from subjects at wave 2 who reported the onset of symptoms as preceding the wave 1 interview. Older and less-educated subjects had significantly higher error rates in reporting onset. The DIS diagnosis of obsessive-compulsive disorder has poor validity, leaving the true incidence and prevalence of the disorder unknown. Older and less-educated subjects require special attention in the design of instruments for use with community samples.


A specific role for Ca2+-dependent adenylyl cyclases in recovery from adaptive presynaptic silencing.
Glutamate generates fast postsynaptic depolarization throughout the CNS. The positive-feedback nature of glutamate signaling likely necessitates flexible adaptive mechanisms that help prevent runaway excitation. We have previously explored presynaptic adaptive silencing, a form of synaptic plasticity produced by ongoing neuronal activity and by strong depolarization. Unsilencing mechanisms that maintain active synapses and restore normal function after adaptation are also important, but mechanisms underlying such presynaptic reactivation remain unexplored. Here we investigate the involvement of the cAMP pathway in the basal balance between silenced and active synapses, as well as the recovery of baseline function after depolarization-induced presynaptic silencing. Activation of the cAMP pathway activates synapses that are silent at rest, and pharmacological inhibition of cAMP signaling silences basally active synapses. Adenylyl cyclase (AC) 1 and AC8, the major Ca2+-sensitive AC isoforms, are not crucial for the baseline balance between silent and active synapses. In cells from mice doubly deficient in AC1 and AC8, the baseline percentage of active synapses was only modestly reduced compared with wild-type synapses, and forskolin unsilencing was similar in the two genotypes. Nevertheless, after strong presynaptic silencing, recovery of normal function was strongly inhibited in AC1/AC8-deficient synapses. The entire recovery phenotype of the double null was reproduced in AC8-deficient but not AC1-deficient cells. We conclude that, under normal conditions, redundant cyclase activity maintains the balance between presynaptically silent and active synapses, but AC8 plays a particularly important role in rapidly resetting the balance of active to silent synapses after adaptation to strong activity.


Aortic insufficiency secondary to aortic root aneurysm or dissection.
Twenty-four patients with aortic regurgitation secondary to aortic root aneurysm (13 patients) or dissection (11 patients) were operated on, utilizing a variety of surgical procedures to cope with the varied pathological findings. These ranged from primary repair of the ascending aorta without any prostheses in patients with acute aortic dissection to replacement of the valve and the entire ascending aorta for aortic root aneurysm. In four patients with Marfan syndrome the right coronary artery was transplanted to the ascending aortic graft, allowing an extension of the graft to the valve anulus and excision of the entire aneurysmal aorta. The immediate and late results have been most encouraging.


Highly Selective Metal-Organic Framework Textile Humidity Sensor.
The increase in demand and popularity of smart textiles brings new and innovative ideas to develop a diverse range of textile-based devices for our daily life applications. Smart textile-based sensors (TEX sensors) become attractive due to the potential to replace current solid-state sensor devices with flexible and wearable devices. We have developed a smart textile sensor for humidity detection using a metal-organic framework (MOF) as an active thin-film layer. We show for the first time the use of the Langmuir-Blodgett (LB) technique for the deposition of a MIL-96(Al) MOF thin film directly onto the fabrics containing interdigitated textile electrodes for the fabrication of a highly selective humidity sensor. The humidity sensors were made from two different types of textiles, namely, linen and cotton, with the linen-based sensor giving the best response due to better coverage of MOF. The TEX sensor showed a reproducible response after multiple cycles of measurements. After 3 weeks of storage, the sensor showed a moderate decrease in response. Moreover, TEX sensors showed a high level of selectivity for the detection of water vapors in the presence of several volatile organic compounds (VOCs). Interestingly, the selectivity is superior to some of the previously reported MOF-coated solid-state interdigitated electrode devices and textile sensors. The method herein described is generic and can be extended to other textiles and coating materials for the detection of toxic gases and vapors.


[Pharmacokinetics of meropenem administered with prolonged infusion time in patients receiving continuous veno-venous hemofiltration].
To demonstrate the pharmacokinetic profile of meropenem when administered by 3-hour infusion in patients undergoing continuous veno-venous hemofiltration (CVVH). The study was conducted in 10 patients, who were treated with CVVH. Each subject received meropenem in 3-hour infusion of 500 mg every 6 hours. Blood samples were collected before infusion (0 hour) and 0.25, 0.5, 1, 1.5, 2, 3, 4, 5, 6 hours (just before the infusion of the next dose) after the beginning of the fourth infusion. The concentrations of meropenem in plasma were measured by high-performance liquid chromatography method, and mean serum meropenem concentration-time curve was plotted. Peak plasma drug concentrations measured 3 hours post-infusion were (25.05 ± 5.64) mg/L, and trough levels after 6 hours of infusion were (13.03 ± 3.01) mg/L. The area under the plasma concentration-time curve (AUC) was (118.42 ± 26.78) mg x h⁻¹ x L⁻². The elimination half-life (T1/2) was (3.74 ± 0.55) hours. The mean residence time (MRT) was (4.99 ± 0.84) hours. The volume of distribution (Vb) was (22.85 ± 9.85) L and clearance of meropenem (CL) was (4.49 ± 1.32) L/h. The percentage of time that the serum drug concentration was above the minimum inhibitory concentration (MIC) accounting for the interval time of infusion (%T>MIC) was 100% (MIC 8 mg/L) in all the 10 patients. Based on these data, we concluded that satisfactory pharmacodynamic parameters could be attained in CVVH patients treated with meropenem by a prolonged infusion time of 3 hours with a dosage of 500 mg for every 6 hours.


Deep sequencing in the management of hepatitis virus infections.
The hepatitis viruses represent a major public health problem worldwide. Procedures for characterization of the genomic composition of their populations, accurate diagnosis, identification of multiple infections, and information on inhibitor-escape mutants for treatment decisions are needed. Deep sequencing methodologies are extremely useful for these viruses since they replicate as complex and dynamic quasispecies swarms whose complexity and mutant composition are biologically relevant traits. Population complexity is a major challenge for disease prevention and control, but also an opportunity to distinguish among related but phenotypically distinct variants that might anticipate disease progression and treatment outcome. Detailed characterization of mutant spectra should permit choosing better treatment options, given the increasing number of new antiviral inhibitors available. In the present review we briefly summarize our experience on the use of deep sequencing for the management of hepatitis virus infections, particularly for hepatitis B and C viruses, and outline some possible new applications of deep sequencing for these important human pathogens.


Infectivity, pathogenicity, and virulence of Trypanosoma cruzi Isolates from sylvatic animals and vectors, and domestic dogs from the United States in ICR strain mice and SD strain rats.
Trypanosoma cruzi, the causative agent of Chagas disease, is widespread in the southern United States. In addition to detection in numerous wildlife host species, cases have been diagnosed in domestic dogs and humans. In the current investigation, groups of laboratory mice [Crl:CD1 (ICR)] were inoculated with one of 18 United States T. cruzi isolates obtained from a wide host range to elucidate their infectivity, pathogenicity, and virulence. In addition, laboratory rats (SD strain) were inoculated with four isolates. Mice and rats were susceptible to infection with all strains, but no morbidity or mortality was noted, which indicates that these T. cruzi isolates from the United States had low virulence for laboratory mice and rats.


C-H functionalisation of aldehydes using light generated, non-stabilised diazo compounds in flow.
The difficulty in accessing and safely utilising non-stabilised diazo species has in the past limited the application of this class of compounds. Here we explore further the use of oxadiazolines, non-stabilised diazo precursors which are bench stable, in direct, non-catalytic, aldehyde C-H functionalisation reactions under UV photolysis in flow and free from additives. Commercially available aldehydes are coupled to afford unsymmetrical aryl-alkyl and alkyl-alkyl ketones while mild conditions and lack of transition metal catalysts allow for exceptional functional group tolerance. Examples are given on small scale and in a larger scale continuous production.


Literature review of the usefulness of repetitive nerve stimulation and single fiber EMG in the electrodiagnostic evaluation of patients with suspected myasthenia gravis or Lambert-Eaton myasthenic syndrome.
A retrospective literature review of the electrodiagnosis of myasthenia gravis (MG) and Lambert--Eaton myasthenic syndrome (LEMS) through July 1998 was performed for the purpose of generating evidence-based practice parameters. There were 545 articles identified, of which 13 articles met at least three of the six criteria set previously by the American Association of Electrodiagnostic Medicine (AAEM). An additional 21 articles were identified from review articles or the references of these first 13 articles leading to a total of 34 articles. Results of studies utilizing repetitive nerve stimulation (RNS) showed that a 10% decrement in amplitude from the first to fourth or fifth intravolley waveform while stimulating at 2--5 HZ is valid for the diagnosis of MG. The degree of increment needed for the diagnosis of LEMS is at least 25% but most accurate when greater than 100%. Abnormal jitter or impulse blocking are the appropriate criteria for diagnosis of neuromuscular junction (NMJ) disorders when using single fiber electromyography (SFEMG). SFEMG is more sensitive than RNS for the diagnosis of disorders of neuromuscular transmission, but may be less specific and may not be available. Therefore, RNS remains the preferred initial test for MG and LEMS.


