Syndromic surveillance for influenza in the emergency department-A systematic review.
The science of surveillance is rapidly evolving due to changes in public health information and preparedness as national security issues, new information technologies and health reform. As the Emergency Department has become a much more utilized venue for acute care, it has also become a more attractive data source for disease surveillance. In recent years, influenza surveillance from the Emergency Department has increased in scope and breadth and has resulted in innovative and increasingly accepted methods of surveillance for influenza and influenza-like-illness (ILI). We undertook a systematic review of published Emergency Department-based influenza and ILI syndromic surveillance systems. A PubMed search using the keywords "syndromic", "surveillance", "influenza" and "emergency" was performed. Manuscripts were included in the analysis if they described (1) data from an Emergency Department (2) surveillance of influenza or ILI and (3) syndromic or clinical data. Meeting abstracts were excluded. The references of included manuscripts were examined for additional studies. A total of 38 manuscripts met the inclusion criteria, describing 24 discrete syndromic surveillance systems. Emergency Department-based influenza syndromic surveillance has been described worldwide. A wide variety of clinical data was used for surveillance, including chief complaint/presentation, preliminary or discharge diagnosis, free text analysis of the entire medical record, Google flu trends, calls to teletriage and help lines, ambulance dispatch calls, case reports of H1N1 in the media, markers of ED crowding, admission and Left Without Being Seen rates. Syndromes used to capture influenza rates were nearly always related to ILI (i.e. fever +/- a respiratory or constitutional complaint), however, other syndromes used for surveillance included fever alone, "respiratory complaint" and seizure. Two very large surveillance networks, the North American DiSTRIBuTE network and the European Triple S system have collected large-scale Emergency Department-based influenza and ILI syndromic surveillance data. Syndromic surveillance for influenza and ILI from the Emergency Department is becoming more prevalent as a measure of yearly influenza outbreaks.


Intrinsically Fluorescent, Stealth Polypyrazoline Nanoparticles with Large Stokes Shift for In Vivo Imaging.
Recent advances in super-resolution microscopy and fluorescence bioimaging allow exploring previously inaccessible biological processes. To this end, there is a need for novel fluorescent probes with specific features in size, photophysical properties, colloidal and optical stabilities, as well as biocompatibility and ability to evade the reticuloendothelial system. Herein, novel fluorescent nanoparticles are introduced based on an inherently fluorescent polypyrazoline (PPy) core and a polyethylene glycol (PEG) shell, which address all aforementioned challenges. Synthesis of the PPy-PEG amphiphilic block copolymer by phototriggered step-growth polymerization is investigated by NMR spectroscopy, size-exclusion chromatography, and mass spectrometry. The corresponding nanoparticles are characterized for their luminescent properties and hydrodynamic size in various aqueous environments (e.g., cell culture media). PPy nanoparticles particularly exhibit a large Stokes shift (Δλ = 160 nm or Δν > 7000 cm-1 ) with visible light excitation and strong colloidal stability. While clearance by macrophages and endothelial cells is minimal, PPy displays good biocompatibility. Finally, PPy nanoparticles prove to be long circulating when injected in zebrafish embryos, as observed by in vivo time-lapse fluorescence microscopy. In summary, PPy nanoparticles are highly promising to be further developed as fluorescent nanodelivery systems with low toxicity and exquisite retention in the blood stream.


Hierarchical porous carbons with layer-by-layer motif architectures from confined soft-template self-assembly in layered materials.
Although various two-dimensional (2D) nanomaterials have been explored as promising capacitive materials due to their unique layered structure, their natural restacking tendency impedes electrolyte transport and significantly restricts their practical applications. Herein, we synthesize all-carbon layer-by-layer motif architectures by introducing 2D ordered mesoporous carbons (OMC) within the interlayer space of 2D nanomaterials. As a proof of concept, MXenes are selected as 2D hosts to design 2D-2D heterostructures. Further removing the metal elements from MXenes leads to the formation of all-carbon 2D-2D heterostructures consisting of alternating layers of MXene-derived carbon (MDC) and OMC. The OMC layers intercalated with the MDC layers not only prevent restacking but also facilitate ion diffusion and electron transfer. The performance of the obtained hybrid carbons as supercapacitor electrodes demonstrates their potential for upcoming electronic devices. This method allows to overcome the restacking and blocking of 2D nanomaterials by constructing ion-accessible OMC within the 2D host material.


Maternal antibodies to infectious agents and risk for non-affective psychoses in the offspring--a matched case-control study.
An increasing number of studies suggest that certain maternal infections are associated with non-affective psychoses in the offspring. Here we investigated if maternal exposure to Toxoplasma gondii, cytomegalovirus (CMV), herpes simplex virus type 1 (HSV-1) or type 2 (HSV-2) prior to delivery was associated with future diagnosis of schizophrenia or other non-affective psychoses in the offspring. This case-control study included 198 individuals born in Sweden 1975-85, diagnosed with schizophrenia (ICD-10, F20) and other non-affective psychoses (ICD-10, F21-29) as in- or outpatients, and 524 matched controls. Specific immunoglobulin G (IgG) levels in archived neonatal dried blood samples from these individuals were determined by immunoassays. Reference levels were determined by prevalences among pregnant women in Sweden 1975-85. Odds ratios (OR) for schizophrenia and other non-affective psychoses were calculated, considering maternal and gestational factors as covariates. Levels of IgG directed at T. gondii corresponding to maternal exposure was associated with subsequent schizophrenia (OR=2.1, 95% CI 1.0-4.5) as were levels of IgG directed at CMV (OR=2.2, 95% CI 1.0-5.1) but not at HSV-1 or -2. There were even stronger associations with higher levels of T. gondii or CMV antibodies. There were no associations between any of the infectious agents and other non-affective psychoses. This study supports findings of maternal exposure to T. gondii and schizophrenia risk in offspring, and extends the risk to also include maternal exposure to CMV. Future studies should confirm the association with CMV exposure and identify mechanisms underlying these associations.


Temporal Trends in Sports Participation among Adolescents between 2001 and 2015: A French School- and Territory-Based Study.
Improving adolescents' levels of sport and physical activity (PA) is an official public health issue. French national government plans were launched in 2001, 2006, and 2011 to improve the participation levels of citizens. These plans should be monitored. To date, information on temporal trends in sports has come from the national population. However, no data are available to measure temporal trends in different territories across the country. Our study aimed to measure these trends among a representative sample of adolescent students of the third biggest French region (Bouches-du-Rhône), but also one of the poorest, between 2001 and 2015. Three surveys were conducted in 2001, 2008, and 2015 in high schools (n = 3218). Logistic regressions adjusted for age were used to determine the impact of socioeconomic status (SES) on sports participation and to measure the changes in sport participation rates. Participation declined among all subgroups of adolescents: from 79.0% to 65.8%. The greatest decrease was observed for boys with a high SES, whilst the lowest was for the high-SES girls. We observed that SES inequalities in access to sport increased among the girls, whilst they reduced among the boys. National government plans seem to have had limited success in this territory. Next to national studies, there is a need to develop territory specific studies which could show important disparities across the national territory.


IKKalpha negatively regulates IRF-5 function in a MyD88-TRAF6 pathway.
Transcription factors of IRF family, IRF-3, IRF-5 and IRF-7 play a critical role in the innate antiviral response. In infected cells, IRF-3 and IRF-7 are activated by TBK-1 and IKK epsilon mediated phosphorylation, while the kinase, phosphorylating IRF-5 in the MyD88 signalling pathway has not yet been identified. We now show that IKK alpha phosphorylates IRF-5 and induces formation of IRF-5 dimers, which have been indicative of IRF-5 activation. However, IKK alpha induced IRF-5 phosphorylation exerts inhibitory effect on the transcriptional activation of type 1 interferon and promoters of the inflammatory cytokines. Addressing the molecular mechanism of IKK alpha mediated inhibition of IRF-5 activity, we show that phosphorylation of IRF-5 by IKK alpha inhibits K63 ubiquitination that is essential for IRF-5 activity. Furthermore, we have identified interaction of IRF-5 with alkaline phosphatase, which causes its de-phosphorylation. The observation that MyD88 activated IRF-5 induces expression of alkaline phosphatase suggests that IRF-5 is under autoregulating loop. Thus these completely new observations identify IKK alpha kinase and alkaline phosphatase as negative regulators of IRF-5 activity in MyD88 pathway and implicate their role in the control of the inflammatory response by attenuation of IRF-5 activity.


The Complete Genome of the Atypical Enteropathogenic Escherichia coli Archetype Isolate E110019 Highlights a Role for Plasmids in Dissemination of the Type III Secreted Effector EspT.
Enteropathogenic Escherichia coli (EPEC) is a leading cause of moderate to severe diarrhea among young children in developing countries, and EPEC isolates can be subdivided into two groups. Typical EPEC (tEPEC) bacteria are characterized by the presence of both the locus of enterocyte effacement (LEE) and the plasmid-encoded bundle-forming pilus (BFP), which are involved in adherence and translocation of type III effectors into the host cells. Atypical EPEC (aEPEC) bacteria also contain the LEE but lack the BFP. In the current report, we describe the complete genome of outbreak-associated aEPEC isolate E110019, which carries four plasmids. Comparative genomic analysis demonstrated that the type III secreted effector EspT gene, an autotransporter gene, a hemolysin gene, and putative fimbrial genes are all carried on plasmids. Further investigation of 65 espT-containing E. coli genomes demonstrated that different espT alleles are associated with multiple plasmids that differ in their overall gene content from the E110019 espT-containing plasmid. EspT has been previously described with respect to its role in the ability of E110019 to invade host cells. While other type III secreted effectors of E. coli have been identified on insertion elements and prophages of the chromosome, we demonstrated in the current study that the espT gene is located on multiple unique plasmids. These findings highlight a role of plasmids in dissemination of a unique E. coli type III secreted effector that is involved in host invasion and severe diarrheal illness.


