Regulation of neurotransmitter release from ocular tissues by isoprostanes.
Isoprostanes are prostaglandin-like compounds formed in vivo primarily by free radical-catalyzed peroxidation of arachidonic acid independent of the cyclooxygenase enzyme. In addition to being utilized as reliable indicators of oxidative stress, 8-isoprostanes exert pharmacological actions on smooth muscles from several tissues and organs, and they play a role in the release of neurotransmitters from the central and peripheral nervous systems. In the anterior uvea of the eye, 8-isoprostanes produce both excitatory and inhibitory effects on sympathetic neurotransmission in isolated mammalian iris ciliary bodies. Thromboxane (TP) receptors mediate the stimulatory action of isoprostanes on norepinephrine (NE) release from sympathetic nerves. In bovine retina, the 8-isoprostanes exhibit a biphasic regulatory effect on potassium-induced [3H]-D-aspartate release, with low concentrations being inhibitory and high concentrations causing an excitatory effect. Excitatory effects of 8-isoprostanes are mediated by TP receptors, while inhibitory responses are mediated by prostaglandin E (EP) receptors. The 8-isoprostanes produce pharmacological actions on sympathetic neurotransmission in mammalian anterior uvea, a response that is species-dependent. In the posterior segment of the eye, 8-isoprostanes elicit a complex response on the retina involving the activation of both prostanoid TP and EP receptors. An effect of isoprostanes on neurotransmitter pools provides new pharmacological target sites for the therapy of some ocular diseases.


Coaxial Mirau interferometer.
We describe a new interferometric configuration for optical coherence tomography that is based on the Mirau interferometer. It uses the photodetector included in a superluminescent diode package, which makes possible a highly miniaturized device. Other advantages of the configuration include its totally coaxial structure, confocal microscope operation, availability of the full working distance of the imaging objective, and no central obscuration. Fundamental characteristics such as resolution and dynamic range are discussed, and the result of measurement on a rough metallic surface is presented.


Toll-Like Receptors and Tissue Remodeling: The Pro/Cons Recent Findings.
The Toll-like Receptor (TLR) family ensures prompt response towards pathogens, protecting the host against infections, and guarantees a realistic balance between protective and detrimental activities. Multiple regulating mechanisms characterize TLR activity that is not limited to innate and adaptive antimicrobial immune responses, as observed in the inflammatory (either infective, allergic, or autoimmune) responses associated with tissue remodeling. Following the insult and the arise of inflammatory response, tissue remodeling takes place and might develop in fibrosis, depending on microenvironment as a result of imbalanced fibroblasts (FBs) and myofibroblasts (myoFBs) activation/survival. The process is driven by an epithelial-fibroblast-immune cell cross-talk. While the main FB function is the matrix metabolism for tissue homeostasis or repair, the myoFB differentiation represents a crucial step in attempting repair of injury. FBs/myoFBs provide more than structural support at site of injury, synthesizing and/or reacting to different cytokines, growth factors, neuromediators and soluble/lipid mediators. TLR-bearing FBs/myoFBs might contribute at the innate immune level, providing a second line of protection/defense as well as being a target/effector cell of tissue remodeling. TLRs might also interfere with acute inflammation as well as with established fibrosis, triggering structural/functional changes in agreement with the genetic background, the site of lesion, the entity of associated infection, the poor blood circulation or the pharmacological treatments, all together strictly influencing tissue repair/remodeling process. This review will focus on the recent findings on TLRs at launch and long-lasting tissue remodeling process, that strongly suggest TLRs as optional targets for future therapies.


Sudden increases in listeriosis rates in England and Wales, 2001 and 2003.
The monthly incidence of listeriosis infections in England and Wales had 2 sudden increases during April 2001 (41%) and March 2003 (48%). Although no causative association is demonstrated, these increases correspond to key dates relating to the onset and aftermath of the 2001 foot and mouth disease outbreak in the United Kingdom.


ERK integrates PKA and PKC signaling in superficial dorsal horn neurons. I. Modulation of A-type K+ currents.
The transient outward potassium currents (also known as A-type currents or IA) are important determinants of neuronal excitability. In the brain, IA is modulated by protein kinase C (PKC), protein kinase A (PKA), and extracellular signal-related kinase (ERK), three kinases that have been shown to be critical modulators of nociception. We wanted to determine the effects of these kinases on IA in superficial dorsal horn neurons. Using whole cell recordings from cultured mouse spinal cord superficial dorsal horn neurons, we found that PKC and PKA both inhibit IA in these cells, and that PKC has a tonic inhibitory action on IA. Further, we provide evidence supporting the hypothesis that PKC and PKA do not modulate IA directly, but rather act as upstream activators of ERKs, which modulate IA. These results suggest that ERKs serve as signal integrators in modulation of IA in dorsal horn neurons and that modulation of A-type potassium currents may underlie aspects of central sensitization mediated by PKC, PKA, and ERKs.


Increased expression of phospholipase D1 in the brains of scrapie-infected mice.
Mitochondrial dysfunction and free radical-induced oxidative damage are critical factors in the pathogenesis of neurodegenerative diseases. Recently, phospholipid breakdown by phospholipase D (PLD) has been recognized as an important signalling pathway in the nervous system. Here, we examined the expression of PLD and alteration of membrane phospholipid in scrapie brain. We have found that protein expression and enzyme activity of PLD1 were increased in scrapie brains compared with controls; in particular, there was an increase in the mitochondrial fraction. PLD1 in mitochondrial membranes from scrapie brains, but not from control brains, was tyrosine phosphorylated. Furthermore, the concentration of mitochondrial phospholipids such as phosphatidylcholine and phosphatidylethanolamine was increased and the content of phosphatidic acid, a product of PLD activity, was up-regulated in the mitochondrial membrane fractions. Immunohistochemically, PLD1 immunoreactivity was significantly increased in activated astrocytes in both cerebral cortex and hippocampus of scrapie brains. Taken together, these results suggest that PLD activation might induce alterations in mitochondrial lipids and, in turn, mediate mitochondrial dysfunction in the brains of scrapie-infected mice.


Monte Carlo model of the penetration depth for polarization gating spectroscopy: influence of illumination-collection geometry and sample optical properties.
Polarization-gating has been widely used to probe superficial tissue structures, but the penetration depth properties of this method have not been completely elucidated. This study employs a polarization-sensitive Monte Carlo method to characterize the penetration depth statistics of polarization-gating. The analysis demonstrates that the penetration depth depends on both the illumination-collection geometry [illumination-collection area (R) and collection angle (θ(c))] and on the optical properties of the sample, which include the scattering coefficient (μ(s)), absorption coefficient (μ(a)), anisotropy factor (g), and the type of the phase function. We develop a mathematical expression relating the average penetration depth to the illumination-collection beam properties and optical properties of the medium. Finally, we quantify the sensitivity of the average penetration depth to changes in optical properties for different geometries of illumination and collection. The penetration depth model derived in this study can be applied to optimizing application-specific fiber-optic probes to target a sampling depth of interest with minimal sensitivity to the optical properties of the sample.


Bowel preparation for CT colonography.
Bowel preparation represents an essential part of CT colonography, as the accuracy of the exam is strongly related to the adequacy of colonic cleansing, and a poor bowel preparation may compromise the diagnostic quality even despite optimization of all other acquisition parameters. Residual stool and fluid in the large bowel may affect the interpretation of the exam and may increase the number of false positives and false negatives. In this regard, the majority of patients having undergone CT colonography state that bowel preparation is the most unpleasant part. Unfortunately, to date no definite consensus has been reached about the ideal bowel preparation technique, and there is great variability in preparation strategies across diagnostic centers. The purpose of this review article is to describe the development and evolution of bowel preparation techniques in order to choose the best approach for optimizing the diagnostic quality of CT colonography in each patient.


Theory of supramolecular co-polymerization in a two-component system.
As a first step to understand the role of molecular or chemical polydispersity in self-assembly, we put forward a coarse-grained model that describes the spontaneous formation of quasi-linear polymers in solutions containing two self-assembling species. Our theoretical framework is based on a two-component self-assembled Ising model in which the chemical bidispersity, i.e., the presence of two distinct chemical entities, is parameterized in terms of the strengths of the binding free energies that depend on the monomer species involved in the pairing interaction. Depending upon the relative values of the binding free energies involved, different morphologies of assemblies that include both components are formed, exhibiting random, blocky or alternating ordering of the two components in the assemblies. Analyzing the model for the case of blocky ordering, which is of most practical interest, we find that the transition from conditions of minimal assembly to those characterized by strong polymerization can be described by a critical concentration that depends on the concentration ratio of the two species. Interestingly, the distribution of monomers in the assemblies is different from that in the original distribution, i.e., the ratio of the concentrations of the two components put into the system. The monomers with a smaller binding free energy are more abundant in short assemblies and monomers with a larger binding affinity are more abundant in longer assemblies. Under certain conditions the two components congregate into separate supramolecular polymeric species and in that sense phase separate. We find strong deviations from the expected growth law for supramolecular polymers even for modest amounts of a second component, provided it is chemically sufficiently distinct from the main one.


Mice deficient in IL-1beta manifest impaired contact hypersensitivity to trinitrochlorobenzone.
Mice rendered deficient in IL-1 beta by gene targeting in embryonic stem cells develop and grow normally in a protected laboratory environment. Endotoxin-stimulated peritoneal macrophages from IL-1beta-deficient mice showed normal synthesis and cellular release of IL-1alpha after treatment with 5 mM ATP demonstrating that IL-1beta is not necessary for expression and release of the IL-1alpha isoform. Mice deficient in IL-1beta showed unaltered sensitivity to endotoxic shock, with or without pretreatment with D-galactosamine. In contrast, IL-1beta-deficient mice showed defective contact hypersensitivity responses to topically applied trinitrochlorobenzene (TNCB). This defect could be overcome either by application of very high doses of sensitizing antigen, or by local intradermal injection of recombinant IL-1beta immediately before antigen application. These data demonstrate an essential role for IL-1beta in contact hypersensitivity and suggest that IL-1beta acts early during the sensitization phase of response. They suggest an important role for IL-1beta in initiation of the host of response at the epidermal barrier.


