Zwitterionic HILIC tandem mass spectrometry with isotope dilution for rapid, sensitive and robust quantification of pyridine nucleotides in biological extracts.
The pyridine nucleotides nicotineamide adenine dinucleotide (NAD) and nicotineamide adenine dinucleotide phosphate (NADP) are conserved coenzymes across all domains of life, and are involved in more than 200 different hydride transfer reactions supporting essential catabolic and anabolic functions. The intracellular levels of these metabolites, and the ratio of their oxidized to reduced forms regulate an extensive network of reactions ranging beyond metabolism. Hence, monitoring their intracellular levels provides information about, but not limited to, the metabolic state of a cell or tissue. Interconversion between oxidized and reduced forms, varying pH liability and varying intracellular concentrations of the different species leaves absolute quantification of the pyridine nucleotides analytically challenging. These polar metabolites are poorly retained on conventional reverseed-phase stationary phases without ion-pair reagents that contaminates the LC-system. Herein we demonstrate that zwitterionic HILIC-tandem mass spectroemtry can be applied to successfully resolve the pyridine nucleotides in biological extracts in a fast, robust and highly sensitive way. The presented method applies isotope dilution to compensate potential loss of these labile metabolites and is validated for low, medium and high biomass samples of two popular biological model systems; Escherichia coli and the human cell line JJN-3. High stability and rapid sample preparation without solvent removal allows for long sequence runs, making this method ideal for high-throughput analysis of biological extracts.


Interdisciplinary treatment for an adult patient with anterior open bite, severe periodontitis, and intellectual disability.
This case report describes the beneficial effects of the interdisciplinary treatment of an adult patient with severe skeletal periodontal problems. A 30-year-old female patient presented with anterior open bite, gummy smile, and facial asymmetry. The patient had chronic generalized severe periodontitis with pathologic maxillary anterior teeth migration and mild intellectual disability. Treatment included 6 months of periodontal treatment, followed by presurgical orthodontic treatment, a Le Fort I osteotomy with anterior segmental osteotomy, a bilateral sagittal split ramus osteotomy, and postsurgical orthodontic treatment. After treatment completion, the patient exhibited functional and aesthetic improvements. Her periodontal condition improved and was maintained after the treatment. Here, we demonstrate a successful treatment outcome in a complicated case following a systematic interdisciplinary approach performed with the correct diagnosis and treatment planning.


Hemodynamic characteristics associated with cerebral aneurysm formation in patients with carotid occlusion.
The pathogenesis of cerebral aneurysms in patients with internal carotid artery (ICA) occlusion is hypothesized to be hemodynamic. For the first time, the authors quantify the hemodynamic characteristics associated with aneurysm formation in patients with ICA occlusion. Records of patients with unilateral ICA stenosis or occlusion ≥ 90% who underwent hemodynamic assessment before treatment using quantitative MR angiography were retrospectively reviewed. The patients were classified into 2 groups based on the presence or absence of aneurysms. The hemodynamic parameters of flow volume rate, flow velocity, and wall shear stress (WSS) were measured in each vessel supplying collateral flow-bilateral A1 segments and bilateral posterior communicating arteries-and then compared between the groups. A total of 36 patients were included (8 with and 28 without aneurysms). The mean flow (72.3 vs 48.9 ml/min, p = 0.10), flow velocity (21.1 vs 12.7 cm/sec, p = 0.006), and WSS (22.0 vs 12.3 dynes/cm2, p = 0.003) were higher in the A1 segment contralateral to the side of the patent ICA in patients with versus without aneurysms. All de novo or growing aneurysms in our cohort were located on the anterior communicating artery (ACoA) or P1 segment. Flow velocity and WSS are significantly higher across the ACoA in patients who harbor an aneurysm, and de novo or growing aneurysms are often located on collateral vessels. Thus, robust primary collaterals after ICA occlusion may be a contributing factor in cerebral aneurysm formation.


Relationship of at Admission Lactate, Unmeasured Anions, and Chloride to the Outcome of Critically Ill Patients.
To investigate the association between the concentration of the causative anions responsible for the main types of metabolic acidosis and the outcome. Prospective observational study. Teaching ICU. All patients admitted from January 2006 to December 2014. None. Four thousand nine hundred one patients were admitted throughout the study period; 1,609 met criteria for metabolic acidosis and 145 had normal acid-base values. The association between at admission lactate, unmeasured anions, and chloride concentration with outcome was assessed by multivariate analysis in the whole cohort and in patients with metabolic acidosis. We also compared the mortality of patients with lactic, unmeasured anions, and hyperchloremic metabolic acidosis with that of patients without acid-base disorders. In the whole population, increased lactate and unmeasured anions were independently associated with increased mortality, even after adjusting for potential confounders (odds ratio [95% CI], 1.14 (1.08-1.20); p < 0.0001 and 1.04 (1.02-1.06); p < 0.0001, respectively). In patients with metabolic acidosis, the results were similar. Patients with lactic and unmeasured anions acidosis, but not those with hyperchloremic acidosis, had an increased mortality compared to patients without alterations (17.7%, 12.7%, 4.9%, and 5.8%, respectively; p < 0.05). In this large cohort of critically ill patients, increased concentrations of lactate and unmeasured anions, but not chloride, were associated with increased mortality. In addition, increased unmeasured anions were the leading cause of metabolic acidosis.


Elucidating the role of Cyclooxygenase-2 in the pathogenesis of oral lichen planus - an immunohistochemical study with supportive histochemical analysis.
Oral lichen planus (OLP) is a chronic, inflammatory disorder that affects the oral mucous membrane. During an inflammatory response, several chemokines and cytokines are released by the cells of the immune system. Activation of MMPs, along with mast cell-derived chymase and tryptase, degrades the basement membrane structural proteins, resulting in basement membrane breaks. To investigate the association between the COX-2 expressions, presence of intact or degranulating mast cells within the connective tissue and the extent of basement membrane discontinuity in OLP cases. This study included a total of 50 formalin-fixed paraffin-embedded specimens (FFPE) of histologically confirmed cases of idiopathic oral lichen planus. A retrospective cross-sectional analysis was carried out by immunohistochemistry to study the epithelial expression of COX-2 and by the use of special stains such as toluidine blue and periodic acid-Schiff (PAS) to study the mast cell count and basement membrane changes in the oral mucosal tissue, respectively. There was a significant (P = 0.03) association between the COX-2 expressions and mast cell count. As the intensity of COX-2 expression increased from mild to moderate or severe, the number of mast cell count almost doubled. Interaction between upregulation of COX-2, mast cell and basement membrane sets a vicious cycle which relates to the chronic nature of the disease. Inhibitors of COX-2 may reduce the inflammatory process preceding the immune dysregulation in OLP.


The effects of clinical aromatherapy for anxiety and depression in the high risk postpartum woman - a pilot study.
The aim of this study was to determine if aromatherapy improves anxiety and/or depression in the high risk postpartum woman and to provide a complementary therapy tool for healthcare practitioners. The pilot study was observational with repeated measures. Private consultation room in a Women's center of a large Indianapolis hospital. 28 women, 0-18 months postpartum. The treatment groups were randomized to either the inhalation group or the aromatherapy hand m'technique. Treatment consisted of 15 min sessions, twice a week for four consecutive weeks. An essential oil blend of rose otto and lavandula angustifolia @ 2% dilution was used in all treatments. The non-randomized control group, comprised of volunteers, was instructed to avoid aromatherapy use during the 4 week study period. Allopathic medical treatment continued for all participants. All subjects completed the Edinburgh Postnatal Depression Scale (EPDS) and Generalized Anxiety Disorder Scale (GAD-7) at the beginning of the study. The scales were then repeated at the midway point (two weeks), and at the end of all treatments (four weeks). Analysis of Variance (ANOVA) was utilized to determine differences in EPDS and/or GAD-7 scores between the aromatherapy and control groups at baseline, midpoint and end of study. No significant differences were found between aromatherapy and control groups at baseline. The midpoint and final scores indicated that aromatherapy had significant improvements greater than the control group on both EPDS and GAD-7 scores. There were no adverse effects reported. The pilot study indicates positive findings with minimal risk for the use of aromatherapy as a complementary therapy in both anxiety and depression scales with the postpartum woman. Future large scale research in aromatherapy with this population is recommended.


Complications following endoscopic submucosal dissection for gastric, esophageal, and colorectal cancer: a review of studies based on nationwide large-scale databases.
Endoscopic submucosal dissection (ESD) is a relatively new procedure used for the treatment of early gastrointestinal cancers regardless of the lesion size and configuration, and it has gradually acquired popularity because of its minimally invasive nature. As compared to conventional endoscopic resection, ESD is a more complex procedure and requires a higher level of technical skill. Therefore, it is associated with a higher complication rate. Many previous studies that investigated the complication rates following ESD analyzed data from a limited number of specialized centers, possibly leading to an underestimation of the complication rates. Further, the relationship between hospital volume and complication rates is poorly understood. In the present study, we searched the MEDLINE and the Cochrane Library databases for studies that have reported on ESD-related complications and the relationship between hospital volume and ESD-related complication rates in a nationwide setting. The complication rates (including perforation, peritonitis, and bleeding) were 3.5% for gastric ESD, 3.3% for esophageal ESD, and 4.6% for colorectal ESD. The studies reviewed showed that ESD-related complication rates were permissibly low, and that there was a linear association between a higher hospital volume and a lower frequency of complications following ESD.


Combined lung and brain ultrasonography for an individualized "brain-protective ventilation strategy" in neurocritical care patients with challenging ventilation needs.
When intracranial hypertension and severe lung damage coexist in the same clinical scenario, their management poses a difficult challenge, especially as concerns mechanical ventilation management. The needs of combined lung and brain protection from secondary damage may conflict, as ventilation strategies commonly used in patients with ARDS are potentially associated with an increased risk of intracranial hypertension. In particular, the use of positive end-expiratory pressure, recruitment maneuvers, prone positioning, and protective lung ventilation can have undesirable effects on cerebral physiology: they may positively or negatively affect intracranial pressure, based on the final repercussions on PaO2 and cerebral perfusion pressure (through changes in cardiac output, mean arterial pressure, venous return, PaO2 and PaCO2), also according to the baseline conditions of cerebral autoregulation. Lung ultrasound (LUS) and brain ultrasound (BUS, as a combination of optic nerve sheath diameter assessment and cerebrovascular Doppler ultrasound) have independently proven their potential in respectively monitoring lung aeration and brain physiology at the bedside. In this narrative review, we describe how the combined use of LUS and BUS on neurocritical patients with demanding mechanical ventilation needs can contribute to ventilation management, with the aim of a tailored "brain-protective ventilation strategy."