Enhanced Oncolytic Activities of the Telomerase-Specific Replication-Competent Adenovirus Expressing Short-Hairpin RNA against Dicer.
Oncolytic viruses have been receiving much attention as potential agents for cancer treatment. Among the various types of oncolytic viruses, the telomerase-specific replication-competent adenovirus (TRAD), which carries the tumor-specific promoter-driven E1 gene expression cassette, exhibits efficient antitumor effects. The development of a novel TRAD that shows higher replication efficiency and antitumor activity would be highly beneficial for safer and more efficient cancer therapy. We recently demonstrated that the endoribonuclease Dicer significantly inhibits the replication of wild-type adenovirus (Ad) via the processing of viral-associated (VA)-RNAs, which are Ad-encoded small noncoding RNAs, and that the knockdown of Dicer leads to enhanced VA-RNA expression and Ad replication after infection with wild-type Ad. Based on these findings, we herein developed a novel TRAD expressing short-hairpin RNA against Dicer (shDicer; TRAD-shDicer). After infection, TRAD-shDicer efficiently induced the knockdown of Dicer. TRAD-shDicer showed significantly higher replication efficiency and tumor cell lysis activity compared with the conventional TRAD in tumor cells. The Dicer expression levels and viabilities of normal cells were not altered by infection with TRAD-shDicer. These results indicate that TRAD-shDicer is a potent antitumor reagent by virtue of its enhanced oncolytic activity. Mol Cancer Ther; 16(1); 251-9. ©2016 AACR.


A comparison of breast muscle characteristics in three broiler great-grandparent lines.
Genetic selection of broiler chickens has led to a gross overdevelopment of the broiler breast muscle pectoralis major. This may have resulted in increased myopathy and detrimental effects on meat quality. The present study examined 3 commercial great-grandparent lines (lines A, B, and C). Lines A and B are female lines, and line C is a male line. The mean BW of line C (2.7 kg) was significantly greater than those of lines A and B (both 2.3 kg). However, the mean breast yield of both lines B and C (8.9 and 8.7%, respectively) was significantly greater than that of line A (6.9%). Line B therefore matched the meat yield of line A while maintaining a high reproductive capacity. The mean breast fillet weight of line A (169 g) was significantly lower than lines B (207 g) and C (235 g). No differences were observed between lines in either mean fiber size or amount of connective tissue. Therefore, additional fibers must provide the additional weight in the breast fillet of lines B and C, compared with A. Plasma creatine kinase activity, a commonly used marker of muscle damage, was significantly higher in line A (1368 IU/L) than in lines B (995 IU/L) and C (982 IU/L). However, qualitative evaluations of muscle pathology revealed no differences among lines. Selection for increased embryonic muscle fiber number, rather than for increased radial fiber growth, could improve growth potential and may also alleviate muscle damage.


Use of computerised general practice data for population surveillance: comparative study of influenza data.
To assess the potential for using routine computerised general practice data for surveillance of illness. Comparison of the incidence of influenza during the 1989 epidemic derived from a computerised database with that derived from the Royal College of General Practitioners's weekly returns service--a well established predominantly manual surveillance system. 433 general practices throughout the United Kingdom that used a commercial computer system linked to a central databank. Incidence of influenza. The slope of the influenza epidemic curve was essentially the same whether derived from the routine computerised data or royal college's weekly returns service data, and the computerised data were geographically consistent. Throughout the study period, however, the computer derived incidence was between one third and one quarter of that derived from the royal college's system (which is served by practitioners trained in surveillance methods). The peak weekly rates were 164 cases per 100,000 for the computerised system and 583 cases per 100,000 for the royal college's surveillance system. The apparent underreporting in the routine computerised data probably reflects lack of motivation and experience in disease surveillance and haphazard computer entry (particularly of consultations that took place outside of the surgery and consultations that did not result in a prescription), along with overestimation of the population under surveillance. Nevertheless, routine computerised surveillance allows rapid data collection from a large number of practices over a wide geographical area and would greatly augment existing methods.


Atp-binding cassette transporter ABC2/ABCA2 in the rat brain: a novel mammalian lysosome-associated membrane protein and a specific marker for oligodendrocytes but not for myelin sheaths.
We recently cloned a full-length cDNA of the rat ATP-binding cassette transporter 2 (ABC2, or ABCA2) protein, a member of the ABC1 (or ABCA) subfamily (-ABC1/ABCA1 is a causal gene for Tangier disease) and found it to be strongly expressed in the rat brain. In this study, we identified ABC2 as a lysosome-associated membrane protein that is being localized specifically in oligodendrocytes. The ABC2-immunolabeled cells were detected mainly in the white matter but were also scattered in gray matter throughout the whole brain. In addition, these cells were found to be colocalized with 2',3'-cyclic nucleotide-3'-phosphodiesterase (CNPase) immunoreactivity when the marker antibody for oligodendrocytes was used. However, no such colocalization was observed with markers for other kinds of glial cells. Unlike the CNP antibody, which also intensely stains myelin sheaths in the white matter, ABC2 immunoreactivity was detected only in the cell bodies of oligodendrocytes. At the ultrastructural level, ABC2 immunoreactivity was detected mostly around lysosome and partly in Golgi apparatus by electron microscopy. This was confirmed by immunocolocalization of ABC2 and lysosomal markers in a neuroblastoma cell line. Immunoblotting analysis of ABC2 from the whole brain and the ABC2-transfected cell line revealed bands at approximately 260 kDa. The result of in situ hybridization with a riboprobe for ABC2 matched the results obtained from immunostaining. These findings strongly suggest that ABC2 is a specific marker for oligodendrocytes but not for myelinsheaths and that it is as a novel mammalian lysosome-associated membrane protein involved in myelinization or other kinds of metabolism in the CNS.


Childhood acute lymphoblastic leukemia with both t(1;19) and t(9;22).
The chromosomal rearrangements t(1;19)(q23;p13.3) and t(9;22) (q34;q11.2) are independent abnormalities commonly observed in the blast cells of children with acute lymphoblastic leukemia (ALL). We report three children whose leukemic cells contained both translocations at diagnosis. The patients, two males aged 3 and 8 years and a female aged 14 years, all presented with central nervous system involvement. One patient exhibited a pre-B leukemic phenotype (cytoplasmic immunoglobulin, cIg, positive), while two had an early pre-B phenotype (cIg negative). All three patients received radiotherapy and multiagent chemotherapy which included an epipodophyllotoxin in two patients. Two patients suffered relapses of ALL, in both cases with disappearance of t(1;19)-containing clones but persistence of t(9;22). The two patients who received an epipodophyllotoxin as part of their chemotherapeutic regimen both developed secondary myeloid leukemia with entirely new cytogenetic findings, including abnormalities of chromosome band 11q23. These patients are the first to be described with this unusual combination of cytogenetic abnormalities.


Progesterone-induced changes in exercise performance and ventilatory response.
To further study the relationship between ventilatory response (VR) and exercise performance, and to investigate to what extent progesterone is responsible for ventilatory changes in the luteal phase of the menstrual cycle, we administered medroxyprogesterone acetate (MPA) to 10 normal males (20 mg three times a day for 5 doses) and compared results with those obtained in a similar study of females. With MPA, there was an increase in the resting VR to hypercapnia; the resting VR to hypoxia was not changed. There was a respiratory alkalosis at rest. During exercise, the PaCO2 remained lower but the pHa was not different because of a tendency toward lower bicarbonate concentration with MPA. Ventilation, when related to CO2 output, was increased at all exercise loads, indicating increased VR to endogenous CO2. However, ventilation was only minimally (3%) increased when related to oxygen uptake or workload. This apparent disparity is because of slightly lower CO2 output at a given oxygen uptake with MPA. As in females, maximal duration of exercise and maximal oxygen uptake were unchanged. Except for degree, MPA induced all the ventilatory changes seen in the menstrual cycle. Increased VR does not adversely affect exercise performance.


The influence of religion on sexual HIV risk.
This systematic review examines the relationship between religion and sexual HIV risk behavior. It focuses primarily on how studies have conceptualized and defined religion, methodologies, and sexual risk outcomes. We also describe regions where studies were conducted and mechanisms by which religion may be associated with sexual risk. We included 137 studies in this review, classifying them as measuring: (1) only religious affiliation (n = 57), (2) only religiosity (n = 48), and (3) both religious affiliation and religiosity (n = 32). A number of studies identified lower levels of sexual HIV risk among Muslims, although many of these examined HIV prevalence rather than specific behavioral risk outcomes. Most studies identified increased religiosity to be associated with lower levels of sexual HIV risk. This finding persists but is weaker when the outcome considered is condom use. The paper reviews ways in which religion may contribute to increase and reduction in sexual HIV risk, gaps in research, and implications for future research on religion and HIV.


Lateral neural model of binocular rivalry.
This article introduces a two-dimensionally extended, neuron-based model for binocular rivalry. The basic block of the model is a certain type of astable multivibrator comprising excitatory and inhibitory neurons. Many of these blocks are laterally coupled on a medium range to provide a two-dimensional layer. Our model, like others, needs noise to reproduce typical stochastic oscillations. Due to its spatial extension, the noise has to be laterally correlated. When the contrast ratio of the pictures varies, their share of the perception time changes in a way that is known from comparable experimental data (Levelt, 1965; Mueller & Blake, 1989). This is a result of the lateral coupling and not a property of the single model block. The presentation of simple and suitable inhomogeneous stimuli leads to an easily describable perception of periodically moving pictures like propagating fronts or breathing spots. This suggests new experiments. Under certain conditions, a bifurcation from static to moving perceptions is predicted and may be checked and employed by future experiments. Recent "paradox" (Logothetis, 1999) observations of two different neuron classes in cortical areas MT (Logothetis & Schall, 1989) and V4 (Leopold & Logothetis, 1996), one that behaves alike under rivaling and nonrivaling conditions and another that drastically changes its behavior, are interpreted as being related to separate inhibitor neurons.


Medications used in overdose and how they are acquired - an investigation of cases attending an inner Melbourne emergency department.
This study aimed to investigate which categories of medication are most commonly implicated in overdose, to compare this information with prescription data and to explore how the medications used in overdoses are typically acquired. A 12-month audit (11/2003-10/2004) of all medication overdose presentations to an inner-Melbourne ED was conducted and the medications compared to published population-based prescription data. Interviews were conducted with 31 patients who attended the ED following a medication overdose and typical stories regarding the acquisition of medications reported. The same broad categories of medications identified in earlier studies were found to contribute to the majority of overdoses in this study, namely benzodiazepines, antidepressants, analgesics and antipsychotics. Two benzodiazepine medications, diazepam and alprazolam, appeared to be over-represented in the overdose data relative to their population rates of prescription. Patient interviews revealed three main reasons for the original acquisition of the medications used in overdose: treatment purposes (77%); recreational use (16%); and overdose (7%). The most common source of medications (68%) used in overdose was prescription by the patient's usual doctor. The high representation of benzodiazepines among medications used in overdose is of ongoing concern. The time of medication prescription and dispensing may be an ideal opportunity for overdose prevention, through judicious prescribing, consideration of treatment alternatives, patient education and encouraging the safe disposal of unused medications.


