Degradation, receptor binding, insulin secreting and antihyperglycaemic actions of palmitate-derivatised native and Ala8-substituted GLP-1 analogues.
The hormone glucagon-like peptide-1(7-36)amide (GLP-1) is released in response to ingested nutrients and acts to promote glucose-dependent insulin secretion ensuring efficient postprandial glucose homeostasis. Unfortunately, the beneficial actions of GLP-1 which give this hormone many of the desirable properties of an antidiabetic drug are short lived due to degradation by dipeptidyl-peptidase IV (DPP IV) and rapid clearance by renal filtration. In this study we have attempted to extend GLP-1 action through the attachment of palmitoyl moieties to the epsilon-amino group in the side chain of the Lys26 residue and to combine this modification with substitutions of the Ala8 residue, namely Val or amino-butyric acid (Abu). In contrast to native GLP-1, which was rapidly degraded, [Lys(pal)26]GLP-1, [Abu8, Lys(pal)26]GLP-1 and [Val8 Lys(pal)26]GLP-1 all exhibited profound stability during 12 h incubations with DPP IV and human plasma. Receptor binding affinity and the ability to increase cyclic AMP in the clonal beta-cell line BRIN-BD11 were decreased by 86- to 167-fold and 15- to 62-fold, respectively compared with native GLP-1. However, insulin secretory potency tested using BRIN-BD11 cells was similar, or in the case of [Val8,Lys(pal)26]GLP-1 enhanced. Furthermore, when administered in vivo together with glucose to diabetic (ob/ob) mice, [Lys(pal)26]GLP-1, [Abu8,Lys(pal)26]GLP-1 and [Val8,Lys(pal)26]GLP-1 did not demonstrate acute glucose-lowering or insulinotropic activity as observed with native GLP-1. These studies support the potential usefulness of fatty acid linked analogues of GLP-1 but indicate the importance of chain length for peptide kinetics and bioavailability.


A daily process examination of the bidirectional relationship between craving and alcohol consumption as measured via interactive voice response.
Craving is a central component to alcohol use disorders, although there are contradictory findings in the literature regarding the importance of craving in alcohol use. The study goal was to examine the bidirectional relationship between craving and alcohol consumption in heavy drinkers. Participants received brief alcohol interventions from their primary care physicians and then were asked to make daily reports of craving and alcohol consumption to an interactive voice response (IVR) telephone system for 180 days. The study sample included 246 participants (166 men) with mean age of 46. Ninety-seven percent were Caucasian and 66% met criteria for alcohol dependence. Analysis used generalized estimating equations (GEE) to evaluate whether craving intensity predicted next day alcohol consumption and whether alcohol consumption predicted next day craving intensity. Significant interactions with gender led to stratified analyses. GEE analyses revealed a significant bidirectional relationship between craving and drinking, where craving intensity predicted next day total drinks consumed (p = 0.001), and total drinks predicted next day craving intensity (p = 0.02). Exploratory analysis found that gender significantly moderated the craving-drinking relationship (p = 0.002) with men increasing next day alcohol use more (b = 0.19) than women (b = 0.08). Findings suggest a bidirectional relationship between craving and drinking may contribute to the development or maintenance of heavy drinking, particularly for men. Based on our findings, we recommend that during brief interventions, physicians address both drinking and craving and provide advice for coping with craving.


Gonadotropin-releasing hormone agonist-induced differences in granulosa cell cycle kinetics are associated with alterations in follicular fluid müllerian-inhibiting substance and androgen content.
We have previously shown that the proliferative index (PI), as determined by flow cytometry of luteinized granulosa cells obtained at oocyte retrieval, is greater in ovulation induction regimens which include the GnRH analog (GnRH-a) leuprolide acetate than those using human menopausal gonadotropin (hMG) only. Specific growth factors or intrafollicular hormones may contribute to this leuprolide acetate-induced difference in cell cycle kinetics. We examined whether differences in the PI of these granulosa cells are associated with the alterations of follicular fluid content of Mullerian-inhibiting substance (MIS) and other intrafollicular hormones including FSH, estradiol, progesterone, androstenedione, and testosterone. The control group consisted of follicular fluid obtained from 18 follicles from 4 women receiving hMG alone. The GnRH-a treated group consisted of follicular fluids obtained from 55 follicles aspirated from 18 women receiving GnRH-a in addition to hMG. One-way analysis of variance using log-transformed data and expressed as geometric means with 95% confidence intervals, demonstrated that the follicles from the control group had a significant 14-fold higher concentration of 2.46 ng/mL MIS, 95% CI (1.8-4.8) vs. 0.18 ng/mL, 95% CI (0.13-0.24) P < 0.0005, a 3-fold higher concentration of 17.55 nmol/L androstenedione, 95% CI (14.6-20.9) vs. 5.76 nmol/L, 95% CI (3.1-10.5) P < 0.02, and a 1.5-fold higher concentration of 29.43 nmol/L testosterone 95% CI (22.5-38.14) vs. 19.3 nmol/L, 95% of CI (11.1-33.9) P < 0.01 than GnRH-a treated follicles, although the PI value in controls was half that of the GnRH-a group. These data demonstrate that GnRH-a induced differences in granulosa cell cycle kinetics are associated with alterations of MIS and androgen intrafollicular fluid content and suggest that MIS may be a mitotic inhibitor of human granulosa cells.


Extracorporeal membrane oxygenation during bronchopulmonary lavage.
Extracorporeal membrane oxygenation (ECMO) in a venoarterial perfusion circuit was used to provide support of gas exchange during bronchopulmonary lavage in a 32-year-old man with pulmonary alveolar proteinosis and severe arterial hypoxemia. Prior to the lavage, Pao2 during mechanical ventilation with 100% oxygen and positive end-expiratory pressure was only 125 mm Hg. Extracorporeal perfusion at a flow rate of 3 liters/min, with oxygen delivery of 244 ml/min, increased the Pao2 to 227 mmHg and lowered the mean pulmonary artery pressure from 28 to 24 mm Hg. During bronchopulmonary lavage and ECMO, the Pao2 ranged between 46 and 96 mm Hg. After the procedure, pulmonary performance decidely improved. By reducing the chances of fatal hypoxemia, ECMO allowed treatment to be instituted for this potentially reversible disorder and proved helpful as a form of support during the management of pulmonary alveolar proteinosis when severe hypoxemia may have other wise precluded bronchopulmonary lavage.