Modification of K-ATP channels in pancreatic beta-cells by trypsin.
The inside-out configuration of the patch-clamp method was used to study the effects of trypsin on the activity of ATP-sensitive potassium (K-ATP) channels from isolated mouse pancreatic beta-cells. Trypsin (20 micrograms/ml) irreversibly enhanced channel activity around twofold by reducing the interburst intervals without altering the burst kinetics. No effect on the single channel conductance or the inward rectification produced by internal Mg2+ was observed: however, the protease did reduce the inhibitory effect of Mg2+ on channel activity. Trypsin both prevented rundown of K-ATP channel activity and reactivated the channels after complete rundown. These effects of trypsin were absent in the presence of trypsin inhibitor. The protease also reduced the inhibitory effect of ATP on channel activity, increasing the dissociation constant from 7 to 49 microM. Trypsin removed the activating effect of ADP (0.1 mmol/l) on channel activity and reduced the inhibitory effect of tolbutamide (0.5 mmol/l). Carboxypeptidase A did not activate K-ATP channels in excised patches, although it was able to slightly reactivate channels after complete rundown, whereas chymotrypsin increased K-ATP channel activity but it did not produce reactivation. The effects of papain were similar to those of trypsin.


Towards an ICF core set for ADHD: a worldwide expert survey on ability and disability.
This is the second in a series of four empirical studies designed to develop International Classification of Functioning, Disability and Health (ICF and Children and Youth version, ICF-CY) core sets for attention deficit hyperactivity disorder (ADHD). The objective of this stage was to gather the opinions from international experts on which ability and disability concepts were considered relevant to functioning in ADHD. An email-based survey was carried out amongst international experts in ADHD. Relevant functional ability and disability concepts were extracted from their responses and linked to the ICF/-CY categories by two independent researchers using a standardised linking procedure. 174 experts from 11 different disciplines and 45 different countries completed the survey. Meaningful concepts identified in their responses were linked to 185 ICF/-CY categories. Of these, 83 categories were identified by at least 5 % of the experts and considered the most relevant to ADHD: 30 of these were related to Body functions (most identified: attention functions, 85 %), 30 to Activities and Participation (most identified: school education, 52 %), 20 to Environmental factors (most identified: support from immediate family, 61 %), and 3 to Body structures (most identified: structure of brain, 83 %). Experts also provided their views on particular abilities related to ADHD, naming characteristics such as high-energy levels, flexibility and resiliency. Gender differences in the expression of ADHD identified by experts pertained mainly to females showing more internalising (e.g. anxiety, low self-esteem) and less externalising behaviours (e.g. hyperactivity), leading to a risk of late- and under-diagnosis in females. Results indicate that the impact of ADHD extends beyond the core symptom domains, into all areas of life and across the lifespan. The current study in combination with three additional preparatory studies (comprehensive scoping review, focus groups, clinical study) will provide the scientific basis to define the ADHD ICF/-CY core sets for multi-purpose use in basic and applied research and every day clinical practice.


A probable case of human neurotrichinellosis in the United States.
Human neurotrichinellosis is seldom reported. This is likely the result of the low incidence of parasites from the genus Trichinella in the United States domestic food supply, as well as difficulties in diagnosing the disease, especially when neither the organism nor the source of the infection are readily available. Although trichinellosis from domestic food supplies has been decreasing for many years, a resurgence has occurred in cases derived from the consumption of wild game. We report a rare case of neurotrichinellosis in the United States and implicate wild game as the source of the infection. These results suggest that clinicians should consider the potential for Trichinella infection in cases where wild game is common in the diets of the patients.


Erythrodermic mycosis fungoides with hypereosinophilic syndrome: a rare presentation.
Mycosis fungoides (MF) is the most common variant of primary cutaneous T-cell lymphoma (CTCL). It is generally associated with an indolent clinical course and characterised by well-defined clinicopathological features. Although rare, CTCLs constitute 65% of all cutaneous lymphoid malignancies, of which 50% are patients with MF. The erythrodermic variants of MF, a malignancy of mature, skin homing and clonal T lymphocytes, usually present in mid to late adulthood. Association with hypereosinophilia is important in prognosis. We report a case of erythrodermic MF with hypereosinophilic syndrome in a 22-year-old female presenting with gradually progressive intractable erythroderma with intensely pruritic multiple papules, plaques, and nodules involving more than 90% of body surface area. Diagnosis was confirmed by histopathological examination and immunophenotyping from multiple skin biopsies.


Disruption of the endoplasmic reticulum by cytotoxins in LLC-PK1 cells.
Prior induction of an endoplasmic reticulum stress response results in protection against reactive cytotoxins in the LLC-PK1 cell line. The purpose of this investigation was to determine therefore if the endoplasmic reticulum was disrupted by iodoacetamide, tert-butylhydroperoxide or sulfamethoxazole hydroxylamine. Toxic concentrations of the three toxins caused a dramatic loss of GRP94 protein within 3-8h of exposure, while induction of GRP78 and calreticulin occurred at 8 and 24h following exposure. There was no evidence of cytosolic elevation of calcium and neither dantrolene nor xestospongin were able to block the cytotoxicity of IDAM and TBHP. Exposure to the toxins led to DNA degradation and cleavage of procaspase-12. There was only evidence of procaspase-3 cleavage after TBHP exposure. These results demonstrate that the ER is disrupted by the reactive cytotoxins examined in LLC-PK1cells and suggest that the cytoprotection against low to moderate concentrations of cytotoxins observed following endoplasmic reticulum stress protein induction is likely due to a mechanism other than maintenance of calcium homeostasis.


Planting exotic relatives has increased the threat posed by Dothistroma septosporum to the Caledonian pine populations of Scotland.
To manage emerging forest diseases and prevent their occurrence in the future, it is essential to determine the origin(s) of the pathogens involved and identify the management practices that have ultimately caused disease problems. One such practice is the widespread planting of exotic tree species within the range of related native taxa. This can lead to emerging forest disease both by facilitating introduction of exotic pathogens and by providing susceptible hosts on which epidemics of native pathogens can develop. We used microsatellite markers to determine the origins of the pathogen Dothistroma septosporum responsible for the current outbreak of Dothistroma needle blight (DNB) on native Caledonian Scots pine (Pinus sylvestris) populations in Scotland and evaluated the role played by widespread planting of two exotic pine species in the development of the disease outbreak. We distinguished three races of D. septosporum in Scotland, one of low genetic diversity associated with introduced lodgepole pine (Pinus contorta), one of high diversity probably derived from the DNB epidemic on introduced Corsican pine (Pinus nigra subsp. laricio) in England and a third of intermediate diversity apparently endemic on Caledonian Scots pine. These races differed for both growth rate and exudate production in culture. Planting of exotic pine stands in the UK appears to have facilitated the introduction of two exotic races of D. septosporum into Scotland which now pose a threat to native Caledonian pines both directly and through potential hybridization and introgression with the endemic race. Our results indicate that both removal of exotic species from the vicinity of Caledonian pine populations and restriction of movement of planting material are required to minimize the impact of the current DNB outbreak. They also demonstrate that planting exotic species that are related to native species reduces rather than enhances the resilience of forests to pathogens.


Gas-phase rearrangements do not affect site localization reliability in phosphoproteomics data sets.
Intramolecular transfer of phosphate during collision-induced dissociation (CID) in ion-trap mass spectrometers has recently been described. Because phosphorylation events are assigned to discrete serine, threonine, and tyrosine residues based on the presence of site-determining ions in MS/MS spectra, phosphate transfer may invalidate or confound site localization in published large-scale phosphorylation data sets. Here, we present evidence for the occurrence of this phenomenon using synthetic phosphopeptide libraries, specifically for doubly charged species. We found, however, that the extent of the transfer reaction was insufficient to cause localization of phosphorylation sites to incorrect residues. We further compared CID to electron-transfer dissociation (ETD) for site localization using synthetic libraries and a large-scale yeast phosphoproteome experiment. The agreement in site localization was >99.5 and 93%, respectively, suggesting that ETD-based site localization is no more reliable than CID. We conclude that intramolecular phosphate transfer does not affect the reliability of current or past phosphorylation data sets.


Endotoxemia-mediated inflammation potentiates aminoglycoside-induced ototoxicity.
The ototoxic aminoglycoside antibiotics are essential to treat severe bacterial infections, particularly in neonatal intensive care units. Using a bacterial lipopolysaccharide (LPS) experimental model of sepsis, we tested whether LPS-mediated inflammation potentiates cochlear uptake of aminoglycosides and permanent hearing loss in mice. Using confocal microscopy and enzyme-linked immunosorbent assays, we found that low-dose LPS (endotoxemia) greatly increased cochlear concentrations of aminoglycosides and resulted in vasodilation of cochlear capillaries without inducing paracellular flux across the blood-labyrinth barrier (BLB) or elevating serum concentrations of the drug. Additionally, endotoxemia increased expression of both serum and cochlear inflammatory markers. These LPS-induced changes, classically mediated by Toll-like receptor 4 (TLR4), were attenuated in TLR4-hyporesponsive mice. Multiday dosing with aminoglycosides during chronic endotoxemia induced greater hearing threshold shifts and sensory cell loss compared to mice without endotoxemia. Thus, endotoxemia-mediated inflammation enhanced aminoglycoside trafficking across the BLB and potentiated aminoglycoside-induced ototoxicity. These data indicate that patients with severe infections are at greater risk of aminoglycoside-induced hearing loss than previously recognized.