Cost-effectiveness of natalizumab vs fingolimod for the treatment of relapsing-remitting multiple sclerosis: analyses in Sweden.
To assess the cost-effectiveness of natalizumab vs fingolimod over 2 years in relapsing-remitting multiple sclerosis (RRMS) patients and patients with rapidly evolving severe disease in Sweden. A decision analytic model was developed to estimate the incremental cost per relapse avoided of natalizumab and fingolimod from the perspective of the Swedish healthcare system. Modeled 2-year costs in Swedish kronor of treating RRMS patients included drug acquisition costs, administration and monitoring costs, and costs of treating MS relapses. Effectiveness was measured in terms of MS relapses avoided using data from the AFFIRM and FREEDOMS trials for all patients with RRMS and from post-hoc sub-group analyses for patients with rapidly evolving severe disease. Probabilistic sensitivity analyses were conducted to assess uncertainty. The analysis showed that, in all patients with MS, treatment with fingolimod costs less (440,463 Kr vs 444,324 Kr), but treatment with natalizumab results in more relapses avoided (0.74 vs 0.59), resulting in an incremental cost-effectiveness ratio (ICER) of 25,448 Kr per relapse avoided. In patients with rapidly evolving severe disease, natalizumab dominated fingolimod. Results of the sensitivity analysis demonstrate the robustness of the model results. At a willingness-to-pay (WTP) threshold of 500,000 Kr per relapse avoided, natalizumab is cost-effective in >80% of simulations in both patient populations. Limitations include absence of data from direct head-to-head studies comparing natalizumab and fingolimod, use of relapse rate reduction rather than sustained disability progression as the primary model outcome, assumption of 100% adherence to MS treatment, and exclusion of adverse event costs in the model. Natalizumab remains a cost-effective treatment option for patients with MS in Sweden. In the RRMS patient population, the incremental cost per relapse avoided is well below a 500,000 Kr WTP threshold per relapse avoided. In the rapidly evolving severe disease patient population, natalizumab dominates fingolimod.


Financing drug discovery for orphan diseases.
Recently proposed 'megafund' financing methods for funding translational medicine and drug development require billions of dollars in capital per megafund to de-risk the drug discovery process enough to issue long-term bonds. Here, we demonstrate that the same financing methods can be applied to orphan drug development but, because of the unique nature of orphan diseases and therapeutics (lower development costs, faster FDA approval times, lower failure rates and lower correlation of failures among disease targets) the amount of capital needed to de-risk such portfolios is much lower in this field. Numerical simulations suggest that an orphan disease megafund of only US$575 million can yield double-digit expected rates of return with only 10-20 projects in the portfolio.


Mandatory parental involvement in minors' abortions: effects of the laws in Minnesota, Missouri, and Indiana.
This study examined the effects of parental involvement laws on the birth rate, in-state abortion rate, odds of interstate travel, and odds of late abortion for minors. Poisson and logistic regression models fitted to vital records compared the periods before and after the laws were enforced. In each state, the in-state abortion rate for minors fell (relative to the rate for older women) when parental involvement laws took effect. Data offered no empirical support for the proposition that the laws drive up birth rates for minors. Although data were incomplete, the laws appeared to increase the odds of a minor's traveling out of state for her abortion. If one judges from the available data, minors who traveled out of state may have accounted for the entire observed decline in the in-state abortion rate, at least in Missouri. The laws appeared to delay minors' abortions past the eighth week, but probably not into the second trimester. Several empirical arguments used against and in support of parental involvement laws do not appear to be substantiated.


Brain microvascular injury and white matter disease provoked by diabetes-associated hyperamylinemia.
The brain blood vessels of patients with type 2 diabetes and dementia have deposition of amylin, an amyloidogenic hormone cosecreted with insulin. It is not known whether vascular amylin deposition is a consequence or a trigger of vascular injury. We tested the hypothesis that the vascular amylin deposits cause endothelial dysfunction and microvascular injury and are modulated by amylin transport in the brain via plasma apolipoproteins. Rats overexpressing amyloidogenic (human) amylin in the pancreas (HIP rats) and amylin knockout (AKO) rats intravenously infused with aggregated amylin were used for in vivo phenotyping. We also carried out biochemical analyses of human brain tissues and studied the effects of the aggregated amylin on endothelial cells ex vivo. Amylin deposition in brain blood vessels is associated with vessel wall disruption and abnormal surrounding neuropil in patients with type 2 diabetes and dementia, in HIP rats, and in AKO rats infused with aggregated amylin. HIP rats have brain microhemorrhages, white matter injury, and neurologic deficits. Vascular amylin deposition provokes loss of endothelial cell coverage and tight junctions. Intravenous infusion in AKO rats of human amylin, or combined human amylin and apolipoprotein E4, showed that amylin binds to plasma apolipoproteins. The intravenous infusion of apolipoprotein E4 exacerbated the brain accumulation of aggregated amylin and vascular pathology in HIP rats. These data identify vascular amylin deposition as a trigger of brain endothelial dysfunction that is modulated by plasma apolipoproteins and represents a potential therapeutic target in diabetes-associated dementia and stroke. Ann Neurol 2017;82:208-222.


The temporal dynamics of differential gene expression in Aspergillus fumigatus interacting with human immature dendritic cells in vitro.
Dendritic cells (DC) are the most important antigen presenting cells and play a pivotal role in host immunity to infectious agents by acting as a bridge between the innate and adaptive immune systems. Monocyte-derived immature DCs (iDC) were infected with viable resting conidia of Aspergillus fumigatus (Af293) for 12 hours at an MOI of 5; cells were sampled every three hours. RNA was extracted from both organisms at each time point and hybridised to microarrays. iDC cell death increased at 6 h in the presence of A. fumigatus which coincided with fungal germ tube emergence; >80% of conidia were associated with iDC. Over the time course A. fumigatus differentially regulated 210 genes, FunCat analysis indicated significant up-regulation of genes involved in fermentation, drug transport, pathogenesis and response to oxidative stress. Genes related to cytotoxicity were differentially regulated but the gliotoxin biosynthesis genes were down regulated over the time course, while Aspf1 was up-regulated at 9 h and 12 h. There was an up-regulation of genes in the subtelomeric regions of the genome as the interaction progressed. The genes up-regulated by iDC in the presence of A. fumigatus indicated that they were producing a pro-inflammatory response which was consistent with previous transcriptome studies of iDC interacting with A. fumigatus germ tubes. This study shows that A. fumigatus adapts to phagocytosis by iDCs by utilising genes that allow it to survive the interaction rather than just up-regulation of specific virulence genes.


Interaction of polycationic antibiotics with Pseudomonas aeruginosa lipopolysaccharide and lipid A studied by using dansyl-polymyxin.
A fluorescent derivative of polymyxin B (dansyl-polymyxin) was used to study the interaction of polycations with lipopolysaccharide (LPS) and lipid A from Pseudomonas aeruginosa. Dansyl-polymyxin became bound to LPS and lipid A sites, including Mg2+-binding sites, resulting in a 20-fold enhancement of fluorescence. A Hill plot of the binding data showed that the binding of dansyl-polymyxin to LPS was cooperative (n = 1.98) and of high affinity (S0.5 = 0.38 microM). The maximal binding capacity of LPS was approximately four molecules of dansyl-polymyxin per mol of LPS. The dansyl-polymyxin interaction with lipid A displayed similar kinetics (n = 2.26; S0.5 = 0.38 microM), and the maximal binding capacity was approximately 2 mol of dansyl-polymyxin per mol of lipid A. A variety of polycationic compounds, including gentamicin, streptomycin, and polymyxin B, as well as Mg2+, were able to displace dansyl-polymyxin bound to LPS or to lipid A. Marked differences both in terms of the degree of displacement and in terms of the amount of competing polycation required to displace a given amount of dansyl-polymyxin were observed. Addition of excess polymyxin B resulted in displacement of all of the dansyl-polymyxin, demonstrating that only polymyxin-binding sites were being probed. Our data demonstrate that polymyxin B binds to multiple sites on LPS, including sites which bind aminoglycoside antibiotics and other polycationic compounds.


Subtraction SPECT co-registered to MRI improves postictal SPECT localization of seizure foci.
To determine whether the detection of focal hypoperfusion by subtraction SPECT co-registered to MRI (SISCOM) improves the sensitivity and specificity of postictal SPECT in intractable partial epilepsy. Postictal SPECT injections are easier to perform than are ictal injections, but the images are more difficult to interpret and have been reported to have lower sensitivity and specificity. Thirty-five consecutive intractable partial epilepsy patients who had postictal SPECT studies were evaluated. The following sets of SPECT images were separately interpreted by three blinded reviewers and classified as either localizing to 1 of 16 possible sites in the brain or as nonlocalizing: unsubtracted postictal and interictal images for conventional side-by-side comparison, SISCOM images of hyperperfusion, SISCOM images of hypoperfusion, and both sets of SISCOM hyperperfusion and hypoperfusion images (combined SISCOM evaluation). Significantly higher proportions of the hyperperfusion SISCOM images (65.7%), the hypoperfusion SISCOM images (74.3%), and the combined SISCOM evaluation (82.9%) were localizing than were the conventional method of side-by-side comparison of unsubtracted images (31.4%; p < 0.0001). Concordance with the discharge diagnosis was higher for the combined SISCOM evaluation than it was for either the hyperperfusion or the hypoperfusion SISCOM images alone (both p < 0.05). For the hypoperfusion SISCOM and the combined SISCOM evaluations, concordance of the localization with the site of epilepsy surgery was associated with a greater probability of an excellent outcome than were nonconcordant/nonlocalizing images (both p < 0.05). The use of SISCOM to detect focal cerebral hypoperfusion, in addition to focal hyperperfusion, improves the sensitivity and specificity of postictal SPECT in intractable partial epilepsy.