Paternal origin of FGFR2 mutations in sporadic cases of Crouzon syndrome and Pfeiffer syndrome.
Crouzon syndrome and Pfeiffer syndrome are both autosomal dominant craniosynostotic disorders that can be caused by mutations in the fibroblast growth factor receptor 2 (FGFR2) gene. To determine the parental origin of these FGFR2 mutations, the amplification refractory mutation system (ARMS) was used. ARMS PCR primers were developed to recognize polymorphisms that could distinguish maternal and paternal alleles. A total of 4,374 bases between introns IIIa and 11 of the FGFR2 gene were sequenced and were assayed by heteroduplex analysis, to identify polymorphisms. Two polymorphisms (1333TA/TATA and 2710 C/T) were found and were used with two previously described polymorphisms, to screen a total of 41 families. Twenty-two of these families were shown to be informative (11 for Crouzon syndrome and 11 for Pfeiffer syndrome). Eleven different mutations in the 22 families were detected by either restriction digest or allele-specific oligonucleotide hybridization of ARMS PCR products. We molecularly proved the origin of these different mutations to be paternal for all informative cases analyzed (P=2. 4x10-7; 95% confidence limits 87%-100%). Advanced paternal age was noted for the fathers of patients with Crouzon syndrome or Pfeiffer syndrome, compared with the fathers of control individuals (34. 50+/-7.65 years vs. 30.45+/-1.28 years, P<.01). Our data on advanced paternal age corroborates and extends previous clinical evidence based on statistical analyses as well as additional reports of advanced paternal age associated with paternal origin of three sporadic mutations causing Apert syndrome (FGFR2) and achondroplasia (FGFR3). Our results suggest that older men either have accumulated or are more susceptible to a variety of germline mutations.


A Revised Classification Scheme for Genetically Diverse Populations of Heterodera glycines.
Heterodera glycines, the soybean cyst nematode, is a major yield-limiting pathogen in most soybean production areas worldwide. Field populations of H. glycines exhibit diversity in their ability to develop on resistant soybean cultivars. Since 1970, this diversity has been characterized by a bioassay used to assign a race classification to a population. The value of the race scheme is reflected in the number and quality of resistant soybean cultivars that have been developed and released by soybean breeders and nematologists working in concert. However, the race scheme also has been misapplied as a means of studying H. glycines genotypes, in part due to the use of the term "race." For fungal and bacterial pathogen species, "race" can theoretically be applied to individuals of a population, thus allowing inference of individual genotypes. Application of a race designation to an individual egg or second-stage juvenile (J2) of H. glycines is not possible because a single J2 cannot be tested on multiple hosts. For other nematode species, "race" is defined by host ranges involving different plant species, whereas the H. glycines race test involves a set of lines of the same plant species. Nonetheless, because H. glycines populations vary in genetic diversity, and this variation has implications for management strategies, a mechanism is needed for documenting and discussing population differences. The HG Type scheme described herein avoids the implication of genetic uniformity or predictability in contrast to the way the race scheme has been used.


Probiotic Lactobacillus rhamnosus GG enhances gastric ulcer healing in rats.
Probiotics are widely used as functional foods which have been advocated for the maintenance of gastrointestinal microflora equilibrium and treatment of gastrointestinal disorders. However, studying the role of probiotics in peptic ulcer disease is limited. The aim of the present study is to investigate the effect of a probiotic strain Lactobacillus rhamnosus GG on gastric ulcer and to elucidate the mechanisms involved. Gastric kissing ulcers were induced in rats by acetic acid (60% v/v). L. rhamnosus GG was given intragastrically at 10(8) cfu/day or 10(9) cfu/day for three consecutive days after ulcer induction. L. rhamnosus GG successfully colonized in the gastric mucosa especially at the ulcer margin. It also significantly and dose-dependently reduced gastric ulcer area. Cell apoptosis to cell proliferation ratio was strongly decreased and accompanied by significant up-regulation of ornithine decarboxylase (ODC) and B-cell lymphoma 2 (Bcl-2) protein expression at the ulcer margin. Angiogenesis was also significantly stimulated together with the induction of vascular endothelial growth factor (VEGF) expression. Furthermore, L. rhamnosus GG up-regulated the phosphorylation level of epidermal growth factor receptor (EGF receptor) without altering the total EGF receptor expression. These findings suggested that L. rhamnosus GG enhanced gastric ulcer healing via the attenuation of cell apoptosis to cell proliferation ratio and increase in angiogenesis. Regulators of these processes such as ODC, Bcl-2, VEGF and EGF receptor are likely to be involved in the healing action of L. rhamnosus GG for gastric ulcer.


Investigation of oncogenic cooperation in simple liver-specific transgenic mouse models using noninvasive in vivo imaging.
Liver cancer is a complex multistep process requiring genetic alterations in multiple proto-oncogenes and tumor suppressor genes. Although hundreds of genes are known to play roles in hepatocarcinogenesis, oncogenic collaboration among these genes is still largely unknown. Here, we report a simple methodology by which oncogenic cooperation between cancer-related genes can be efficiently investigated in the liver. We developed various non-germline transgenic mouse models using hydrodynamics-based transfection which express HrasG12V, SmoM2, and a short-hairpin RNA down-regulating p53 (shp53) individually or in combination in the liver. In this transgenic system, firefly luciferase was co-expressed with the oncogenes as a reporter, allowing tumor growth in the liver to be monitored over time without an invasive procedure. Very strong bioluminescence imaging (BLI) signals were observed at 4 weeks post-hydrodynamic injection (PHI) in mice co-expressing HrasG12V and shp53, while only background signals were detected in other double or single transgenic groups until 30 weeks PHI. Consistent with the BLI data, tumors were observed in the HrasG12V plus shp53 group at 4 weeks PHI, while other transgenic groups failed to exhibit a hyperplastic nodule at 30 weeks PHI. In the HrasG12V plus shp53 transgenic group, BLI signals were well-correlated with actual tumor growth in the liver, confirming the versatility of BLI-based monitoring of tumor growth in this organ. The methodology described here is expected to accelerate and facilitate in vivo studies of the hepatocarcinogenic potential of cancer-related genes by means of oncogenic cooperation.


The reproducibility of colour Doppler in chronic cerebrospinal venous insufficiency associated with multiple sclerosis.
Chronic cerebrospinal venous insufficiency (CCSVI) is a syndrome described in multiple sclerosis (MS) patients, characterized by stenosis of the main extracranial veins with hampered cerebral venous outflow. In the original description echo-colour Doppler demonstrated to be an ideal non invasive tool for screening CCSVI patients, but the reproducibility was not assessed. Aim of this study is to assess the variability coefficient between trained and in not trained echo-colour Doppler operators. Thirty-six (36) subjects, matched for age and gender, were subset in 3 groups (group A, 12 healthy controls, HC; group B, 12 multiple sclerosis patients, MS; group C, 12 patients with other neurological disease, OND) underwent echo-colour Doppler screening for CCSVI according to an original protocol previously described. The inter observer variability rate was assessed by comparing respectively trained vs not trained operators, and trained vs trained operators, by using the same echo-colour Doppler equipment. In addition, by scanning 15 subjects after one month from the first session, intra observer coefficient was also assessed in trained operator. The inter observer variability rate between trained and not trained echo-colour Doppler operators, were not completely satisfactory (K coefficient 0.47 95% CI 0.27-0.68). To the contrary the inter observer agreement between trained operators was much more reliable (K coefficient 0.80 95% CI 0.59-1.01). Finally, the intra observer variability rate in trained operators was 0.93, (95% CI 0.80-1.06) confirming a highly satisfactory agreement. Echo-colour Doppler is a powerful, non-invasive and reproducible tool for screening CCSVI-MS but it needs special training.