Indian Society of Critical Care Medicine Experts Committee Consensus Statement on ICU Planning and Designing, 2020.
Indian Society of Critical Care Medicine (ISCCM) guidelines on Planning and Designing Intensive care (ICU) were first developed in 2001 and later updated in 2007. These guidelines were adopted in India, many developing Nations and major Institutions including NABH. Various international professional bodies in critical care have their own position papers and guidelines on planning and designing of ICUs; being the professional body of intensivists in India ISCCM therefore addresses the subject in contemporary context relevant to our clinical practice, its variability according to specialty and subspecialty, quality, resource limitation, size and location of the institution. Aim: To have a consensus document reflecting the philosophy of ISCCM to deliver safe & quality Critical Care in India, taking into consideration the requirement of regulatory agencies (national & international) and need of people at large, including promotion of training, education and skill upgradation. It also aiming to promote leadership and development and managerial skill among the critical care team. Material and Methods: Extensive review of literature including search of databases in English language, resources of regulatory bodies, guidelines and recommendations of international critical care societies. National Survey of ISCCM members and experts to understand their viewpoints on respective issues. Visiting of different types and levels of ICUs by team members to understand prevailing practices, aspiration and Challenges. Several face to face meetings of the expert committee members in big and small groups with extensive discussions, presentations, brain storming and development of initial consensus draft. Discussion on draft through video conferencing, phone calls, Emails circulations, one to one discussion Result: Based upon extensive review, survey and input of experts' ICUs were categorized in to three levels suitable in Indian setting. Level III ICUs further divided into sub category A and B. Recommendations were grouped in to structure, equipment and services of ICU with consideration of variation in level of ICU of different category of hospitals. Conclusion: This paper summarizes consensus statement of various aspect of ICU planning and design. Defined mandatory and desirable standards of all level of ICUs and made recommendations regarding structure and layout of ICUs. Definition of intensive care and intensivist, planning for strength of ICU and requirement of manpower were also described. Rungta N, Zirpe KG, Dixit SB, Mehta Y, Chaudhry D, Govil D, et al. Indian Society of Critical Care Medicine Experts Committee Consensus Statement on ICU Planning and Designing, 2020. Indian J Crit Care Med 2020;24(Suppl 1):S43-S60.


SU-E-I-57: CT Dose Metrics: What Are We Tracking.
Recent guidance by The Joint Commission and CRCPD recommendations require establishing CT reference dose levels (RDLs) for clinical protocols and recording CT dose metrics in the patient's medical record. This presentation addresses which dose values in the CT report should be recorded for the purposes of monitoring patient dose and determining RDLs effectively. CT dose reports are commonly recorded in the PACS as screen captures, although structured reporting is becoming available on current CT scanners and PACS systems. The goal is to obtain data that correctly reflects the patient's dose, but the dose information captured is not standardized across vendors and can be difficult to compare. Multi-phase studies, deviations from established protocols and dynamic scanning present problems when recording numbers to establish RDLs because of the lack of information on the anatomy scanned. The cumulative DLP and CTDIvol manually entered by radiologic technologists into the electronic medical record were compared with more detailed dose metrics compiled from PACS images. Analysis of this data showed that simple cumulative metrics are a poor indicator of patient dose. Major problems are 1) the inclusion of dynamic scan doses associated with bolus tracking, which can skew protocol CTDIvol values by a factor of up to 20, and 2) add-on scans of non-overlapping anatomy which can inaccurately increase apparent patient dose. Recording CTDIvol to monitor patient doses is not straightforward, since details of the actual anatomy scanned are lacking without image-based review. More granular dose reporting which identifies individual acquisitions is required; however, current RIS systems do not provide the flexibility necessary to capture all this information.


Pharmaceutical Optimization of Peptide Toxins for Ion Channel Targets: Potent, Selective, and Long-Lived Antagonists of Kv1.3.
To realize the medicinal potential of peptide toxins, naturally occurring disulfide-rich peptides, as ion channel antagonists, more efficient pharmaceutical optimization technologies must be developed. Here, we show that the therapeutic properties of multiple cysteine toxin peptides can be rapidly and substantially improved by combining direct chemical strategies with high-throughput electrophysiology. We applied whole-molecule, brute-force, structure-activity analoging to ShK, a peptide toxin from the sea anemone Stichodactyla helianthus that inhibits the voltage-gated potassium ion channel Kv1.3, to effectively discover critical structural changes for 15× selectivity against the closely related neuronal ion channel Kv1.1. Subsequent site-specific polymer conjugation resulted in an exquisitely selective Kv1.3 antagonist (>1000× over Kv1.1) with picomolar functional activity in whole blood and a pharmacokinetic profile suitable for weekly administration in primates. The pharmacological potential of the optimized toxin peptide was demonstrated by potent and sustained inhibition of cytokine secretion from T cells, a therapeutic target for autoimmune diseases, in cynomolgus monkeys.


Mechanism of downregulation of photosystem I content under high-light conditions in the cyanobacterium Synechocystis sp. PCC 6803.
Downregulation of photosystem I (PSI) content is an essential process for cyanobacteria to grow under high-light (HL) conditions. In a pmgA (sll1968) mutant of Synechocystis sp. PCC 6803, the levels of PSI content, chlorophyll and transcripts of the psaAB genes encoding reaction-centre subunits of PSI could not be maintained low during HL incubation, although the causal relationship among these phenotypes remains unknown. In this study, we modulated the activity of psaAB transcription or that of chlorophyll synthesis to estimate their contribution to the regulation of PSI content under HL conditions. Analysis of the psaAB-OX strain, in which the psaAB genes were overexpressed under HL conditions, revealed that the amount of psaAB transcript could not affect PSI content by itself. Suppression of chlorophyll synthesis by an inhibitor, laevulinic acid, in the pmgA mutant revealed that chlorophyll availability could be a determinant of PSI content under HL. It was also suggested that chlorophyll content under HL conditions is mainly regulated at the level of 5-aminolaevulinic acid synthesis. We conclude that, upon the shift to HL conditions, activities of psaAB transcription and of 5-aminolaevulinic acid synthesis are strictly downregulated by regulatory mechanism(s) independent of PmgA during the first 6 h, and then a PmgA-mediated regulatory mechanism becomes active after 6 h onward of HL incubation to maintain these activities at a low level.


Karyotype analysis of Leishmania species and its use in classification and clinical diagnosis.
Chromosomes of four species of Leishmania represented by ten different geographic isolates were analyzed by pulsed field gradient gel electrophoresis (PFG) to assess chromosome stability in these parasitic protozoans. Among different geographic isolates of the same subspecies, more than two-thirds of chromosomes had similar sizes, ethidium bromide staining intensities, and locations of alpha,beta-tubulin genes. However, among New World Leishmania, members of different species or subspecies have fewer than one-third of their chromosomes in common. Therefore, PFG karyotypes of Leishmania exhibit intraspecific variability similar to that reported for other parasitic protozoans. The greater similarities of the karyotypes of members of the same Leishmania subspecies may indicate that they represent valid taxa. These similarities also allowed the use of PFG in clinical diagnosis for rapid and accurate typing of patient isolates.


All-or-none activation of CRAC channels by agonist elicits graded responses in populations of mast cells.
In nonexcitable cells, receptor stimulation evokes Ca(2+) release from the endoplasmic reticulum stores followed by Ca(2+) influx through store-operated Ca(2+) channels in the plasma membrane. In mast cells, store-operated entry is mediated via Ca(2+) release-activated Ca(2+) (CRAC) channels. In this study, we find that stimulation of muscarinic receptors in cultured mast cells results in Ca(2+)-dependent activation of protein kinase Calpha and the mitogen activated protein kinases ERK1/2 and this is required for the subsequent stimulation of the enzymes Ca(2+)-dependent phospholipase A(2) and 5-lipoxygenase, generating the intracellular messenger arachidonic acid and the proinflammatory intercellular messenger leukotriene C(4). In cell population studies, ERK activation, arachidonic acid release, and leukotriene C(4) secretion were all graded with stimulus intensity. However, at a single cell level, Ca(2+) influx was related to agonist concentration in an essentially all-or-none manner. This paradox of all-or-none CRAC channel activation in single cells with graded responses in cell populations was resolved by the finding that increasing agonist concentration recruited more mast cells but each cell responded by generating all-or-none Ca(2+) influx. These findings were extended to acutely isolated rat peritoneal mast cells where muscarinic or P2Y receptor stimulation evoked all-or-none activation of Ca(2+)entry but graded responses in cell populations. Our results identify a novel way for grading responses to agonists in immune cells and highlight the importance of CRAC channels as a key pharmacological target to control mast cell activation.