Ultrastructural and Single-Cell-Level Characterization Reveals Metabolic Versatility in a Microbial Eukaryote Community from an Ice-Covered Antarctic Lake.
The McMurdo Dry Valleys (MCM) of southern Victoria Land, Antarctica, harbor numerous ice-covered bodies of water that provide year-round liquid water oases for isolated food webs dominated by the microbial loop. Single-cell microbial eukaryotes (protists) occupy major trophic positions within this truncated food web, ranging from primary producers (e.g., chlorophytes, haptophytes, and cryptophytes) to tertiary predators (e.g., ciliates, dinoflagellates, and choanoflagellates). To advance the understanding of MCM protist ecology and the roles of MCM protists in nutrient and energy cycling, we investigated potential metabolic strategies and microbial interactions of key MCM protists isolated from a well-described lake (Lake Bonney). Fluorescence-activated cell sorting (FACS) of enrichment cultures, combined with single amplified genome/amplicon sequencing and fluorescence microscopy, revealed that MCM protists possess diverse potential metabolic capabilities and interactions. Two metabolically distinct bacterial clades (Flavobacteria and Methylobacteriaceae) were independently associated with two key MCM lake microalgae (Isochrysis and Chlamydomonas, respectively). We also report on the discovery of two heterotrophic nanoflagellates belonging to the Stramenopila supergroup, one of which lives as a parasite of Chlamydomonas, a dominate primary producer in the shallow, nutrient-poor layers of the lake. Single-cell eukaryotes called protists play critical roles in the cycling of organic matter in aquatic environments. In the ice-covered lakes of Antarctica, protists play key roles in the aquatic food web, providing the majority of organic carbon to the rest of the food web (photosynthetic protists) and acting as the major consumers at the top of the food web (predatory protists). In this study, we utilized a combination of techniques (microscopy, cell sorting, and genomic analysis) to describe the trophic abilities of Antarctic lake protists and their potential interactions with other microbes. Our work reveals that Antarctic lake protists rely on metabolic versatility for their energy and nutrient requirements in this unique and isolated environment.


SU-E-I-107: Suitability of Various Radiation Detectors Used in Radiation Therapy for X-Ray Dosimetry in Computed Tomography.
Assessment of suitability for X-ray dosimetry in computed tomography of various ionization chambers, diodes and two-dimensional detector arrays primarily used in radiation therapy. An Oldelft X-ray simulation unit was used to irradiate PTW 60008, 60012 dosimetry diodes, PTW 23332, 31013, 31010, 31006 axial symmetrical ionization chambers, PTW 23343, 34001 plane parallel ionization chambers and PTW Starcheck and 2D-Array seven29 as well as a prototype Farmer chamber with a copper wall. Peak potential was varied from 50 kV up to 125 kV and beam qualities were quantified through half-value-layer measurements. Energy response was investigated free in air as well as in 2 cm depth in a solid water phantom and refers to a manufacturer calibrated PTW 60004 diode for kV-dosimetry. The thimble ionization chambers PTW 31010, 31013, the uncapsuled diode PTW 60012 and the PTW 2D-Array seven29 exhibit an energy response deviation in the investigated energy region of approximately 10% or lower thus proving good usability in X-ray dosimetry if higher spatial resolution is needed or rotational irradiations occur. It could be shown that in radiation therapy routinely used detectors are usable in a much lower energy region. The rotational symmetry is of advantage in computed tomography dosimetry and enables dose profile as well as point dose measurements in a suitable phantom for estimation of organ doses. Additional the PTW 2D-Array seven29 can give a quick overview of radiation fields in non-rotating tasks.


COVID-19 Special Column: Principles Behind the Technology for Detecting SARS-CoV-2, the Cause of COVID-19.
Nationwide shortages of tests that detect severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and diagnose coronavirus disease 2019 (COVID-19) have led the US Food and Drug Administration (FDA) to significantly relax regulations regarding COVID-19 diagnostic testing. To date the FDA has given emergency use authorization (EUA) to 48 COVID-19 in vitro diagnostic tests and 21 high complexity molecular-based laboratory developed tests, as well as implemented policies that give broad authority to clinical laboratories and commercial manufacturers in the development, distribution, and use of COVID-19 diagnostic tests. Currently, there are 2 types of diagnostic tests available for the detection of SARS-CoV-2: (1) molecular and (2) serological tests. Molecular detection of nucleic acid (RNA or DNA) sequences relating to the suspected pathogen is indicative of an active infection with the suspected pathogen. Serological tests detect antibodies against the suspected pathogen, which are produced by an individual's immune system. A positive serological test result indicates recent exposure to the suspected pathogen but cannot be used to determine if the individual is actively infected with the pathogen or immune to reinfection. In this article, the SARS-CoV-2 diagnostic tests currently approved by the FDA under EUA are reviewed, and other diagnostic tests that researchers are developing to detect SARS-CoV-2 infection are discussed.


Roles of NMDARs in maintenance of the mouse cerebrovascular endothelial cell-constructed tight junction barrier.
Glutamate can activate NMDA receptor (NMDAR) and subsequently induces excitotoxic neuron loss. However, roles of NMDARs in the blood-brain barrier (BBB) are little known. This study used a mouse cerebrovascular endothelial cell (MCEC) model to evaluate the effects of NMDAR activation on maintenance of the BBB and its possible mechanisms. Analysis of confocal microscopy revealed expressions of NMDAR subunits, GluN1 and GLUN2B, in MCECs. An immunoblot assay further showed the existence of GluN1 in plasma membranes of MCECs. In brain tissues, a confocal microscopic analysis demonstrated co-localization of GluN1 and factor VIII, a biomarker of MCECs. In addition, GluN1 mRNA was detected in MCECs and the brain. Functional assays showed that exposure of MCECs to NMDA increased calcium influx. Separately, NMDA suppressed transendothelial electrical resistance values, levels of occludin, and occludin tight junctions. As to the mechanism, NMDA stimulated sequential phosphorylations of extracellular signal-regulated kinase (ERK)1/2 and mitogen-activated ERK (MEK)1. Interestingly, amounts of matrix metalloproteinase (MMP)2 and MMP9 in MCECs were augmented by NMDA. The NMDA-induced alterations in ERK1/2 phosphorylation and occludin levels were reversed by pretreatment with PD98059, a MEK inhibitor, and MK-801, a NMDAR antagonist, respectively. Therefore, this study shows the functional presence of NMDARs in MCECs, and NMDAR activation can disrupt the MCEC-constructed tight junction barrier via activation of the MEK1/2-ERK1/2 signaling pathway and upregulation of MMP2/9 expressions.


A systems biology approach to explore the impact of maple tree dormancy release on sap variation and maple syrup quality.
Maple sap is a complex nutrient matrix collected during spring to produce maple syrup. The characteristics of sap change over the production period and its composition directly impacts syrup quality. This variability could in part be attributed to changes in tree metabolism following dormancy release, but little is known about these changes in deciduous trees. Therefore, understanding the variation in sap composition associated with dormancy release could help pinpoint the causes of some defects in maple syrup. In particular, a defect known as "buddy", is an increasing concern for the industry. This off-flavor appears around the time of bud break, hence its name. To investigate sap variation related to bud break and the buddy defect, we monitored sap variation with respect to a dormancy release index (Sbb) and syrup quality. First, we looked at variation in amino acid content during this period. We observed a shift in amino acid relative proportions associated with dormancy release and found that most of them increase rapidly near the point of bud break, correlating with changes in syrup quality. Second, we identified biological processes that respond to variation in maple sap by performing a competition assay using the barcoded Saccharomyces cerevisiae prototroph deletion collection. This untargeted approach revealed that the organic sulfur content may be responsible for the development of the buddy off-flavor, and that dormancy release is necessary for the appearance of the defect, but other factors such as microbial activity may also be contributing.


Options for a Health System Researcher to Choose in Meta Review (MR) Approaches-Meta Narrative (MN) and Meta Triangulation (MT).
Two new approaches in systematic reviewing i.e. Meta-narrative review(MNR) (which a health researcher can use for topics which are differently conceptualized and studied by different types of researchers for policy decisions) and Meta-triangulation review(MTR) (done to build theory for studying multifaceted phenomena characterized by expansive and contested research domains) are ready for penetration in an arena of health system research. So critical look at which approach in Meta-review is better i.e. Meta-narrative review or Meta-triangulation review, can give new insights to a health system researcher. A systematic review on 2 key words-"meta-narrative review" and "meta-triangulation review" in health system research, were searched from key search engines, such as Pubmed, Cochrane library, Bio-med Central and Google Scholar etc till 21st March 2014 since last 20 years. Studies from both developed and developing world were included in any form and scope to draw final conclusions. However unpublished data from thesis was not included in systematic review. Meta-narrative review is a type of systematic review which can be used for a wide range of topics and questions involving making judgments and inferences in public health. On the other hand Meta-triangulation review is a three-phased, qualitative meta-analysis process which can be used to explore variations in the assumptions of alternative paradigms, gain insights into these multiple paradigms at one point of time and addresses emerging themes and the resulting theories.


A structural model of the anaphase promoting complex co-activator (Cdh1) and in silico design of inhibitory compounds.
Anaphase promoting complex (APC) controls cell cycle and chromosome segregation. The APC activation occurs after binding of co-activators, cdh1 and cdc20. Cdh1 plays a role in cancer pathogenesis and is known as a potential drug target. The main aim of this study was prediction of 3D structure of cdh1 and designing the inhibitory compounds based on the structural model. First, 3D structure of cdh1 was predicted by means of homology modelling and molecular dynamics tools, MODELLER and Gromacs package, respectively. Then, inhibitory compounds were designed using virtual screening and molecular docking by means AutoDock package. The overall structure of cdh1 is propeller like and each DW40 repeat contains four anti-parallel beta-sheets. Moreover, binding pocket of the inhibitory compounds was determined. The results might be helpful in finding a suitable cdh1 inhibitor for the treatment of cancer.