Diagnostic accuracy study of anorectal manometry for diagnosis of dyssynergic defecation.
The diagnostic accuracy of anorectal manometry (AM), which is necessary to diagnose functional defecatory disorders (FDD), is unknown. Using blinded analysis and standardised reporting of diagnostic accuracy, we evaluated whether AM could discriminate between asymptomatic controls and patients with functional constipation (FC). Derived line plots of anorectal pressure profiles during simulated defecation were independently analysed in random order by three expert observers blinded to health status in 85 women with FC and 85 age-matched asymptomatic healthy volunteers (HV). Using accepted criteria, these pressure profiles were characterised as normal (ie, increased rectal pressure coordinated with anal relaxation) or types I-IV dyssynergia. Interobserver agreement and diagnostic accuracy were determined. Blinded consensus-based assessment disclosed a normal pattern in 16/170 (9%) of all participants and only 11/85 (13%) HV. The combined frequency of dyssynergic patterns (I-IV) was very similar in FC (80/85 (94%)) and HV (74/85 (87%)). Type I dyssynergia ('paradoxical' contraction) was less prevalent in FC (17/85 (20%) than in HV (31/85 (36.5%), p=0.03). After statistical correction, only type IV dyssynergia was moderately useful for discriminating between FC (39/85 (46%)) and HV (17/85 (20%)) (p=0.001, positive predictive value=70.0%, positive likelihood ratio=2.3). Interobserver agreement was substantial or moderate for identifying a normal pattern, dyssynergia types I and IV, and FDD, and fair for types II and III. While the interpretation of AM patterns is reproducible, nearly 90% of HV have a pattern that is currently regarded as 'abnormal' by AM. Hence, AM is of limited utility for distinguishing between FC and HV.


Polyclonality of concurrent natural populations of Alteromonas macleodii.
We have analyzed a natural population of the marine bacterium, Alteromonas macleodii, from a single sample of seawater to evaluate the genomic diversity present. We performed full genome sequencing of four isolates and 161 metagenomic fosmid clones, all of which were assigned to A. macleodii by sequence similarity. Out of the four strain genomes, A. macleodii deep ecotype (AltDE1) represented a different genome, whereas AltDE2 and AltDE3 were identical to the previously described AltDE. Although the core genome (~80%) had an average nucleotide identity of 98.51%, both AltDE and AltDE1 contained flexible genomic islands (fGIs), that is, genomic islands present in both genomes in the same genomic context but having different gene content. Some of the fGIs encode cell surface receptors known to be phage recognition targets, such as the O-chain of the lipopolysaccharide, whereas others have genes involved in physiological traits (e.g., nutrient transport, degradation, and metal resistance) denoting microniche specialization. The presence in metagenomic fosmids of genomic fragments differing from the sequenced strain genomes, together with the presence of new fGIs, indicates that there are at least two more A. macleodii clones present. The availability of three or more sequences overlapping the same genomic region also allowed us to estimate the frequency and distribution of recombination events among these different clones, indicating that these clustered near the genomic islands. The results indicate that this natural A. macleodii population has multiple clones with a potential for different phage susceptibility and exploitation of resources, within a seemingly unstructured habitat.


Genistein Has Antiviral Activity against Herpes B Virus and Acts Synergistically with Antiviral Treatments to Reduce Effective Dose.
Herpes B virus is a deadly zoonotic agent that can be transmitted to humans from the macaque monkey, an animal widely used in biomedical research. Currently, there is no cure for human B virus infection and treatments require a life-long daily regimen of antivirals, namely acyclovir and ganciclovir. Long-term antiviral treatments have been associated with significant debilitating side effects, thus, there is an ongoing search for alternative efficacious antiviral treatment. In this study, the antiviral activity of genistein was quantified against B virus in a primary cell culture model system. Genistein prevented plaque formation of B virus and reduced virus production with an IC50 value of 33 and 46 μM for human and macaque fibroblasts, respectively. Genistein did not interfere directly with viral entry, but instead targeted an event post-viral replication. Finally, we showed that genistein could be used at its IC50 concentration in conjunction with both acyclovir and ganciclovir to reduce their effective dose against B virus with a 93% and 99% reduction in IC50 values, respectively. The results presented here illuminate the therapeutic potential of genistein as an effective antiviral agent against B virus when used alone or in combination with current antiviral therapies.


Endocrinological studies on TAP-144-SR, a sustained-release formulation of a potent GnRH agonist (D-Leu6-[des-Gly10-NH2]-GnRH ethylamide), in male rats.
The paradoxical effects of TAP-144-SR, a biodegradable sustained-release formulation of a potent GnRH agonist (TAP-144, leuprolide acetate) were evaluated in male rats by comparing its potency with that of TAP-144 solution. A single sc injection of TAP-144-SR (equivalent to 0.1 mg/kg/day as TAP-144), prepared by encapsulating the agonist in microcapsules of copoly (DL-lactic/glycolic acid), suppressed serum levels of androgens, and the levels remained suppressed for 4 weeks. The potency of the paradoxical effects of TAP-144-SR was evaluated 4 weeks after treatment by comparing it with that of TAP-144 solution administered daily for 4 weeks. Both daily injections of TAP-144 solution and a single injection of TAP-144-SR (equivalent to 0.02, 0.2 or 2 mg/kg/day as TAP-144) decreased the weight of the testes, prostates and seminal vesicles in a dose-dependent manner in a 4-week assay in male rats. TAP-144-SR was more effective than TAP-144 solution in reducing these organ weights. Serum and pituitary concentrations of LH and FSH and serum testosterone levels were also lower in TAP-144-SR-treated than in TAP-144 solution-treated rats. These results indicate that the paradoxical effects were more extensive upon TAP-144-SR treatment, suggesting that maintaining constant serum TAP-144 levels results in more extensive desensitization of the pituitary and testes. These results also suggest advantages of TAP-144-SR over TAP-144 solution in both efficacy and convenience as an anti-prostatic tumor agent.


Lens and cornea lesions of rats fed corn syrup and the protective effects of alpha lipoic acid.
To examine the pathological findings that occurred in the lens and cornea and biochemical findings in the lens of rats fed with corn syrup and the protective effects of alpha lipoic acid (ALA). Twenty-four rats were randomly divided into three groups. Group I served as the control group. Group II was used as the study group; the rats were treated with 30% corn sugar solution for 10 weeks. Group III was the treatment group. Corn syrup was given by the oral route to the rats during the study, and ALA (100 mg/kg) was added to the treatment 4 weeks after the study began. At the end of the experiment, central corneal thickness (CCT) was measured in all rats with an ultrasonic pachymeter. Then the right eyes of the rats were enucleated for histopathological examination of the cornea and lens. The left lenses were homogenized for biochemical analyses. The lenses of the rats treated with corn syrup revealed severe damage; many lens fibers appeared swollen and ruptured with large vacuoles near the lens epithelium. In addition, malondialdehyde (MDA) levels, a parameter of oxidative stress, increased but not significantly in Group II; however. ALA treatment decreased MDA levels significantly. Antioxidant enzyme and catalase (CAT) activities were significantly decreased in Group II, and ALA treatment increased these activities; however, the increase was not significant. Changes were observed in the cornea such as epithelial alterations, subepithelial vacuolizations, collagen fibers loss in the stromal layer, interruptions in the subepithelial basement membrane and central corneal thickening. Corn syrup can cause severe damage in rat lenses and corneas. However, ALA ameliorates the effect of corn syrup-related lesions on the cornea and lens.


Alteration of clonal profile. III. T15 ontogenetic advantages are not sufficient for establishing idiotypic dominance in adoptive transfer.
We have examined the ontogeny of BALB/c plaque-forming cell (PFC) responses to phosphorylcholine (PC) from fetal and neonatal liver by using the (CBA/N x BALB/c)F1 transplantation model. In this system, thymus-dependent (PC-keyhole limpet hemocyanin) and thymus-independent class 1 (PC-Brucella abortus, PC-lipopolysaccharide) PC antigens stimulate B cell subpopulations, which functionally emerge early after transfer. Responsiveness to a thymus-independent class 2 antigen, C-polysaccharide extract of a Streptococcus pneumoniae mutant, is acquired later. The response to PC antigens tested initially exhibited T15 dominance. Non-T15 clones, which are not expressed to a great degree in normal BALB/c mice, are inherently slow in their rate of maturation; in adoptive transfer, however, they eventually comprise much of the transplanted anti-PC PFC response. Obviously, the advantages the T15 subset has in ontogeny do not result in idiotypic dominance once the immature cells are removed from the intact BALB/c environment. We discuss possible regulatory mechanisms involved in the alteration of the T15+:T15- ratio.


Statin use and its effect on all-cause mortality of melanoma patients: a population-based Dutch cohort study.
Preclinical data showed anticancer effects of statins in melanoma, but meta-analyses could not demonstrate a reduced melanoma incidence in statin users. Rather than preventing occurrence, statins might reduce growth and metastatic spread of melanomas and ultimately improve survival. In this population-based study, we investigated the relationship between statin use and survival of melanoma patients. Patients ≥18 years who were diagnosed with cutaneous melanoma (Breslow thickness >1 mm) and registered in the Eindhoven Cancer Registry and in PHARMO Database Network between 1 January 1998 and 31 December 2010 were eligible. The hazard ratio (HR) of all-cause mortality was calculated by employing adjusted time-dependent and time-fixed Cox proportional hazard models. Disease-specific survival was estimated by means of 3-year relative survival rates (RSR). A control cohort of randomly selected patients using statins from PHARMO Database Network matched on age and gender was used to compare RSR of statin users to the general population. After melanoma diagnosis, 171 of 709 patients used statins. Use of statins showed a nonsignificantly decreased hazard of death (adjusted HR 0.76, 95% confidence interval [CI] 0.50-1.61). After stratification for gender, male but not female statin users showed a favorable outcome compared to nonusers (HR 0.57, 95% CI 0.32-0.99; HR 1.22, 95% CI 0.62-2.38, respectively). Three-year RSR for male statin users tended to be higher than for nonusers (91% vs. 80.5%, P = 0.06), no differences were observed in women (87.1% vs. 92.5%, P = 0.76). Statin use was not associated with an improved survival of melanoma patients. The trend for better survival of male in contrast to female statin users warrants further research.