Neurofilament Light Chain: Blood Biomarker of Neonatal Neuronal Injury.
Background: Neurofilament light chain (NfL) is a highly promising biomarker of neuroaxonal injury that has mainly been studied in adult neurodegenerative disease. Its involvement in neonatal disease remains largely unknown. Our aim was to establish NfL plasma concentrations in preterm and term infants in the first week of life. Methods: Plasma NfL was measured by single molecule array immunoassay in two neonatal cohorts: cohort 1 contained 203 term and preterm infants, median gestational age (GA) 37.9 weeks (interquartile range [IQR] 31.9-39.4), in whom venous and arterial umbilical cord blood was sampled at birth and venous blood at day of life (DOL) 3; cohort 2 contained 98 preterm infants, median GA 29.3 weeks (IQR 26.9-30.6), in whom venous blood was sampled at DOL 7. Results: Median NfL concentrations in venous blood increased significantly from birth (18.2 pg/mL [IQR 12.8-30.8, cohort 1]) to DOL 3 (50.9 pg/mL [41.3-100, cohort 1]) and DOL 7 (126 pg/mL [78.8-225, cohort 2]) (p < 0.001). In both cohorts NfL correlated inversely with birth weight (BW, Spearman's rho -0.403, p < 0.001, cohort 1; R -0.525, p < 0.001, cohort 2) and GA (R -0.271, p < 0.001, cohort 1; R -0.487, p < 0.001, cohort 2). Additional significant correlations were found for maternal age at delivery, preeclampsia, delivery mode, 5-min Apgar, duration of oxygen supplementation, sepsis, and brain damage (intraventricular hemorrhage or periventricular leukomalacia). Multivariable logistic regression analysis identified the independent predictors of NfL in cohort 1 as BW (beta = -0.297, p = 0.003), delivery mode (beta = 0.237, p = 0.001) and preeclampsia (beta = 0.183, p = 0.022) and in cohort 2 as BW (beta = -0.385, p = 0.001) and brain damage (beta = 0.222, p = 0.015). Conclusion: Neonatal NfL levels correlate inversely with maturity and BW, increase during the first days of life, and relate to brain injury factors such as intraventricular hemorrhage and periventricular leukomalacia, and also to vaginal delivery.


A brief measure of predeath grief in dementia caregivers: the Caregiver Grief Questionnaire.
The study of predeath grief is hampered by measures that are often lengthy and not clearly differentiated from other caregiving outcomes, most notably burden. We aimed to validate a new 11-item Caregiver Grief Questionnaire (CGQ) assessing two dimensions of predeath grief, namely relational deprivation and emotional pain. Cross-sectional survey. Community and psychogeriatric clinics. 173 Alzheimer (AD) caregivers who cared for relatives with different degrees of severity (63 mild, 60 moderate, and 50 severe). Besides the CGQ, measures of caregiver burden and depressive symptoms, and care-recipients' neuropsychiatric symptoms and functional impairment were assessed. Confirmatory factor analysis supported the hypothesized 2-factor over the 1-factor model, and both subscales were only moderately correlated with burden. Two-week test-retest reliabilities were excellent. Caregivers for mild AD reported less grief than those caring for more severe relatives. Z tests revealed significantly different correlational patterns for the two dimensions, with emotional pain more related to global burden and depressive symptoms, and relational deprivation more related to care-recipients' functional impairment. Both dimensions were mildly correlated with neuropsychiatric symptoms (especially disruptive behaviors and psychotic symptoms) of the care-recipient. Results supported the reliability and validity of the two-dimensional measure of predeath grief. As a brief measure, it can be readily added to research instruments to facilitate study of this important phenomenon along with other caregiving outcomes.


Short-term administration of disulfiram for reversal of latent HIV infection: a phase 2 dose-escalation study.
In vitro, disulfiram activated HIV transcription in a primary T-cell model of HIV latency and in a pilot clinical study increased plasma HIV RNA in individuals with adequate drug exposure. We assessed the effect of disulfiram on HIV transcription in a dose-escalation study. In this prospective dose-escalation study, to optimise disulfiram exposure we included adults with HIV on suppressive antiretroviral therapy, with plasma HIV RNA of less than 50 copies per mL and a CD4 cell count greater than 350 cells per μL. Participants were allocated sequentially to one of three dosing groups (500 mg, 1000 mg, and 2000 mg) and received disulfiram daily for 3 days. Only the staff who did laboratory assays were masked to group assignment. The primary endpoint was change in cell-associated unspliced HIV RNA in CD4 cells. The primary analysis method was a negative binomial regression, with the number of copies as the outcome variable and the input total RNA or plasma volume as an exposure variable, which is equivalent to modelling copies or input. We used these models to estimate changes from before disulfiram to timepoints during and after disulfiram administration. This study is registered with ClinicalTrials.gov, number NCT01944371. Of 34 participants screened for eligibility at The Alfred Hospital (Melbourne, VIC, Australia), and San Francisco General Hospital (San Francisco, CA, USA), 30 people were enrolled between Sept 24, 2013, and March 31, 2014. The estimated fold increases in cell-associated unspliced HIV RNA from baseline were 1·7 (95% CI 1·3-2·2; p<0·0001) to the timepoint during disulfiram treatment and 2·1 (1·5-2·9; p<0·0001) to the timepoint after disulfiram in the 500 mg group; 1·9 (1·6-2·4; p<0·0001) and 2·5 (1·9-3·3; p<0·0001) in the 1000 mg group; and 1·6 (1·2-2·1; p=0·0026) and 2·1 (1·5-3·1; p=0·0001) in the 2000 mg group. No deaths occurred, and no serious adverse events were noted. Disulfiram was well tolerated at all doses. Short-term administration of disulfiram resulted in increases in cell-associated unspliced HIV RNA at all doses, consistent with activating HIV latency. Disulfiram may be suited for future studies of combination and prolonged therapy to activate latent HIV. The Foundation for AIDS Research (amfAR); National Institute of Allergy and Infectious Diseases, National Institutes of Health; Australian National Health and Medical Research Council.


Low frequency pulsed electromagnetic field--a viable alternative therapy for arthritis.
Arthritis refers to more than 100 disorders of the musculoskeletal system. The existing pharmacological interventions for arthritis offer only symptomatic relief and they are not definitive and curative. Magnetic healing has been known from antiquity and it is evolved to the present times with the advent of electromagnetism. The original basis for the trial of this form of therapy is the interaction between the biological systems with the natural magnetic fields. Optimization of the physical window comprising the electromagnetic field generator and signal properties (frequency, intensity, duration, waveform) with the biological window, inclusive of the experimental model, age and stimulus has helped in achieving consistent beneficial results. Low frequency pulsed electromagnetic field (PEMF) can provide noninvasive, safe and easy to apply method to treat pain, inflammation and dysfunctions associated with rheumatoid arthritis (RA) and osteoarthritis (OA) and PEMF has a long term record of safety. This review focusses on the therapeutic application of PEMF in the treatment of these forms of arthritis. The analysis of various studies (animal models of arthritis, cell culture systems and clinical trials) reporting the use of PEMF for arthritis cure has conclusively shown that PEMF not only alleviates the pain in the arthritis condition but it also affords chondroprotection, exerts antiinflammatory action and helps in bone remodeling and this could be developed as a viable alternative for arthritis therapy.


Flexible high-order QAM transmitter using tandem IQ modulators for generating 16/32/36/64-QAM with balanced complexity in electronics and optics.
In order to adapt to the dynamics in the future optical networks, we propose a flexible high-order QAM transmitter using a tandem in-phase/quadrature (IQ) modulators to synthesize different high-order quadrature amplitude modulation (QAM) formats, such as 16QAM, 32 or 36QAM and 64QAM. To generate high-order QAMs, an offset-QAM is firstly generated using an IQ modulator driven by electronics with reduced modulation-level, and then mapped to other quadrants through another following IQ modulator configured as a standard quadrature phase-shift keying (QPSK) modulator. All of the embedded sub-Mach-Zehnder modulators are operated in push-pull configurations to avoid introducing excess phase chirp. In contrast with the schemes based on a single IQ modulator driven by multilevel electronics or a highly-integrated parallel modulator, by deploying commercially-available optical modulators and driving electronics with reduced modulation-level, the transmitter complexity in optics and electronics is well-balanced. In the case of generating optical 64QAM, different from another tandem scheme deploying dual-drive IQ modulator driven by independent four binary streams, less phase chirp is observed in our proposed scheme, and comparable implementation penalty is obtained even without applying additional specific compensation algorithm in the coherent receiver. Moreover, thanks to the tandem structure and the deployment of QPSK modulator, the obtained high-order QAM is naturally differentially coded, which is helpful to solve the phase ambiguity at coherent receiver. We experimentally demonstrate the generations of these high-order QAMs including 16QAM, 32/36QAM and 64QAM, and confirm the error-free operations with comparable BER performance to the "electrical" approach based on a single IQ modulator.


derrière: a TGF-beta family member required for posterior development in Xenopus.
TGF-beta signaling plays a key role in induction of the Xenopus mesoderm and endoderm. Using a yeast-based selection scheme, we isolated derrière, a novel TGF-beta family member that is closely related to Vg1 and that is required for normal mesodermal patterning, particularly in posterior regions of the embryo. Unlike Vg1, derrière is expressed zygotically, with RNA localized to the future endoderm and mesoderm by late blastula, and to the posterior mesoderm by mid-gastrula. The derrière expression pattern appears to be identical to the zygotic expression domain of VegT (Xombi, Brat, Antipodean), and can be activated by VegT as well as fibroblast growth factor (FGF). In turn, derrière activates expression of itself, VegT and eFGF, suggesting that a regulatory loop exists between these genes. derrière is a potent mesoderm and endoderm inducer, acting in a dose-dependent fashion. When misexpressed ventrally, derrière induces a secondary axis lacking a head, an effect that is due to dorsalization of the ventral marginal zone. When misexpressed dorsally, derrière suppresses head formation. derrière can also posteriorize neurectoderm, but appears to do so indirectly. Together, these data suggest that derrière expression is compatible only with posterior fates. In order to assess the in vivo function of derrière, we constructed a dominant interfering Derrière protein (Cm-Derrière), which preferentially blocks Derrière activity relative to that of other TGFbeta family members. Cm-derrière expression in embryos leads to posterior truncation, including defects in blastopore lip formation, gastrulation and neural tube closure. Normal expression of anterior and hindbrain markers is observed; however, paraxial mesodermal gene expression is ablated. This phenotype can be rescued by wild-type derrière and by VegT. Our findings indicate that derrière plays a crucial role in mesodermal patterning and development of posterior regions in Xenopus.