Gambling experiences, problems, research and policy: gambling in Germany.
The objective of this paper is to present an overview of gambling in Germany, including historical development, legislative and economic changes as well as treatment options and their effectiveness. The available scientific literature and research reports on gambling in Germany were reviewed to obtain relevant information on history, commercialization, legislation, treatment and research agenda. Gambling in Germany is characterized by compromises between protective and economic efforts. At present, gambling is illegal in Germany, and provision is subject to the state monopoly. Mere gaming machines (specific slot machines) are not classified as gambling activity, permitting commercial providers. In recent years, implementing regulations for state gambling and gaming machines have been changed. Concerning the treatment of pathological gambling, various options exist; treatment costs have been covered by health and pension insurance since 2001. Information on the effectiveness of treatment in Germany is limited. Similarly, the number of peer-reviewed publications on gambling is small. German gambling legislation was subject to major changes in the past years. Based on the available body of research (longitudinal), studies on risk and protective factors and the aetiology of pathological gambling are needed. The effectiveness of pathological gambling treatment in Germany and the impact of gambling regulations on gambling behaviour also need to be investigated.


Spermless males elicit large-scale female responses to mating in the malaria mosquito Anopheles gambiae.
Anopheles gambiae sensu stricto is the major vector of malaria, a disease with devastating consequences for human health. Given the constant spread of the disease, alternative approaches to the use of insecticides are urgently needed to control vector populations. Females of this species undergo large behavioral changes after mating, which include a life-long refractoriness to further insemination and the induction of egg laying in blood-fed individuals. Genetic control strategies aimed at impacting Anopheles fertility through the release of sterile males are being advocated to reduce the size of mosquito field populations. Such strategies depend on the ability of the released sterile males to mate successfully with wild females and to switch off the female receptivity to further copulation. Here we evaluate the role of sperm in regulating female behavioral responses after mating in An. gambiae. We developed spermless males by RNAi silencing of a germ cell differentiation gene. These males mated successfully and preserved standard accessory gland functions. Females mated to spermless males exhibited normal postcopulatory responses, which included laying large numbers of eggs upon blood feeding and becoming refractory to subsequent insemination. Moreover, spermless males induced transcriptional changes in female reproductive genes comparable to those elicited by fertile males. Our data demonstrate that, in contrast to Drosophila, targeting sperm in An. gambiae preserves normal male and female reproductive behavior for the traits and time frame analyzed and validate the use of approaches based on incapacitation or elimination of sperm for genetic control of vector populations to block malaria transmission.


Latino men and familial risk communication about prostate cancer.
To investigate how familial communication about prostate cancer (PCa) risk and screening affects sons of men with PCa. Qualitative grounded theory. Southern California. 17 Latino sons of PCa survivors. The team conducted semistructured interviews and follow-up interviews. Therefore, the sample includes 25 transcripts. Data were analyzed with a mix of a priori topical codes and grounded theory techniques. Sons were in need of information about familial risk and screening options. They became sensitized to PCa, desired information, and held protective intentions. Hopeful intentions came up against cultural taboos around sex, reproductive health, and intimacy that limited discussions between fathers and sons. Fathers were a valued source of information but play various roles, which affect sons' screening intentions. Open communication between father and son promoted awareness of screening and familial risk. Uncertainty about familial risk and screening options, especially early detection strategies, was exacerbated by cultural taboos around PCa. Fathers could have been primary and credible advocates for shared decision making, but sons had difficulty learning from their fathers' experience. FINDINGS from the study can help inform community-based interventions with Latino families, help to culturally tailor health messaging, and sensitize clinicians to a group that needs concerted counseling about PCa risk and screening.


High-resolution matrix-assisted laser desorption ionization-imaging mass spectrometry of lipids in rodent optic nerve tissue.
To develop a method for generating high spatial resolution (10 µm) matrix-assisted laser desorption ionization (MALDI) images of lipids in rodent optic nerve tissue. Ice-embedded optic nerve tissue from rats and mice were cryosectioned across the coronal and sagittal axes of the nerve fiber. Sections were thaw mounted on gold-coated MALDI plates and were washed with ammonium acetate to remove biologic salts before being coated in 2,5-dihydroxybenzoic acid by sublimation. MALDI images were generated in positive and negative ion modes at 10 µm spatial resolution. Lipid identification was performed with a high mass resolution Fourier transform ion cyclotron resonance mass spectrometer. Several lipid species were observed with high signal intensity in MALDI images of optic nerve tissue. Several lipids were localized to specific structures including in the meninges surrounding the optic nerve and in the central neuronal tissue. Specifically, phosphatidylcholine species were observed throughout the nerve tissue in positive ion mode while sulfatide species were observed in high abundance in the meninges surrounding the optic nerve in negative ion mode. Accurate mass measurements and fragmentation using sustained off-resonance irradiation with a high mass resolution Fourier transform ion cyclotron resonance mass spectrometer instrument allowed for identification of lipid species present in the small structure of the optic nerve directly from tissue sections. An optimized sample preparation method provides excellent sensitivity for lipid species present within optic nerve tissue. This allowed the laser spot size and fluence to be reduced to obtain a high spatial resolution of 10 µm. This new imaging modality can now be applied to determine spatial and molecular changes in optic nerve tissue with disease.


Metabolomic Response to Acute Hypoxic Exercise and Recovery in Adult Males.
Metabolomics is a relatively new "omics" approach used to characterize metabolites in a biological system at baseline and following a diversity of stimuli. However, the metabolomic response to exercise in hypoxia currently remains unknown. To examine this, 24 male participants completed 1 h of exercise at a workload corresponding to 75% of pre-determined O2max in hypoxia (Fio2 = 0.16%), and repeated in normoxia (Fio2 = 0.21%), while pre- and post-exercise and 3 h post-exercise metabolites were analyzed using a LC ESI-qTOF-MS untargeted metabolomics approach in serum samples. Exercise in hypoxia and in normoxia independently increased metabolism as shown by a change in a combination of twenty-two metabolites associated with lipid metabolism (p < 0.05, pre vs. post-exercise), though hypoxia per se did not induce a greater metabolic change when compared with normoxia (p > 0.05). Recovery from exercise in hypoxia independently decreased seventeen metabolites associated with lipid metabolism (p < 0.05, post vs. 3 h post-exercise), compared with twenty-two metabolites in normoxia (p < 0.05, post vs. 3 h post-exercise). Twenty-six metabolites were identified as responders to exercise and recovery (pooled hypoxia and normoxia pre vs. recovery, p < 0.05), including metabolites associated with purine metabolism (adenine, adenosine and hypoxanthine), the amino acid phenylalanine, and several acylcarnitine molecules. Our novel data provides preliminary evidence of subtle metabolic differences to exercise and recovery in hypoxia and normoxia. Specifically, exercise in hypoxia activates metabolic pathways aligned to purine and lipid metabolism, but this effect is not selectively different from exercise in normoxia. We also show that exercise per se can activate pathways associated with lipid, protein and purine nucleotide metabolism.