Lysis of colonic epithelial cells by allogeneic mononuclear and lymphokine activated killer cells derived from peripheral blood and intestinal mucosa: evidence against a pathogenic role in inflammatory bowel disease.
A sensitive 4 h 51Cr-release cytotoxicity assay has been developed using as targets colonic epithelial cells obtained by Dispase-collagenase digestion of resected mucosa or colonoscopic biopsies. Peripheral blood mononuclear cells (MNC) from most healthy donors showed low, but significant levels of cytotoxicity for normal epithelial cell target cells of 8.7 (4.4) % (mean (SD] and similar levels were found in 14 ulcerative colitis (6.5 (4.4) %) and 16 Crohn's disease (6.2 (5.2) %) patients. Neither drug therapy nor disease activity influenced the results. The sensitivity of colonic epithelial cells isolated from inflamed and histologically normal mucosa to lysis by peripheral blood MNC from a single donor was not affected by the underlying disease. Anti-epithelial cell activity did not correlate with anti-K562 activity and the cytotoxic cell was plastic non-adherent and Leu-11b-. None of 15 MNC populations isolated from mucosa of normal, tumour bearing, or chronically inflamed intestine exhibited significant lysis of colonic epithelial cells despite killing of K562 target cells in 10. Lymphokine activated killer (LAK) cells, generated by interleukin-2 stimulation in vitro of nine intestinal and seven peripheral blood MNC populations, exhibited high levels of lysis of K562 cells but, on every occasion, failed to lyse colonic epithelial cells. These data indicate that spontaneously cytotoxic or LAK cells are unlikely to play a role in the generation of colonic epithelial cell injury by direct cytotoxicity in inflammatory bowel disease.


Inhibition of TGF-β signaling at the nuclear envelope: characterization of interactions between MAN1, Smad2 and Smad3, and PPM1A.
Signaling by transforming growth factor-β (TGF-β) is critical for various developmental processes and culminates in the activation of the transcription factors Smad2 and Smad3. MAN1, an integral protein of the inner nuclear membrane, inhibits TGF-β signaling by binding to Smad2 and Smad3. Depletion of the gene LEMD3 encoding MAN1 leads to developmental anomalies in mice, and heterozygous loss-of-function mutations in LEMD3 in humans cause sclerosing bone dysplasia. We modeled the three-dimensional structure of the MAN1-Smad2 complex from nuclear magnetic resonance and small-angle x-ray scattering data. As predicted by this model, we found that MAN1 competed in vitro and in cells with the transcription factor FAST1 (forkhead activin signal transducer 1) for binding to Smad2. The model further predicted that MAN1 bound to activated Smad2-Smad4 or Smad3-Smad4 complexes, which was confirmed by in vitro experiments; however, in cells, MAN1 bound only to Smad2 and Smad3 and not to the Smad4-containing complexes. Overexpression of MAN1 led to dephosphorylation of Smad2 and Smad3, thus hindering their recognition by Smad4, and MAN1 bound directly in vitro to the phosphatase PPM1A, which catalyzes the dephosphorylation of Smad2/3. These results demonstrate a nuclear envelope-localized mechanism of inactivating TGF-β signaling in which MAN1 competes with transcription factors for binding to Smad2 and Smad3 and facilitates their dephosphorylation by PPM1A.


Boosting the Quantitative Inorganic Surface-Enhanced Raman Scattering Sensing to the Limit: The Case of Nitrite/Nitrate Detection.
A high-performance ionic-sensing platform has been developed by an interdisciplinary approach, combining the classical colorimetric Griess reaction and new concepts of nanotechnology, such as plasmonic coupling of nanoparticles and surface-enhanced Raman scattering (SERS) spectroscopy. This approach exploits the advantages of combined SERS/surface-enhanced resonant Raman Scattering (SERRS) by inducing the formation of homogeneous hot spots and a colored complex in resonance with the laser line, to yield detection limits for nitrite down to the subpicomolar level. The performance of this new method was compared with the classical Griess reaction and ionic chromatography showing detection limits about 6 and 3 orders of magnitude lower, respectively.


A review of the epidemiologic literature on the role of environmental arsenic exposure and cardiovascular diseases.
Cardiovascular disease is the leading cause of mortality worldwide. Arsenic is a ubiquitous metalloid in the crust of the earth. Chronic arsenic poisoning is becoming an emerging epidemic in Asia. Epidemiological studies have shown that chronic arsenic poisoning through ingestion of arsenic-contaminated water is associated with various cardiovascular diseases in dose-response relationships. These cardiovascular disorders include carotid atherosclerosis detected by ultrasonography, impaired microcirculation, prolonged QT interval and increased QT dispersion in electrocardiography, and clinical outcomes such as hypertension, blackfoot disease (a unique peripheral vascular disease endemic in southwestern Taiwan), coronary artery disease and cerebral infarction. Chronic arsenic poisoning is an independent risk factor for cardiovascular disease. The adverse cardiovascular effects of long-term arsenic exposure may be persistent and/or irreversible. Arsenic-induced cardiovascular diseases in human population may result from the interaction among genetic, environment and nutritional factors. The major adverse cardiovascular effect of chronic arsenic poisoning has been established qualitatively and quantitatively in the high arsenic exposure areas, but the low-dose effect of arsenic on cardiovascular diseases remains to be explored. Cardiovascular death is the major cause of mortality worldwide, and a small increased risk may imply a large quantity of excess mortality.


Allosteric control of RNA polymerase by a site that contacts nascent RNA hairpins.
DNA, RNA, and regulatory molecules control gene expression through interactions with RNA polymerase (RNAP). We show that a short alpha helix at the tip of the flaplike domain that covers the RNA exit channel of RNAP contacts a nascent RNA stem-loop structure (hairpin) that inhibits transcription, and that this flap-tip helix is required for activity of the regulatory protein NusA. Protein-RNA cross-linking, molecular modeling, and effects of alterations in RNAP and RNA all suggest that a tripartite interaction of RNAP, NusA, and the hairpin inhibits nucleotide addition in the active site, which is located 65 angstroms away. These findings favor an allosteric model for regulation of transcript elongation.


Lack of grading agreement among international hemostasis external quality assessment programs.
: Laboratory quality programs rely on internal quality control and external quality assessment (EQA). EQA programs provide unknown specimens for the laboratory to test. The laboratory's result is compared with other (peer) laboratories performing the same test. EQA programs assign target values using a variety of methods statistical tools and performance assessment of 'pass' or 'fail' is made. EQA provider members of the international organization, external quality assurance in thrombosis and hemostasis, took part in a study to compare outcome of performance analysis using the same data set of laboratory results. Eleven EQA organizations using eight different analytical approaches participated. Data for a normal and prolonged activated partial thromboplastin time (aPTT) and a normal and reduced factor VIII (FVIII) from 218 laboratories were sent to the EQA providers who analyzed the data set using their method of evaluation for aPTT and FVIII, determining the performance for each laboratory record in the data set. Providers also summarized their statistical approach to assignment of target values and laboratory performance. Each laboratory record in the data set was graded pass/fail by all EQA providers for each of the four analytes. There was a lack of agreement of pass/fail grading among EQA programs. Discordance in the grading was 17.9 and 11% of normal and prolonged aPTT results, respectively, and 20.2 and 17.4% of normal and reduced FVIII results, respectively. All EQA programs in this study employed statistical methods compliant with the International Standardization Organization (ISO), ISO 13528, yet the evaluation of laboratory results for all four analytes showed remarkable grading discordance.


Effects of platelet-sparing leukocyte reduction and agitation methods on in vitro measures of hemostatic function in cold-stored whole blood.
Agitation of platelet units stored at room temperature is performed routinely to maintain platelet function, and leukoreduction of blood products is the standard of care in many countries to reduce immune consequences of transfusion. The effect of agitation and leukoreduction on whole blood stored at 4°C requires investigation, as reductions in hemostatic capacity of whole blood may reduce its efficacy in treating trauma-induced coagulopathy and platelet dysfunction. We hypothesize that agitation of whole blood will not affect hemostatic function and that leukoreduction will reduce hemostatic function of whole blood. In this in vitro randomized controlled study, 21 units of leukoreduced and 20 nonleukoreduced whole blood units were each randomly assigned into four agitation groups. Hemostatic parameters were measured using viscoelastic assays (rotational thromboelastometry-Extrinsic Screening Test (ROTEM-EXTEM) and thromboelastography (TEG) platelet mapping), impedance aggregometry (agonists-adenosine phosphate, arachidonic acid, thrombin receptor activating peptide, and collagen), and a thrombin generation assay from these whole blood units before and after filtration and on 0, 5, 10, and 15 days of storage at 4°C. Leukoreduction compared to nonleukoreduction reduced platelet concentration on Day 0. Viscoelastic measures and thrombin generation parameters revealed significant reduction in hemostatic function between the leukoreduced units and the nonleukoreduced units at a few time points. Leukoreduced units consistently demonstrated reduced platelet aggregation compared to the nonleukoreduced units. Agitation methods did not significantly affect any of the hemostatic parameters examined. Leukoreduction of whole blood with a platelet-sparing filter caused a moderate but significant reduction in some measures of whole blood hemostatic function most evident early in storage. The benefits of leukoreduction should be weighed against the potential reduced hemostatic function of leukoreduced units. Agitation of whole blood is not required to maintain hemostatic function. In vitro randomized controlled trial, level 1.


Transfusions and long-term functional outcomes in traumatic brain injury.
In this paper, the authors' goal was to examine the relationship between transfusion and long-term functional outcomes in moderately anemic patients (lowest hematocrit [HCT] level 21-30%) with traumatic brain injury (TBI). While evidence suggests that transfusions are associated with poor hospital outcomes, no study has examined transfusions and long-term functional outcomes in this population. The preferred transfusion threshold remains controversial. The authors performed a retrospective review of patients who were admitted with TBI between September 2005 and November 2007, extracting data such as HCT level, status of red blood cell transfusion, admission Glasgow Coma Scale (GCS) score, serum glucose, and length of hospital stay. Outcome measures assessed at 6 months were Glasgow Outcome Scale-Extended score, Functional Status Examination score, and patient death. A multivariate generalized linear model controlling for confounding variables was used to assess the association between transfusion and outcome. During the study period, 292 patients were identified, and 139 (47.6%) met the criteria for moderate anemia. Roughly half (54.7%) underwent transfusions. Univariate analyses showed significant correlations between outcome score and patient age, admission GCS score, head Abbreviated Injury Scale score, number of days with an HCT level < 30%, highest glucose level, number of days with a glucose level > 200 mg/dl, length of hospital stay, number of patients receiving a transfusion, and transfusion volume. In multivariate analysis, admission GCS score, receiving a transfusion, and transfusion volume were the only variables associated with outcome (F = 2.458, p = 0.007; F = 11.694, p = 0.001; and F = 1.991, p = 0.020, respectively). There was no association between transfusion and death. Transfusions may contribute to poor long-term functional outcomes in anemic patients with TBI. Transfusion strategies should be aimed at patients with symptomatic anemia or physiological compromise, and transfusion volume should be minimized.