Position of the Academy of Nutrition and Dietetics: nutrition guidance for healthy children ages 2 to 11 years.
It is the position of the Academy of Nutrition and Dietetics that children ages 2 to 11 years should achieve optimal physical and cognitive development, maintain healthy weights, enjoy food, and reduce the risk of chronic disease through appropriate eating habits and participation in regular physical activity. Rapid increases in the prevalence of childhood obesity during the 1980s and 1990s focused attention on young children's overconsumption of energy-dense, nutrient-poor foods and beverages and lack of physical activity. While recent data suggest a stabilization of obesity rates, several public health concerns remain. These include the most effective ways to promote healthy weights, the number of children living in food insecurity, the under-consumption of key nutrients, and the early development of diet-related risks for chronic diseases, such as cardiovascular disease, type 2 diabetes, cancer, obesity, and osteoporosis. This Position Paper reviews what children 2 to 11 years old in the United States are reportedly eating, explores trends in food and nutrient intakes, and examines the impact of federal nutrition programs on child nutrition. Current dietary recommendations and guidelines for physical activity are also discussed. The roles of parents and caregivers in influencing the development of life-long healthy eating behaviors are highlighted. The Academy of Nutrition and Dietetics works with other allied health and food industry professionals to translate dietary recommendations and guidelines into positive, practical health messages. Specific recommendations and sources of science-based nutrition messages to improve the nutritional well-being of children are provided for food and nutrition practitioners.


Lapatinib, a Dual Inhibitor of Epidermal Growth Factor Receptor (EGFR) and HER-2, Enhances Radiosensitivity in Mouse Bladder Tumor Line-2 (MBT-2) Cells In Vitro and In Vivo.
BACKGROUND The aim of this study was to evaluate the effect of lapatinib, a dual inhibitor of epidermal growth factor receptor (EGFR) and HER-2, on the radiosensitivity of murine bladder tumor line-2 (MBT-2) cells in vitro and in vivo. MATERIAL AND METHODS MBT-2 cells were pretreated with lapatinib at doses ranging from 200-1,000 nM for 30 min followed by radiation at doses ranging from 2.5-10 Gy for 30 min. A clonogenic assay (colony formation assay) assessed cell survival. Western blot measured phosphorylated epidermal growth factor receptor (p-EGFR), phosphorylated AKT (p-AKT), and phosphorylated HER-2 (p-HER2) and the apoptosis marker, PARP. The C3H/HeN mouse tumor xenograft model underwent subcutaneous injection of MBT-2 cells; mice were divided into four groups, treated with lapatinib (200 mg/kg), radiation (15 Gy), a combination of both, and with vehicle (control). RESULTS Lapatinib pretreatment, combined with radiation, decreased MBT-2 cell survival, and suppressed radiation-activated levels of p-EGFR and p-HER-2. MBT-2 cells treated with a 10 Gy dose of radiation and 1000 nM of lapatinib showed combination index (CI) values of <1 indicating synergy. Increased expression of γ-H2AX, indicated increased apoptosis. In mice with tumor xenografts, a daily dose of lapatinib (200 mg/kg/day) for seven days combined with radiation on the fourth day suppressed tumor growth to a greater degree than radiation alone. CONCLUSIONS Lapatinib treatment enhanced the radiation sensitivity in an in vitro and in vivo murine bladder cancer model by decreasing radiation-mediated EGFR and HER-2 activation, and by causing DNA damage leading to cell apoptosis.


Septic shock: a multidisciplinary response team and weekly feedback to clinicians improve the process of care and mortality.
To evaluate the impact of weekly feedback to clinicians and the activation of a sepsis response team on the process of care and hospital mortality in patients with severe sepsis or septic shock. Prospective, interventional cohort study. The medical intensive care unit of a tertiary, academic medical center. Patients with severe sepsis or septic shock consecutively treated in a medical intensive care unit. Daily auditing and weekly feedback, and sepsis response team activation. During a 33-month study period, from January 2007 through September 2009, we performed daily screening of patients for severe sepsis or septic shock. Study periods were divided into baseline (screening only), daily auditing with weekly feedback, and sepsis response team activation. Comparisons among the three periods were made by using univariate and multiple logistic regression analyses. Compliance with the overall sepsis resuscitation bundle and its individual elements and hospital mortality were used as outcome measures. A total of 984 episodes of severe sepsis and septic shock were identified during the study periods, severe sepsis in 52 (5.3%) and septic shock in 932 (94.7%). The compliance rate with all elements of the sepsis resuscitation bundle increased from 12.7% at baseline to 37.7% and 53.7% during the weekly feedback and sepsis response team activation periods, respectively (p < .001). Overall hospital mortality rate was 30.3%, 28.3%, and 22.0% during baseline, weekly feedback, and sepsis response team periods, respectively (p = .029). Multiple logistic regression analysis showed that the sepsis response team was associated with reduced risk of hospital death (odds ratio, 0.657; 95% confidence interval, 0.456-0.945; p = .023) whereas hepatic cirrhosis, hepatic failure, leukemia, multiple myeloma, transfer from the same hospital ward, do-not-resuscitate status at the recognition of severe sepsis/septic shock, and lactate level were associated with increased risk of death. In septic shock, the activation of the sepsis response team in combination with weekly feedback increases the compliance with the process of care and reduces hospital mortality rate.


Classification and characterisation of brain network changes in chronic back pain: A multicenter study.
Background. Chronic pain is a common, often disabling condition thought to involve a combination of peripheral and central neurobiological factors. However, the extent and nature of changes in the brain is poorly understood. Methods. We investigated brain network architecture using resting-state fMRI data in chronic back pain patients in the UK and Japan (41 patients, 56 controls), as well as open data from USA. We applied machine learning and deep learning (conditional variational autoencoder architecture) methods to explore classification of patients/controls based on network connectivity. We then studied the network topology of the data, and developed a multislice modularity method to look for consensus evidence of modular reorganisation in chronic back pain. Results. Machine learning and deep learning allowed reliable classification of patients in a third, independent open data set with an accuracy of 63%, with 68% in cross validation of all data. We identified robust evidence of network hub disruption in chronic pain, most consistently with respect to clustering coefficient and betweenness centrality. We found a consensus pattern of modular reorganisation involving extensive, bilateral regions of sensorimotor cortex, and characterised primarily by negative reorganisation - a tendency for sensorimotor cortex nodes to be less inclined to form pairwise modular links with other brain nodes. Furthermore, these regions were found to display increased connectivity with the pregenual anterior cingulate cortex, a region known to be involved in endogenous pain control. In contrast, intraparietal sulcus displayed a propensity towards positive modular reorganisation, suggesting that it might have a role in forming modules associated with the chronic pain state. Conclusion. The results provide evidence of consistent and characteristic brain network changes in chronic pain, characterised primarily by extensive reorganisation of the network architecture of the sensorimotor cortex.


Improved graft survival after renal transplantation in the United States, 1988 to 1996.
The introduction of cyclosporine has resulted in improvement in the short-term outcome of renal transplantation, but its effect on the long-term survival of kidney transplants is not known. We analyzed the influence of demographic characteristics (age, sex, and race), transplant-related variables (living or cadaveric donor, panel-reactive antibody titer, extent of HLA matching, and cold-ischemia time), and post-transplantation variables (presence or absence of acute rejection, delayed graft function, and therapy with mycophenolate mofetil and tacrolimus) on graft survival for all 93,934 renal transplantations performed in the United States between 1988 and 1996. A regression analysis adjusted for these variables was used to estimate the risk of graft failure within the first year and more than one year after transplantation. From 1988 to 1996, the one-year survival rate for grafts from living donors increased from 88.8 to 93.9 percent, and the rate for cadaveric grafts increased from 75.7 to 87.7 percent. The half-life for grafts from living donors increased steadily from 12.7 to 21.6 years, and that for cadaveric grafts increased from 7.9 to 13.8 years. After censoring of data for patients who died with functioning grafts, the half-life for grafts from living donors increased from 16.9 years to 35.9 years, and that for cadaveric grafts increased from 11.0 years to 19.5 years. The average yearly reduction in the relative hazard of graft failure after one year was 4.2 percent for all recipients (P<0.001), 0.4 percent for those who had acute rejection (P=0.57), and 6.3 percent for those who did not have acute rejection (P<0.001). Since 1988, there has been a substantial increase in short-term and long-term survival of kidney grafts from both living and cadaveric donors.


Beneficial effects of simulated gastro-intestinal digests of fried egg and its fractions on blood pressure, plasma lipids and oxidative stress in spontaneously hypertensive rats.
We have previously characterized several antihypertensive peptides in simulated digests of cooked eggs and showed blood pressure lowering property of fried whole egg digest. However, the long-term effects of this hydrolysate and its fractions on blood pressure are not known. Therefore, the objectives of the study were to determine the effects of long term administration of fried whole egg hydrolysate and its fractions (i.e. egg white and egg yolk) on regulation of blood pressure and associated factors in cardiovascular disease such as plasma lipid profile and tissue oxidative stress. We used spontaneously hypertensive rats (SHR), an animal model of essential hypertension. Hydrolysates of fried egg and its fractions were prepared by simulated gastro-intestinal digestion with pepsin and pancreatin. 16-17 week old male SHRs were orally administered fried whole egg hydrolysate, non-hydrolyzed fried whole egg, egg white hydrolysate or egg yolk hydrolysates (either defatted, or not) daily for 18 days. Blood pressure (BP) and heart rate were monitored by telemetry. Animals were sacrificed at the end of the treatment for vascular function studies and evaluating plasma lipid profile and tissue oxidative stress. BP was reduced by feeding fried whole egg hydrolysate but not by the non-hydrolyzed product suggesting a critical role for in vitro digestion in releasing anti-hypertensive peptides. Egg white hydrolysate and defatted egg yolk hydrolysate (but not egg yolk hydrolysate) also had similar effects. Reduction in BP was accompanied by the restoration of nitric oxide (NO) dependent vasorelaxation and reduction of plasma angiotensin II. Fried whole egg hydrolysate also reduced plasma levels of triglyceride although it was increased by the non-hydrolyzed sample. Additionally the hydrolyzed preparations attenuated tissue oxidative stress. Our results demonstrate that fried egg hydrolysates exert anti-hypertensive effects, improve plasma lipid profile and attenuate tissue oxidative stress in vivo.