An exon 4 mutation identified in the majority of South African familial hypercholesterolaemics.
The prevalence of familial hypercholesterolaemia (FH) is significantly higher in the Afrikaans speaking population (Afrikaners) of South Africa than reported in most other populations. A founder gene effect has been proposed to explain the high FH frequency, implying that the same low density lipoprotein (LDL) receptor gene defect is present in the majority of affected Afrikaners. By using DNA amplification and sequence determination, we have detected a point mutation in DNA from two Afrikaner FH homozygotes. A cytosine to guanine base substitution at nucleotide position 681 of the LDL receptor cDNA results in an amino acid change from aspartic acid to glutamic acid at residue 206 in the cysteine rich ligand binding domain of the LDL receptor. Since three previously mapped transport deficient alleles of the LDL receptor were also traced to cysteine rich repeats of the protein, these results suggest that the mutation is responsible for the receptor defective mutation predominantly found in Afrikaner FH homozygotes. The mutation gives rise to an additional DdeI restriction site in DNA of affected subjects and segregation of the mutation with the disease was confirmed in five large Afrikaner FH families. We predict that 65% of affected South African Afrikaners carry this particular base substitution. Amplification of genomic DNA, using the polymerase chain reaction method, and restriction enzyme analysis now permit accurate diagnosis of the mutation in subjects with FH.


Hibernating bears conserve muscle strength and maintain fatigue resistance.
Black bears spend several months each winter confined to a small space within their den without food or water. In nonhibernating mammals, these conditions typically result in severe muscle atrophy, causing a loss of strength and endurance. However, an initial study indicated that bears appeared to conserve strength while denning. We conducted an in vivo, nonsubjective measurement of strength, resistance to fatigue, and contractile properties on the tibialis anterior muscle of six hibernating bears during both early and late winter using a rigid leg brace and foot force plate. After 110 d of anorexia and confinement, skeletal muscle strength loss in hibernating bears was about one-half that in humans confined to bed rest. Bears lost 29% of muscle strength over 110 d of denning without food, while humans on a balanced diet but confined to bed for 90 d have been reported to lose 54% of their strength. Additionally, muscle contractile properties, including contraction time, half-relaxation time, half-maximum value time, peak rate of development and decay, time to peak force development, and time to peak force decay did not change, indicating that no small-scale alterations in whole-muscle function occurred over the winter. This study further supports our previous findings that black bears have a high resistance to atrophy despite being subjected to long-term anorexia and limited mobility.


Immunogenicity and safety of an acellular pertussis, diphtheria, tetanus, inactivated poliovirus, Hib-conjugate combined vaccine (Pentaxim) and monovalent hepatitis B vaccine at 6, 10 and 14 weeks of age in infants in South Africa.
To assess the immunogenicity and safety data for a pentavalent combination vaccine containing acellular pertussis, inactivated poliovirus, and Haemophilus influenzae (Hib) polysaccharide-conjugate antigens. A DTaP-IPV//PRP T vaccine (Pentaxim) was given at 6, 10 and 14 weeks of age to 212 infants in South Africa. Monovalent hepatitis B vaccine was given concomitantly. Immunogenicity was assessed using seroprotection and seroconversion rates; safety was assessed by monitoring for solicited injection site and systemic adverse events, and follow-up monitoring for unsolicited adverse events and serious adverse events. Immunogenicity was high for each vaccine antigen, and similar to a reference study done in France using a similar (2, 3 and 4 months of age) administration schedule. After the third dose, 94.6% of participants had anti-PRP > or = 0.15 microg/ml. The anti-PRP geometric mean antibody titre (GMT) was 2.0 microg/ml. The seroprotection rates for diphtheria and tetanus (> or = 0.01 IU/ml), poliovirus types 1, 2 and 3 (> or = 8 1/dil U) and hepatitis B were all 100%. Anti-polio GMTs were very high, 1 453, 1 699 and 2 398 (1/dil U) for types 1, 2 and 3, respectively. The seroconversion/vaccine response rates to pertussis antigens (4-fold increase in antibody concentration) were 97.5% for PT and 83.9% for FHA. The DTaP-IPV//PRP T vaccine was highly immunogenic at 6, 10 and 14 weeks of age in infants in South Africa, was compatible with the monovalent hepatitis B vaccine, and was well tolerated.


Platelet aggregation inhibitors in hot water extract of green tea.
The effect of hot water extract of green tea on the collagen-induced aggregation of washed rabbit platelets was examined. The extract lowered submaximal aggregation and prolonged the lag time in a dose-dependent manner. After fractionation of the extract, it was revealed that the tea catechins (tannins) are active principles for inhibition and that ester-type catechins are more effective than free-type catechins. One of the ester type catechins, epigallocatechin gallate (EGCG), suppressed the collagen-induced platelet aggregation completely at the concentration of 0.2 mg/ml (= 0.45 mM). Comparing IC50 values of EGCG and aspirin it was found that the potency of EGCG is comparable to that of aspirin. Thrombin- and platelet activating factor (PAF)-induced aggregation was also inhibited by EGCG. The elevation of cyclic adenosine 3',5'-monophosphate (cAMP) level was not observed in EGCG treated platelets.


The epidemiology of nematode infections in sheep in a cool tropical environment.
The epidemiology of nematode infections in Menz sheep was studied in the highlands of Ethiopia at the International Livestock Research Institute (ILRI), Debre Berhan Research Station, using a series of tracer lambs grazing contaminated pasture for either 4, 16, 32 or 48 weeks from July 1992 to June 1994. The basic nematode seasonal infectivity pattern was expressed in terms of relative numbers of third-stage larvae (L3) available on pasture for different months. Data from faecal nematode egg counts, pasture larval recoveries and worm counts from the tracer lambs were used to investigate the infectivity pattern. Four nematode species of economic importance: Longistrongylus (Pseudomarshallagia) elongata, Trichostronglylus colubriformis, Haemonchus contortus, and Dictyocaulus filaria, were recovered from sheep. The largest numbers were recovered during the wet season (i.e. July to November) with peaks in late August and early September. During this 2 year study period, the seasonal pattern of sheep gastrointestinal nematodes was clearly defined. An important finding was that conditions during the short rainy season (i.e. March-May) were not conducive to the development and survival of nematode eggs and the free-living stages, hence little or no transmission occurred. Rainfall and humidity seemed to be the most important factors for the development of eggs and free-living stages. The period of acquisition of third-stage larvae from pasture was found to be relatively short, suggesting that a strategic control programme is feasible with minimal anthelmintic use. Results from the studies are discussed in relation to control strategies.


Reliability and validity of the Alcohol Marketing Assessment Rating Tool (AMART).
Current national responses to the monitoring and prevention of alcohol marketing to vulnerable populations, such as youth, typically rely on partial bans, co-regulation and industry self-regulation. Efforts to evaluate compliance with existing regulations are often cumbersome and resource intensive. We sought to develop a short questionnaire to assess compliance of alcohol advertising to existing alcohol industry self-regulated marketing codes. Questions for the Alcohol Marketing Assessment Rating Tool (AMART) were taken from a longer rating instrument that was originally developed to detect violations of a self-regulated alcohol marketing code. Secondary analysis of data from three studies of alcohol advertisements was performed to determine the reliability and validity of AMART at detecting code violations, using the longer form as the criterion. One sample was used as an exploratory set. Two samples were used to validate the questionnaire. In the exploratory set, the reliability of AMART was considered near perfect (kappa = 0.92). Sensitivity was 97%, and specificity was 100%. Positive predictive value (PPV) was perfect and negative predictive value (NPV) was approximately 90%. In the validation sets, reliability was considered substantial to near perfect (kappa = 0.71-0.94). Specificity and PPV remained perfect, and NPV was 86%-90%. The AMART is a reliable tool to detect violations of a self-regulated marketing code in alcohol advertising. It significantly decreases the amount of resources needed to evaluate a finite number of advertisements.


An exploration of the concept of comfort.
This paper explores the concept of comfort within conceptual frameworks that identify comfort as a key component. The paper focuses on two main issues: firstly, descriptions of comfort and how comfort is defined; secondly, the relationship between comfort and other core concepts and how it is similar to, or differs from these concepts. The descriptions of comfort identify the difficulty of defining comfort and note that historical interpretations of comfort have changed over time. Key authors provide different perspectives of comfort and hence a uniform definition is not evident. The degree to which patients' perspectives are represented varies within the frameworks which suggests that further clarity is needed in this area. The relationship between comfort and other concepts considers the boundaries between comfort and two concepts: nursing as therapy and caring. Comfort tends to be seen as a minor aspect of these concepts focusing on relief of discomfort or actions that demonstrate caring. The paper concludes that there is a lack of clarity around the use of the term comfort which fits with a view of comfort as an emerging concept. Descriptions of comfort are clearly evident in the literature but differ in relation to the authors' views of nursing. The relationship between comfort and nursing as therapy and caring is also complex and highlights the debate in relation to comfort as a part of caring or as the overall focus of care.


Import of ADP/ATP carrier into mitochondria: two receptors act in parallel.
We have identified the yeast homologue of Neurospora crassa MOM72, the mitochondrial import receptor for the ADP/ATP carrier (AAC), by functional studies and by cDNA sequencing. Mitochondria of a yeast mutant in which the gene for MOM72 was disrupted were impaired in specific binding and import of AAC. Unexpectedly, we found a residual, yet significant import of AAC into mitochondria lacking MOM72 that occurred via the receptor MOM19. We conclude that both MOM72 and MOM19 can direct AAC into mitochondria, albeit with different efficiency. Moreover, the precursor of MOM72 apparently does not require a positively charged sequence at the extreme amino terminus for targeting to mitochondria.


Macrophage migration inhibitory factor is secreted by rhabdomyosarcoma cells, modulates tumor metastasis by binding to CXCR4 and CXCR7 receptors and inhibits recruitment of cancer-associated fibroblasts.
The overexpression of macrophage migration inhibitory factor (MIF) has been observed in many tumors and is implicated in oncogenic transformation and tumor progression. MIF activates CXCR2 and CD74 receptors and, as recently reported, may also bind to the stromal-derived factor-1 (SDF-1)-binding receptor CXCR4. Here, we report that human rhabdomyosarcoma (RMS) cell lines secrete MIF and that this chemokine (a) induces phosphorylation of mitogen-activated protein kinase (MAPK) p42/44 and AKT, (b) stimulates RMS cell adhesion, (c) enhances tumor vascularization, but surprisingly (d) decreases recruitment of cancer-associated fibroblasts (CAF). Because RMS cells used in our studies do not express CXCR2 and CD74 receptors, the biological effects of MIF on RMS cells depend on its interaction with CXCR4, and as we report here for the first time, MIF may also engage another SDF-1-binding receptor (CXCR7) as well. Interestingly, downregulation of MIF in RMS cells inoculated into immunodeficient mice led to formation of larger tumors that displayed higher stromal cell support. Based on these observations, we postulate that MIF is an important autocrine/paracrine factor that stimulates both CXCR4 and CXCR7 receptors to enhance the adhesiveness of RMS cells. We also envision that when locally secreted by a growing tumor, MIF prevents responsiveness of RMS to chemoattractants secreted outside the growing tumor (e.g., SDF-1) and thereby prevents release of cells into the circulation. On the other hand, despite its obvious proangiopoietic effects, MIF inhibits in CXCR2/CD74-dependent manner recruitment of CAFs to the growing tumor. Our data indicate that therapeutic inhibition of MIF in RMS may accelerate metastasis and tumor growth.