Transferable skills of incoming medical students and their development over the first academic year: The United Arab Emirates experience.
Increasingly, it is being recognised in higher and medical education that learners should be adequately prepared for the unpredictable nature of professional practice. Several generic or transferable skills or capabilities (e.g., communication, information handling) that will enable graduates to function in an ever-changing professional world have been identified. Using a validated inventory comprising six categories of transferable skills, three cohorts of incoming male and female medical students at a Gulf university documented their level of practice and confidence for 31 skills. The exercise was repeated a year later. New medical students identified computer and organisational skills and the ability to manage their learning as strengths, but scores for technical and numeracy, information handling and presentation and communication skills suggested that learners generally required guidance. A year later, despite considerable self-reported information handling and communication skills development, learners generally did not consider themselves self-sufficient. A significant gender difference emerged, with incoming males reporting less experience and confidence in many skills. This gap was reduced but did not disappear over the first academic year. An audit such as this may be useful for identifying individual skills levels as well as providing insight into shortcomings in the academic programme in terms of opportunities for transferable skills development.


Retention in care and adherence to ART are critical elements of HIV care interventions.
Retention in care and adherence to antiretroviral treatment (ART) are critical elements of HIV care interventions and are closely associated with optimal individual and public health outcomes and cost effectiveness. This literature review was conducted to analyse how the roles of clients in HIV care and treatment are discussed, from terminology used to measurement methods to consequences of a wide range of patient-related factors impacting client adherence to ART and retention in care. Unfortunately, data suggests that clients find it hard to follow recommended behaviour. For HIV, the greatest loss to follow-up occurs before starting treatment, though each step of the continuum of care is affected. Measurement approaches can be divided into 'direct' and 'indirect' methods; in practice, a combination is often considered the best strategy. Inadequate retention and adherence lead to decreased health outcomes (morbidity, mortality, drug resistances, risk of transmission) and cost effectiveness (increased costs and lower productivity).


Folic acid conjugated Fe3O4 magnetic nanoparticles for targeted delivery of doxorubicin.
The interfacial engineering of magnetic nanoparticles (MNPs) with specific functional groups or targeting ligands is important for their in vivo applications. We report here the preparation and characterization of bifunctional magnetic nanoparticles (BMNPs) which contain a carboxylic moiety for drug binding and an amine moiety for folate mediated drug targeting. BMNPs were prepared by introducing bioactive cysteine molecules onto the surface of undecenoic acid coated Fe3O4 magnetic nanoparticles (UMNPs) via a thiol-ene click reaction and then, folic acid was conjugated with these BMNPs through an EDC-NHS coupling reaction. X-ray diffraction (XRD) and transmission electron microscopy (TEM) analysis indicate the formation of highly crystalline single-phase Fe3O4 nanostructures. The changes in the interfacial characteristics of the nanoparticles and the presence of an organic coating are evident from Fourier transform infrared (FTIR) spectroscopy, dynamic light scattering (DLS), zeta-potential measurement, and thermogravimetric analysis (TGA). These nanocarriers have an average size of 10 nm, and have a pH dependent charge conversional feature and protein resistance characteristic in physiological medium. These nanoparticles also show high loading affinity for an anticancer drug, doxorubicin hydrochloride (DOX) and its pH dependent release. This is highly beneficial for cancer therapy as the relatively low pH in tumors will specifically stimulate the drug release at the site of interest. Furthermore, our fluorescence microscopy and flow cytometry studies confirmed the higher cellular internalization capability of these folic acid conjugated nanoparticles in cancer cells over-expressing folate receptors.


The hunter and the pianist: two hepatic encephalopathy tales.
Details of 2 patients with cirrhosis and hepatic encephalopathy whose hobby or job were possibly responsible for a selectively enhanced performance in 1 neuropsychiatric test are reported. Clinicians should be alert to the fact that personal inclinations and habits may impinge on both neuropsychological and psychophysic performance, thus producing a mismatch between the results of different mental status tests. A prospective study with accurate history taking, use of comprehensive assessment protocols, and modeling/critical interpretation of the test results is required to confirm this hypothesis.


Immunomodulators and immunosuppressants for relapsing-remitting multiple sclerosis: a network meta-analysis.
Different therapeutic strategies are available for the treatment of people with relapsing-remitting multiple sclerosis (RRMS), including immunomodulators, immunosuppressants and biologics. Although there is consensus that these therapies reduce the frequency of relapses, their relative benefit in delaying new relapses or disability worsening remains unclear due to the limited number of direct comparison trials. To compare the benefit and acceptability of interferon beta-1b, interferon beta-1a (Avonex, Rebif), glatiramer acetate, natalizumab, mitoxantrone, fingolimod, teriflunomide, dimethyl fumarate, alemtuzumab, pegylated interferon beta-1a, daclizumab, laquinimod, azathioprine and immunoglobulins for the treatment of people with RRMS and to provide a ranking of these treatments according to their benefit and acceptability, defined as the proportion of participants who withdrew due to any adverse event. We searched the Cochrane Multiple Sclerosis and Rare Diseases of the CNS Group Trials Register, which contains trials from CENTRAL (2014, Issue 9), MEDLINE (1966 to 2014), EMBASE (1974 to 2014), CINAHL (1981 to 2014), LILACS (1982 to 2014), clinicaltrials.gov and the WHO trials registry, and US Food and Drug Administration (FDA) reports. We ran the most recent search in September 2014. Randomised controlled trials (RCTs) that studied one or more of the 15 treatments as monotherapy, compared to placebo or to another active agent, for use in adults with RRMS. Two authors independently identified studies from the search results and performed data extraction. We performed data synthesis by pairwise meta-analysis and network meta-analysis. We assessed the quality of the body of evidence for outcomes within the network meta-analysis according to GRADE, as very low, low, moderate or high. We included 39 studies in this review, in which 25,113 participants were randomised. The majority of the included trials were short-term studies, with a median duration of 24 months. Twenty-four (60%) were placebo-controlled and 15 (40%) were head-to-head studies.Network meta-analysis showed that, in terms of a protective effect against the recurrence of relapses in RRMS during the first 24 months of treatment, alemtuzumab, mitoxantrone, natalizumab, and fingolimod outperformed other drugs. The most effective drug was alemtuzumab (risk ratio (RR) versus placebo 0.46, 95% confidence interval (CI) 0.38 to 0.55; surface under the cumulative ranking curve (SUCRA) 96%; moderate quality evidence), followed by mitoxantrone (RR 0.47, 95% CI 0.27 to 0.81; SUCRA 92%; very low quality evidence), natalizumab (RR 0.56, 95% CI 0.47 to 0.66; SUCRA 88%; high quality evidence), and fingolimod (RR 0.72, 95% CI 0.64 to 0.81; SUCRA 71%; moderate quality evidence).Disability worsening was based on a surrogate marker, defined as irreversible worsening confirmed at three-month follow-up, measured during the first 24 months in the majority of included studies. Both direct and indirect comparisons revealed that the most effective treatments were mitoxantrone (RR versus placebo 0.20, 95% CI 0.05 to 0.84; SUCRA 96%; low quality evidence), alemtuzumab (RR 0.35, 95% CI 0.26 to 0.48; SUCRA 94%; low quality evidence), and natalizumab (RR 0.64, 95% CI 0.49 to 0.85; SUCRA 74%; moderate quality evidence).Almost all of the agents included in this review were associated with a higher proportion of participants who withdrew due to any adverse event compared to placebo. Based on the network meta-analysis methodology, the corresponding RR estimates versus placebo over the first 24 months of follow-up were: mitoxantrone 9.92 (95% CI 0.54 to 168.84), fingolimod 1.69 (95% CI 1.32 to 2.17), natalizumab 1.53 (95% CI 0.93 to 2.53), and alemtuzumab 0.72 (95% CI 0.32 to 1.61).Information on serious adverse events (SAEs) was scanty, characterised by heterogeneous results and based on a very low number of events observed during the short-term duration of the trials included in this review. Conservative interpretation of these results is warranted, since most of the included treatments have been evaluated in few trials. The GRADE approach recommends providing implications for practice based on moderate to high quality evidence. Our review shows that alemtuzumab, natalizumab, and fingolimod are the best choices for preventing clinical relapses in people with RRMS, but this evidence is limited to the first 24 months of follow-up. For the prevention of disability worsening in the short term (24 months), only natalizumab shows a beneficial effect on the basis of moderate quality evidence (all of the other estimates were based on low to very low quality evidence). Currently, therefore, insufficient evidence is available to evaluate treatments for the prevention of irreversible disability worsening.There are two additional major concerns that have to be considered. First, the benefit of all of these treatments beyond two years is uncertain and this is a relevant issue for a disease with a duration of 30 to 40 years. Second, short-term trials provide scanty and poorly reported safety data and do not provide useful evidence in order to obtain a reliable risk profile of treatments. In order to provide long-term information on the safety of the treatments included in this review, it will be necessary also to evaluate non-randomised studies and post-marketing reports released from the regulatory agencies. Finally, more than 70% of the studies included in this review were sponsored by pharmaceutical companies and this may have influenced the results.There are three needs that the research agenda should address. First, randomised trials of direct comparisons between active agents would be useful, avoiding further placebo-controlled studies. Second, follow-up of the original trial cohorts should be mandatory. Third, more studies are needed to assess the medium and long-term benefit and safety of immunotherapies and the comparative safety of different agents.