Independent interactions of ubiquitin-binding domains in a ubiquitin-mediated ternary complex.
Ubiquitin (Ub) modifications are transduced by receptor proteins that use Ub-binding domains (UBDs) to recognize distinct interaction faces on the Ub surface. We report the nuclear magnetic resonance (NMR) solution structures of the A20-like zinc finger (A20 Znf) UBD of the Ub receptor ZNF216, and its complex with Ub, and show that the binding surface on Ub centered on Asp58 leaves the canonical hydrophobic Ile44 patch free to participate in additional interactions. We have modeled ternary complexes of the different families of UBDs and show that while many are expected to bind competitively to the same Ile44 surface or show steric incompatibility, other combinations (in particular, those involving the A20 Znf domain) are consistent with a single Ub moiety simultaneously participating in multiple interactions with different UBDs. We subsequently demonstrate by NMR that the A20 Znf domain of ZNF216 and the UBA domain of the p62 protein (an Ile44-binding UBD), which function in the same biological pathways, are able to form such a Ub-mediated ternary complex through independent interactions with a single Ub. This work supports an emerging concept of Ub acting as a scaffold to mediate multiprotein complex assembly.


Tirofiban. A review of its use in acute coronary syndromes.
Tirofiban is an intravenously administered nonpeptide glycoprotein IIb/IIIa receptor antagonist which specifically inhibits fibrinogen-dependent platelet aggregation and prolongs bleeding times in patients with acute coronary syndromes. Adenosine diphosphate (ADP)-induced platelet aggregation returns to near-baseline levels within 4 to 8 hours after cessation of a tirofiban infusion, a finding consistent with the drug's elimination half-life of approximately 2 hours. Three large clinical trials have shown that, when administered with a standard heparin and aspirin regimen, tirofiban reduces the risk of ischaemic complications in patients with unstable angina/non-Q-wave myocardial infarction (MI) and in patients undergoing percutaneous revascularisation. In PRISM-PLUS, a study involving 1915 patients with unstable angina/non-Q-wave MI, administration of intravenous tirofiban (0.4 microgram/kg/min loading dose for 30 minutes followed by a 0.10 microgram/kg/min infusion) with heparin for at least 48 (mean 71.3) hours reduced the 7-day risk of the composite end-point of MI, death and refractory ischaemia by 32% compared with heparin alone. The between-group risk reduction remained significant at 30 days (22%) and 6 months (19%). Similarly, in high-risk patients undergoing coronary angioplasty in RESTORE, the addition of tirofiban (10 micrograms/kg bolus in the 3 minutes prior to intervention followed by 0.15 microgram/kg/min for 36 hours) to a standard heparin regimen significantly reduced the risk of ischaemic complications by 38% on day 2 and 27% on day 7 compared with heparin alone. Although interim analysis in PRISM-PLUS showed that the use of tirofiban without heparin increased the 7-day risk of death compared with heparin alone, this finding was inconsistent with the effects of tirofiban on the risk of death in PRISM, a study involving 3232 patients with unstable angina/non-Q-wave MI. Tirofiban is generally well tolerated. Bleeding complications were the most commonly reported events associated with tirofiban in clinical trials, but the rate of major bleeding in tirofiban recipients was not significantly different from that reported with heparin. Thrombocytopenia (platelet count < 90,000 cells/microliter) occurred slightly more frequently with tirofiban (with or without heparin) than with heparin alone. Tirofiban reduces the risk of ischaemic complications in patients with unstable angina/non-Q-wave MI and high-risk patients undergoing revascularisation when used against a background of heparin and aspirin. Furthermore, the drug has an acceptable tolerability profile. Therefore, intravenous tirofiban is likely to be used as an adjunct to heparin and aspirin in patients with acute coronary syndromes including high-risk patients undergoing revascularisation.


Pre-test of questions on health-related resource use and expenditure, using behaviour coding and cognitive interviewing techniques.
Validated instruments collecting data on health-related resource use are lacking, but required, for example, to investigate predictors of healthcare use or for health economic evaluation.The objective of the study was to develop, test and refine a questionnaire collecting data on health-related resource use and expenditure in patients with diabetes. The questionnaire was tested in 43 patients with diabetes mellitus types 1 and 2 in Germany. Response behaviour suggestive of problems with questions (item non-response, request for clarification, comments, inadequate answer, "don't know") was systematically registered. Cognitive interviews focusing on information retrieval and comprehension problems were carried out. Many participants had difficulties answering questions pertaining to frequency of visits to the general practitioner (26%), time spent receiving healthcare services (39%), regular medication currently taken (35%) and out of pocket expenditure on medication (42%). These difficulties seem to result mainly from poor memory. A number of comprehension problems were established and relevant questions were revised accordingly. The questionnaire on health-related resource use and expenditure for use in diabetes research in Germany was developed and refined after careful testing. Ideally, the questionnaire should be externally validated for different modes of administration and recall periods within a variety of populations.


Association of Elevations of Specific T Cell and Monocyte Subpopulations in Rheumatoid Arthritis With Subclinical Coronary Artery Atherosclerosis.
Coronary artery disease (CAD) is the leading cause of excess deaths in rheumatoid arthritis (RA). However, identification of features denoting patients with a risk of developing CAD is lacking. The composition of circulating peripheral blood mononuclear cell (PBMC) subsets in RA patients differs markedly from that in healthy controls with regard to the extent of T cell activation, with clonal expansion and differentiation to effector memory status, and presence of inflammatory monocytes. In this study, we sought to evaluate whether elevations in these PBMC subpopulations in RA patients could denote those with an increased risk of subclinical CAD, as determined by the presence of coronary artery calcification (CAC). The study cohort comprised 72 patients with RA who underwent cardiac computed tomography to assess CAC. PBMC subsets were determined by multiparameter flow cytometry. Multivariable logistic regression was used to determine the associations between PBMC subpopulations and the presence of CAC. Among the 72 patients with RA, 33% had CAC and exhibited significant increases in the levels of circulating CD4 T cell subsets denoting activation and differentiation to the effector memory phenotypes. Analogous increases in the levels of CD8 T cell subsets, as well as in the CD14(high)CD16+ intermediate monocyte subset, were also present in these patients, as compared to those without CAC. The increases in the CD4 and CD8 T cell subsets were highly intercorrelated, whereas the increases in CD14(high)CD16+ monocytes were independent of elevations in the CD4 T cell subsets. After adjustments for relevant confounders, the levels of CD4+CD56+CD57+ T cells and CD14(high)CD16+ monocytes remained associated with the presence of CAC. These findings indicate that PBMC subsets are markers for the presence of CAC and suggest that mechanisms of atherogenesis in RA may operate in part through the elevations in these subsets, raising further questions about the mechanisms underlying the presence of such alterations in cell composition in patients with RA and the potential for shared etiologic pathways between RA and cardiovascular disease.


Cost-effectiveness analysis for Pap smear screening and human papillomavirus DNA testing and vaccination.
As the effectiveness of cytology-based screening programme for cervical cancer in mortality reduction has reached a plateau, various preventive strategies have been considered, including intensive Pap smear screening and the supplemental use of human papillomavirus (HPV) DNA test or HPV vaccination. Cost and effectiveness of these various preventive strategies are therefore of great concern for health policy makers. We intended to assess whether the combination of HPV DNA testing or HPV vaccination with Pap smear screening programme or the sole annual Pap smear screening is more effective and cost-effective in prevention of cervical cancer than the existing triennial Pap smear screening programme. A Markov decision model was constructed to compare total costs and effectiveness between different preventive strategies (including annual Pap smear, HPV DNA testing or HPV vaccination together with Pap smear screening programme) as opposed to the triennial Pap smear screening alone (the comparator). Probabilistic cost-effectiveness (C-E) analysis was adopted to plot a series of simulated incremental C-E ratios scattered over C-E plane and also to yield the acceptability curve for different comparisons of strategies. The threshold of vaccine cost and the influence of attendance rate were also investigated. Compared with triennial Pap smear screening programme, most of preventive strategies cost more but gain additional life years (quadrant I of C-E plane) except HPV DNA testing with Pap smear every 5 years dominated by triennial Pap smear screening programme. The most cost-effective strategy was annual Pap smear (incremental C-E ratio = $31 698), followed by HPV DNA testing with Pap smear every 3 years ($36 627), and vaccination programme with triennial Pap smear screening ($44 688) with the corresponding cost-effective probabilities by the acceptability curve being 65.52%, 52.08% and 35.84% given the threshold of $40 000 of willingness to pay. Vaccination combined with triennial Pap smear would be as cost-effective as annual Pap smear provided the cost of vaccination was lowered to $250 per full course of injection. Among various preventive strategies annual Pap smear screening programme is still the most cost-effective and additional HPV DNA testing is a cost-effective choice under a reasonable threshold of willingness to pay. Vaccination programme in combination with triennial screening would be cost-effective if vaccine cost can be greatly reduced in a large economic scale.


Dinuclear copper(I) complexes containing cyclodiphosphazane derivatives and pyridyl ligands: synthesis, structural studies, and antiproliferative activity toward human cervical and breast cancer cells.
Several mixed-ligand copper(I) complexes of cyclodiphosphazanes, [(t)BuNP(NC(4)H(8)X)](2) (1, X = O; 2, X = NMe), were synthesized by reacting the octanuclear copper(I) complexes [Cu(8)(μ(2)-I)(8){[(t)BuNP(NC(4)H(8)X)](2)}(4)] (3, X = O; 4, X = NMe) with various pyridyl ligands. Interaction of the metallomacrocyclic complex 3 or 4 with pyridine, 2,2'-bipyridine, and 1,10-phenanthroline afforded the neutral dinuclear complexes [(C(5)H(5)N)(4)Cu(2)I(2){[(t)BuNP(NC(4)H(8)X)](2)}] (5, X = O; 6, X = NMe), [(2,2'-bpy)(2)Cu(2)I(2){[(t)BuNP(NC(4)H(8)X)](2)}] (7, X = O; 8, X = NMe), and [(1,10-phen)(2)Cu(2)I(2){[(t)BuNP(NC(4)H(8)X)](2)}] (9, X = O; 10, X = NMe), respectively, in good yield. The new dinuclear complexes 3, 5, and 7-9 were tested for their cytotoxic properties against human cervical cancer (HeLa) cells. The results indicated that all of the copper complexes have in vitro antitumor activity either similar to or better than that of cisplatin, a widely used anticancer drug. Among the compounds tested, complex 9 showed the most potent inhibitory activity in HeLa cells. In addition, complex 9 was found to potently inhibit proliferation of human breast cancer cells (MCF-7), highly metastatic breast cancer cells (MDA-MB 231), and nontransformed Chinese hamster ovary (CHO) cells. Complex 9 inhibited proliferation of these cells in culture more potently than cisplatin; for example, complex 9 was found to inhibit proliferation of HeLa and MCF-7 cells 3 and 5 times more efficiently than cisplatin. Complex 9 treatment damaged the DNA integrity, blocked the cells in the G1 phase of the cell cycle, and induced apoptosis via a p53-dependent pathway. The molecular structures of complexes 9 and 10 were confirmed by single-crystal X-ray diffraction studies.