Optimizing the beam pattern of a forward-viewing ring-annular ultrasound array for intravascular imaging.
Intravascular ultrasound (IVUS) imaging systems using circumferential arrays mounted on cardiac catheter tips fire beams orthogonal to the principal axis of the catheter. The system produces high resolution cross-sectional images but must be guided by conventional angioscopy. A real-time forward-viewing array, integrated into the same catheter, could greatly reduce radiation exposure by decreasing angiographic guidance. Unfortunately, the mounting requirement of a catheter guide wire prohibits a full-disk imaging aperture. Given only an annulus of array elements, prior theoretical investigations have only considered a circular ring of point transceivers and focusing strategies using all elements in the highly dense array, both impractical assumptions. In this paper, we consider a practical array geometry and signal processing architecture for a forward-viewing IVUS system. Our specific design uses a total of 210 transceiver firings with synthetic reconstruction for a given 3-D image frame. Simulation results demonstrate this design can achieve side-lobes under -40 dB for on-axis situations and under -30 dB for steering to the edge of a 80 degrees cone.


The role of patients' explanatory models and daily-lived experience in hypertension self-management.
Uncontrolled hypertension remains a significant problem for many patients. Few interventions to improve patients' hypertension self-management have had lasting effects. Previous work has focused largely on patients' beliefs as predictors of behavior, but little is understood about beliefs as they are embedded in patients' social contexts. This study aims to explore how patients' "explanatory models" of hypertension (understandings of the causes, mechanisms or pathophysiology, course of illness, symptoms and effects of treatment) and social context relate to their reported daily hypertension self-management behaviors. Semi-structured qualitative interviews with a diverse group of patients at two large urban Veterans Administration Medical centers. PARTICIPANTS (OR PATIENTS OR SUBJECTS): African-American, white and Latino Veterans Affairs (VA) primary care patients with uncontrolled blood pressure. We conducted thematic analysis using tools of grounded theory to identify key themes surrounding patients' explanatory models, social context and hypertension management behaviors. Patients' perceptions of the cause and course of hypertension, experiences of hypertension symptoms, and beliefs about the effectiveness of treatment were related to different hypertension self-management behaviors. Moreover, patients' daily-lived experiences, such as an isolated lifestyle, serious competing health problems, a lack of habits and routines, barriers to exercise and prioritizing lifestyle choices, also interfered with optimal hypertension self-management. Designing interventions to improve patients' hypertension self-management requires consideration of patients' explanatory models and their daily-lived experience. We propose a new conceptual model - the dynamic model of hypertension self-management behavior - which incorporates these key elements of patients' experiences.


Critical analysis of antibacterial agents in clinical development.
The antibacterial agents currently in clinical development are predominantly derivatives of well-established antibiotic classes and were selected to address the class-specific resistance mechanisms and determinants that were known at the time of their discovery. Many of these agents aim to target the antibiotic-resistant priority pathogens listed by the WHO, including Gram-negative bacteria in the critical priority category, such as carbapenem-resistant Acinetobacter, Pseudomonas and Enterobacterales. Although some current compounds in the pipeline have exhibited increased susceptibility rates in surveillance studies that depend on geography, pre-existing cross-resistance both within and across antibacterial classes limits the activity of many of the new agents against the most extensively drug-resistant (XDR) and pan-drug-resistant (PDR) Gram-negative pathogens. In particular, cross-resistance to unrelated classes may occur by co-selection of resistant strains, thus leading to the rapid emergence and subsequent spread of resistance. There is a continued need for innovation and new-class antibacterial agents in order to provide effective therapeutic options against infections specifically caused by XDR and PDR Gram-negative bacteria.


[Subtelomeric deletion 9qter: definition of the syndrome and parental origin in 2 patients].
Subtelomeric chromosome imbalances are increasingly known as a cause for mental retardation. New phenotypes associated with specific rearrangements are also being delineated, such as 9q microdeletion syndrome. Here we define the major phenotypic features and the parental origin of 9q deletion. We present 2 children with a phenotype that is characterized by mental retardation, distinctive facial features and congenital anomalies. Both patients showed a chromosome 9q subtelomeric deletion detected by MLPA (multiplex ligation-dependent probe amplification), and confirmed by FISH (fluorescent in situ hybridization). In order to delimit the size and the parental origin of 9q deletion, we performed microsatellite segregation analyses. We identified 2 patients with a de novo terminal deletion of the chromosome region 9q34. The deleted region spanned less than 0.8 and 1.5 Mb, respectively, affecting in both cases the paternal chromosome. 9q34 deletion syndrome appears as a clinically recognizable phenotype characterised by moderate-severe mental retardation, hypotonia, flat face with hyperthelorism, synophrys, anteverted nares, carp-shaped mouth with protruding tongue and conotruncal heart defects. Most de novo deletions arise in the chromosomes of paternal origin.


Analysis of high-throughput sequencing and annotation strategies for phage genomes.
Bacterial viruses (phages) play a critical role in shaping microbial populations as they influence both host mortality and horizontal gene transfer. As such, they have a significant impact on local and global ecosystem function and human health. Despite their importance, little is known about the genomic diversity harbored in phages, as methods to capture complete phage genomes have been hampered by the lack of knowledge about the target genomes, and difficulties in generating sufficient quantities of genomic DNA for sequencing. Of the approximately 550 phage genomes currently available in the public domain, fewer than 5% are marine phage. To advance the study of phage biology through comparative genomic approaches we used marine cyanophage as a model system. We compared DNA preparation methodologies (DNA extraction directly from either phage lysates or CsCl purified phage particles), and sequencing strategies that utilize either Sanger sequencing of a linker amplification shotgun library (LASL) or of a whole genome shotgun library (WGSL), or 454 pyrosequencing methods. We demonstrate that genomic DNA sample preparation directly from a phage lysate, combined with 454 pyrosequencing, is best suited for phage genome sequencing at scale, as this method is capable of capturing complete continuous genomes with high accuracy. In addition, we describe an automated annotation informatics pipeline that delivers high-quality annotation and yields few false positives and negatives in ORF calling. These DNA preparation, sequencing and annotation strategies enable a high-throughput approach to the burgeoning field of phage genomics.


Ovine fetal malformations induced by in utero inoculation with Main Drain, San Angelo, and LaCrosse viruses.
The teratogenic potential of three bunyaviruses, two California serogroup bunyaviruses, LaCrosse virus and San Angelo virus, and a Bunyamwera serogroup member, Main Drain virus, in sheep was studied following in utero inoculation of ewes in early gestation. Although Main Drain virus appeared to be most teratogenic, all three viruses induced a range of lesions including arthrogryposis, hydrocephalus, fetal death, axial skeletal deviations, anasarca, and oligohydramnios. The teratogenic effects of these viruses are identical to those described in ovine infections by Cache Valley and Akabane viruses. Demonstration of a common bunyaviral tropism for fetal tissue infection that results in congenital brain and musculoskeletal malformations provides evidence that human in utero infection by bunyaviruses could result in similar malformations in human infants.


Acne and risk of prostate cancer.
In a recent study, prostatectomy specimens from which Propionibacterium acnes was cultured were more likely to have inflammation than culture-negative specimens or specimens positive for other bacteria, leading the authors to hypothesize that P. acnes-mediated inflammation may contribute to prostate carcinogenesis. To indirectly explore associations between P. acnes and prostate cancer, we investigated severe acne, as measured by tetracycline use for 4 or more years, in relation to incident prostate cancer in the Health Professionals Follow-up Study. On the 1992 follow-up questionnaire, participants were asked whether they had ever used "tetracycline for at least 2 months at a time (e.g., for acne or other reason)" and their duration of use. Prostate cancer diagnoses were ascertained on each subsequent biennial questionnaire and confirmed by medical record review. Between 1992 and 2002, 2,147 cases of prostate cancer were reported among 34,629 eligible participants. Men who used tetracycline for 4 or more years had a significantly higher risk of prostate cancer (16 cases, 1,569 person-years) than men who did not use tetracycline (2,071 cases, 304,822 person-years, multivariable-adjusted RR = 1.70, 95% CI: 1.03-2.80). Although intriguing, this finding should be viewed cautiously because of the small number of exposed cases, indirect assessment of severe acne, and complex etiology of acne, which is not limited to P. acnes infection. Therefore, additional biologic and epidemiologic studies are necessary to determine and elucidate the possible role of P. acnes infection in prostate carcinogenesis.


A new haemocyanin in cuttlefish (Sepia officinalis) eggs: sequence analysis and relevance during ontogeny.
Haemocyanin is the respiratory protein of most of the Mollusca. In cephalopods and gastropods at least two distinct isoforms are differentially expressed. However, their physiological purpose is unknown. For the common cuttlefish Sepia officinalis, three isoforms are known so far, whereas for only two of them the complete mRNA sequences are available. In this study, we sequenced the complete mRNA of the third haemocyanin isoform and measured the relative expression of all three isoforms during embryogenesis to reveal a potential ontogenetic relevance. The cDNA of isoform 3 clearly correlates to the known Sepia officinalis haemocyanin subunits consisting of eight functional units and an internal duplicated functional unit d. Our molecular phylogenetic analyses reveal the third isoform representing a potentially ancestral haemocyanin isoform, and the analyses of the expression of haemocyanin type 3 reveal that haemocyanin type 3 only can be observed within eggs and during early development. Isoforms 1 and 2 are absent at these stages. After hatching, isoform 3 is downregulated, and isoform 1 and 2 are upregulated. Our study clearly shows an embryonic relevance of the third isoform, which will be further discussed in the light of the changes in the physiological function of haemocyanin during ontogeny. Taken together with the fact that it could also be the isoform closest related to the common ancestor of cuttlefish haemocyanin, the phylogeny of cuttlefish haemocyanin may be recapitulated during its ontogeny.