Adrenal insufficiency in critically ill emergency department patients: a Taiwan preliminary study.
Unrecognized adrenal insufficiency can have serious consequences in critically ill emergency department (ED) patients. This prospective pilot study of adrenal function in patients with severe illness was undertaken to determine the prevalence of adrenal dysfunction and any relation to prior herbal drug use. In a high-volume urban tertiary care ED, adult patients with sepsis or acute myocardial infarction (AMI) were eligible for the study. Over a two-month period, a convenience sample was enrolled by the authors on arrival to the ED. Inclusion criteria were systemic inflammatory response syndrome (SIRS) criteria plus evidence of at least one organ dysfunction or cardiac marker plus electrocardiogram-proven AMI. Exclusion criteria included known corticosteroid use. Serum cortisol was measured on arrival and for those patients with a level of <15 microg/dL (<414 nmol/L), an adrenocorticotropic hormone (ACTH) stimulation test was performed. Of the 30 enrolled patients, 23 (77%) were suffering from severe sepsis and the other seven (23%) had an AMI. Thirteen of the 30 patients (43%; 95% CI = 25% to 65%) had serum cortisol levels of <15 microg/dL, consistent with adrenal insufficiency, nine with severe sepsis and four with an AMI. Eight (62%; 95% CI = 32% to 86%) of the 13 patients with low cortisol levels reported using herbal medications, while only two (12%; 95% CI = 1% to 36%) of the 17 with normal cortisol levels reported taking herb drugs (p = 0.01). Only two (15%; 95% CI = 2% to 45%) of the patients with low cortisol levels failed their corticotropin stimulation test, suggestive of true adrenocortical insufficiency. Both reported using herbal preparations. These results indicate that adrenal dysfunction is common among a group of critically ill patients seen in this Taiwanese ED. Moreover, the use of herbal drugs was high in the patients with low serum cortisols. Further studies are required to both confirm these findings and clarify whether a number of herbal medications contain corticosteroids.


Thrombotic complications and thromboprophylaxis across all three stages of single ventricle heart palliation.
To describe the incidence of thrombotic complications across all 3 stages of single ventricle palliation and the association between thromboprophylaxis use and thrombotic risk. Two separate cross-sectional studies were performed that included 195 patients born between 2003-2008 and 162 patients who underwent Fontan after 2000. The incidence of thrombotic complications was 40% and 28% after initial palliation and superior cavopulmonary connection (SCPC), respectively; 5-year freedom from thrombotic complications after Fontan was 79%. Thromboprophylaxis was initiated for 70%, 46%, and 94% of patients after initial palliation, SCPC, and Fontan, respectively. Thromboprophylaxis with enoxaparin (vs no thromboprophylaxis) was associated with a reduction in risk of thrombotic complications after initial palliation (hazard ratio [HR] 0.5, P = .05) and SCPC (HR 0.2, P = .04). Thromboprophylaxis with warfarin was associated with a reduction in thrombotic complications after Fontan (HR 0.27, P = .05 vs acetylsalicylic acid; HR 0.18, P = .02 vs no thromboprophylaxis). Thrombotic complications were associated with increased mortality after initial palliation (HR 5.5, P < .001) and SCPC (HR 12.5, P < .001). Three patients experienced major bleeding complications without permanent sequelae (2 enoxaparin, 1 warfarin). Given the negative impact of thrombotic complications on survival, the low risk of serious bleeding complications, and the association between thromboprophylaxis and lowered thrombotic complication risk across all 3 palliative stages, routine use of thromboprophylaxis from the initial palliation to the early post-Fontan period in this population may be indicated.


A monoclonal antibody to guanine nucleotide binding protein inhibits the light-activated cyclic GMP pathway in frog rod outer segments.
A monoclonal antibody that blocks the light-activated cyclic GMP (cGMP) pathway in frog photoreceptor outer segments (ROS) has been obtained. The antibody (4A) inhibits guanine nucleotide binding to G-protein, the intermediate that links rhodopsin excitation to cGMP phosphodiesterase (PDE), inhibiting light-induced PDE activity as a consequence. Antibody inhibition of the light-activated cGMP pathway is complete at a stoichiometry of approximately one antibody per G-protein in the mixture, which indicates high specificity of the inhibition. Inhibition is more pronounced than that caused by PDE inhibitors such as isobutylmethylxanthine (IBMX) or Ro 20-1724. Antibody 4A has the further effect of inhibiting the phosphorylation of two low molecular weight proteins, components I and II, whose phosphorylation normally can be stimulated by raising cGMP levels. The inhibition is not overridden by adding cGMP, which suggests that the G-protein influences these phosphorylations by a pathway distinct from its action on cGMP concentration. Antibody 4A may prove useful as a probe of the relevance of the cGMP pathway to visual transduction in living photoreceptors. Six other monoclonal antibodies to G-protein, as well as six monoclonal antibodies to rhodopsin and one to PDE, do not block light-activated guanine nucleotide binding, PDE activity, or ROS protein phosphorylations.


Enantioseparation by crystallization using magnetic substrates.
Enantiospecific crystallization of the three amino acids asparagine (Asn), glutamic acid hydrochloride (Glu·HCl) and threonine (Thr), induced by ferromagnetic (FM) substrates, is reported. The FM substrates were prepared by evaporating nickel capped with a thin gold layer on standard silicon wafers. Magnets were positioned underneath the substrate with either their North (N) or South (S) poles pointing up. Asymmetric induction, controlled by the magnetic substrates, was demonstrated for the crystallization of the pure enantiomers and was then extended for the racemic mixtures of Asn and Glu·HCl. In the case of the solution of the pure enantiomers, the l enantiomer was crystallized preferentially at one pole of the magnet and the d enantiomer at the other. Consequently, the racemates of Asn and Glu·HCl undergo separation under the influence of the magnetic substrate. With Thr, however, despite the enantiospecific interactions of the pure enantiomers with the FM, no separation of the emerging crystals could be achieved with the racemates, although they crystallize as conglomerates, implying differences taking place in the crystallization step. The results reported here are not directly related to the magnetic field, but rather to the aligned spins within the ferromagnets. The findings provide a novel method for resolving enantiomers by crystallization and offer a new perspective for a possible role played by magnetic substrates regarding the origin of chirality in nature.


Atomistic insight into orthoborate-based ionic liquids: force field development and evaluation.
We have developed an all-atomistic force field for a new class of halogen-free chelated orthoborate-phosphonium ionic liquids. The force field is based on an AMBER framework with determination of force field parameters for phosphorus and boron atoms, as well as refinement of several available parameters. The bond and angle force constants were adjusted to fit vibration frequency data derived from both experimental measurements and ab initio calculations. The force field parameters for several dihedral angles were obtained by fitting torsion energy profiles deduced from ab initio calculations. To validate the proposed force field parameters, atomistic simulations were performed for 12 ionic liquids consisting of tetraalkylphosphonium cations and chelated orthoborate anions. The predicted densities for neat ionic liquids and the [P6,6,6,14][BOB] sample, with a water content of approximately 2.3-2.5 wt %, are in excellent agreement with available experimental data. The potential energy components of 12 ionic liquids were discussed in detail. The radial distribution functions and spatial distribution functions were analyzed and visualized to probe the microscopic ionic structures of these ionic liquids. There are mainly four high-probability regions of chelated orthoborate anions distributed around tetraalkylphosphonium cations in the first solvation shell, and such probability distribution functions are strongly influenced by the size of anions.


Use of mechanical circulatory support in patients undergoing percutaneous coronary intervention: insights from the National Cardiovascular Data Registry.
Little is known about the contemporary use of intra-aortic balloon pump (IABP) and other mechanical circulatory support (O-MCS) devices in patients undergoing percutaneous coronary intervention (PCI) in the setting of cardiogenic shock. We identified 76 474 patients who underwent PCI in the setting of cardiogenic shock at one of 1429 National Cardiovascular Data Registry CathPCI participating hospitals from 2009 to 2013. Temporal trends and hospital-level variation in the use of IABP and O-MCS were evaluated. No mechanical circulatory support was used in 41 286 (54%) patients, 29 730 (39%) received IABP only, 2711 (3.5%) received O-MCS only, and 2747 (3.6%) received both IABP and O-MCS. At the start of the study period, 45% of patients undergoing PCI in the setting of cardiogenic shock received an IABP and 6.7% received O-MCS. The proportion of patients receiving IABP declined at an average rate of 0.3% per quarter, whereas the rate of O-MCS use was unchanged over the study period. The predicted probability of IABP use varied significantly by site (hospital median 42%, interquartile range 33% to 51%, range 8% to 85%). The probability of O-MCS use was <5% for half of hospitals and >20% in less than one-tenth of hospitals. In this large national registry, the use of IABP in the setting of PCI for cardiogenic shock decreased over time without a concurrent increase in O-MCS use. The probability of IABP and O-MCS use varied across hospitals, and the use of O-MCS was clustered at a small number of hospitals.


The College of American Pathologists Biorepository Accreditation Program: Results from the First 5 Years.
The College of American Pathologists (CAP) developed the Biorepository Accreditation Program (BAP) in 2012. This program integrates best practices from the International Society for Biological and Environmental Biorepositories, the National Cancer Institute, the Organisation for Economic Cooperation and Development, the Center for Medicare and Medicaid Services, and the CAP Laboratory Accreditation Program. The goal of this elective program is to provide requirements for standardization in biorepository processes that will result in high-quality specimens that can be used to support research, drug discovery, and personalized medicine. CAP uses a peer inspection model to ensure the inspectors have proper expertise and to promote educational efforts through information sharing. Lead inspectors are comprised of pathologists, PhDs, and managers of biorepositories and they are often supported by CAP staff inspectors. Accreditation is a 3-year continuous cycle of quality with a peer inspection occurring at the start of year 1 and a self-inspection and CAP desk assessment at the start of year 2 and 3. At this time 53 biorepositories are fully CAP BAP accredited and 13 are in the process of obtaining accreditation. There are currently 273 established standards with requirement lists customized based on the scope of activities performed by a biorepository. A total of 90 inspections were completed between May 2012 and December 2016. Sixty-one were initial inspections and 29 were reinspections. A total of 527 deficiencies were identified in the areas of Equipment/Instrumentation (22%), Information Technology (18%), Specimen Handling and QC (15%), Quality Management (16%), Personnel (11%), Safety (10%), Facilities (6%), and Regulatory (2%). Assessment of common deficiencies identifies areas of focus for continuous improvement and educational opportunities. Overall success of the program is high based on the current enrollment of 66 biorepositories, anecdotal participant feedback and increasing national recognition of the BAP in federal documents.