Integration of over 9,000 mass spectrometry experiments builds a global map of human protein complexes.
Macromolecular protein complexes carry out many of the essential functions of cells, and many genetic diseases arise from disrupting the functions of such complexes. Currently, there is great interest in defining the complete set of human protein complexes, but recent published maps lack comprehensive coverage. Here, through the synthesis of over 9,000 published mass spectrometry experiments, we present hu.MAP, the most comprehensive and accurate human protein complex map to date, containing > 4,600 total complexes, > 7,700 proteins, and > 56,000 unique interactions, including thousands of confident protein interactions not identified by the original publications. hu.MAP accurately recapitulates known complexes withheld from the learning procedure, which was optimized with the aid of a new quantitative metric (k-cliques) for comparing sets of sets. The vast majority of complexes in our map are significantly enriched with literature annotations, and the map overall shows improved coverage of many disease-associated proteins, as we describe in detail for ciliopathies. Using hu.MAP, we predicted and experimentally validated candidate ciliopathy disease genes in vivo in a model vertebrate, discovering CCDC138, WDR90, and KIAA1328 to be new cilia basal body/centriolar satellite proteins, and identifying ANKRD55 as a novel member of the intraflagellar transport machinery. By offering significant improvements to the accuracy and coverage of human protein complexes, hu.MAP (http://proteincomplexes.org) serves as a valuable resource for better understanding the core cellular functions of human proteins and helping to determine mechanistic foundations of human disease.


Proteasomal interference prevents zona pellucida penetration and fertilization in mammals.
The ubiquitin-proteasome pathway has been implicated in the penetration of ascidian vitelline envelope by the fertilizing spermatozoon (Sawada et al., Proc Natl Acad Sci U S A 2002; 99:1223-1228). The present study provides experimental evidence demonstrating proteasome involvement in the penetration of mammalian zona pellucida (ZP). Using porcine in vitro fertilization as a model, penetration of ZP was completely inhibited by specific proteasomal inhibitors MG-132 and lactacystin. Three commercial rabbit sera recognizing 20S proteasomal core subunits beta-1i, beta-2i, alpha-6, and beta-5 completely blocked fertilization at a very low concentration (i.e., diluted 1/2000 to 1/8000 in fertilization medium). Neither proteasome inhibitors nor antibodies had any effects on sperm-ZP binding and acrosome exocytosis in zona-enclosed oocytes or on fertilization rates in zona-free oocytes, which were highly polyspermic. Consistent with a possible role of ubiquitin-proteasome pathway in ZP penetration, ubiquitin and various alpha and beta type proteasomal subunits were detected in boar sperm acrosome by specific antibodies, immunoprecipitated and microsequenced by MALDI-TOF from boar sperm extracts. Antiubiquitin-immunoreactive substrates were detected on the outer face of ZP by epifluorescence microscopy. This study therefore provides strong evidence implicating the ubiquitin-proteasome pathway in mammalian fertilization and zona penetration. This finding opens a new line of acrosome/ZP research because further studies of the sperm acrosomal proteasome can provide new tools for the management of polyspermia during in vitro fertilization and identify new targets for contraceptive development.


Integration is required for productive infection of monocyte-derived macrophages by human immunodeficiency virus type 1.
Certain human immunodeficiency virus type 1 (HIV-1) isolates are able to productively infect nondividing cells of the monocyte/macrophage lineage. We have used a molecular genetic approach to construct two different HIV-1 integrase mutants that were studied in the context of an infectious, macrophage-tropic HIV-1 molecular clone. One mutant, HIV-1 delta D(35)E, containing a 37-residue deletion within the central, catalytic domain of integrase, was noninfectious in both peripheral blood mononuclear cells and monocyte-derived macrophages. The HIV-1 delta D(35)E mutant, however, exhibited defects in the assembly and/or release of progeny virions in transient transfection assays, as well as defects in entry and/or viral DNA synthesis during the early stages of monocyte-derived macrophage infection. The second mutant, HIV-1D116N/8, containing a single Asp-to-Asn substitution at the invariant Asp-116 residue of integrase, was also noninfectious in both peripheral blood mononuclear cells and monocyte-derived macrophages but, in contrast to HIV-1 delta D(35)E, was indistinguishable from wild-type virus in reverse transcriptase production. PCR analysis indicated that HIV-1D116N/8 entered monocyte-derived macrophages efficiently and reverse transcribed its RNA but was unable to complete its replication cycle because of a presumed block to integration. These data are consistent with the hypothesis that integration is an obligate step in productive HIV-1 infection of activated peripheral blood mononuclear cells and primary human macrophage cultures.


Three-dimensional, in vivo MRI with self-gating and image coregistration in the mouse.
Motion during magnetic resonance imaging (MRI) scans routinely results in undesirable image artifact or blurring. Since high-resolution, three-dimensional (3D) imaging of the mouse requires long scan times for satisfactory signal-to-noise ratio (SNR) and image quality, motion-related artifacts are likely over much of the body and limit applications of mouse MRI. In this investigation, we explored the use of self-gated imaging methods and image coregistration for improving image quality in the presence of motion. Self-gated signal results from a modified 3D gradient-echo sequence showed detection of periodic respiratory and cardiac motion in the adult mouse-with excellent comparison to traditional measurements, sensitivity to respiration-induced tissue changes in the brain, and even detection of embryonic cardiac motion in utero. Serial image coregistration with rapidly-acquired, low-SNR volumes further enabled detection and correction of bulk changes in embryo location during in utero imaging sessions and subsequent reconstruction of high-quality images. These methods, in combination, are shown to expand the range of applications for 3D mouse MRI, enabling late-stage embryonic heart imaging and introducing the possibility of longitudinal developmental studies from embryonic stages through adulthood.


Conditional reimbursement within the Dutch drug policy.
In The Netherlands, conditional reimbursement is considered to be a promising approach to achieving more effective and efficient pharmaceutical care. Because of its formal status and nationwide regulation, conditional reimbursement may allow governments to better control medical decision-making. To evaluate the effects of conditional reimbursement on medicine use and its performance as a policy tool, we compared observed volumes of medicine use with expected volumes. In addition, we mapped the annual growth by analysing trends in the volumes of use of all conditionally reimbursed drugs; starting with the year the drug entered the market (using macro-level data). Next we explored five cases in depth (using micro-level data) in order to explore what fraction of individual prescriptions met the requirements. We also performed qualitative research (document analysis, interviews (N=65)) in order to obtain the stakeholders' perspectives on how the measure functions, as well as to interpret the case studies data further. The findings suggest that conditional reimbursement may be an effective policy instrument, but that several changes are needed to optimize its impact. These changes are predominantly related to transparency (e.g. conditions are set following clear procedures and criteria), legitimacy (conditions should be consistent with criteria for prioritization), feasibility of procedures to control appropriate use, and timely and appropriate commitment of the stakeholders.