Haemophilus influenzae serotype B (Hib) seroprevalence in England and Wales in 2009.
A national seroprevalence study was performed to determine the prevalence of Haemophilus influenzae type b (Hib) antibodies in England and Wales in 2009, when Hib disease incidence was the lowest ever recorded. A total of 2,693 anonymised residual sera from routine diagnostic testing submitted by participating National Health Service hospital laboratories were tested for Hib anti-polyribosyl-ribitol phosphate (PRP) IgG antibodies using a fluorescent bead assay. Median anti-PRP IgG concentrations were highest in toddlers aged 1–4 years (2.65 μg/ml), followed by children aged 5–9 years (1.95 μg/ml). Antibody concentrations were significantly lower after this age, but were still significantly higher among 10–19 year-olds (0.54 μg/ml) compared with adults aged >20 years (0.16 μg/ ml; p<0.0001). Half of the adults (51%) did not have Hib antibody concentrations ≥0.15 μg/ml, the level considered to confer short-term protection. Thus, the current excellent Hib control appears to be the result of high anti-PRP antibody concentrations in children aged up to 10 years, achieved through the various childhood vaccination campaigns offering booster immunisation. The lack of seroprotection in adults emphasises the importance of maintaining control of the disease and, most probably carriage, in children, therefore raising the question as to whether long-term routine boosting of either pre-school children or adolescents may be required.


The road not taken: retreat and diverge in local search for simplified protein structure prediction.
Given a protein's amino acid sequence, the protein structure prediction problem is to find a three dimensional structure that has the native energy level. For many decades, it has been one of the most challenging problems in computational biology. A simplified version of the problem is to find an on-lattice self-avoiding walk that minimizes the interaction energy among the amino acids. Local search methods have been preferably used in solving the protein structure prediction problem for their efficiency in finding very good solutions quickly. However, they suffer mainly from two problems: re-visitation and stagnancy. In this paper, we present an efficient local search algorithm that deals with these two problems. During search, we select the best candidate at each iteration, but store the unexplored second best candidates in a set of elite conformations, and explore them whenever the search faces stagnation. Moreover, we propose a new non-isomorphic encoding for the protein conformations to store the conformations and to check similarity when applied with a memory based search. This new encoding helps eliminate conformations that are equivalent under rotation and translation, and thus results in better prevention of re-visitation. On standard benchmark proteins, our algorithm significantly outperforms the state-of-the art approaches for Hydrophobic-Polar energy models and Face Centered Cubic Lattice.


Involvement of SARA in Axon and Dendrite Growth.
SARA (Smad Anchor for Receptor Activation) plays a crucial role in Rab5-mediated endocytosis in cell lines localizing to early endosomes where it regulates morphology and function. Here, we analyzed the role of SARA during neuronal development and tested whether it functions as a regulator of endocytic trafficking of selected axonal and membrane proteins. Suppression of SARA perturbs the appearance of juxtanuclear endocytic recycling compartments and the neurons show long axons with large growth cones. Furthermore, surface distribution of the cell adhesion molecule L1 in axons and the fusion of vesicles containing transferring receptor (TfR) in dendrites were increased in neurons where SARA was silenced. Conversely, SARA overexpression generated large early endosomes and reduced neurite outgrowth. Taken together, our findings suggest a significant contribution of SARA to key aspects of neuronal development, including neurite formation.


Conception rates in women desiring pregnancy after levonorgestrel 52 mg intrauterine system (Liletta®) discontinuation.
Evaluate reproductive function in nulligravid and gravid women after levonorgestrel 52 mg intrauterine system (IUS) discontinuation based on time to pregnancy. We evaluated women participating in the ACCESS IUS multicenter, Phase 3, open-label clinical trial of the Liletta(®) levonorgestrel 52 mg IUS who discontinued the IUS within 60 months of use and desired pregnancy. Study staff contacted participants every three months after IUS discontinuation for up to 12 months to determine whether pregnancy occurred. We excluded women who opted to stop attempting to conceive before 12 months. We evaluated 12-month conception rates in participants 16-35 years at IUS placement, comparing dichotomous outcomes using Fisher's exact test. We performed a multivariable analysis to assess the association of baseline characteristics, age at discontinuation, duration of IUS use, and positive sexually transmitted infection testing during IUS use with conception. Among 165 women who attempted to conceive, 142 (86.1%) did so within 12 months with a median time to conception of 92 days. The 12-month conception rates did not differ between nulligravid (66/76 [86.8%]) and gravid (76/89 [85.4%]) women (p = 0.83) and nulliparous (78/90 [86.7%]) and parous (64/75 [85.3%]) women (p = 0.83). In multivariable analysis, only obesity (aOR 0.3 [95% CI 0.1-0.8]) was associated with ability to conceive. After levonorgestrel 52 mg IUS discontinuation, women have rapid return of fertility in the year post-removal. Fertility rates after IUS removal do not vary based on gravidity, parity, age at discontinuation, or duration of IUS use. This contemporary IUS study included a large population of nulligravid and nulliparous women. IUS use over many years does not effect spontaneous fertility after IUS discontinuation, regardless of gravidity or parity. Providers and patients should have no concern about the impact of IUS use on future fertility.


The DEK oncoprotein binds to highly and ubiquitously expressed genes with a dual role in their transcriptional regulation.
The DEK gene is highly expressed in a wide range of cancer cells, and a recurrent translocation partner in acute myeloid leukemia. While DEK has been identified as one of the most abundant proteins in human chromatin, its function and binding properties are not fully understood. We performed ChIP-seq analysis in the myeloid cell line U937 and coupled it with epigenetic and gene expression analysis to explore the genome-wide binding pattern of DEK and its role in gene regulation. We show that DEK preferentially binds to open chromatin, with a low degree of DNA methylation and scarce in the heterochromatin marker H3K9me(3) but rich in the euchromatin marks H3K4me(2/3), H3K27ac and H3K9ac. More specifically, DEK binding is predominantly located at the transcription start sites of highly transcribed genes and a comparative analysis with previously established transcription factor binding patterns shows a similarity with that of RNA polymerase II. Further bioinformatic analysis demonstrates that DEK mainly binds to genes that are ubiquitously expressed across tissues. The functional significance of DEK binding was demonstrated by knockdown of DEK by shRNA, resulting in both significant upregulation and downregulation of DEK-bound genes. We find that DEK binds to transcription start sites with a dual role in activation and repression of highly and ubiquitously expressed genes.


Oxygen accessibility to ribonuclease a: quantitative interpretation of nuclear spin relaxation induced by a freely diffusing paramagnet.
The nuclear spin relaxation induced by a freely diffusing paramagnetic center provides a direct measure of intermolecular accessibility. A number of factors are involved in a quantitative interpretation of relaxation data including excluded volume effects, solvation differences, and the details of the electron spin relaxation in the paramagnetic center. In the case where the electron relaxation time is short compared with correlation times describing the electron-nuclear coupling, the nuclear spin relaxation rates may be related to the effective local concentration of the paramagnetic center at different locations about the solute of interest. The local concentrations may in turn be related to differences in the local free energies of interaction between the diffusing paramagnet and the cosolute. We demonstrate this approach for the case of ribonuclease A and deduce surface free energy differences for a large number of protein proton sites. We find that the oxygen accessibility is poorly represented by hard-sphere models such as computed solvent or steric accessibility. There is a distribution of local intermolecular interactions with a width of the order of RT that dominates the report of the intermolecular exploration of the protein by this simple solute.


Are primary care services a substitute or complement for specialty and inpatient services?
To determine whether strategies designed to increase members' use of primary care services result in decreases (substitution) or increases (complementation) in the use and cost of other types of health services. Encounter and cost data were extracted from the Department of Veterans Affairs (VA) administrative data sources for the period 1995-1999. This timeframe captures the VA's natural experiment of increasing geographic access to primary care by establishing new satellite primary care clinics, known as Community-Based Outpatient Clinics (CBOCs). We exploited this natural experiment to estimate the substitutability of primary care for other health services and its impact on cost. Hypotheses were tested using ordinary least squares (OLS) regression, which was potentially subject to endogeneity bias. Endogeneity bias was assessed using a Hausman test. Endogeneity bias was accounted for by using instrumental variables analysis, which capitalized on the establishment of CBOCs to provide an exogenous identifier (change in travel distance to primary care). Demographic, encounter, and cost data were collected for all veterans using VA health services who resided in the catchment areas of new CBOCs and for a matched group of veterans residing outside CBOC catchment areas. Change in distance to primary care was a significant and substantial predictor of change in primary care visits. OLS analyses indicated that an increase in primary care service use was associated with increases in the use of all specialty outpatient services and inpatient services, as well as increases in inpatient and outpatient costs. Hausman tests confirmed that OLS results for specialty mental health encounters and mental health admissions were unbiased, but that results for specialty medical encounters, physical health admissions, and outpatient costs were biased. Instrumental variables analyses indicated that an increase in primary care encounters was associated with a decrease in specialty medical encounters and was not associated with an increase in physical health admissions, or outpatient costs. Results provide evidence that health systems can implement strategies to encourage their members to use more primary care services without driving up physical health costs.