Homing properties of adipose-derived stem cells to intracerebral glioma and the effects of adenovirus infection.
The inevitable clinical recurrence of high grade gliomas after standard treatment is due to the highly diffuse infiltrating parts of these tumors, which remain after surgery and respond poorly to radiation and chemotherapy. It has been proposed to employ the homing capacity of neural stem cells (NSCs) to different types of intracerebral pathology for selective targeting of glioma cells, and delivery of transgenic expressed therapeutics. This approach has been successful in a number of preclinical experimental studies, however, a major drawback for clinical translation has been the limitation of harvesting and ex vivo expansion of NSCs in patients. Here we demonstrate that adipose derived stem cells (ASCs), which are easily harvested in relatively large quantities in humans, display the same tropism for gliomas as NSCs in vitro and in vivo. Both ipsilateral as well as contralateral injection of these cells in brains of glioma-bearing mice, led to extensive homing to the tumor by the ASCs. The potential of loading these cellular vehicles with transgenes was assessed using adenoviral vectors. ASCs could be infected with adenoviral vectors, albeit at very high MOI. Insertion of the arg-gly-asp (RGD) motif into the adenovirus fiber knob, thereby redirecting primary attachment of the virus to integrins, resulted in a striking 7000-fold increase in infection efficiency. However, in vivo migration of adenovirus-infected ASCs was not observed, most likely due to an inflammatory response to these cells which was not observed with control non-infected ASCs. These results indicate that ASCs are an interesting candidate for further development for cell-based therapy of gliomas, however adenoviruses are not appropriate vectors for delivery of transgenes in this context.


Emotion perception, but not affect perception, is impaired with semantic memory loss.
For decades, psychologists and neuroscientists have hypothesized that the ability to perceive emotions on others' faces is inborn, prelinguistic, and universal. Concept knowledge about emotion has been assumed to be epiphenomenal to emotion perception. In this article, we report findings from 3 patients with semantic dementia that cannot be explained by this "basic emotion" view. These patients, who have substantial deficits in semantic processing abilities, spontaneously perceived pleasant and unpleasant expressions on faces, but not discrete emotions such as anger, disgust, fear, or sadness, even in a task that did not require the use of emotion words. Our findings support the hypothesis that discrete emotion concept knowledge helps transform perceptions of affect (positively or negatively valenced facial expressions) into perceptions of discrete emotions such as anger, disgust, fear, and sadness. These findings have important consequences for understanding the processes supporting emotion perception.


Death in the United States, 2007.
Data from the National Vital Statistics System, Mortality In 2007, the age-adjusted death rate for the United States reached a record low of 760.3 per 100,000 population. Life expectancy at birth reached a record high of 77.9 years. States in the southeast region have higher death rates than those in other regions of the country. In 2007, the five leading causes of death were heart disease, cancer, stroke, chronic lower respiratory diseases, and accidents. These accounted for over 64 percent of all deaths in the United States. White females have the longest life expectancy (80.7 years), followed by black females (77.0 years). The gap in life expectancy between white persons and black persons declined by 35 percent between 1989 and 2007. The race differential was 4.6 years in 2007.


Modifying action sounds influences people's emotional responses and bodily sensations.
We report an experiment designed to investigate the effect of modifying the sound of high-heeled shoes on women's self-reported valence, arousal, and dominance scores, as well as any changes to a variety of measures of bodily sensation. We also assessed whether self-evaluated personality traits and the enjoyment associated with wearing heels were correlated with these effects. Forty-eight women walked down a "virtual runway" while listening to four interaction sounds (leather- and polypropylene-soled high-heeled shoes contacting ceramic flooring or carpet). Analysis of the questionnaires that the participants completed indicated that the type of sonic interaction impacted valence, arousal, and dominance scores, as well as the evaluated bodily sensations. There were also correlations between these scores and both self-evaluated personality traits and the reported enjoyment associated with wearing high heels. These results demonstrate the effect that the sound of a woman's physical interaction with the environment can have, especially when her contact with the ground while walking makes a louder sound. More generally, these results demonstrate that the manipulation of product extrinsic sounds can modify people's evaluation of their emotional outcomes (valence, arousal, and dominance), as well as their bodily sensations.


Colonization of a Central Venous Catheter by the Hyaline Fungus Fusarium solani Species Complex: A Case Report and SEM Imaging.
The incidence of opportunistic infections by filamentous fungi is increasing partly due to the widespread use of central venous catheters (CVC), indwelling medical devices, and antineoplastic/immunosuppressive drugs. The case of a 13-year-old boy under treatment for acute lymphoblastic leukemia is presented. The boy was readmitted to the Pediatric Ward for intermittent fever of unknown origin. Results of blood cultures drawn from peripheral venous sites or through the CVC were compared. CVC-derived bottles (but not those from peripheral veins) yielded hyaline fungi that, based on morphology, were identified as belonging to the Fusarium solani species complex. Gene amplification and direct sequencing of the fungal ITS1 rRNA region and the EF-1alpha gene confirmed the isolate as belonging to the Fusarium solani species complex. Portions of the CVC were analyzed by scanning electron microscopy. Fungi mycelia with long protruding hyphae were seen into the lumen. The firm adhesion of the fungal formation to the inner surface of the catheter was evident. In the absence of systemic infection, catheter removal and prophylactic voriconazole therapy were followed by disappearance of febrile events and recovery. Thus, indwelling catheters are prone to contamination by environmental fungi.


Automated Adherence Reminders for High Risk Children With Asthma: A Research Protocol.
The use of inhaled corticosteroid (ICS) medications has been shown to improve asthma control and reduce asthma-related morbidity and mortality. Two recent randomized trials demonstrated dramatic improvements in ICS adherence by monitoring adherence with electronic sensors and providing automated reminders to participants to take their ICS medications. Given their lower levels of adherence and higher levels of asthma-related emergency department (ED) visits, hospitalizations, and death, urban minority populations could potentially benefit greatly from these types of interventions. The principal objective of this study will be to evaluate the feasibility, acceptability, and limited efficacy of a text message (short message service, SMS) reminder intervention to enhance ICS adherence in an urban minority population of children with asthma. We will also assess trajectories of ICS adherence in the 2 months following asthma hospitalization. Participants will include 40 children aged 2-13 years, who are currently admitted to the Children's Hospital of Philadelphia (CHOP) for asthma, and their parent or legal guardian. Participants will be assigned to intervention and control arms using a 1:1 randomization scheme. The intervention arm will receive daily text message reminders for a 30-day intervention phase following hospitalization. This will be followed by a 30-day follow-up phase, in which all participants may choose whether or not to receive the text messages. Feasibility will be assessed by measuring (1) retention of the participants through the study phases and (2) perceived usefulness, acceptability, and preferences regarding the intervention components. Limited efficacy outcomes will include percent adherence to prescribed ICS regimen measured using Propeller Health sensors and change in parent-reported asthma control. We will perform an exploratory analysis to assess for discrete trajectories of adherence using group-based trajectory modeling (GBTM). Study enrollment began in December 2015 and the intervention and follow-up phases are ongoing. Results of the data analysis are expected to be available by December 2016. This study will add to the literature by providing foundational feasibility data on which elements of a mobile health text-message reminder intervention may need to be modified to suit the needs and constraints of high-risk urban minority populations. Clinicaltrials.gov NCT02615743; https://www.clinicaltrials.gov/ct2/show/study/NCT02615743 (Archived with WebCite at http://www.webcitation.org/6ji59rAXN).


Epidemiology of neonatal hyperbilirubinemia.
Interview and record review data from 12,023 singleton deliveries were analyzed to determine the relationships between neonatal hyperbilirubinemia (10 mg/dL or greater) and maternal characteristics. Confounding variables were controlled by multiple logistic regression analysis. There was a statistically significant positive relationship between hyperbilirubinemia and low birth weight, Oriental race, premature rupture of membranes, breast-feeding, neonatal infection, use of the "pill" at time of conception, instrumental delivery, and history of first trimester bleeding. Maternal smoking and black race were negatively related to hyperbilirubinemia and statistically significant. In this study, other previously suspected etiologic factors such as epidural anesthesia, parity, use of oxytocin in labor, and white race were not associated with hyperbilirubinemia.


Assessing the potential for intraguild predation among taxonomically disparate micro-carnivores: marsupials and arthropods.
Interspecific competition may occur when resources are limited, and is often most intense between animals in the same ecological guild. Intraguild predation (IGP) is a distinctive form of interference competition, where a dominant predator selectively kills subordinate rivals to gain increased access to resources. However, before IGP can be identified, organisms must be confirmed as members of the same guild and occur together in space and time. The lesser hairy-footed dunnart (Sminthopsis youngsoni, Dasyuridae) is a generalist marsupial insectivore in arid Australia, but consumes wolf spiders (Lycosa spp., Lycosidae) disproportionately often relative to their availability. Here, we test the hypothesis that this disproportionate predation is a product of frequent encounter rates between the interactants due to high overlap in their diets and use of space and time. Diet and prey availability were determined using direct observations and invertebrate pitfall trapping, microhabitat use by tracking individuals of both species-groups, and temporal activity using spotlighting and camera traps. Major overlap (greater than 75% similarity) was found in diet and temporal activity, and weaker overlap in microhabitat use. Taken together, these findings suggest reasonable potential, for the first time, for competition and intraguild predation to occur between taxa as disparate as marsupials and spiders.


Technology trends in antibody purification.
This article reviews technology trends in antibody purification. Section 1 discusses non-chromatography methods, including precipitation, liquid-liquid extraction, and high performance tangential flow filtration. The second addresses chromatography methods. It begins with discussion of fluidized and fixed bed formats. It continues with stationary phase architecture: diffusive particles, perfusive particles, membranes and monoliths. The remainder of the section reviews recent innovations in size exclusion, anion exchange, cation exchange, hydrophobic interaction, immobilized metal affinity, mixed-mode, and bioaffinity chromatography. Section 3 addresses an emerging trend of formulating process buffers to prevent or correct anomalies in the antibodies being purified. Methods are discussed for preventing aggregate formation, dissociating antibody-contaminant complexes, restoring native antibody from aggregates, and conserving or restoring native disulfide pairing.