Chlamydiales in guinea-pigs and their zoonotic potential.
The aim was to detect and characterize chlamydial infections in guinea-pigs (GP) with ocular disease, study their pathogenicity and zoonotic potential and to test for the presence of Acanthamoebae spp. in GP eyes and to investigate whether they could act as vectors for Chlamydia-like organisms. Overall 126 GP, of which 77 were symptomatic, were screened by clinical examination, cytology, gross pathology, histology, immunohistochemistry, polymerase chain reaction (PCR) and bacteriology. A new Chlamydiaceae-specific intergenic spacer rRNA gene PCR, designed to amplify this segment linking the 16S and 23S regions, was performed. DNA samples were also received from one owner including samples of his cat and rabbit. Guinea-pigs: 48 of 75 symptomatic, but only 11 of 48 asymptomatic GP were positive by PCR for Chlamydophila caviae guinea-pig inclusion conjunctivitis (GPIC) (P < 0.0001). Eighteen of 75 or 15/48, respectively, were positive for DNA from Chlamydia-like organisms. Acanthamoebae-DNA could be found in two GP, of which one was symptomatic. Owner, cat and rabbit: Samples of all three species were positive by PCR for C. caviae GPIC and the owner's one-day disposable contact lenses showed a positive PCR result for the Chlamydia-like organism Parachlamydia acanthamoebae. No Acanthamoebae-DNA could be detected. This study is the first to describe Chlamydia-like organisms in GP and to detect C. caviae GPIC in human, cat and rabbit. Therefore, C. caviae GPIC could pose a zoonotic potential. We believe that the finding of C. caviae GPIC in species other than GP is probably not unique.


Using Security Questions to Link Participants in Longitudinal Data Collection.
Anonymous data collection systems are often necessary when assessing sensitive behaviors but can pose challenges to researchers seeking to link participants over time. To assist researchers in anonymously linking participants, we outlined and tested a novel security question linking (security question linking; SEEK) method. The SEEK method includes four steps: (1) data management and standardization, (2) many-to-many matching, (3) fuzzy matching, and (4) rematching and verification. The method is demonstrated in SAS with two samples from a longitudinal study of adolescent dating violence. After an initial assessment during a laboratory visit, participants were asked to complete an online assessment either (a) once, 3 months later (Sample 1, n = 60), or (b) three times at 1-month intervals (Sample 2, n = 140). Demographics, eye color, and responses to nine security questions were used as key variables to link responses from the laboratory and online follow-up assessments. The rates of matched cases were 100% in Sample 1 and from 94.3 to 98.3% in Sample 2. To quantify the confidence in the data quality of successfully matched pairs, we reported the means and standard deviations of the number of matched security questions. In addition, we reported the rank order and counts of the mismatched components in key variables. Results indicate that the SEEK method provides a feasible and reliable solution to link responses in longitudinal studies with sensitive questions.


Large difference in carbon emission – burial balances between boreal and arctic lakes.
Lakes play an important role in the global carbon (C) cycle by burying C in sediments and emitting CO2 and CH4 to the atmosphere. The strengths and control of these fundamentally different pathways are therefore of interest when assessing the continental C balance and its response to environmental change. In this study, based on new high-resolution estimates in combination with literature data, we show that annual emission:burial ratios are generally ten times higher in boreal compared to subarctic - arctic lakes. These results suggest major differences in lake C cycling between biomes, as lakes in warmer boreal regions emit more and store relatively less C than lakes in colder arctic regions. Such effects are of major importance for understanding climatic feedbacks on the continental C sink - source function at high latitudes. If predictions of global warming and northward expansion of the boreal biome are correct, it is likely that increasing C emissions from high latitude lakes will partly counteract the presumed increasing terrestrial C sink capacity at high latitudes.


Effect of polymer brush architecture on antibiofouling properties.
Polymer brushes show great promise in next-generation antibiofouling surfaces. Here, we have studied the influence of polymer brush architecture on protein resistance. By carefully optimizing reaction conditions, we were able to polymerize oligoglycerol-based brushes with sterically demanding linear or dendronized side chains on gold surfaces. Protein adsorption from serum and plasma was analyzed by surface plasmon resonance. Our findings reveal a pronounced dependence of biofouling on brush architecture. Bulky yet flexible side chains as in dendronized brushes provide an ideal environment to repel protein-possibly through formation of a hydration layer, which can be further enhanced by presenting free hydroxyl groups on the polymer brushes. A deeper understanding of how brush architecture influences protein resistance will ultimately enable fabrication of surface coatings tailored to specific requirements in biomedical applications.


Preventing childhood obesity: what works?
Rates of overweight in North American children and adolescents have increased dramatically since the 1970s. Childhood obesity has reached epidemic proportions and calls for prevention and treatment programs to reverse this trend have been made. However, the evidence base needed for effective action is still incomplete, especially for childhood obesity prevention programs. This paper focuses on primary prevention of childhood obesity and has three aims: (1) to briefly describe current primary prevention approaches for childhood obesity and the evidence for their impact; (2) to elucidate promising, but untested intervention strategies using an ecological framework and evidence from experimental and epidemiological research on factors influencing children's eating and weight status; and (3) to introduce a multiphase strategy for screening intervention components and building and evaluating potent interventions for childhood obesity. Most childhood obesity prevention programs have focused on school-aged children and have had little success. We suggest that, given these findings, prevention efforts should be expanded to explore other contexts in which children live as possible settings for intervention efforts, including the family and childcare settings. Given that 25% of preschool children are already overweight, intervening with children before school entry should be a priority. A review of experimental research on the developing controls of food intake in infancy and childhood suggests possible intervention strategies, focusing on parenting and aspects of the feeding environment. Epidemiological findings point to even earlier modifiable risk factors, including gestational weight gain, maternal prepregnancy weight, and formula feeding. However, the potential impact of altering these risk factors remains to be evaluated. In response to this problem, we suggest a new, multiphase method for accomplishing this, including screening intervention components, refining intervention designs and confirming component efficacy to build and evaluate potent, optimized interventions.


An Outbreak of Synthetic Cannabinoid-Associated Coagulopathy in Illinois.
In March and April 2018, more than 150 patients presented to hospitals in Illinois with coagulopathy and bleeding diathesis. Area physicians and public health organizations identified an association between coagulopathy and synthetic cannabinoid use. Preliminary tests of patient serum samples and drug samples revealed that brodifacoum, an anticoagulant, was the likely adulterant. We reviewed physician-reported data from patients admitted to Saint Francis Medical Center in Peoria, Illinois, between March 28 and April 21, 2018, and included in a case series adult patients who met the criteria used to diagnose synthetic cannabinoid-associated coagulopathy. A confirmatory anticoagulant poisoning panel was ordered at the discretion of the treating physician. A total of 34 patients were identified as having synthetic cannabinoid-associated coagulopathy during 45 hospitalizations. Confirmatory anticoagulant testing was performed in 15 of the 34 patients, and superwarfarin poisoning was confirmed in the 15 patients tested. Anticoagulant tests were positive for brodifacoum in 15 patients (100%), difenacoum in 5 (33%), bromadiolone in 2 (13%), and warfarin in 1 (7%). Common symptoms at presentation included gross hematuria in 19 patients (56%) and abdominal pain in 16 (47%). Computed tomography was performed to evaluate abdominal pain and revealed renal abnormalities in 12 patients. Vitamin K1 (phytonadione) was administered orally in all 34 patients and was also administered intravenously in 23 (68%). Red-cell transfusion was performed in 5 patients (15%), and fresh-frozen plasma infusion in 19 (56%). Four-factor prothrombin complex concentrate was used in 1 patient. One patient died from complications of spontaneous intracranial hemorrhage. Our data indicate that superwarfarin adulterants of synthetic cannabinoids can lead to clinically significant coagulopathy. In our series, in most of the cases in which the patient presented with bleeding diathesis, symptoms were controlled with the use of vitamin K1 replacement therapy. The specific synthetic cannabinoid compounds are not known.


Comparing the experience of outpatient therapy in home and day hospital settings after traumatic brain injury: patient, significant other and therapist perspectives.
To explore how therapy in a home and day hospital setting impacts on rehabilitation processes and outcomes from the perspective of the patients, their significant others and their treating occupational therapists. Fourteen participants with severe traumatic brain injury received a one-to-one, goal-directed, client-centred outpatient occupational therapy programme (a) in their home for 6 weeks and (b) in a day hospital clinic for 6 weeks. The experience of rehabilitation in both settings was explored using semi-structured interviews with the participants, their significant others and their treating occupational therapists. Participants and their significant others described the two environments as disparate with home-based therapy perceived as more relaxing, normal, satisfying and effective. The approach to therapy at home was commonly described as 'real-life' whereas the therapy approach in day hospital was characterised as 'simulation of real life tasks' and 'remedial exercises'. Participants' experience of therapy relationships at home was characterised as 'friendship', in which the therapist was a 'visitor', whereas in the hospital, participants were characterised as 'patients' and therapists as 'bosses' and 'teachers'. The experience of home-based therapy was perceived as more convenient, positive and preferred by patients and their family members. Therapists described more therapeutic benefits and the ability to work more effectively on activity and participation level goals in the client's real-life environment.