Global participation in sport and leisure-time physical activities: A systematic review and meta-analysis.
This review aimed to determine the most popular physical activities performed by children, adolescents, and adults globally. Statistic bureau websites and article databases Scopus, ProQuest, SPORTDiscus, and Science Direct were searched between November 17th, 2014 and April 31st, 2015. Eligible studies were published in the last 10years with participation rates for specific physical activities among individuals five years or older. Data extraction for included articles (n=64) was assessed independently and agreed upon by two authors. A random-effects model was used to calculate participation rates in specific activities for each age group and region. In total 73,304 articles were retrieved and 64 articles representing 47 countries were included in the final meta-analysis. Among adults, walking was the most popular activity in the Americas (18.9%; 95% CI 10.2 to 32.5), Eastern Mediterranean (15.0%; 95% CI 5.8 to 33.6), Southeast Asia (39.3%; 95% CI 0.9 to 98.0) and Western Pacific (41.8%; 95% CI 25.2 to 60.6). In Europe and Africa, soccer (10.0%; 95% CI 6.5 to 15.1) and running (9.3%; 95% CI 0.9 to 53.9), respectively, were top activities. Child and adolescent participation results were highly dependent upon region. American youth team sport participation was high, while youth from the Eastern Mediterranean and Western Pacific were more likely to report participation in lifelong physical activities. Global data for adults reflects a consistent pattern of participation in running and walking. Among all age groups and regions soccer was popular. In children and adolescents, preferences were variable between regions.


Randomized trial comparing the safety and antibody responses to live attenuated versus inactivated influenza vaccine when administered to breastfeeding women.
Live attenuated influenza vaccine (LAIV) and inactivated influenza vaccine (IIV) are both licensed for administration to nursing mothers. Little is known about the potential for transmission of LAIV viruses from the mother to the infant and the comparative breast milk antibody responses to LAIV and IIV. We performed a randomized, double-blind study comparing the immunogenicity of LAIV to IIV when administered to nursing mothers. The safety of LAIV to IIV in women and their infants was also compared. Women received LAIV + intramuscular placebo, or IIV + intranasal placebo on Day 0. Breast milk and nasal swabs (from women and infants) were collected on Days 0, 2, and 8 for detection of LAIV. Breast milk and serum antibody responses were measured at Days 0 and 28. The primary hypothesis was that LAIV would provide superior induction of breast milk IgA responses to influenza as compared to IIV when administered to nursing mothers. Breast milk IgG, breast milk IgA (H1N1 only), serum hemagglutination inhibition (HAI), and serum IgG responses were significantly higher following administration of IIV compared to LAIV. Receipt of either LAIV or IIV was safe in women and their infants. One (1%) LAIV recipient transmitted vaccine virus to her infant who remained well. No influenza virus was detected in breast milk. Breast milk and serum antibody responses were higher for IIV compared to LAIV. LAIV and IIV were safe for nursing women but there was one (1%) possible transmission of LAIV to an infant. This study suggests that IIV may be the preferred vaccine for nursing mothers.


Using STED and ELSM confocal microscopy for a better knowledge of fused silica polished glass interface.
Characteristics and nature of close surface defects existing in fused silica polished optical surfaces were explored. Samples were deliberately scratched using a modified polishing process in presence of different fluorescent dyes. Various techniques including Epi-fluorescence Laser Scanning Mode (ELSM) or STimulated Emission Depletion (STED) confocal microscopy were used to measure and quantify scratches that are sometimes embedded under the polished layer. We show using a non-destructive technique that depth of the modified region extends far below the surface. Moreover cracks of 120 nm width can be present ten micrometers below the surface.


Mice lacking cyclin D1 are small and show defects in eye and mammary gland development.
Using homologous recombination, mice lacking cyclin D1 were generated by replacing most of the first exon of the Cyl-1 gene with sequences encoding neomycin resistance. Cyl-1(-1-) mice were viable and fertile but consistently smaller than their heterozygous or wild-type littermates. The nullizygous animals also showed two distinctive abnormalities: a severe retinopathy caused by impaired development of all layers of the retina and, in the mammary gland during pregnancy, a marked reduction in acinar development accompanied by a failure to lactate. Approximately 50% of animals also had a malformation of the jaw that manifested itself as a misalignment of the incisor teeth. Mouse embryo fibroblasts isolated from 14 day nullizygous, heterozygous, or wild-type embryos and grown under standard conditions showed similar cell-cycle and growth characteristics. Thus although cyclin D1 kinase activity may facilitate G1 progression, it is not essential for the development of most tissues and organs, and only a few specialized cell lineages are demonstrably sensitive to its absence.


Association of Airflow Limitation With Carotid Atherosclerosis in a Japanese Community - The Hisayama Study.
There has been no large-scale observational study examining the association between chronic obstructive pulmonary disease (COPD) or airflow limitation and carotid atherosclerosis in the general population across a wide range of generations in Asia. In the present study we assessed the association between airflow limitation and carotid intima-media thickness (IMT) in a general Japanese population, with consideration of a comprehensive array of cardiovascular risk factors.Methods and Results:In all, 2,099 community-dwelling Japanese subjects were included in the study. Airflow limitation was defined by spirometry. Maximum and mean IMT values were measured using carotid ultrasonography. Among the subjects, 352 (16.8%) had airflow limitation. The geometric mean values of maximum IMT and mean IMT were significantly higher in subjects with than without airflow limitation (1.27 vs. 1.18 mm, respectively, for maximum IMT; 0.73 mm vs. 0.72 mm, respectively, for mean IMT) and increased with the severity of airflow limitation after adjustment for conventional risk factors, including smoking habits and serum high-sensitivity C-reactive protein. It should be noted that the magnitude of these associations was greater in the middle-aged (40-64 years) than elderly (≥65 years) subgroup. The findings of the present study suggest that airflow limitation is a significant risk factor for carotid atherosclerosis, especially in midlife, in the general Japanese population.


Life history changes in coral fluorescence and the effects of light intensity on larval physiology and settlement in Seriatopora hystrix.
Fluorescence is common in both coral adult and larval stages, and is produced by fluorescent proteins that absorb higher energy light and emit lower energy light. This study investigated the changes of coral fluorescence in different life history stages and the effects of parental light environment on larval fluorescence, larval endosymbiotic dinoflagellate abundance, larval size and settlement in the brooding coral Seriatopora hystrix. Data showed that coral fluorescence changed during development from green in larvae to cyan in adult colonies. In larvae, two green fluorescent proteins (GFPs) co-occur where the peak emission of one GFP overlaps with the peak excitation of the second GFP allowing the potential for energy transfer. Coral larvae showed great variation in GFP fluorescence, dinoflagellate abundance, and size. There was no obvious relationship between green fluorescence intensity and dinoflagellate abundance, green fluorescence intensity and larval size, or dinoflagellate abundance and larval size. Larvae of parents from high and low light treatments showed similar green fluorescence intensity, yet small but significant differences in size, dinoflagellate abundance, and settlement. The large variation in larval physiology combined with subtle effects of parental environment on larval characteristics seem to indicate that even though adult corals produce larvae with a wide range of physiological capacities, these larvae can still show small preferences for settling in similar habitats as their parents. These data highlight the importance of environmental conditions at the onset of life history and parent colony effects on coral larvae.


Microglial TLR4-dependent autophagy induces ischemic white matter damage via STAT1/6 pathway.
Rationale: Ischemic white matter damage frequently results in myelin loss, accompanied with microglial activation. We previously found that directing microglia towards an anti-inflammatory phenotype provided a beneficial microenvironment and helped maintain white matter integrity during chronic cerebral hypoperfusion. However, the molecular mechanisms underlying microglial polarization remain elusive. Methods: Hypoperfusion induced white matter damage mice model and lipopolysaccharide (LPS) induced primary cultured microglia were established. Autophagy activation in microglia was detected both in vivo and in vitro by immunofluorescence, Western blot and electron microscopy. Autophagy inhibitors/agonist were administrated to investigate the role of autophagic process in modulating microglial phenotypes. Quantitative real time-polymerase chain reaction and Western blot were carried out to investigate the possible pathway. Results: We identified rapid accumulation of autophagosomes in primary cultured microglia exposed to LPS and within activated microglia during white matter ischemic damage. Autophagy inhibitors switched microglial function from pro-inflammatory to anti-inflammatory phenotype. Furthermore, we found TLR4, one of the major receptors binding LPS, was most highly expressed on microglia in corpus callosum during white matter ischemic damage, and TLR4 deficiency could mimic the phenomenon in microglial functional transformation, and exhibit a protective activity in chronic cerebral hypoperfusion. Whereas, the anti-inflammatory phenotype of microglia in TLR4 deficiency group was largely abolished by the activation of autophagic process. Finally, our transcriptional analysis confirmed that the up-regulation of STAT1 and down-regulation of STAT6 in microglia exposure to LPS could be reversed by autophagy inhibition. Conclusion: These results indicated that TLR4-dependent autophagy regulates microglial polarization and induces ischemic white matter damage via STAT1/6 pathway.