Safety of the APOLLO Onyx delivery microcatheter for embolization of brain arteriovenous malformations: results from a prospective post-market study.
Catheter retention and difficulty in retrieval have been observed during embolization of brain arteriovenous malformations (bAVMs) with the Onyx liquid embolic system (Onyx). The Apollo Onyx delivery microcatheter (Apollo) is a single lumen catheter designed for controlled delivery of Onyx into the neurovasculature, with a detachable distal tip to aid catheter retrieval. This study evaluates the safety of the Apollo for delivery of Onyx during embolization of bAVMs. This was a prospective, non-randomized, single-arm, multicenter, post-market study of patients with a bAVM who underwent Onyx embolization with the Apollo between May 2015 and February 2018. The primary endpoint was any catheter-related adverse event (AE) at 30 days, such as unintentional tip detachment or malfunction with clinical sequelae, or retained catheter. Procedure-related AEs (untoward medical occurrence, disease, injury, or clinical signs) and serious AEs (life threatening illness or injury, permanent physiological impairment, hospitalization, or requiring intervention) were also recorded. A total of 112 patients were enrolled (mean age 44.1±17.6 years, 56.3% men), and 201 Apollo devices were used in 142 embolization procedures. The mean Spetzler-Martin grade was 2.38. The primary endpoint was not observed (0/112, 0%). The catheter tip detached during 83 (58.5%) procedures, of which 2 (2.4%) were unintentional and did not result in clinical sequelae. At 30 days, procedure related AEs occurred in 26 (23.2%) patients, and procedure-related serious AEs in 12 (10.7%). At 12 months, there were 3 (2.7%) mortalities, including 2 (1.8%) neurological deaths, none of which were device-related. This study demonstrates the safety of Apollo for Onyx embolization of bAVMs. CNCT02378883.


Global gene expression profiling of pleural mesotheliomas: overexpression of aurora kinases and P16/CDKN2A deletion as prognostic factors and critical evaluation of microarray-based prognostic prediction.
Most gene expression profiling studies of mesothelioma have been based on relatively small sample numbers, limiting their statistical power. We did Affymetrix U133A microarray analysis on 99 pleural mesotheliomas, in which multivariate analysis showed advanced-stage, sarcomatous histology and P16/CDKN2A homozygous deletion to be significant independent adverse prognostic factors. Comparison of the expression profiles of epithelioid versus sarcomatous mesotheliomas identified many genes significantly overexpressed among the former, including previously unrecognized ones, such as uroplakins and kallikrein 11, both confirmed by immunohistochemistry. Examination of the gene expression correlates of survival showed that more aggressive mesotheliomas expressed higher levels of Aurora kinases A and B and functionally related genes involved in mitosis and cell cycle control. Independent confirmation of the negative effect of Aurora kinase B was obtained by immunohistochemistry in a separate patient cohort. A role for Aurora kinases in the aggressive behavior of mesotheliomas is of potential clinical interest because of the recent development of small-molecule inhibitors. We then used our data to develop microarray-based predictors of 1 year survival; these achieved a maximal accuracy of 68% in cross-validation. However, this was inferior to prognostic prediction based on standard clinicopathologic variables and P16/CDNK2A status (accuracy, 73%), and adding the microarray model to the latter did not improve overall accuracy. Finally, we evaluated three recently published microarray-based outcome prediction models, but their accuracies ranged from 63% to 67%, consistently lower than reported. Gene expression profiling of mesotheliomas is an important discovery tool, but its power in clinical prognostication has been overestimated.


All-guided stellar interferometer with an integrated optics recombiner.
We report laboratory tests of an all-guided stellar interferometer used for optical aperture synthesis. In anticipation of use of the interferometer in space missions, this research is focused especially on compactness of the recombining device. The coherent transport and delay lines are implemented with polarization-maintaining fiber. Beam recombination is achieved by means of an integrated optics component. This two-arm interferometer operates at 670-nm mean wavelength and allows for a 24-cm correction for the differential air path.


Rapid versus conventional initiation of quetiapine in the treatment of schizophrenia: a randomized, parallel-group trial.
The primary objective of this study was to compare the safety and tolerability of a rapid initiation of quetiapine with the conventional initiation approved by the U.S. Food and Drug Administration (FDA). The secondary objectives included assessment of the efficacy of a rapid initiation of quetiapine compared with a conventional initiation approved by the FDA. Patients with acute schizophrenia were randomly assigned in a 3:1 ratio to the rapid-initiation group (200 mg on day 1, 400 mg on day 2, 600 mg on day 3, and 800 mg on day 4) or to the conventional-initiation group (50 mg on day 1, 100 mg on day 2, and increased in 100 mg/day increments to reach 400 mg on day 5). The tolerability measures were Barnes Akathisia Scale (BAS) and Simpson-Angus Scale (SAS) as well as all adverse events at day 1, 2, 3, 4, 5, 6, and 7 and at day 14. Standard efficacy measures were administered at baseline, day 1, day 4, day 5, day 7, and day 14. These measures consisted of the Positive and Negative Syndrome Scale (PANSS), PANSS-Excited Component (EC), and Clinical Global Impressions-Severity of Illness (CGI-S) scale. Forty patients were randomly assigned to treatment. The mean (SD) dose of quetiapine at study end point was 763.3 (106.6) and 600.0 (249.4) mg/day in the rapid-initiation group and conventional-initiation group, respectively. The most common side effects were sedation and dizziness, with no significant differences in frequency between groups. Only 2/30 patients from the rapid-initiation group discontinued treatment due to an adverse event (both for sedation), and 1/10 patients from the conventional-initiation group discontinued before receiving quetiapine. Neither serious adverse events nor differences between groups in vital signs, laboratory assessments, ECG measures, or weight changes were reported. Rapid initiation of quetiapine was generally well-tolerated and was associated with a faster onset of action than conventional initiation as measured by improvement in psychotic symptoms at days 4 and 5. This study may offer preliminary evidence for tolerability and effectiveness in rapid dose initiation of quetiapine in the treatment of schizophrenia.


Do clinical guidelines reduce clinician dependent costs?
Clinician dependent costs are the costs of care that are under the discretion of the healthcare provider. These costs include the costs of drugs, tests and investigations, and discretionary outpatient visits and impatient stays. The purpose of this review was to summarize recent evidence, relevant to both developed and developing countries on whether evidence based clinical guidelines can change hospitals variable costs which are clinician dependent, and the degree of financial savings achieved at hospital level. Potential studies for inclusion were identified using structured searches of Econlit, J-Stor, and Pubmed databases. Two reviewers independently evaluated retrieved studies for inclusion. The methodological quality of the selected articles was assessed using the Oxford Centre for Evidence- Based Medicine (CEBM) levels of evidence. The results suggest that 10 of the 11 interventions were successful reducing financial costs. Most of the interventions, either in modeling studies or real interventions generate significant financial saving, although the former reported higher savings because the studies assumed 100 percent compliance.