Mothers' and their daughters' use of preventive measures against cervical cancer.
Vaccination against human papillomavirus (HPV) and screening are complementary preventive measures against cervical cancer. In Denmark, screening and vaccination are free of charge for the women. In total, 75% of women are screened and about 90% of girls are vaccinated with at least one dose. Our aim was to determine whether, in Denmark, daughters of unscreened mothers are less likely to be vaccinated against HPV than are daughters of screened mothers. We used population-based data from the Danish Patient Register, Health Service Registration, Pathology Data Bank, and Civil Registration System. Individual-level data on screening, vaccination, and vital status until 31 December 2010 were retrieved. Daughters were linked to their mothers through the link provided in the Civil Registration System. The study population included 149,147 girls born in 1993-1997 and their mothers. We calculated birth cohort-specific relative risks (RRs) of non-initiation of HPV vaccination in daughters depending on their mothers' screening status. In total, 8% of girls did not receive any vaccination, and 35% of their mothers were unscreened. Among the 92% of girls receiving at least one vaccine dose, 14% of mothers were unscreened. The birth cohort-specific RRs of non-initiation of vaccination given an unscreened mother varied between 2.16 (95% CI: 2.00-2.33) and 2.83 (95% CI: 2.63-3.05). The observed association between screening and vaccination suggest that it will be difficult to increase the vaccination coverage by, for example, counselling at the mother's cervical screening appointment. Other measures to increase the coverage with vaccination will be important.


Factors affecting disruption in families of adults with mental illness.
This study examined relationships between vulnerability/risk and protective factors, and family functioning in women family members of adults with serious mental illness. Using a descriptive, correlational design, this secondary analysis examined characteristics of the family member with mental illness (e.g., diagnosis, level of care) and measures of caregiver stigma and strain, client dependence, family disruption, sense of coherence, and resourcefulness. Family disruption was greatest in women who provided direct care and whose family member had major depression, followed by bipolar disorder, schizophrenia, and panic disorder. Sense of coherence and resourcefulness were associated with lower family disruption, but did not mediate the effects of caregiver strain. Interventions restricted to one family member may be insufficient for improving the family functioning.


Neurologic manifestations of thyroid disease.
Animals with polyneuropathy associated with primary hypothyroidism have clinical neurologic signs that range from peripheral vestibular signs, lower motor neuronal deficits, laryngeal paralysis, to megaesophagus; however, a few affected animals also show evidence of a more generalized polyneuropathy with cranial (facial more than vestibular nerve) and spinal nerves being affected most commonly. Confirmation of the diagnosis will depend on neurologic signs, clinicopathologic results including an abnormally low T4 response to TSH and electrodiagnostic findings. Successful treatment is based on thyroxine supplementation therapy. In severe cases with laryngeal paralysis, additional surgical treatment is indicated. Some animals with megaesophagus may develop permanent neurologic disabilities.


Sharing resources to create a district drug formulary: a countywide controlled trial.
Creating a drug formulary takes considerable time, but merely adopting one lacks local perspective and ownership. Sharing resources between several practices treads a middle path between these extremes, but is it effective? The aim of the study was to audit the influence of a district primary care drug formulary on prescribing by general practitioners. A controlled trial was carried out to compare prescribing by 50 general practitioners from 11 urban and semirural practices in south Bedfordshire that participated in creating a district drug formulary with prescribing by all other general practitioners in the county. The proportion of prescription items that were for drugs listed in the formulary rose significantly in three therapeutics groups: cardiovascular (by 7-12% above control practice values); musculoskeletal (by 1-11% above control practice values); and obstetrics and gynaecology (by 6-9% above control practice values). The number of items prescribed per prescribing unit fell significantly in three therapeutic groups: musculoskeletal (by 1-7% below control practice values); nervous (by 7-12% below control practice values); and nutrition and blood (by 15-21% below control practice values). The estimated saving resulting from the creation of the formulary was 150,000 pounds (3000 pounds per doctor) per year. Sharing resources between practices to create a district-wide primary care drug formulary can lead to changes in prescribing and reduce costs sustained over 3 years.


Extracting DNA from FFPE Tissue Biospecimens Using User-Friendly Automated Technology: Is There an Impact on Yield or Quality?
DNA extracted from formalin-fixed, paraffin-embedded (FFPE) tissue blocks is amenable to analytical techniques, including sequencing. DNA extraction protocols are typically long and complex, often involving an overnight proteinase K digest. Automated platforms that shorten and simplify the process are therefore an attractive proposition for users wanting a faster turn-around or to process large numbers of biospecimens. It is, however, unclear whether automated extraction systems return poorer DNA yields or quality than manual extractions performed by experienced technicians. We extracted DNA from 42 FFPE clinical tissue biospecimens using the QiaCube (Qiagen) and ExScale (ExScale Biospecimen Solutions) automated platforms, comparing DNA yields and integrities with those from manual extractions. The QIAamp DNA FFPE Spin Column Kit was used for manual and QiaCube DNA extractions and the ExScale extractions were performed using two of the manufacturer's magnetic bead kits: one extracting DNA only and the other simultaneously extracting DNA and RNA. In all automated extraction methods, DNA yields and integrities (assayed using DNA Integrity Numbers from a 4200 TapeStation and the qPCR-based Illumina FFPE QC Assay) were poorer than in the manual method, with the QiaCube system performing better than the ExScale system. However, ExScale was fastest, offered the highest reproducibility when extracting DNA only, and required the least intervention or technician experience. Thus, the extraction methods have different strengths and weaknesses, would appeal to different users with different requirements, and therefore, we cannot recommend one method over another.


Comparative antimicrobial efficacy of alcohol-based hand rub and conventional surgical scrub in a medical center.
Hand hygiene is the cornerstone of aseptic techniques to reduce surgical site infection. Conventional surgical scrub is effective for disinfecting a surgeon's hands. However, the compliance of conventional scrub may be hindered by skin damage, allergy, and time. Alcohol-based hand rub has a satisfactory antimicrobial effect, but mostly in laboratory settings. Our aim was to compare a conventional surgical scrub with an alcohol-based hand rub to evaluate antimicrobial efficacy. From June 1, 2010 to July 31, 2011, 128 healthcare workers were enrolled in the study. They used an alcohol-based hand rub or a conventional surgical scrub as preoperative hand antisepsis during their routine practice. Hand sampling for cultures were performed before and after operations. Positive culture plates were further processed for pathogen identification. The culture positive rate of the alcohol-based hand rub was 6.2% before operations and 10.8% after operations. Both rates were lower than the conventional surgical scrub [47.6% before operations (p < 0.001) and 25.4% after operations (p = 0.03)]. The most identified pathogens were Gram-positive with coagulase-negative staphylococci being the major pathogen. Multivariate analysis showed that prior hand condition (p = 0.21) and type of surgery such as cardiovascular surgery (p = 0.12) were less relevant, but the alcohol-based hand rub was a significant protective factor for positive hand cultures. The alcohol-based hand rub was more efficacious for surgical antisepsis and had sustained efficacy, compared to conventional surgical scrub. We suggest that alcohol-based hand rubs could be an alternative surgical antiseptic in the operative theater.


Genetic aberrations in glioblastoma multiforme: translocation of chromosome 10 in an O-2A-like cell line.
We have examined the genetic aberrations in two near-diploid glioblastoma multiforme cell lines that appear to have arisen from different glial lineages. One cell line, Hu-O-2A/Gb1, expresses antigens and metabolic profiles characteristic of the oligodendrocyte-type-2 astrocyte (0-2A) lineage of the rat central nervous system. This line generates, in vitro, cells with characteristics of 0-2A progenitor cells, oligodendrocytes and astrocytes. The second cell line, IN1434, is derived from an astrocyte or a precursor cell restricted to astrocytic differentiation. In Hu-O-2A/Gb1 the sole homologue of chromosome 10 is disrupted at band 10p11-12.1 by translocation with chromosomes X and 15. The translocation breakpoint is localized between genetic markers D10S2103 and [D10S637, D10S1962, D10S355]. Other aberrations include a 5;14 translocation, deletion of the long and short arms of chromosome 16 and loss of one copy of the CDKN2 gene. IN1434 cells share some cytogenetic abnormalities with Hu-O-2A/Gb1 cells, despite their apparent derivation from a different biological origin, but also have translocations involving the long and short arms of chromosome 1 and the long arm of chromosome 7, and deletion of chromosome 13 at bands 13q12-21.


Health status measurement in Parkinson's disease: validity of the PDQ-39 and Nottingham Health Profile.
We assessed the feasibility and psychometric properties of two commonly used health status questionnaires in Parkinson's disease (PD): the generic Nottingham Health Profile (NHP) and the disease-specific 39-item Parkinson's disease Questionnaire (PDQ-39), from a cross-sectional postal survey of PD patients (N = 81), using traditional and Rasch measurement methodologies. Overall response rate was 88%. Both questionnaires were found feasible, although the NHP performed less well. The PDQ-39 had fewer floor effects and was better able to separate respondents into distinct groups than the NHP, whereas the latter exhibited less ambiguous dimensionality and better targeting of respondents with non-extreme scores. Reliability and validity indices were similar, and potential differential item functioning by age and gender groups was found for both questionnaires. PDQ-39 response alternatives indicated ambiguity. With few exceptions, questionnaire scales were unable to meet recommended standards fully. While preliminary, this study illustrates the need for thorough evaluation of outcome measures and has implications beyond the questionnaires used here. Although promising, both questionnaires warrant further developmental work and stronger support of measurement validity before they could be considered fully suitable for valid use in PD, in particular in earlier stages of the disease.


First Report of Clover Proliferation Group Phytoplasmas (16SrVI-A) Associated with Purple Top Diseased Potatoes (Solanum tuberosum) in China.
Phytoplasma diseases on potatoes are not well understood and have gone largely undetected in China. During the growing seasons of 2005 through 2010, potato disease surveys were conducted in seed and commercial fields in Yunnan Province. Samples were also harvested from three seed potato production areas in the Inner Mongolia Autonomous Region in 2007 and 2010. Disease incidence in these fields ranged from 15 to 85%. Plants displayed symptoms of branch proliferation, aerial tuber formation, upward rolling yellowish and purplish apical leaves, and extremely short stolen or chain tubers (irregular-shaped tubers). Total DNA from 250 samples was extracted from the leaves, stems, and roots of symptomatic and asymptomatic plants. A nested PCR was performed by using primer pair P1/P7 followed by R16F2n/R16R2 to detect the presence of phytoplasmas (1,3). An approximate 1.2-kb PCR product was amplified from symptomatic plants but not from asymptomatic plants. Restriction fragment length polymorphism (RFLP) patterns were analyzed by digesting the 1.2-kb amplicon singly with restriction enzymes AluI, BfaI, MseI, HhaI, HinfI, HpaII, KpnI, RsaI, and TaqI. The RFLP patterns of 120 of the 250 samples matched patterns of the clover proliferation (CP) group (16SrVI) subgroup A (16SrVI-A) phytoplasma (1). In addition, the nested PCR product of P1A/P7A (2) following P1/P7 amplification was cloned and sequenced (GenBank Accession No. HQ609490). Nucleotide sequences were analyzed by iPhyClassifier software (4), confirming the relationship of this phytoplasma to 'Candidatus Phytoplasma trifolii' with RFLP patterns identical to group 16SrVI-A. To our knowledge, this is the first report of the CP group phytoplasmas associated with purple top diseased potatoes in China. References: (1) I.-M. Lee et al. Int. J. Syst. Bacteriol. 48:1153, 1998. (2) I.-M. Lee et al. Int. J. Syst. Evol. Microbiol 54:337, 2004. (3) C. D. Smart et al. Appl. Environ. Microbiol. 62:2988, 1996. (4) Y. Zhao et al. Int. J. Syst. Evol. Microbiol. 59:2582, 2009.