[Insula and disgust].
The disgust emotion is elicited by a variety of stimuli ranging from rotten food to immoral persons. When we encounter such disgusting stimuli, whether they are physical or social, we commonly experience rejection responses by the body such as nausea and revolt. In fact, since the time of Darwin, it has been argued that disgust has its origins in a rejection response to offensive food, and that the sensations of tastes and odors play a crucial role in the experience of disgust. This view predicts that the insula is closely related to disgust because it serves both gustatory and visceral motor functions including the control of vomiting. Indeed, the insula is activated by a broad range of disgust-related stimuli such as disgusted facial expressions, unpleasant odors, pictures of rotten food, and unfair acts. However, increasing evidence indicates that the insula plays an important role in the experience of not only unpleasant but also pleasant bodily feelings. In brief, the insula seems to be involved in the conscious perception of emotional bodily feelings in general, or somatic markers, and assist in our decisions as to approach vs. avoidance.


Live endoscopy events (LEEs): European Society of Gastrointestinal Endoscopy Position Statement - Update 2021.
The European Society of Gastrointestinal Endoscopy (ESGE) is dedicated to improving the quality of gastrointestinal endoscopy, including through educational activities such as live endoscopy events (LEEs). The primary goal of LEEs should be to facilitate the improvement of endoscopic patient care through the acquisition of best endoscopic practice. Patients should not expect additional benefit from being treated during a LEE compared to a routine setting. There is limited available evidence on LEE safety but to date there is no indication that patients are at increased risk from participation. Pre-recorded cases with live facilitation can also be used to fulfill learning outcomes. Establishing an endoscopic curriculum with clear learning outcomes is important to structure attendees' learning, assess course outcomes, and allow appropriate targeting of courses to learner experience. Increasingly, LEEs are streamed online and therefore the necessary measures should be taken to ensure that patients have given appropriate consent and that their anonymity has been safeguarded. ESGE recommends that an endoscopist who is not participating in the live demonstrations is named as patient advocate, and that patient safety should must be prioritized throughout. In all ESGE-organized LEEs the intended learning outcomes, procedural indications and descriptions, attendee feedback, and adverse events should be recorded and submitted in a post-event report to ESGE.


Human tankyrases are aberrantly expressed in colon tumors and contain multiple epitopes that induce humoral and cellular immune responses in cancer patients.
Tankyrases 1 and 2 are telomere-associated poly(ADP-ribose) polymerases (PARP) that can positively regulate telomere elongation and interact with multiple cellular proteins. Recent reports implicated tankyrases as tumor antigens and potential targets of anticancer treatment. We examined expression of tankyrases in colon tumors and immune response to these enzymes in patients with different types of cancer. mRNA and protein expression was evaluated by quantitative real-time RT-PCR and Western blotting, respectively. Humoral immune response to recombinant tankyrases was investigated by modified enzyme-linked immunoassays. Cellular immune response was analysed by ELISPOT and (51)Cr release assays. We found that both mRNA and protein levels of tankyrase 2 (TNKL) are upregulated in colon tumors. In contrast, protein level of tankyrase 1 (TNKS) is downregulated, while mRNA level shows variable changes. More than a quarter of colon cancer patients develop humoral immune response to at least one of the two tankyrases. In this study we mapped common and unique B-cell epitopes located in different domains of the two proteins. Additionally, we present evidence for T-cell responses both to epitopes that are unique for TNKL and to those shared between TNKL and TNKS. Our study favors a biomarker usage of antibody response to tankyrases. Spontaneous CD8(+) T-cell responses to these enzymes are rare and further investigation is needed to evaluate tankyrases as potential targets for cancer immunotherapy.


Selection against PUMA gene expression in Myc-driven B-cell lymphomagenesis.
The p53 tumor suppressor pathway limits oncogenesis by inducing cell cycle arrest or apoptosis. A key p53 target gene is PUMA, which encodes a BH3-only proapoptotic protein. Here we demonstrate that Puma deletion in the Emu-Myc mouse model of Burkitt lymphoma accelerates lymphomagenesis and that approximately 75% of Emu-Myc lymphomas naturally select against Puma protein expression. Furthermore, approximately 40% of primary human Burkitt lymphomas fail to express detectable levels of PUMA and in some tumors this is associated with DNA methylation. Burkitt lymphoma cell lines phenocopy the primary tumors with respect to DNA methylation and diminished PUMA expression, which can be reactivated following inhibition of DNA methyltransferases. These findings establish that PUMA is silenced in human malignancies, and they suggest PUMA as a target for the development of novel chemotherapeutics.


A smart and versatile theranostic nanomedicine platform based on nanoporphyrin.
Multifunctional nanoparticles with combined diagnostic and therapeutic functions show great promise towards personalized nanomedicine. However, attaining consistently high performance of these functions in vivo in one single nanoconstruct remains extremely challenging. Here we demonstrate the use of one single polymer to develop a smart 'all-in-one' nanoporphyrin platform that conveniently integrates a broad range of clinically relevant functions. Nanoporphyrins can be used as amplifiable multimodality nanoprobes for near-infrared fluorescence imaging (NIRFI), magnetic resonance imaging (MRI), positron emission tomography (PET) and dual modal PET-MRI. Nanoporphyrins greatly increase the imaging sensitivity for tumour detection through background suppression in blood, as well as preferential accumulation and signal amplification in tumours. Nanoporphyrins also function as multiphase nanotransducers that can efficiently convert light to heat inside tumours for photothermal therapy (PTT), and light to singlet oxygen for photodynamic therapy (PDT). Furthermore, nanoporphyrins act as programmable releasing nanocarriers for targeted delivery of drugs or therapeutic radio-metals into tumours.


Human gut microbes impact host serum metabolome and insulin sensitivity.
Insulin resistance is a forerunner state of ischaemic cardiovascular disease and type 2 diabetes. Here we show how the human gut microbiome impacts the serum metabolome and associates with insulin resistance in 277 non-diabetic Danish individuals. The serum metabolome of insulin-resistant individuals is characterized by increased levels of branched-chain amino acids (BCAAs), which correlate with a gut microbiome that has an enriched biosynthetic potential for BCAAs and is deprived of genes encoding bacterial inward transporters for these amino acids. Prevotella copri and Bacteroides vulgatus are identified as the main species driving the association between biosynthesis of BCAAs and insulin resistance, and in mice we demonstrate that P. copri can induce insulin resistance, aggravate glucose intolerance and augment circulating levels of BCAAs. Our findings suggest that microbial targets may have the potential to diminish insulin resistance and reduce the incidence of common metabolic and cardiovascular disorders.


A decrease in cardiovascular risk factors in healthy 40-year-old Swedish men between 1980-1983 and 1991-1992.
Cardiovascular risk factors were compared between two samples of urban middle-aged healthy men investigated in 1980-1983 (n = 106) and 1991-1992 (n = 118), respectively. All subjects, who served as controls in an ongoing study on mechanisms behind myocardial infarction, were randomly selected from a register that contains all the inhabitants (1.65 million) in the Stockholm Metropolitan Area. The study programme included recordings of weight, height, smoking habits, blood pressure and blood sampling. Blood and lipoprotein lipid levels, glucose concentrations before and during an oral glucose tolerance test and fibrinogen levels were determined. The 1991-1992 sample had lower systolic blood pressure and lower concentrations of total blood cholesterol, fasting blood glucose and fibrinogen than the 1980-1983 sample. The lower total blood cholesterol level in the 1991-1992 sample was due to a decrease in low- and high-density lipoprotein cholesterol concentrations. In addition, a tendency was seen towards a decrease in prevalence of smoking in the latter sample. No differences were noted in body mass index, diastolic blood pressure, oral glucose tolerance or total triglycerides between the two samples.


Gene flow and population structure of a common agricultural wild species (Microtus agrestis) under different land management regimes.
The impact of landscape structure and land management on dispersal of populations of wild species inhabiting the agricultural landscape was investigated focusing on the field vole (Microtus agrestis) in three different areas in Denmark using molecular genetic markers. The main hypotheses were the following: (i) organic farms act as genetic sources and diversity reservoirs for species living in agricultural areas and (ii) gene flow and genetic structure in the agricultural landscape are influenced by the degree of landscape complexity and connectivity. A total of 443 individual voles were sampled within 2 consecutive years from two agricultural areas and one relatively undisturbed grassland area. As genetic markers, 15 polymorphic microsatellite loci (nuclear markers) and the central part of the cytochrome-b (mitochondrial sequence) were analysed for all samples. The results indicate that management (that is, organic or conventional management) was important for genetic population structure across the landscape, but that landscape structure was the main factor shaping gene flow and genetic diversity. More importantly, the presence of organically managed areas did not act as a genetic reservoir for conventional areas, instead the most important predictor of effective population size was the amount of unmanaged available habitat (core area). The relatively undisturbed natural area showed a lower level of genetic structuring and genetic diversity compared with the two agricultural areas. These findings altogether suggest that political decisions for supporting wildlife friendly land management should take into account both management and landscape structure factors.