Genome sequencing of strain Cellulosimicrobium sp. TH-20 with ginseng biotransformation ability.
Biotransformation for increasing the pharmaceutical effect of ginsenosides is getting more and more attractions. Strain Cellulosimicrobium sp. TH-20 isolated from ginseng soil samples was identified to produce enzymes contributing to its excellent biotransformation activity against ginsenosides, the main active components of ginseng. Based on phylogenetic tree and homology analysis, the strain can be designated as Cellulosimicrobium sp. Genome sequencing was performed using the Illumina Miseq to explore the functional genes involved in ginsenoside transformation. The draft genome of Cellulosimicrobium sp. TH-20 encoded 3450 open reading frames, 51 tRNA, and 9 rRNA. All ORFs were annotated using NCBI BLAST with non-redundant proteins, Gene Ontology, Cluster of Orthologous Gene, and Kyoto Encyclopedia of Genes and Genomes databases. A total of 11 genes were selected based on the functional annotation analysis. These genes are relevant to ginsenoside biotransformation, including 6 for beta-glucosidase, 1 for alpha-N-arabinofuranosidase, 1 for alpha-1,6-glucosidase, 1 for endo-1,4-beta-xylanase, 1 for alpha-L-arabinofuranosidase, and 1 for beta-galactosidase. These glycosidases were predicted to catalyze the hydrolysis of sugar moieties attached to the aglycon of ginsenosides and led to the transformation of PPD-type and PPT-type ginsenosides.


Optical control of field-emission sites by femtosecond laser pulses.
We have investigated field-emission patterns from a clean tungsten tip apex induced by femtosecond laser pulses. Strongly asymmetric modulations of the field-emission intensity distributions are observed depending on the polarization of the light and the laser incidence direction relative to the azimuthal orientation of tip apex. In effect, we have realized an ultrafast pulsed field-emission source with site selectivity. Simulations of local fields on the tip apex and of electron emission patterns based on photoexcited nonequilibrium electron distributions explain our observations quantitatively.


Advancing Drug Development in Gynecologic Malignancies.
Gynecologic malignancies continue to be a major cause of morbidity and mortality in the United States despite recent advances in oncologic therapies. To realize the promise of immunotherapy and biomarker-driven approaches to improve clinical outcomes for patients, better communication among stakeholders in the drug development and approval pathways is needed. To this end, the FDA-AACR-SGO Drug Development in Gynecologic Malignancies Workshop brought together clinicians, patient advocates, researchers, industry representatives, and regulators in June 2018, to review the state of the science in gynecologic cancers and explore how scientific advances impact approval processes. Topics of discussion and key takeaways are summarized in this Perspectives in Regulatory Science and Policy article. Single-agent immunotherapies have demonstrated variable and often modest response rates among gynecologic cancers. Combination therapies and other novel approaches, such as cell-based therapies, may show improved efficacy compared with single-agent immunotherapies; however, utilizing innovative clinical trial designs will be necessary to progress further. Companion and complementary diagnostics inform physicians of potential benefits of specific therapeutics for patients; however, they serve different functions that have important regulatory implications, thus trialists should understand the distinctions between diagnostic types. PARP inhibitors hold great promise for treating ovarian cancers, both as monotherapies and in combination with chemotherapeutics, other targeted agents, and immunotherapies. Rare gynecologic cancers often exhibit unique molecular characteristics that can serve as effective targets to which novel therapeutics can be developed. This workshop highlighted the importance of future open discussions on scientific and regulatory challenges in drug development for gynecologic malignancies.


Spectrofluorometric measurement of the binding of ethidium to superhelical DNA from cell nuclei.
Structures retaining many of the morphological features of nuclei may be released by lysing HeLa cells in solutions containing non-ionic detergents and high concentrations of salt. These nucleoids contain few chromatin proteins. We have shown that the DNA of nucleoids is quasicircular and supercoiled by measure spectrofluorometrically the amount of the intercalating dye, ethidium, bound to unirradiated and gamma-irradiated nucleoids. Ethidium binds to nucleoids in the manner characteristic of the binding to superhelical DNA: at low concentrations more ethidium binds to unirradiated nucleoids than to their gamma-irradiated counterparts with broken DNA, and at higher concentrations less ethidium binds to the unirradiated nucleoids. The quasi-circles in nucleoids are 22 times less sensitive to gamma-irradiation than are circles of pure PM2 DNA: they must contain about 2.2 X 10(5) base pairs. The constraints that maintain the quasi-circularity of nucleoid DNA are very resistant to extremes of temperature and alkali; some remain under conditions in which the duplex is denatured. The constraints are destabilised by ethidium suggesting that they are stabilised by free energy of supercoiling. Proteolytic enzymes, but not ribonucleases, remove the constraints. Possible structures for the constraining mechanism are discussed.


Local variations in the timing of RSV epidemics.
Respiratory syncytial virus (RSV) is a primary cause of hospitalizations in children worldwide. The timing of seasonal RSV epidemics needs to be known in order to administer prophylaxis to high-risk infants at the appropriate time. We used data from the Connecticut State Inpatient Database to identify RSV hospitalizations based on ICD-9 diagnostic codes. Harmonic regression analyses were used to evaluate RSV epidemic timing at the county level and ZIP code levels. Linear regression was used to investigate associations between the socioeconomic status of a locality and RSV epidemic timing. 9,740 hospitalizations coded as RSV occurred among children less than 2 years old between July 1, 1997 and June 30, 2013. The earliest ZIP code had a seasonal RSV epidemic that peaked, on average, 4.64 weeks earlier than the latest ZIP code. Earlier epidemic timing was significantly associated with demographic characteristics (higher population density and larger fraction of the population that was black). Seasonal RSV epidemics in Connecticut occurred earlier in areas that were more urban (higher population density and larger fraction of the population that was). These findings could be used to better time the administration of prophylaxis to high-risk infants.


Car seat test for preterm infants: comparison with polysomnography.
To monitor preterm infants in a cot and a car seat and compare an observed car seat trial with polysomnography (PSG). Non-randomised controlled trial. Regional neonatal unit. Preterm infants before discharge. Nap PSG respiratory and sleep variables were measured including gastro-oesophageal pH. Nurse observations included respiratory distress, apnoea measured by apnoea alarm, oxygen saturation and heart rate. Infants were studied supine in a cot and then in a car seat. Nursing observations were compared with PSG during the car seat trial only. Criteria for failure of the PSG and observed tests were predefined. Difference in respiratory instability between cot and car seat. Concurrence regarding failure of the car seat trial between nurse-observed data and PSG. 20 infants (median gestation 33 weeks (range 28-35 weeks; median postmenstrual age (PMA) at study 36.5 weeks (range 35-38 weeks)) were studied. There were sufficient car seat data on 18 infants for comparison. There were fewer central apnoeas and arousals in the cot than the car seat (p = 0.047 and p = 0.024, respectively). Airway obstruction was not more common in the car seat. Younger PMA at time of study predicted failure in both car seat (p = 0.022) and cot (p = 0.022). The nurse-observed test had low sensitivity for predicting PSG failure but more accurately predicted airway obstruction on PSG. Immature infants exhibit respiratory instability in cots and car seats. A car seat test does not accurately detect all adverse events during sleep in the seat.