The morphostatic limit for a model of skeletal pattern formation in the vertebrate limb.
A recently proposed mathematical model of a "core" set of cellular and molecular interactions present in the developing vertebrate limb was shown to exhibit pattern-forming instabilities and limb skeleton-like patterns under certain restrictive conditions, suggesting that it may authentically represent the underlying embryonic process (Hentschel et al., Proc. R. Soc. B 271, 1713-1722, 2004). The model, an eight-equation system of partial differential equations, incorporates the behavior of mesenchymal cells as "reactors," both participating in the generation of morphogen patterns and changing their state and position in response to them. The full system, which has smooth solutions that exist globally in time, is nonetheless highly complex and difficult to handle analytically or numerically. According to a recent classification of developmental mechanisms (Salazar-Ciudad et al., Development 130, 2027-2037, 2003), the limb model of Hentschel et al. is "morphodynamic," since differentiation of new cell types occurs simultaneously with cell rearrangement. This contrasts with "morphostatic" mechanisms, in which cell identity becomes established independently of cell rearrangement. Under the hypothesis that development of some vertebrate limbs employs the core mechanism in a morphostatic fashion, we derive in an analytically rigorous fashion a pair of equations representing the spatiotemporal evolution of the morphogen fields under the assumption that cell differentiation relaxes faster than the evolution of the overall cell density (i.e., the morphostatic limit of the full system). This simple reaction-diffusion system is unique in having been derived analytically from a substantially more complex system involving multiple morphogens, extracellular matrix deposition, haptotaxis, and cell translocation. We identify regions in the parameter space of the reduced system where Turing-type pattern formation is possible, which we refer to as its "Turing space." Obtained values of the parameters are used in numerical simulations of the reduced system, using a new Galerkin finite element method, in tissue domains with nonstandard geometry. The reduced system exhibits patterns of spots and stripes like those seen in developing limbs, indicating its potential utility in hybrid continuum-discrete stochastic modeling of limb development. Lastly, we discuss the possible role in limb evolution of selection for increasingly morphostatic developmental mechanisms.


Sporadic and radiation-associated papillary thyroid cancers can be distinguished using routine immunohistochemistry.
We investigated protein abundance in order to differentiate radiation-associated papillary thyroid cancers (PTC) from other etiologies for e.g. forensic purposes. Proteins were extracted from frozen tissues originating from 91 sporadic PTCs and 86 post-Chernobyl PTCs. Proteins were separated gel-electrophoretically, gels were silver stained, spots scanned and their intensity quantified. After excision of spots from the gel and protein digestion, MALDI-TOF mass spectrometry was performed followed by correlation of these results to human proteins using appropriate software and database. After this screening approach, altogether 20 candidate proteins were selected and measured semiquantitatively (Remmele score) using immunohistochemistry. Logistic regression modeling was performed for discriminating the groups. NTRK1, metalloproteinases (MMP-1, MMP-9 and MMP-13) and Cathepsins (-W and -X) proved to be of highest significance for discriminating the groups irrespective of the regression model utilized. When considering age and gender, each of 3 proteins by itself made possible a complete separation of the groups otherwise a combination of 2 of the 5 proteins mentioned was needed. In conclusion, abundance of proteins known to be associated with a more aggressive tumor type (MMPs and Cathepsins) appeared increased in post-Chernobyl PTC compared to sporadic PTC, thus underlining the known aggressiveness of radiation-associated PTC. These proteins make it possible to completely distinguish post-Chernobyl from sporadic PTC using routine immunohistology.


From monkey-like action recognition to human language: an evolutionary framework for neurolinguistics.
The article analyzes the neural and functional grounding of language skills as well as their emergence in hominid evolution, hypothesizing stages leading from abilities known to exist in monkeys and apes and presumed to exist in our hominid ancestors right through to modern spoken and signed languages. The starting point is the observation that both premotor area F5 in monkeys and Broca's area in humans contain a "mirror system" active for both execution and observation of manual actions, and that F5 and Broca's area are homologous brain regions. This grounded the mirror system hypothesis of Rizzolatti and Arbib (1998) which offers the mirror system for grasping as a key neural "missing link" between the abilities of our nonhuman ancestors of 20 million years ago and modern human language, with manual gestures rather than a system for vocal communication providing the initial seed for this evolutionary process. The present article, however, goes "beyond the mirror" to offer hypotheses on evolutionary changes within and outside the mirror systems which may have occurred to equip Homo sapiens with a language-ready brain. Crucial to the early stages of this progression is the mirror system for grasping and its extension to permit imitation. Imitation is seen as evolving via a so-called simple system such as that found in chimpanzees (which allows imitation of complex "object-oriented" sequences but only as the result of extensive practice) to a so-called complex system found in humans (which allows rapid imitation even of complex sequences, under appropriate conditions) which supports pantomime. This is hypothesized to have provided the substrate for the development of protosign, a combinatorially open repertoire of manual gestures, which then provides the scaffolding for the emergence of protospeech (which thus owes little to nonhuman vocalizations), with protosign and protospeech then developing in an expanding spiral. It is argued that these stages involve biological evolution of both brain and body. By contrast, it is argued that the progression from protosign and protospeech to languages with full-blown syntax and compositional semantics was a historical phenomenon in the development of Homo sapiens, involving few if any further biological changes.


Relationship between nosocomial Acinetobacter species occurring in two geographical areas (Greece and the UK).
Fifty-two isolates of Acinetobacter spp. obtained from three Greek and one UK hospital, were studied using partial 16 S ribosomal DNA sequence analysis, repetitive extragenic palindromic sequence-based polymerase chain reaction (REP-PCR) mediated fingerprinting and DNA macro-restriction analysis. The aim was twofold: first, to discern the major differences in the population of Acinetobacter spp. between the two countries. Second, to compare a simple PCR-based typing scheme with pulsed-field gel electrophoresis (PFGE). The multi-resistant Greek isolates were within DNA groups 2 and TU13, and clustered into three types both by REP-PCR and PFGE. By contrast, the more susceptible Oxford isolates were heterogeneous on 16 S RNA sequence analysis and distinguishable on typing. The need for studies that elucidate the phylogeny of Acinetobacter spp. inside and outside hospitals are important, as this will help clarify the relationship between organisms that are increasingly recognized as causes of severe infections.


Reindeer habitat use in relation to two small wind farms, during preconstruction, construction, and operation.
Worldwide there is a rush toward wind power development and its associated infrastructure. In Fennoscandia, large-scale wind farms comprising several hundred windmills are currently built in important grazing ranges used for Sámi reindeer husbandry. In this study, reindeer habitat use was assessed using reindeer fecal pellet group counts in relation to two relatively small wind farms, with 8 and 10 turbines, respectively. In 2009, 1,315 15-m2 plots were established and pellet groups were counted and cleaned from the plots. This was repeated once a year in May, during preconstruction, construction, and operation of the wind farms, covering 6 years (2009-2014) of reindeer habitat use in the area. We modeled the presence/absence of any pellets in a plot at both the local (wind farm site) and regional (reindeer calving to autumn range) scale with a hierarchical logistic regression, where spatial correlation was accounted for via random effects, using vegetation type, and the interaction between distance to wind turbine and time period as predictor variables. Our results revealed an absolute reduction in pellet groups by 66% and 86% around each wind farm, respectively, at local scale and by 61% at regional scale during the operation phase compared to the preconstruction phase. At the regional, scale habitat use declined close to the turbines in the same comparison. However, at the local scale, we observed increased habitat use close to the wind turbines at one of the wind farms during the operation phase. This may be explained by continued use of an important migration route close to the wind farm. The reduced use at the regional scale nevertheless suggests that there may be an overall avoidance of both wind farms during operation, but further studies of reindeer movement and behavior are needed to gain a better understanding of the mechanisms behind this suggested avoidance.


Perception of Melodic Contour and Intonation in Autism Spectrum Disorder: Evidence From Mandarin Speakers.
Tone language experience benefits pitch processing in music and speech for typically developing individuals. No known studies have examined pitch processing in individuals with autism who speak a tone language. This study investigated discrimination and identification of melodic contour and speech intonation in a group of Mandarin-speaking individuals with high-functioning autism. Individuals with autism showed superior melodic contour identification but comparable contour discrimination relative to controls. In contrast, these individuals performed worse than controls on both discrimination and identification of speech intonation. These findings provide the first evidence for differential pitch processing in music and speech in tone language speakers with autism, suggesting that tone language experience may not compensate for speech intonation perception deficits in individuals with autism.


Oxidative stress underlies the mechanism for Ca(2+)-induced permeability transition of mitochondria.
Recent studies demonstrated that the generation of intracellular reactive oxygen species (ROS) was enhanced prior to the onset of mitochondrial membrane permeability transition (MPT), a critical step for the induction of DNA fragmentation and apoptosis. Although Ca2+ induces typical MPT that involves depolarization and swelling of mitochondria and finally releases cytochrome c into cytosol, the mechanism by which ROS induce MPT remains unclear. In the presence of inorganic phosphate, Ca2+ increased the oxygen consumption and ROS production by isolated mitochondria as determined by a chemiluminescence (CHL) method using L-012. Ca2+ increased the generation of H2O2 by some mechanism that was inhibited by cyclosporin A but not by superoxide dismutase (SOD) and trifluoperazine. Ca2+ decreased the content of free thiols in adenine nucleotide translocase (ANT) in mitochondrial membranes with concomitant increase in ROS generation. The presence of cyclosporin A, trifluoperazine, or SOD inhibited the Ca(2+)-induced increase of L-012 CHL and decrease in the free thiols of ANT. These results indicate that Ca2+ increases the generation of ROS which oxidize the free thiol groups in mitochondrial ANT, thereby inducing MPT to release cytochrome c.


Preventing Plasmon Coupling between Gold Nanorods Improves the Sensitivity of Photoacoustic Detection of Labeled Stem Cells in Vivo.
Gold nanorods are excellent contrast agents for imaging technologies which rely on near-infrared absorption such as photoacoustic imaging. For cell tracking applications, the cells of interest are labeled with the contrast agent prior to injection. However, after uptake into cells by endocytosis, the confinement and high concentration in endosomes leads to plasmon band broadening and reduced absorbance. This would limit the potential of multispectral optoacoustic tomography in terms of spectral processing and, consequently, sensitivity. Here, we show that steric hindrance provided by silica coating of the nanorods leads to the preservation of their spectral properties and improved photoacoustic sensitivity. This strategy allowed the detection and monitoring of as few as 2 × 10(4) mesenchymal stem cells in mice over a period of 15 days with a high spatial resolution. Importantly, the silica-coated nanorods did not affect the viability or differentiation potential of the transplanted mesenchymal stem cells.