Using diffusion weighted MR imaging in a lipomatous chest wall lesion.
Hibemoma is a rare benign soft tissue tumor derived from brown fat.The tumor is also known as "fetal lipoma, lipoma of embryogenic fat and lipoma of immature adipose tissue. Hibemomas are slow-growing, painless soft tissue tumors which do not recur after surgical resection. Preferred locations are brown fat containing sites as thigh, interscapular region, shoulder, axilla, and mediastinum.The tumor occurs most commonly in adults, with a mean age of 38 years (age range, 2-75 years). We present a rare case of hibernoma with radiological and pathological findings.


Cell-mediated immunity and lymphocyte populations in experimental Argentine hemorrhagic fever (Junín Virus).
Guinea pigs infected with the XJ prototype strain of Junín virus reproduce the main features of Argentine hemorrhagic fever, showing hemorrhages, leukothrombocytopenia, and focal lymphoid tissue necrosis. Viral lymphotropism is shown by the presence of viral antigens, severe cytopathic effect, and high virus titers in lymphoid organs. A pronounced depression of humoral immune response to sheep erythrocytes as well as to the virus is described. This study was carried out to determine whether cellular immune response was also modified and which cell populations were affected. Delayed hypersensitivity skin reaction to purified protein derivative was found to be markedly depressed after infection. A noticeable decrease in both percentages and absolute T lymphocyte numbers, detected by E rosettes, in spleen and lymph nodes, together with a low absolute T cell number in peripheral blood, were observed. Total cell counts in spleen, lymph nodes, and peripheral blood were also reduced. On the contrary, no modification in percentages of B lymphocytes, as measured by EAC rosettes, was found. These results indicate that cell-mediated immunity is markedly impaired in guinea pigs infected with the XJ strain of Junín virus. Its relationship with the pathogenesis of the disease is discussed.


Biological characteristics of human-urine-derived stem cells: potential for cell-based therapy in neurology.
Stem cells in human urine have gained attention in recent years; however, urine-derived stem cells (USCs) are far from being well elucidated. In this study, we compared the biological characteristics of USCs with adipose-derived stem cells (ASCs) and investigated whether USCs could serve as a potential cell source for neural tissue engineering. USCs were isolated from voided urine with a modified culture medium. Through a series of experiments, we examined the growth rate, surface antigens, and differentiation potential of USCs, and compared them with ASCs. USCs showed robust proliferation ability. After serial propagation, USCs retained normal karyotypes. Cell surface antigen expression of USCs was similar to ASCs. With lineage-specific induction factors, USCs could differentiate toward the osteogenic, chondrogenic, adipogenic, and neurogenic lineages. To assess the ability of USCs to survive, differentiate, and migrate, they were seeded onto hydrogel scaffold and transplanted into rat brain. The results showed that USCs were able to survive in the lesion site, migrate to other areas, and express proteins that were associated with neural phenotypes. The results of our study demonstrate that USCs possess similar biological characteristics with ASCs and have multilineage differentiation potential. Moreover USCs can differentiate to neuron-like cells in rat brain. The present study shows that USCs are a promising cell source for tissue engineering and regenerative medicine.


Dual mechanisms of regulation of transcription of luteinizing hormone receptor gene by nuclear orphan receptors and histone deacetylase complexes.
The luteinizing hormone receptor (LHR), a member of the G protein-coupled, seven transmembrane receptor family, is essential for normal sexual development and reproductive function. LHR are expressed primarily in the gonads, but also are found in non-gonadal and cancer tissues. LH acts through LH receptors in Leydig cells to maintain general metabolic processes and steroidogenic enzymes, and in the ovary enhances follicular development and steroidogenesis in granulosa and luteal cells. The major transcriptional start sites of the LHR gene are located within the 176bp promoter domain. In the rat, the LHR gene is constitutively inhibited by upstream sequences (-176/-2056bp) in several cell systems, while in the human only a minor inhibitory effect was observed in JAR and HeLa cells (>20%). The TATA-less human promoter is driven by Sp1 and Sp3 transactivators that bind to two Sp1 domains at -79bp [Sp1(I)] and -119bp [Sp1(II)] (from ATG) with additive effects. An imperfect estrogen receptor half-site response element direct-repeat within the LHR promoter is an inhibitory locus. Endogenous orphan receptors, EAR2 and EAR3/COUP-TFI, bind this motif and repress promoter activity by 70%. TR4 also binds this motif and stimulates promoter activity (up to 2.5-fold). This is reversed by coexpression of EAR2 or EAR3/COUP-TFI through competitive binding to this site. Comparative studies of hDR and rDR orphan receptors binding and function revealed sequence-specific requirements. The A/C mismatch between hDR and rDR is responsible for the lack of TR4 binding and function in the rat. The G 3'-adjacent to the hDR core is important for EAR2/EAR3-COUP-TFI high-affinity binding. The Sp1-1 site is critical for EAR3/COUP-TFI repression, with minor participation for EAR2, and is not involved in the TR4 effect. Interaction of EAR3/COUP-TFI with Sp1 perturbs association of TFIIB with Sp1, independently of HDACs, and caused impairment of LHR transcription. Other aspect of control is through HDAC/mSin3A mechanism. Inhibition of HDACs by TSA increases LHR promoter activity in JAR cells (40-fold), association of acetylated H3/H4 with the LHR promoter, recruitment of Pol II to the promoter, and LHR mRNA levels. A multiprotein complex is recruited to the hLHR promoter via interaction with Sp1/Sp3: HDACs dock directly to Sp1-1 bound DNA and indirectly to Sp3-1 bound DNA through RbAp48, while mSin3A interacts HDACs and potentiates HDAC1-mediated repression. Our studies have demonstrated that orphan receptor-ERE complexes, and the HDAC1-HDAC2-mSin3A complex have important roles in the regulation of LHR gene transcription by interaction with Sp1/Sp3, and by region-specific changes in histone acetylation and Pol II recruitment within the LHR promoter.


Expression of functional bovine cholesterol side chain cleavage cytochrome P450 (P450scc) in Escherichia coli.
Escherichia coli expression vectors containing the trc promoter and the complete DNA sequence of either the precursor or the mature form of bovine adrenocortical cholesterol side chain cleavage cytochrome P450 (P450scc) were transformed into E. coli strain JM109 and transcription induced with isopropyl-beta-D-thiogalactopyranoside (IPTG). Immunoreactive cytochrome P450scc was produced using the plasmid containing the mature P450scc sequence but not with the plasmid containing the sequence of the precursor form of P450scc, even though P450scc RNA was detectable in both cases. The mature form of P450scc was detected spectrophotometrically in a reduced CO-difference spectrum in E. coli (40-60 nmol/liter culture). Cholesterol and hydroxylated derivatives (22-hydroxycholesterol and 25-hydroxycholesterol) produce a type 1 substrate-binding spectrum in IPTG-induced, transformed E. coli. The P450scc was found to be associated with the E. coli membrane fraction and the enzymatic activity of side chain cleavage of 25-hydroxycholesterol was reconstituted using solubilized membranes, in the presence of purified bovine adrenocortical adrenodoxin and NADPH-adrenodoxin reductase (turnover number; 15.4 nmol/min/nmol P450). This bacterial expression system provides functional P450scc, in the absence of other forms of P450, which can be used for evaluation of enzymatic and spectral properties of this mitochondrial P450 by site-directed mutagenesis.


Amino acid incorporation into rat brain proteins during spreading cortical depression.
Unilateral spreading cortical depression was elicited by applying potassium chloride solutions to the dura of conscious, freely moving rats. Incorporation of (3)H-leucine into soluble cortical proteins was decreased in the depressed hemisphere relative to the control side, while soluble brainstem proteins from both sides had the same specific activity. Various subfractions of soluble cortical proteins were affected to equal degrees.


Evaluation of clinical and metabolic changes after non surgical periodontal treatment of type 2 diabetes mellitus patients: A clinico biochemical study.
To evaluate the clinical and metabolic changes following nonsurgical periodontal therapy of type-2 diabetic patients and nondiabetic patients. Fifteen nondiabetics and fifteen type-2 diabetics with moderate to severe periodontitis were selected after meeting inclusion and exclusion criteria. Periodontal pocket probing was performed using a Williams graduated periodontal probe and comparisons of the clinical and biochemical parameters like plaque index, gingival index, probing depth, glycated hemoglobin, and lipid profile (total cholesterol, high density lipoprotein, low density lipoprotein, triglycerides and very low density lipoprotein) were done between and within two groups at baseline and 3 month. A correlation was observed between clinical and biochemical parameters between and within two groups at baseline and 3 month. The P value of diabetic group was<0.001 in all clinical parameters (plaque index PI, Gingival index GI, and Probing Pocket Depth PPD) in diabetic and nondiabetic group, showed statistically highly significant difference in GI, PI, PPD, <.01 in Glycated Hemoglobulin (HbAlc1) in test group and<0.05 in control group showed statistically significant difference. The P value of test group was >0.05 in Lipid profile (total cholesterol, low density lipoprotein, high density lipoprotein, serum triglyceride, very low density lipoprotein) that showed no significant difference and in control group<0.05 in total Cholesterol, high density lipoprotein and <0.01 in Low density lipoprotein, Very Low Density Lipoprotein, and Serum Triglyceride, respectively, showed statistically significant in Total cholesterol, Low density lipoprotein, Very Low Density Lipoprotein, and Serum Triglyceride, not significant in high density lipoprotein. The P value >0.05 showed no statistical significance of correlation between the test and control groups. Both groups of patients showed an improvement in clinical and metabolic parameters assessed at 3 months after nonsurgical periodontal therapy.