Change in Renal Function among HIV-Infected Koreans Receiving Tenofovir Disoproxil Fumarate-Backbone Antiretroviral Therapy: A 3-Year Follow-Up Study.
Tenofovir disoproxil fumarate (TDF) is commonly prescribed as a fixed-dose, co-formulated antiretroviral drug for HIV-1 infection. The major concern of long-term TDF use is renal dysfunction. However, little is known about the long-term patterns of changes in renal function in HIV-infected Koreans receiving TDF. We prospectively followed 50 HIV-infected Koreans, performing laboratory tests every 3 months during the first year and every 6 months for the next 2 years. Urine N-acetyl-β-D-glucosaminidase (NAG) and plasma cystatin-C were measured using samples collected in the first year. Data on renal function were retrospectively collected on HIV-infected patients receiving first-line TDF (n=40) and in antiretroviral therapy (ART)-naïve patients (n=24) for 3 years. Renal function was evaluated as estimated glomerular filtration rate (eGFR) from serum creatinine [Modification of Diet in Renal Disease (MDRD)] and cystatin-C. The eGFR (cystatin-C) showed significant changes from 0 to 48 wks (p=0.002), with the lowest levels at 24 wks (84.3±18.8 mL/min vs. 90.3±22.5 mL/min, p=0.021 by post hoc test). Urine NAG levels did not differ at 0, 12, 24, and 48 wks, although eGFR (MDRD) significantly decreased from 0 (98.7±18.9 mL/min/1.73 m²) to 144 wks (89.0±14.7 mL/min/1.73 m²) (p=0.010). The first-line TDF group had significantly lower eGFR (MDRD) than the ART-naïve group at 144 wks (89.7 mL/min/1.73 m² vs. 98.4 mL/min/1.73 m², p=0.036). Thirteen (26%) participants experienced a decrease in renal impairment of 10 mL/min/1.73 m² in eGFR (MDRD) at 144 wks. These data suggest that clinically meaningful renal injury can develop in HIV-infected Koreans receiving long-term TDF.


Does changing pain-related knowledge reduce pain and improve function through changes in catastrophizing?
Evidence from randomized controlled studies shows that reconceptualizing pain improves patients' knowledge of pain biology, reduces catastrophizing thoughts, and improves pain and function. However, causal relationships between these variables remain untested. It is hypothesized that reductions in catastrophizing could mediate the relationship between improvements in pain knowledge and improvements in pain and function. To test this causal mechanism, we conducted longitudinal mediation analyses on a cohort of 799 patients who were exposed to a pain education intervention. Patients provided responses to the neurophysiology of pain questionnaire, catastrophic thoughts about pain scale, visual analogue pain scale, and the patient specific functional scale, at baseline, 1-month, 6-month, and 12-month follow-up. With adjustment for potential confounding variables, an improvement in pain biology knowledge was significantly associated with a reduction in pain intensity (total effect = -2.20, 95% confidence interval [CI] = -2.96 to -1.44). However, this effect was not mediated by a reduction in catastrophizing (indirect effect = -0.16, 95% CI = -0.36 to 0.02). This might be due to a weak, nonsignificant relationship between changes in catastrophizing and pain intensity (path b = 0.19, 95% CI = -0.03 to 0.41). Similar trends were found in models with function as the outcome. Our findings indicate that change in catastrophizing did not mediate the effect of pain knowledge acquisition on change in pain or function. The strength of this conclusion is moderated, however, if patient-clinician relational factors are conceptualized as a consequence of catastrophizing, rather than a cause.


Frequency of kidney rejection in diabetic patients undergoing simultaneous kidney and pancreatic islet cell transplantation.
An increased frequency of kidney rejection has been reported in diabetic patients who have simultaneous pancreas and kidney transplantation compared with patients who have a kidney transplant alone. Kidney graft outcome is similar in the two groups. The mechanism for increased kidney graft rejection with a simultaneous pancreas graft is not clear. It is ascribed to the immunogenicity of the exocrine pancreas that initiates migration of activated cells from the peripheral blood that are entrapped in the kidney. Since the volume of the transplanted tissue is less in islet transplantation (usually < 2 ml) than in pancreas transplantation, one might not expect an increased frequency of kidney rejection in islet cell recipients. We looked at biopsy-proven kidney rejection episodes in patients who had combined kidney and islet transplants and compared this with the frequency of rejection in diabetic and nondiabetic patients who underwent a kidney transplant alone under the same immunosuppression. Diabetic patients who had kidney islet transplants (n = 9) had a higher frequency of rejection (100%) compared with diabetic patients (n = 107, 55.1%) and nondiabetic patients (n = 327, 65%) who had a kidney transplant alone. The 1-year graft and patient survival rates were not different among the groups. Although the number of patients is small, it would appear that transplantation of a low volume of islet cells with high purity can lead to an increased frequency of kidney rejection. This is unlikely to be explained solely on the basis of fewer antigen matches in these recipients but may reflect the inherent immunogenicity of the purified islet preparations. Alternatively, there may be an effect of their direct infusion into the portal vein.


Neutron stimulated emission computed tomography: a Monte Carlo simulation approach.
A Monte Carlo simulation has been developed for neutron stimulated emission computed tomography (NSECT) using the GEANT4 toolkit. NSECT is a new approach to biomedical imaging that allows spectral analysis of the elements present within the sample. In NSECT, a beam of high-energy neutrons interrogates a sample and the nuclei in the sample are stimulated to an excited state by inelastic scattering of the neutrons. The characteristic gammas emitted by the excited nuclei are captured in a spectrometer to form multi-energy spectra. Currently, a tomographic image is formed using a collimated neutron beam to define the line integral paths for the tomographic projections. These projection data are reconstructed to form a representation of the distribution of individual elements in the sample. To facilitate the development of this technique, a Monte Carlo simulation model has been constructed from the GEANT4 toolkit. This simulation includes modeling of the neutron beam source and collimation, the samples, the neutron interactions within the samples, the emission of characteristic gammas, and the detection of these gammas in a Germanium crystal. In addition, the model allows the absorbed radiation dose to be calculated for internal components of the sample. NSECT presents challenges not typically addressed in Monte Carlo modeling of high-energy physics applications. In order to address issues critical to the clinical development of NSECT, this paper will describe the GEANT4 simulation environment and three separate simulations performed to accomplish three specific aims. First, comparison of a simulation to a tomographic experiment will verify the accuracy of both the gamma energy spectra produced and the positioning of the beam relative to the sample. Second, parametric analysis of simulations performed with different user-defined variables will determine the best way to effectively model low energy neutrons in tissue, which is a concern with the high hydrogen content in biological tissue. Third, determination of the energy absorbed in tissue during neutron interrogation in order to estimate the dose. Results from these three simulation experiments demonstrate that GEANT4 is an effective simulation platform that can be used to facilitate the future development and optimization of NSECT.