A peptide sequence confers retention and rapid degradation in the endoplasmic reticulum.
A nonlysosomal pathway exists for the degradation of newly synthesized proteins retained within the endoplasmic reticulum (ER). This pathway is extremely selective: whereas some proteins are rapidly degraded, others survive for long periods in the ER. The question of whether this selectivity is due to the presence within the sensitive proteins of definable peptide sequences that are sufficient to target them for degradation has been addressed. Deletion of a carboxyl-terminal sequence, comprising the transmembrane domain and short cytoplasmic tail of the alpha chain of the T cell antigen receptor (TCR-alpha), prevented the rapid degradation of this polypeptide. Fusion of this carboxyl-terminal sequence to the extracellular domain of the Tac antigen, a protein that is normally transported to the cell surface where it survives long-term, resulted in the retention and rapid degradation of the chimeric protein in the ER. Additional mutagenesis revealed that the transmembrane domain of TCR-alpha alone was sufficient to cause degradation within the ER. This degradation was not a direct consequence of retention in the ER, as blocking transport of newly synthesized proteins out of the ER with brefeldin A did not lead to degradation of the normal Tac antigen. It is proposed that a 23-amino acid sequence, comprising the transmembrane domain of TCR-alpha, contains information that determines targeting for degradation within the ER system.


Molecular detection, identification and drug resistance detection in Mycobacterium tuberculosis.
This minireview presents recent developments in molecular methods for the diagnosis of tuberculosis, including detection, identification and determination of drug resistance of Mycobacterium tuberculosis. Tuberculosis remains one of the major causes of global death from a single infectious agent. This situation is worsened by the HIV/AIDS pandemic because one-third of HIV/AIDS patients are coinfected with M. tuberculosis. Also of great concern is the emergence of drug-resistant tuberculosis because there are almost no treatment options available for patients affected by highly resistant strains of M. tuberculosis. Advances in molecular biology techniques and a better knowledge of the molecular mechanisms of drug resistance have provided new tools for the rapid diagnosis of tuberculosis. Several nucleic acid amplification technologies have been developed and evaluated. New molecular approaches are being introduced continuously. This minireview will also comment on the future perspectives for the molecular diagnosis of tuberculosis and the feasibility for the implementation of these newer techniques in the clinical diagnostic laboratory.


Effect of taxol on first and second meiotic spindle formation in oocytes of the surf clam, Spisula solidissima.
The effect of taxol on first and second meiotic spindle formation was examined in oocytes of the surf clam, Spisula solidissima, by immunofluorescence staining with anti-tubulin antibody. The first meiotic spindle appeared to form as in untreated control cells. However, the spindle did not migrate toward the periphery of taxol-activated oocytes, resulting in blockage of the formation of the first polar body. In spite of inhibited microtubule depolymerization and failure of spindle disappearance, the pole separation in telophase that is typical of this material began at the same time as in untreated cells. Polymerization of the second spindle microtubules onto the spindle persisting from the first meiosis led to the formation of a triple form of spindle connected at the poles of each other. The subsequent emergence of ring-shaped microtubule-containing structures in mature activated eggs was not affected by taxol. The mechanism of meiotic spindle formation thus seemsto be different from that in mitosis, where taxol has been shown to block spindle formation completely.


Comparison of histological grading methods in mucoepidermoid carcinoma of minor salivary glands.
Mucoepidermoid carcinoma (MEC) is the most common salivary gland malignancy and its grading is greatly consequential in the management and prognosis of patients with the disease. To compare histologic grading systems in MEC of minor salivary glands. Two qualitative (modified Healy and Memorial Sloan-Kettering Cancer Center [MSKCC] methods) and two quantitative (Armed Forces Institute of Pathology [AFIP] and Brandwein methods) were evaluated. Diagnostics slides of 19 patients including one recurrent case were evaluated using the four grading systems. Percentages and proportions were used. Agreement across all grading system was found to be very low (32%) while there was a better agreement between AFIP and MSKCC methods (84%) between modified Healy and Brandwein (58%). The method that gave the poorest agreement with all the others was the Brandwein grading. In general, the AFIP and MSKCC methods tended to grade the tumors lower while the Brandwein and modified Healy methods seemed to grade them higher. Most MEC of minor salivary glands appear to be low-grade tumors. It is conceivable that some grading methods (Brandwein and modified Healy) may lead to an unnecessary escalation of management methods in these tumors. The MSKCC method may have emphasized some parameters which may not have much importance in minor salivary gland MEC. The AFIP method appears to be the most appropriate to use for the grading of minor salivary gland MEC. Further studies are required to confirm or disprove this finding.


High-purity preparation of HSV-2 vaccine candidate ACAM529 is immunogenic and efficacious in vivo.
Genital herpes is a sexually transmitted infection (STI) caused by herpes simplex virus 2 (HSV-2) and to a lesser extent herpes simplex virus 1 (HSV-1). Infection by HSV-2 is life-long and is associated with significant cost to healthcare systems and social stigma despite the highly prevalent nature of the disease. For instance, the proportion of HSV-2 seropositive to seronegative adults is approximately 1 in 5 in the US and greater than 4 in 5 in some areas of sub-Saharan Africa. The replication-defective vaccine strain virus dl5-29 was re-derived using cells appropriate for GMP manufacturing and renamed ACAM529. Immunization with dl5-29 was previously reported to be protective both in mice and in guinea pigs, however these studies were performed with vaccine that was purified using methods that cannot be scaled for manufacturing of clinical material. Here we describe methods which serve as a major step towards preparation of ACAM529 which may be suitable for testing in humans. ACAM529 can be harvested from infected cell culture of the trans-complementing cell line AV529 clone 19 (AV529-19) without mechanical cell disruption. ACAM529 may then be purified with respect to host cell DNA and proteins by a novel purification scheme, which includes a combination of endonuclease treatment, depth filtration, anion-exchange chromatography and ultrafiltration/diafiltration (UF/DF). The resultant virus retains infectivity and is ∼ 200-fold more pure with respect to host cell DNA and proteins than is ACAM529 purified by ultracentrifugation. Additionally, we describe a side-by-side comparison of chromatography-purified ACAM529 with sucrose cushion-purified ACAM529, which shows that both preparations are equally immunogenic and protective when tested in vivo.


Resonance in the human medial gastrocnemius muscle during cyclic ankle bending exercise.
We investigated the behavior of the muscle tendon unit (MTU) of the medial gastrocnemius muscle during cyclic ankle bending exercise at eight different frequencies (ranging from 1.33 to 3.67 Hz). The changes in the length of fascicle in the muscle during the exercises were determined by real-time ultrasound imaging. The coordinates of anatomical references and the ground reaction force were determined from video recording and a force plate, respectively. The length change of the MTU (the distance from the origin to insertion of the muscle) was calculated from changes in the knee and ankle joint angles. It was found that the amplitude ratio and phase difference between the fascicle and MTU lengths were both dependent on the movement frequency. At lower frequencies, the fascicle lengths varied almost in phase with the MTU length, whereas they varied out of phase at the higher frequencies. At intermediate frequency, the amplitude of the fascicle became very small compared with that of the MTU, which is considered resonance. We constructed a mechanical model of the MTU based on a notion of forced oscillation in a mass-spring system. The obtained data were well explained by the model. It was concluded that the behavior of the MTU highly depends on the movement frequency due to the viscoelasticity of the MTU.


A coastal perspective on security.
This paper examines security issues from the unique perspective of our nation's coastlines and associated infrastructure. It surveys ongoing efforts to secure offshore shipping lanes, as well as the transportation systems and huge capital investments on the narrow strip of land intersecting with coastal waters. The paper recounts the extraordinary demands recently placed on the Coast Guard, port authorities and other agencies charged with offshore security. New federal requirements such as port assessments continue to be mandated, while solutions to finding are still unfolding. An up-to-date summary of maritime security functions is provided. Those requirements are compared and contrasted with security guidelines and regulatory demands placed upon mobile and fixed assets of the Chemical Process Industry (CPI) in coastal environs. These span the gamut from recommendations by industry groups and professional organizations, to federal and state requirements, to insurance demands, to general duty obligations.


Alpha-1,3-galactosyltransferase-deficient miniature pigs produced by serial cloning using neonatal skin fibroblasts with loss of heterozygosity.
Production of alpha-1,3-galactosyltransferase (αGT)-deficient pigs is essential to overcome xenograft rejection in pig-to-human xenotransplantation. However, the production of such pigs requires a great deal of cost, time, and labor. Heterozygous αGT knockout pigs should be bred at least for two generations to ultimately obtain homozygote progenies. The present study was conducted to produce αGT-deficient miniature pigs in much reduced time using mitotic recombination in neonatal ear skin fibroblasts. Miniature pig fibroblasts were transfected with αGT gene-targeting vector. Resulting gene-targeted fibroblasts were used for nuclear transfer (NT) to produce heterozygous αGT gene-targeted piglets. Fibroblasts isolated from ear skin biopsies of these piglets were cultured for 6 to 8 passages to induce loss of heterozygosity (LOH) and treated with biotin-conjugated IB4 that binds to galactose-α-1,3-galactose, an epitope produced by αGT. Using magnetic activated cell sorting, cells with monoallelic disruption of αGT were removed. Remaining cells with LOH carrying biallelic disruption of αGT were used for the second round NT to produce homozygous αGT gene-targeted piglets. Monoallelic mutation of αGT gene was confirmed by polymerase chain reaction in fibroblasts. Using these cells as nuclear donors, three heterozygous αGT gene-targeted piglets were produced by NT. Fibroblasts were collected from ear skin biopsies of these piglets, and homozygosity was induced by LOH. The second round NT using these fibroblasts resulted in production of three homozygous αGT knockout piglets. The present study demonstrates that the time required for the production of αGT-deficient miniature pigs could be reduced significantly by postnatal skin biopsies and subsequent selection of mitotic recombinants. Such procedure may be beneficial for the production of homozygote knockout animals, especially in species, such as pigs, that require a substantial length of time for breeding.