The Role of Microbiome and Genotype in Daphnia magna upon Parasite Re-Exposure.
Recently, it has been shown that the community of gut microorganisms plays a crucial role in host performance with respect to parasite tolerance. Knowledge, however, is lacking on the role of the gut microbiome in mediating host tolerance after parasite re-exposure, especially considering multiple parasite infections. We here aimed to fill this knowledge gap by studying the role of the gut microbiome on tolerance in Daphnia magna upon multiple parasite species re-exposure. Additionally, we investigated the role of the host genotype in the interaction between the gut microbiome and the host phenotypic performance. A microbiome transplant experiment was performed in which three germ-free D. magna genotypes were exposed to a gut microbial inoculum and a parasite community treatment. The gut microbiome inocula were pre-exposed to the same parasite communities or a control treatment. Daphnia performance was monitored, and amplicon sequencing was performed to characterize the gut microbial community. Our experimental results showed that the gut microbiome plays no role in Daphnia tolerance upon parasite re-exposure. We did, however, find a main effect of the gut microbiome on Daphnia body size reflecting parasite specific responses. Our results also showed that it is rather the Daphnia genotype, and not the gut microbiome, that affected parasite-induced host mortality. Additionally, we found a role of the genotype in structuring the gut microbial community, both in alpha diversity as in the microbial composition.


The safety and efficacy of {+/-}3,4-methylenedioxymethamphetamine-assisted psychotherapy in subjects with chronic, treatment-resistant posttraumatic stress disorder: the first randomized controlled pilot study.
Case reports indicate that psychiatrists administered ±3,4-methylenedioxymethamphetamine (MDMA) as a catalyst to psychotherapy before recreational use of MDMA as 'Ecstasy' resulted in its criminalization in 1985. Over two decades later, this study is the first completed clinical trial evaluating MDMA as a therapeutic adjunct. Twenty patients with chronic posttraumatic stress disorder, refractory to both psychotherapy and psychopharmacology, were randomly assigned to psychotherapy with concomitant active drug (n = 12) or inactive placebo (n = 8) administered during two 8-h experimental psychotherapy sessions. Both groups received preparatory and follow-up non-drug psychotherapy. The primary outcome measure was the Clinician-Administered PTSD Scale, administered at baseline, 4 days after each experimental session, and 2 months after the second session. Neurocognitive testing, blood pressure, and temperature monitoring were performed. After 2-month follow-up, placebo subjects were offered the option to re-enroll in the experimental procedure with open-label MDMA. Decrease in Clinician-Administered PTSD Scale scores from baseline was significantly greater for the group that received MDMA than for the placebo group at all three time points after baseline. The rate of clinical response was 10/12 (83%) in the active treatment group versus 2/8 (25%) in the placebo group. There were no drug-related serious adverse events, adverse neurocognitive effects or clinically significant blood pressure increases. MDMA-assisted psychotherapy can be administered to posttraumatic stress disorder patients without evidence of harm, and it may be useful in patients refractory to other treatments.


Differing calcification processes in cultured vascular smooth muscle cells and osteoblasts.
Arterial medial calcification (AMC) is the deposition of calcium phosphate mineral, often as hydroxyapatite, in the medial layer of the arteries. AMC shares some similarities to skeletal mineralisation and has been associated with the transdifferentiation of vascular smooth muscle cells (VSMCs) towards an osteoblast-like phenotype. This study used primary mouse VSMCs and calvarial osteoblasts to directly compare the established and widely used in vitro models of AMC and bone formation. Significant differences were identified between osteoblasts and calcifying VSMCs. First, osteoblasts formed large mineralised bone nodules that were associated with widespread deposition of an extracellular collagenous matrix. In contrast, VSMCs formed small discrete regions of calcification that were not associated with collagen deposition and did not resemble bone. Second, calcifying VSMCs displayed a progressive reduction in cell viability over time (≤7-fold), with a 50% increase in apoptosis, whereas osteoblast and control VSMCs viability remained unchanged. Third, osteoblasts expressed high levels of alkaline phosphatase (TNAP) activity and TNAP inhibition reduced bone formation by to 90%. TNAP activity in calcifying VSMCs was ∼100-fold lower than that of bone-forming osteoblasts and cultures treated with β-glycerophosphate, a TNAP substrate, did not calcify. Furthermore, TNAP inhibition had no effect on VSMC calcification. Although, VSMC calcification was associated with increased mRNA expression of osteoblast-related genes (e.g. Runx2, osterix, osteocalcin, osteopontin), the relative expression of these genes was up to 40-fold lower in calcifying VSMCs versus bone-forming osteoblasts. In summary, calcifying VSMCs in vitro display some limited osteoblast-like characteristics but also differ in several key respects: 1) their inability to form collagen-containing bone; 2) their lack of reliance on TNAP to promote mineral deposition; and, 3) the deleterious effect of calcification on their viability.


Prophylactic postoperative measures to minimize surgical site infections in spine surgery: systematic review and evidence summary.
There are three phases in prophylaxis of surgical site infections (SSI): preoperative, intraoperative and postoperative. There is lack of consensus and paucity of evidence with SSI prophylaxis in the postoperative period. To systematically evaluate the literature, and provide evidence-based summaries on postoperative measures for SSI prophylaxis in spine surgery. Systematic review, meta-analysis, evidence synthesis. A systematic review conforming to PRIMSA guidelines was performed utilizing PubMed (MEDLINE), EMBASE, and the Cochrane Database from inception to January 2019. The GRADE approach was used for quality appraisal and synthesis of evidence. Six postoperative care domains with associated key questions were identified. Included studies were extracted into evidence tables, data synthesized quantitatively and qualitatively, and evidence appraised per GRADE approach. Forty-one studies (nine RCT, 32 cohort studies) were included. In the setting of preincisional antimicrobial prophylaxis (AMP) administration, use of postoperative AMP for SSI reduction has not been found to reduce rate of SSI in lumbosacral spine surgery. Prolonged administration of AMP for more than 48 hours postoperatively does not seem to reduce the rate of SSI in decompression-only or lumbar spine fusion surgery. Utilization of wound drainage systems in lumbosacral spine and adolescent idiopathic scoliosis corrective surgery does not seem to alter the overall rate of SSI in spine surgery. Concomitant administration of AMP in the presence of a wound drain does not seem to reduce the overall rate of SSI, deep SSI, or superficial SSI in thoracolumbar fusion performed for degenerative and deformity spine pathologies, and in adolescent idiopathic scoliosis corrective surgery. Enhanced-recovery after surgery clinical pathways and infection-specific protocols do not seem to reduce rate of SSI in spine surgery. Insufficient evidence exists for other types of spine surgery not mentioned above, and also for non-AMP pharmacological measures, dressing type and duration, suture and staple management, and postoperative nutrition for SSI prophylaxis in spine surgery. Despite the postoperative period being key in SSI prophylaxis, the literature is sparse and without consensus on optimum postoperative care for SSI prevention in spine surgery. The current best evidence is presented with its limitations. High quality studies addressing high risk cohorts such as the elderly, obese, and diabetic populations, and for traumatic and oncological indications are urgently required.


Evidence-based patient information programme in early multiple sclerosis: a randomised controlled trial.
To evaluate the efficacy of an evidence-based patient information programme aiming to increase informed choice in patients with early multiple sclerosis (MS). Patients with early MS face a number of uncertainties concerning diagnosis, prognosis and effectiveness of immunotherapy. Prior studies suggest that evidence-based patient information combined with group education can promote informed choice in MS patients. A 12-month, six-centre, double-blind randomised controlled clinical trial with 192 patients with a diagnosis of confirmed relapsing-remitting MS or clinical isolated syndrome in Germany. A 4-h interactive evidence-based educational programme was compared with a 4-h MS-specific stress management programme. The primary endpoint was informed choice after 6 months comprising risk knowledge and congruency between attitude towards immunotherapy and actual immunotherapy uptake. Secondary endpoints included autonomy preference, decision autonomy, decisional conflict and satisfaction, anxiety and depression, and number of immunotherapies. For the primary endpoint, a significant difference was shown with 50 of 85 (59%) participants in the intervention group achieving informed choice after 6 months compared with 18 of 89 (20%) in the control group (OR 0.2 (95% CI 0.1 to 0.4), p<0.001). Four weeks after the intervention, more participants in the intervention group showed good risk knowledge (difference between groups 39% (95% CI 26% to 53%), p<0.001). There were no significant differences between groups for attitude towards immunotherapy and for immunotherapy uptake. There were trends towards increased autonomy preference after the intervention and increased adherence to immunotherapies in the intervention group. The intervention significantly increased informed choice and relevant risk knowledge without negative side effects.


Male infertility testing: reactive oxygen species and antioxidant capacity.
Reactive oxygen species (ROS) are an integral component of sperm developmental physiology, capacitation, and function. Elevated ROS levels, from processes such as infection or inflammation, can be associated with aberrations of sperm development, function, and fertilizing capacity. We review the impact of ROS on sperm physiology, its place in infertility evaluation, the implications for reproductive outcomes, and antioxidant therapy. Our systematic review of PubMed literature from the last 3 decades focuses on the physiology and etiology of ROS and oxidative stress (OS), evaluation of ROS, and antioxidants. ROS is normally produced physiologically and is used to maintain cellular processes such as sperm maturation, capacitation, and sperm-oocyte interaction. When ROS production exceeds the buffering capacity of antioxidants, OS occurs and can have a negative impact on sperm and fertility. ROS and antioxidant capacity testing can potentially add additional prognostic information to standard laboratory testing for the infertile male, although its role as standard part of an evaluation has yet to be determined. Elevated ROS levels have been implicated with abnormal semen parameters and male infertility, but the impact of ROS on fertilization rates and pregnancy is controversial. This is partly because of the lack of consensus on what type of patients may be suitable for ROS testing and assay standardization. Routine ROS testing for the infertile male is not currently recommended.