Merkel polyomavirus-specific T cells fluctuate with merkel cell carcinoma burden and express therapeutically targetable PD-1 and Tim-3 exhaustion markers.
The persistent expression of Merkel cell polyomavirus (MCPyV) oncoproteins in Merkel cell carcinoma (MCC) provides a unique opportunity to characterize immune evasion mechanisms in human cancer. We isolated MCPyV-specific T cells and determined their frequency and functional status. Multiparameter flow cytometry panels and HLA/peptide tetramers were used to identify and characterize T cells from tumors (n = 7) and blood (n = 18) of patients with MCC and control subjects (n = 10). PD-1 ligand (PD-L1) and CD8 expression within tumors were determined using mRNA profiling (n = 35) and immunohistochemistry (n = 13). MCPyV-specific CD8 T cells were detected directly ex vivo from the blood samples of 7 out of 11 (64%) patients with MCPyV-positive tumors. In contrast, 0 of 10 control subjects had detectable levels of these cells in their blood (P < 0.01). MCPyV-specific T cells in serial blood specimens increased with MCC disease progression and decreased with effective therapy. MCPyV-specific CD8 T cells and MCC-infiltrating lymphocytes expressed higher levels of therapeutically targetable PD-1 and Tim-3 inhibitory receptors compared with T cells specific to other human viruses (P < 0.01). PD-L1 was present in 9 of 13 (69%) MCCs and its expression was correlated with CD8-lymphocyte infiltration. MCC-targeting T cells expand with tumor burden and express high levels of immune checkpoint receptors PD-1 and Tim-3. Reversal of these inhibitory pathways is therefore a promising therapeutic approach for this virus-driven cancer.


Dehalogenation of polybrominated diphenyl ethers and polychlorinated biphenyl by bimetallic, impregnated, and nanoscale zerovalent iron.
Nanoscale zerovalent iron particles (nZVI), bimetallic nanoparticles (nZVI/Pd), and nZVI/Pd impregnated activated carbon (nZVI/Pd-AC) composite particles were synthesized and investigated for their effectiveness to remove polybrominated diphenyl ethers (PBDEs) and/or polychlorinated biphenyls (PCBs). Palladization of nZVI promoted the dehalogenation kinetics for mono- to tri-BDEs and 2,3,4-trichlorobiphenyl (PCB 21). Compared to nZVI, the iron-normalized rate constants for nZVI/Pd were about 2-, 3-, and 4-orders of magnitude greater for tri-, di-, and mono-BDEs, respectively, with diphenyl ether as a main reaction product. The reaction kinetics and pathways suggest an H-atom transfer mechanism. The reaction pathways with nZVI/Pd favor preferential removal of para-halogens on PBDEs and PCBs. X-ray fluorescence mapping of nZVI/Pd-AC showed that Pd mainly deposits on the outer part of particles, while Fe was present throughout the activated carbon particles. While BDE 21 was sorbed onto activated carbon composites quickly, debromination was slower compared to reaction with freely dispersed nZVI/Pd. Our XPS and chemical data suggest about 7% of the total iron within the activated carbon was zerovalent, which shows the difficulty with in-situ synthesis of a significant fraction of zerovalent iron in the microporous material. Related factors that likely hinder the reaction with nZVI/Pd-AC are the heterogeneous distribution of nZVI and Pd on activated carbon and/or immobilization of hydrophobic organic contaminants at the adsorption sites thereby inhibiting contact with nZVI.


Concurrent brain radiotherapy and EGFR-TKI may improve intracranial metastases control in non-small cell lung cancer and have survival benefit in patients with low DS-GPA score.
Epidermal growth factor receptor-tyrosine kinase inhibitor (EGFR-TKI) has intracranial activity in EGFR-mutant Non-Small Cell Lung Cancer (NSCLC). The optimal timing of brain radiotherapy (RT) and appropriate patients who need early brain RT remains undetermined. This is a retrospective study of EGFR-mutant NSCLC patients with newly diagnosed brain metastases (BMs) before EGFR-TKI initiation. Intra-cranial progression free survival (IC-PFS) and overall survival (OS) were measured from the date of EGFR-TKI treatment. A total of 113 patients were eligible, 49 received concurrent early brain RT with EGFR-TKI and 64 were treated with EGFR-TKI alone as initial therapy, including 27 with salvage RT upon BM progression. The patients with early brain RT had superior IC-PFS than those without early brain RT (21.4 vs 15.0 months, P=0.001), which remained significant in multivariate analysis (HR 0.30, P<0.001). The median overall survival (OS) for early RT, EGFR-TKI alone and salvage RT groups was 28.1, 24.5, and 24.6 months, respectively (P=0.604). Similar IC-PFS (23.6 vs 21.4 months, P=0.253) and OS (24.6 vs 28.1 months, P=0.385) were observed between salvage RT and early RT groups. For patients with Diagnosis-Specific Graded Prognostic Assessment (DS-GPA) score of 0 to 2, early brain RT was the independent factor for improved OS (HR 0.33, P=0.025). In conclusion, concurrent early brain RT with EGFR-TKI may improve intracranial disease control in EGFR-mutant NSCLC with BM and have survival benefit in patients with low DS-GPA score. Salvage brain RT upon BM progression may be acceptable in some patients.


17beta-Estradiol upregulates and activates WOX1/WWOXv1 and WOX2/WWOXv2 in vitro: potential role in cancerous progression of breast and prostate to a premetastatic state in vivo.
Human WWOX gene encodes a proapoptotic WW domain-containing oxidoreductase WOX1 (also named WWOX, FOR2 or WWOXv1). Apoptotic and stress stimuli activate WOX1 via Tyr33 phosphorylation and nuclear translocation. WOX1 possesses a tetrad NSYK motif in the C-terminal short-chain alcohol dehydrogenase/reductase (SDR) domain, which may bind estrogen and androgen. Here, we determined that 17beta-estradiol (E(2)) activated WOX1, p53 and ERK in COS7 fibroblasts, primary lung epithelial cells, and androgen receptor (AR)-negative prostate DU145 cells, but not in estrogen receptor (ER)-positive breast MCF7 cells. Androgen also activated WOX1 in the AR-negative DU145 cells. These observations suggest that sex hormone-mediated Tyr33 phosphorylation and nuclear translocation of WOX1 is independent of ER and AR. Stress stimuli increase physical binding of p53 with WOX1 in vivo. We determined here that E(2) increased the formation of p53/WOX1 complex and their nuclear translocation in COS7 cells; however, nuclear translocation of this complex could not occur in MCF7 cells. By immunohistochemistry, we determined that progression of prostate from normal to hyperplasia, cancerous and metastatic stages positively correlate with upregulation and activation of WOX1 and WOX2 (FOR1/WWOXv2). In contrast, breast cancer development to a premetastatic state is associated with upregulation and Tyr33 phosphorylation of cytosolic WOX1 and WOX2, followed by significant downregulation or absent expression during metastasis. These Tyr33-phosphorylated proteins are mostly located in the mitochondria without translocating to the nuclei, which is comparable to those findings in cultured breast cancer cells. Together, sex steroid hormone-induced activation of WOX1 and WOX2 is independent of ER and AR, and this activation positively correlates with cancerous progression of prostate and breast to a premetastatic state.


The p38 pathway, a major pleiotropic cascade that transduces stress and metastatic signals in endothelial cells.
By gating the traffic of molecules and cells across the vessel wall, endothelial cells play a central role in regulating cardiovascular functions and systemic homeostasis and in modulating pathophysiological processes such as inflammation and immunity. Accordingly, the loss of endothelial cell integrity is associated with pathological disorders that include atherosclerosis and cancer. The p38 mitogen-activated protein kinase (MAPK) cascades are major signaling pathways that regulate several functions of endothelial cells in response to exogenous and endogenous stimuli including growth factors, stress and cytokines. The p38 MAPK family contains four isoforms p38α, p38β, p38γ and p38δ that are encoded by four different genes. They are all widely expressed although to different levels in almost all human tissues. p38α/MAPK14, that is ubiquitously expressed is the prototype member of the family and is referred here as p38. It regulates the production of inflammatory mediators, and controls cell proliferation, differentiation, migration and survival. Its activation in endothelial cells leads to actin remodeling, angiogenesis, DNA damage response and thereby has major impact on cardiovascular homeostasis, and on cancer progression. In this manuscript, we review the biology of p38 in regulating endothelial functions especially in response to oxidative stress and during the metastatic process.


Fortuitous diagnosis of iliac osteomyelitis: septic arthritis of the hip in the neonate.
The neonate is unable to relate specific complaints of pain and may not exhibit the usual signs of illness or infection. Septic arthritis of the hip is a surgical emergency in the neonate, and it should be considered in any irritable or ill child who has a high index of suspicion. Prompt diagnosis and immediate treatment are necessary. Nearly all babies undergo routine examination of the hip for dysplasia. It is recognized that limitation of abduction of the hip in the neonate may not represent developmental dysplasia of the hip but may represent other etiologies, such as fracture, infection, congenital anomaly, or tumor. The following case report illustrates the importance of careful clinical evaluation of an apparent asymptomatic neonate.


Long-read sequencing reveals a 4.4 kb tandem repeat region in the mitogenome of Echinococcus granulosus (sensu stricto) genotype G1.
Echinococcus tapeworms cause a severe helminthic zoonosis called echinococcosis. The genus comprises various species and genotypes, of which E. granulosus (sensu stricto) represents a significant global public health and socioeconomic burden. Mitochondrial (mt) genomes have provided useful genetic markers to explore the nature and extent of genetic diversity within Echinococcus and have underpinned phylogenetic and population structure analyses of this genus. Our recent work indicated a sequence gap (> 1 kb) in the mt genomes of E. granulosus genotype G1, which could not be determined by PCR-based Sanger sequencing. The aim of the present study was to define the complete mt genome, irrespective of structural complexities, using a long-read sequencing method. We extracted high molecular weight genomic DNA from protoscoleces from a single cyst of E. granulosus genotype G1 from a sheep from Australia using a conventional method and sequenced it using PacBio Sequel (long-read) technology, complemented by BGISEQ-500 short-read sequencing. Sequence data obtained were assembled using a recently-developed workflow. We assembled a complete mt genome sequence of 17,675 bp, which is > 4 kb larger than the complete mt genomes known for E. granulosus genotype G1. This assembly includes a previously-elusive tandem repeat region, which is 4417 bp long and consists of ten near-identical 441-445 bp repeat units, each harbouring a 184 bp non-coding region and adjacent regions. We also identified a short non-coding region of 183 bp, which includes an inverted repeat. We report what we consider to be the first complete mt genome of E. granulosus genotype G1 and characterise all repeat regions in this genome. The numbers, sizes, sequences and functions of tandem repeat regions remain to be studied in different isolates of genotype G1 and in other genotypes and species. The discovery of such 'new' repeat elements in the mt genome of genotype G1 by PacBio sequencing raises a question about the completeness of some published genomes of taeniid cestodes assembled from conventional or short-read sequence datasets. This study shows that long-read sequencing readily overcomes the challenges of assembling repeat elements to achieve improved genomes.