Measles virus transmembrane fusion protein synthesized de novo or presented in immunostimulating complexes is endogenously processed for HLA class I- and class II-restricted cytotoxic T cell recognition.
The routes used by antigen-presenting cells (APC) to convert the transmembrane fusion glycoprotein (F) of measles virus (MV) to HLA class I and class II presentable peptides have been examined, using cloned cytotoxic T lymphocytes in functional assays. Presentation by Epstein-Barr virus-transformed B lymphoblastoid cell lines was achieved using live virus, ultraviolet light-inactivated virus, and purified MV-F delivered either as such or incorporated in immunostimulating complexes (MV-F-ISCOM). Only live virus and MV-F-ISCOM allow presentation by class I molecules, while all antigen preparations permit class II-restricted presentation. We observe presentation of MV-F from live virus and as MV-F-ISCOM by class II molecules in a fashion that is not perturbed by chloroquine. Our studies visualize novel presentation pathways of type I transmembrane proteins.


High-throughput screening of tick-borne pathogens in Europe.
Due to increased travel, climatic, and environmental changes, the incidence of tick-borne disease in both humans and animals is increasing throughout Europe. Therefore, extended surveillance tools are desirable. To accurately screen tick-borne pathogens (TBPs), a large scale epidemiological study was conducted on 7050 Ixodes ricinus nymphs collected from France, Denmark, and the Netherlands using a powerful new high-throughput approach. This advanced methodology permitted the simultaneous detection of 25 bacterial, and 12 parasitic species (including; Borrelia, Anaplasma, Ehrlichia, Rickettsia, Bartonella, Candidatus Neoehrlichia, Coxiella, Francisella, Babesia, and Theileria genus) across 94 samples. We successfully determined the prevalence of expected (Borrelia burgdorferi sensu lato, Anaplasma phagocytophilum, Rickettsia helvetica, Candidatus Neoehrlichia mikurensis, Babesia divergens, Babesia venatorum), unexpected (Borrelia miyamotoi), and rare (Bartonella henselae) pathogens in the three European countries. Moreover we detected Borrelia spielmanii, Borrelia miyamotoi, Babesia divergens, and Babesia venatorum for the first time in Danish ticks. This surveillance method represents a major improvement in epidemiological studies, able to facilitate comprehensive testing of TBPs, and which can also be customized to monitor emerging diseases.


Bilateral tactile hypersensitivity and neuroimmune responses after spared nerve injury in mice lacking vasoactive intestinal peptide.
Vasoactive intestinal peptide (VIP) is one of the neuropeptides showing the strongest up-regulation in the nociceptive pathway after peripheral nerve injury and has been proposed to support neuropathic pain. Nevertheless, the story may be more complicated considering the known suppressive effects of the peptide on the immune reactivity of microglial cells, which have been heavily implicated in the onset and maintenance of pain after nerve injury. We here used mice deficient in VIP and the model of spared nerve injury, characterized by persistent tactile hypersensitivity. While tactile hypersensitivity developed similarly to wild type mice for the ipsilateral hindpaw, only transgenic mice showed a mirror-image tactile hypersensitivity in the contralateral hindpaw. This exacerbated neuropathic pain phenotype appeared to be mediated through a local mechanism acting at the level of the lumbar spinal cord as a distant nerve lesion in the front limb did not lead to hindpaw hypersensitivity in VIP-deficient mice. Innocuous tactile hindpaw stimulation was found to increase a neuronal activation marker in the bilateral superficial laminae of the lumbar dorsal horn of VIP-deficient, but not wild type mice, after SNI. A deeper study into the immune responsiveness to the nerve lesion also proved that VIP-deficient mice had a stronger early pro-inflammatory cytokine response and a more pronounced microglial reactivity compared to wild type controls. The latter was also observed at four weeks after spared nerve injury, a time at which bilateral tactile hypersensitivity persisted in VIP-deficient mice. These data suggest an action of VIP in neuropathic states that is more complicated than previously assumed. Future research is now needed for a deeper understanding of the relative contribution of receptors and fiber populations involved in the VIP-neuropathic pain link.


Biosimilars - terms of use.
The impending expiry of the patent on a number of leading biologic drugs has led to a surge in the development of 'biosimilar' or 'follow-on' products. However, in contrast to generic small-molecule medicines, biosimilars are not identical to their reference products. The differences and complexities surrounding both the molecular structure and the manufacturing process for biologics and biosimilars have resulted in a lack of clarity regarding the terms used in different parts of the world to define various aspects of development and utilization such as regulatory approval, pharmacovigilance, interchangeability and treatment-naivety. This makes quantitative evaluation of biosimilars a great challenge to both the scientific community and regulatory agencies. This manuscript attempts to clarify the terms used and address an important knowledge gap which is currently resulting in an increasing rush to position biosimilars for certain indications and patients in the absence of agreed upon definitions.


Branching morphogenesis of human mammary epithelial cells in collagen gels.
To study the morphogenesis of human epithelial cells in vitro we have used a three-dimensional collagen matrix and a newly developed mammary epithelial cell line, 1-7 HB2. In standard medium 1-7 HB2 cells formed compact balls/spheres inside collagen type I gels, while cocultivation with various fibroblast cell lines or growth in fibroblast-conditioned media resulted in the appearance of branching structures. At least two different soluble factors secreted by fibroblasts were found to be implicated in the branching morphogenesis. Firstly, hepatocyte growth factor/scatter factor could induce branching in a concentration-dependent manner. Moreover, a polyclonal serum against hepatocyte growth factor/scatter factor completely inhibited the branching morphogenesis induced by medium conditioned by MRC-5 fibroblast cells. In contrast, a morphogenetic activity secreted by human foreskin fibroblasts was identified that appears to be different from hepatocyte growth factor/scatter factor and from a number of other well-characterized growth factors or cytokines. This model system has been used to examine the role of integrins in mammary morphogenesis. The expression of the alpha 2 beta 1, alpha 3 beta 1 and alpha 6 beta 4 integrins was decreased when cells were plated on collagen gels. The addition of specific blocking monoclonal antibodies directed to the alpha 2- and beta 1-integrin subunits to growth media impaired cell-cell interactions and interfered with the formation of compact structures inside collagen gels, suggesting that the alpha 2 beta 1 integrin can control intercellular adhesion in mammary morphogenesis. In contrast one of the blocking monoclonal antibodies against the alpha 3-integrin subunit (P1B5) mimicked the effect of soluble 'morphogens'. Our results suggest that the modulation of alpha 3 beta 1 activity may represent an important event in the induction of branching morphogenesis of human mammary epithelial cells.


CikA, an Input Pathway Component, Senses the Oxidized Quinone Signal to Generate Phase Delays in the Cyanobacterial Circadian Clock.
The circadian clock is a timekeeping system in most organisms that keeps track of the time of day. The rhythm generated by the circadian oscillator must be constantly synchronized with the environmental day/night cycle to make the timekeeping system truly advantageous. In the cyanobacterial circadian clock, quinone is a biological signaling molecule used for entraining and fine-tuning the oscillator, a process in which the external signals are transduced into biological metabolites that adjust the phase of the circadian oscillation. Among the clock proteins, the pseudo-receiver domain of KaiA and CikA can sense external cues by detecting the oxidation state of quinone, a metabolite that reflects the light/dark cycle, although the molecular mechanism is not fully understood. Here, we show the antagonistic phase shifts produced by the quinone sensing of KaiA and CikA. We introduced a new cyanobacterial circadian clock mixture that includes an input component in vitro. KaiA and CikA cause phase advances and delays, respectively, in this circadian clock mixture in response to the quinone signal. In the entrainment process, oxidized quinone modulates the functions of KaiA and CikA, which dominate alternatively at day and night in the cell. This in turn changes the phosphorylation state of KaiC-the central oscillator in cyanobacteria-ensuring full synchronization of the circadian clock. Moreover, we reemphasize the mechanistic input functionality of CikA, contrary to other reports that focus only on its output action.


Longitudinal trends in physical activity patterns in selected urban south Indian school children.
There are very few studies describing the pattern of physical activity of children in India. This study was carried out to document patterns of physical activity in south Indian school children aged 8 to 15 yr and examine changes over a one year period. Physical activity was assessed using interviewer-administered questionnaires at baseline (n=256) and at follow up (n=203) in 2006 and 2007. Frequency and duration of each activity was recorded and metabolic equivalents (MET) assigned. Sedentary activity included activities with MET < 1.5, and moderate-to- vigorous physical activity (MVPA) with >3.0. For each activity, daily duration, intensity (MET), and the product of the two (MET-minutes) were computed. Children were categorized by age group, gender and socio-economic status. Height and weight were measured. At baseline, sedentary activity was higher in children aged >11 yr, while intensity of MVPA was higher in boys than girls. Over one year, physical activity at school significantly decreased (P<0.001). There was also a significant decrease in MVPA MET-min (P<0.001) with interaction effects of age group (P<0.001) and gender (P<0.001). There was a significant decline in moderate-to-vigorous physical activity over a single year follow up, largely due to a decrease in physical activity at school. There appears to be a gap between State educational policies that promote physical well-being of school-going children and actual practice.