Histone monoubiquitylation position determines specificity and direction of enzymatic cross-talk with histone methyltransferases Dot1L and PRC2.
It is well established that chromatin is a destination for signal transduction, affecting many DNA-templated processes. Histone proteins in particular are extensively post-translationally modified. We are interested in how the complex repertoire of histone modifications is coordinately regulated to generate meaningful combinations of "marks" at physiologically relevant genomic locations. One important mechanism is "cross-talk" between pre-existing histone post-translational modifications and enzymes that subsequently add or remove modifications on chromatin. Here, we use chemically defined "designer" nucleosomes to investigate novel enzymatic cross-talk relationships between the most abundant histone ubiquitylation sites, H2AK119ub and H2BK120ub, and two important histone methyltransferases, Dot1L and PRC2. Although the presence of H2Bub in nucleosomes greatly stimulated Dot1L methylation of H3K79, we found that H2Aub did not influence Dot1L activity. In contrast, we show that H2Aub inhibited PRC2 methylation of H3K27, but H2Bub did not influence PRC2 activity. Taken together, these results highlight how the position of nucleosome monoubiquitylation affects the specificity and direction of cross-talk with enzymatic activities on chromatin.


The endocannabinoid system in the regulation of emotions throughout lifespan: a discussion on therapeutic perspectives.
Alterations in emotion regulation processes may form the basis of psychopathologies. The endocannabinoid (eCB) system, composed of endogenous ligands, the enzymatic machinery in charge of their metabolism and the specific metabotropic receptors, has emerged as a major neuromodulatory system critically involved in the control of emotional homeostasis and stress responsiveness. Data from animal models indicate that the eCB system plays a key role in brain development, and is probably involved in the control of emotional states from early developmental stages. The present review summarizes the latest information on the role of the eCB system in emotionality and anxiety-related disorders throughout the lifespan. Putative therapeutic strategies based on the pharmacological modulation of this system will be discussed. Given the fact that the pharmacological modulation of the eCB system has recently arisen as a promising strategy in the management of anxiety and mood disorders, the potential efficacy of this pharmacological approach (i.e. blockers of the catabolic pathway) will be discussed, as well as pharmacological alternatives such as modulators of cannabinoid receptors other than the classical CB1 receptor, or administration of other plant-derived compounds (e.g. cannabidiol).


The temporal dynamics of visual working memory guidance of selective attention.
The biased competition model proposes that there is top-down directing of attention to a stimulus matching the contents of working memory (WM), even when the maintenance of a WM representation is detrimental to target relevant performance. Despite many studies elucidating that spatial WM guidance can be present early in the visual processing system, whether visual WM guidance also influences perceptual selection remains poorly understood. Here, we investigated the electrophysiological correlates of early guidance of attention by WM in humans. Participants were required to perform a visual search task while concurrently maintaining object representations in their visual WM. Behavioral results showed that response times (RTs) were longer when the distractor in the visual search task was held in WM. The earliest WM guidance effect was observed in the P1 component (90-130 ms), with match trials eliciting larger P1 amplitude than mismatch trials. A similar result was also found in the N1 component (160-200 ms). These P1 and N1 effects could not be attributed to bottom-up perceptual priming from the presentation of a memory cue, because there was no significant difference in early event-related potential (ERP) component when the cue was merely perceptually identified but not actively held in WM. Standardized Low Resolution Electrical Tomography Analysis (sLORETA) showed that the early WM guidance occurred in the occipital lobe and the N1-related activation occurred in the parietal gyrus. Time-frequency data suggested that alpha-band event-related spectral perturbation (ERSP) magnitudes increased under the match condition compared with the mismatch condition only when the cue was held in WM. In conclusion, the present study suggests that the reappearance of a stimulus held in WM enhanced activity in the occipital area. Subsequently, this initial capture of attention by WM could be inhibited by competing visual inputs through attention re-orientation, reflecting by the alpha-band rhythm.


The cullin 4B-based UV-damaged DNA-binding protein ligase binds to UV-damaged chromatin and ubiquitinates histone H2A.
By removing UV-induced lesions from DNA, the nucleotide excision repair (NER) pathway preserves the integrity of the genome. The UV-damaged DNA-binding (UV-DDB) protein complex is involved in the recognition of chromatin-embedded UV-damaged DNA, which is the least understood step of NER. UV-DDB consists of DDB1 and DDB2, and it is a component of the cullin 4A (CUL4A)-based ubiquitin ligase, DDB1-CUL4A(DDB2). We previously showed that DDB1-CUL4A(DDB2) ubiquitinates histone H2A at the sites of UV lesions in a DDB2-dependent manner. Mutations in DDB2 cause a cancer prone syndrome, xeroderma pigmentosum group E (XP-E). CUL4A and its paralog, cullin 4B (CUL4B), copurify with the UV-DDB complex, but it is unclear whether CUL4B has a role in NER as a separate E3 ubiquitin ligase. Here, we present evidence that CUL4A and CUL4B form two individual E3 ligases, DDB1-CUL4A(DDB2) and DDB1-CUL4B(DDB2). To investigate CUL4B's possible role in NER, we examined its subcellular localization in unirradiated and irradiated cells. CUL4B colocalizes with DDB2 at UV-damaged DNA sites. Furthermore, CUL4B binds to UV-damaged chromatin as a part of the DDB1-CUL4B(DDB2) E3 ligase in the presence of functional DDB2. In contrast to CUL4A, CUL4B is localized in the nucleus and facilitates the transfer of DDB1 into the nucleus independently of DDB2. Importantly, DDB1-CUL4B(DDB2) is more efficient than DDB1-CUL4A(DDB2) in monoubiquitinating histone H2A in vitro. Overall, this study suggests that DDB1-CUL4B(DDB2) E3 ligase may have a distinctive function in modifying the chromatin structure at the site of UV lesions to promote efficient NER.


The human immunodeficiency virus type 1 envelope confers higher rates of replicative fitness to perinatally transmitted viruses than to nontransmitted viruses.
Selection of a minor viral genotype during perinatal transmission of human Immunodeficiency virus type 1 (HIV-1) has been observed, but there is a lack of information on the correlation of the restrictive transmission with biological properties of the virus, such as replicative fitness. Recombinant viruses expressing the enhanced green fluorescent protein or the Discosoma sp. red fluorescent (DsRed2) protein carrying the V1 to V5 regions of env from seven mother-infant pairs (MIPs) infected by subtype C HIV-1 were constructed, and competition assays were carried out to compare the fitness between the transmitted and nontransmitted viruses. Flow cytometry was used to quantify the frequency of infected cells, and the replicative fitness was determined based on a calculation that takes into account replication of competing viruses in a single infection versus dual infections. Transmitted viruses from five MIPs with the mothers chronically infected showed a restrictive env genotype, and all the recombinant viruses carrying the infants' Env had higher replicative fitness than those carrying the Env from the mothers. This growth fitness is lineage specific and can be observed only within the same MIP. In contrast, in two MIPs where the mothers had undergone recent acute infection, the viral Env sequences were similar between the mothers and infants and showed no further restriction in quasispecies during perinatal transmission. The recombinant viruses carrying the Env from the infants' viruses also showed replication fitness similar to those carrying the mothers' Env proteins. Our results suggest that newly transmitted viruses from chronically infected mothers have been selected to have higher replicative fitness to favor transmission, and this advantage is conferred by the V1 to V5 region of Env of the transmitted viruses. This finding has important implications for vaccine design or development of strategies to prevent HIV-1 transmission.


A practical guideline for the release of patients treated by I-131 based on Monte Carlo dose calculations for family members.
We recently published effective doses per time-integrated activity (mSv MBq(-1) s(-1)) for paediatric and adult family members exposed to an adult patient released from hospital following I-131 therapy. In the present study, we intend to provide medical physicists with a methodology to estimate family member effective dose in daily clinical practice because the duration of post-radiation precautions for the patient-family member exposure scenario has not been explicitly delineated based on the effective dose. Four different exposure scenarios are considered in this study including (1) a patient and a family member standing face to face, (2) a patient and a family member lying side by side, (3) an adult female patient holding a newborn child to her chest and (4) a one-year-old child standing on the lap of an adult female patient following her I-131 therapy. The results of this study suggest that an adult female hyperthyroidism (HT) patient who was administered with 740 MBq should keep a distance of 100 cm from a 15-year-old child for six days and the same distance from other adults for seven days. The HT female patient should avoid holding a newborn against her chest for at least 16 days following hospital discharge, and a female patient treated with 5550 MBq for differentiated thyroid cancer should not hold her newborn child for at least 15 days following hospital discharge. This study also gives dose coefficients allowing one to predict age-specific effective doses to family members given the measured dose rate (mSv h(-1)) of the patient. In conclusion, effective dose-based patient release criteria with a modified NRC two-component model provide a site medical physicist with less restrictive and age-specific radiation precaution guidance as they fully consider a patient's iodine biokinetics and photon attenuation within both the patient and the exposed family members.


Progress in combating cigarette smuggling: controlling the supply chain.
The illicit tobacco trade results in huge losses of revenue to governments, estimated at $US40-50 billion in 2006, and in increased consumption and thus health problems because it makes tobacco available more cheaply. On 20 October 2008 the second meeting of the International Negotiating Body (INB2) on the illicit trade protocol of WHO's Framework Convention on Tobacco Control (FCTC) will discuss measures to tackle the illicit trade in tobacco products. This paper presents the experience over the last decade of three countries, Italy, Spain and the United Kingdom, which shows that tobacco smuggling can be successfully tackled. The evidence strongly suggests that the key to controlling smuggling is controlling the supply chain, and that the supply chain is controlled to a great extent by the tobacco industry.


Dissecting the binding mechanism of the linker histone in live cells: an integrated FRAP analysis.
The linker histone H1 has a fundamental role in DNA compaction. Although models for H1 binding generally involve the H1 C-terminal tail and sites S1 and S2 within the H1 globular domain, there is debate about the importance of these binding regions and almost nothing is known about how they work together. Using a novel fluorescence recovery after photobleaching (FRAP) procedure, we have measured the affinities of these regions individually, in pairs, and in the full molecule to demonstrate for the first time that binding among several combinations is cooperative in live cells. Our analysis reveals two preferred H1 binding pathways and we find evidence for a novel conformational change required by both. These results paint a complex, highly dynamic picture of H1-chromatin binding, with a significant fraction of H1 molecules only partially bound in metastable states that can be readily competed against. We anticipate the methods we have developed here will be broadly applicable, particularly for deciphering the binding kinetics of other nuclear proteins that, similar to H1, interact with and modify chromatin.