Breeding and Genetics Symposium: really big data: processing and analysis of very large data sets.
Modern animal breeding data sets are large and getting larger, due in part to recent availability of high-density SNP arrays and cheap sequencing technology. High-performance computing methods for efficient data warehousing and analysis are under development. Financial and security considerations are important when using shared clusters. Sound software engineering practices are needed, and it is better to use existing solutions when possible. Storage requirements for genotypes are modest, although full-sequence data will require greater storage capacity. Storage requirements for intermediate and results files for genetic evaluations are much greater, particularly when multiple runs must be stored for research and validation studies. The greatest gains in accuracy from genomic selection have been realized for traits of low heritability, and there is increasing interest in new health and management traits. The collection of sufficient phenotypes to produce accurate evaluations may take many years, and high-reliability proofs for older bulls are needed to estimate marker effects. Data mining algorithms applied to large data sets may help identify unexpected relationships in the data, and improved visualization tools will provide insights. Genomic selection using large data requires a lot of computing power, particularly when large fractions of the population are genotyped. Theoretical improvements have made possible the inversion of large numerator relationship matrices, permitted the solving of large systems of equations, and produced fast algorithms for variance component estimation. Recent work shows that single-step approaches combining BLUP with a genomic relationship (G) matrix have similar computational requirements to traditional BLUP, and the limiting factor is the construction and inversion of G for many genotypes. A naïve algorithm for creating G for 14,000 individuals required almost 24 h to run, but custom libraries and parallel computing reduced that to 15 m. Large data sets also create challenges for the delivery of genetic evaluations that must be overcome in a way that does not disrupt the transition from conventional to genomic evaluations. Processing time is important, especially as real-time systems for on-farm decisions are developed. The ultimate value of these systems is to decrease time-to-results in research, increase accuracy in genomic evaluations, and accelerate rates of genetic improvement.


Lotus japonicus gene Ljsbp is highly conserved among plants and animals and encodes a homologue to the mammalian selenium-binding proteins.
We have isolated and characterized a Lotus japonicus gene (Ljsbp) encoding a putative polypeptide with striking homology to the mammalian 56-kDa selenium-binding protein (SBP). cDNA clones homologous to LjSBP were also isolated from soybean, Medicago sativa, and Arabidopsis thaliana. Comparative expression studies in L japonicus and A. thaliana showed that sbp transcripts are present in various tissues and at different levels. Especially in L japonicus nodules and seedpods and A. thaliana siliques, sbp expression appears to be developmentally up-regulated. sbp Gene transcripts were localized by in situ hybridization in the infected cells and vascular bundles of young nodules, while in mature nodules, low levels of expression were only detected in the parenchymatous cells. Expression of sbp transcripts in young seedpods and siliques was clearly visible in vascular tissues and embryos, while in embryos, low levels of expression were detected in the root epidermis and the vascular bundles. Polyclonal antibodies raised against a truncated LjSBP recombinant protein recognized a polypeptide of about 60 kDa in nodule extracts. Immunohistochemical experiments showed that accumulation of LjSBP occurred in root hairs, in the root epidermis above the nodule primordium, in the phloem of the vasculature, and abundantly in the infected cells of young nodules. Irrespective of the presence of rhizobia, expression of SBP was also observed in root tips, where it was confined in the root epidermis and protophloem cells. We hypothesize that LjSBP may have more than one physiological role and can be implicated in controlling the oxidation/reduction status of target proteins, in vesicular Golgi transport, or both.


Myocarditis in the Setting of Cancer Therapeutics: Proposed Case Definitions for Emerging Clinical Syndromes in Cardio-Oncology
Recent developments in cancer therapeutics have improved outcomes but have also been associated with cardiovascular complications. Therapies harnessing the immune system have been associated with an immune-mediated myocardial injury described as myocarditis. Immune checkpoint inhibitors are one such therapy with an increasing number of case and cohort reports describing a clinical syndrome of immune checkpoint inhibitor–associated myocarditis. Although the full spectrum of immune checkpoint inhibitor–associated cardiovascular disease still needs to be fully defined, described cases of myocarditis range from syndromes with mild signs and symptoms to fatal events. These observations in the clinical setting stand in contrast to outcomes from randomized clinical trials in which myocarditis is a rare event that is investigator reported and lacking in a specific case definition. The complexities associated with diagnosis, as well as the heterogeneous clinical presentation of immune checkpoint inhibitor–associated myocarditis, have made ascertainment and identification of myocarditis with high specificity challenging in clinical trials and other data sets, limiting the ability to better understand the incidence, outcomes, and predictors of these rare events. Therefore, establishing a uniform definition of myocarditis for application in clinical trials of cancer immunotherapies will enable greater understanding of these events. We propose an operational definition of cancer therapy-associated myocarditis that may facilitate case ascertainment and report and therefore may enhance the understanding of the incidence, outcomes, and risk factors of this novel clinical syndrome.


Health-related quality of life 1 year after allogeneic or autologous stem-cell transplantation: a prospective study.
To evaluate health-related quality of life (HRQOL) in adults treated with high-dose chemotherapy followed by allogeneic (SCT) and autologous (ASCT) stem-cell transplantation 1 year after transplantation, using data from concurrent lymphoma patients receiving combination chemotherapy (CT) as a reference. Forty-one leukemia patients (SCT group), 51 lymphoma patients (ASCT group), and 85 CT patients completed the European Organization for Research and Treatment of Cancer QLQ-C30 questionnaire at baseline and after 1 year. The SCT group (median age, 36 years) had better functioning scores and less symptomatology at baseline compared with the ASCT (median age, 41 years) and CT (median age, 37 years) groups. Statistically significant differences of 10 or more points on the 0 to 100 scales were found for 10 of 15 scales and items (P< or =.01) between the SCT and ASCT groups. Global quality of life (79 v 58, P<.0001), role function (83 v 65, P = .001), sleep disturbances (6 v 28, P<.0001), and fatigue (25 v 44, P = .0001) deviated most. The differences were 10 or more points for seven of 15 scales and items comparing the SCT and CT groups, with sleep disturbances (6 v 35, P<.0001) and pain (11 v 29, P<.01) deviating most. Differences across groups were smaller after 1 year; cognitive function was the only scale with a statistically significant difference (ASCT 80 v CT 89; P = .002). Patterns of change in HRQOL scores were different between groups during follow-up. A great improvement was found in the ASCT group (P<.01 for emotional and role function, fatigue, appetite, and constipation), whereas no significant changes were observed for the SCT group. Prospective studies with extended follow-up periods are necessary to separate a slow recovery process from more permanently reduced HRQOL after transplantation and to examine the late side effects from previous treatment.


Carcinoma of the penis--appraisal of a modified tumour-staging system.
To evaluate variables for the prediction of lymph node metastases in carcinoma of the penis, using a recently proposed modified tumour-staging system that combines the histological degree of differentiation and extent of local invasion of the primary tumour. Thirty-five patients with squamous carcinoma of the penis and histo- or cytological staging of the inguinal lymph nodes were reviewed. A clinical TNM staging system was used in which the size (diameter) of the primary tumour and the clinical extent of invasion were considered. Subsequently, the tumours were also staged according to a modified T-system in which the histological degree of differentiation and pathological extent of tumour invasion were combined. Penectomy was performed in 34 patients (partial amputation in 20 and radical penectomy in 17). Inguinal lymphadenectomy was performed in 31 patients and in four the presence of lymph node metastases was confirmed by aspiration cytology. Using the clinical TNM staging system, lymph node metastases were histo- or cytologically present in no patients with T1, in five of 19 with T2, in 10 of 13 with T3 and in both patients with T4 tumours. Lymph node metastases were present in two of eight patients without clinically palpable inguinal nodes, in three of 14 with nodes clinically thought to be infective and in 11 of 12 nodes clinically considered to be malignant. Lymph node metastases were present in five of 17 patients with grade 1, in nine of 13 with grade 2 and in three of five with grade 3 tumours. Using the modified histological T-staging system (T1 = grade 1-2, invasive through dermis; T2 = any grade, invasion of corpus spongiosum or cavernosum; T3 = any grade, invasion of urethra; T4 = grade 3, regardless of invasion) lymph node metastases were present in one of nine patients with T1, in eight of 16 with T2, in all five with T3 and in three of five with T4 tumours. The modified T-staging system, which combines histological differentiation with pathological extent of invasion, provided the best predictive distinction between T1 and T2-4 tumours, indicating that lymphadenectomy can be avoided in T1 tumours, but should be performed in all patients with T2-4 tumours. We recommend bilateral inguinal lymphadenectomy 6-8 weeks after penectomy in such patients.


Severity of alcohol use and problem behaviors among school-based youths in Puerto Rico.
The present study sought to: (a) categorize youths into groups based on their level of alcohol use and number of symptoms of alcohol abuse and dependence defined in the Diagnostic and Statistical Manual of Mental Disorders (DSM-IV), and (b) examine whether these categories were associated with other problem behaviors in which youths engage (marijuana use, sexual intercourse, and having been arrested or having trouble with the law). The study is based on a cross-sectional survey administered to 972 school-based youths from one middle school and one high school in San Juan, Puerto Rico. Youths were categorized based on their alcohol use and alcohol problems. These categories were then examined for associations with lifetime marijuana use, lifetime sexual intercourse, and having been arrested or having had trouble with the law in the past year. The original eight categories of alcohol use were collapsed into six categories based on the results. For virtually every group characterized by higher severity of alcohol use and alcohol problems, researchers found an increasing prevalence of marijuana use in their lifetimes, increasing odds of sexual intercourse in their lifetimes, and having had trouble with the law in the past year. Knowing about variations in alcohol use and alcohol problems may be instrumental in measuring the degree to which youths may also be engaging in a range of other elevated risk behaviors and a progression to more serious forms of alcohol and drug use.