Recombinant human epidermal growth factor (rhEGF) protects radiation-induced intestine injury in murine system.
This study was to investigate whether rhEGF protects radiation induced intestine injury without compromising antitumor effect of radiation in murine system. A radiation induced intestinal injury model was established in mice by whole body irradiation. Using this model, 4 groups were set; control, rhEGF (100 µg/kg intraperitoneally), radiation (10 Gy), and a combination (rhEGF and radiation). The level of apoptosis and proliferation were analyzed by TUNEL assay and proliferation cell nuclear antigen (PCNA) immunohistochemical staining, respectively, as well as observation of survival and body weight change. A tumor growth delay assay was performed using murine syngeneic tumors; one radioresistant tumor, HCa-I and one radiosensitive tumor, MCa-K. In the radiation induced intestinal injury model, the 10 Gy group had significantly more weight loss with less number of crypt cells and higher apoptosis than the 8 Gy group. Using 10 Gy model, radioprotective effect of rhEGF was tested. Addition of rhEGF improved not only the body weight loss but also survival following radiation. It also induced suppression of apoptosis as well as increase of PCNA expression and recovery of villi. rhEGF did not enhance the tumor growth after radiation exposure in the tested tumors. These findings suggest that combination of exogenous rhEGF and radiation can be a new anticancer strategy by protecting radiation-induced intestinal injury without alleviating antitumor effect of radiation.


A chorismate mutase from the soybean cyst nematode Heterodera glycines shows polymorphisms that correlate with virulence.
Parasitism genes from phytoparasitic nematodes are thought to be essential for nematode invasion of the host plant, to help the nematode establish feeding sites, and to aid nematodes in the suppression of host plant defenses. One gene that may play several roles in nematode parasitism is chorismate mutase (CM). This secreted enzyme is produced in the nematode's esophageal glands and appears to function within the plant cell to manipulate the plant's shikimate pathway, which controls plant cell growth, development, structure, and pathogen defense. Using degenerate polymerase chain reaction primers, we amplified and cloned a chorismate mutase (Hg-cm-1) from Heterodera glycines, the soybean cyst nematode (SCN), and showed it had CM activity. RNA in situ hybridization of Hg-cm-1 cDNA to SCN sections confirms that it is specifically expressed in the nematodes' esophageal glands. DNA gel blots of genomic DNA isolated from SCN inbred lines that have differing virulence on SCN resistant soybean show Hg-cm-1 is a member of a polymorphic gene family. Some Hg-cm family members predominate in SCN inbred lines that are virulent on certain SCN resistant soybean cultivars. The same polymorphisms and correlation with virulence are seen in the Hg-cm-1 expressed in the SCN second-stage juveniles. Based on the enzymatic activity of Hg-cm-1 and the observation that different forms of the mutase are expressed in virulent nematodes, we hypothesize that the Hg-cm-1 is a virulence gene, some forms of which allow SCN to parasitize certain resistant soybean plants.


The effectiveness and cost-effectiveness of lay counsellor-delivered psychological treatments for harmful and dependent drinking and moderate to severe depression in primary care in India: PREMIUM study protocol for randomized controlled trials.
The leading mental health causes of the global burden of disease are depression in women and alcohol use disorders in men. A major hurdle to the implementation of evidence-based psychological treatments in primary care in developing countries is the non-availability of skilled human resources. The aim of these trials is to evaluate the effectiveness and cost-effectiveness of two psychological treatments developed for the treatment of depression and alcohol use disorders in primary care in India. This study protocol is for parallel group, randomized controlled trials (Healthy Activity Program for moderate to severe depression, Counselling for Alcohol Problems for harmful and dependent drinking) in eight primary health centres in Goa, India. Adult primary care attendees will be screened with the Patient Health Questionnaire for depression and, in men only, the Alcohol Use Disorders Identification Test for drinking problems. Screen-positive attendees will be invited to participate; men who screen positive for both disorders will be invited to participate in the Counselling for Alcohol Problems trial. Those who consent will be allocated in a 1:1 ratio to receive either the respective psychological treatment plus enhanced usual care or enhanced usual care only using a computer generated allocation sequence, stratified by primary health centre and, for depression, by sex. The enhanced usual care comprises providing primary health centre doctors with contextualized World Health Organization guidelines and screening results. Psychological treatments will be delivered by lay counsellors, over a maximum period of three months. Primary outcomes are severity of disorder and remission rates at three months post-enrolment and, for the Counselling for Alcohol Problems trial, drinking and the impact of drinking on daily lives. Secondary outcomes include severity of disorder and remission rates at 12 months, disability scores, suicidal behaviour and economic impact, and cost-effectiveness at three and 12 months. 500 participants with depression and 400 participants with harmful drinking will be recruited. Primary analyses will be intention-to-treat. These trials may offer a new approach for the treatment of moderate-severe depression and drinking problems in primary care that is potentially scalable as it relies on delivery by a single pool of lay counsellors. Both trials are registered with the International Society for the Registration of Clinical Trials (Healthy Activity Programme registration number ISRCTN95149997; Counselling for Alcohol Problems registration number ISRCTN76465238).


Depolarization block of neurons during maintenance of electrographic seizures.
Epileptic seizures are associated with neuronal hyperactivity. Here, however, we investigated whether continuous neuronal firing is necessary to maintain electrographic seizures. We studied a class of "low-Ca2+" ictal epileptiform bursts, induced in rat hippocampal slices, that are characterized by prolonged (2-15 s) interruptions in population spike generation. We found that, during these interruptions, neuronal firing was suppressed rather than desynchronized. Intracellular current injection, application of extracellular uniform electric fields, and antidromic stimulation showed that the source of action potential disruption was depolarization block. The duration of the extracellular potassium transients associated with each ictal burst was not affected by disruptions in neuronal firing. Application of phenytoin or veratridine indicated a critical role for the persistent sodium current in maintaining depolarization block. Our results show that continuous neuronal firing is not necessary for the maintenance of experimental electrographic seizures.


World distribution, population genetics, and health burden of the hemoglobinopathies.
Although information about the precise world distribution and frequency of the inherited hemoglobin disorders is still limited, there is no doubt that they are going to pose an increasing burden on global health resources in the future. Their high frequency is a reflection of natural selection combined with a high frequency of consanguineous marriages in many countries, together with an epidemiological transition; whereby, as public health measures improve in the poorer countries of the world, more babies with these disorders are surviving to present for treatment.


Gene transfer to the tendon-bone insertion site.
This study investigated whether gene transfer to the tendon-bone insertion site is possible during early tendon-transplant healing using viral vectors. In addition, we evaluated the optimal gene delivery technique for an in vivo adenoviral gene transfer to a tendon-bone insertion site in a bone tunnel. Twenty-six rabbits underwent a bilateral transfer of the flexor digitorum longus tendon into a bone canal in the calcaneus. The animals were divided into two groups. The first group (n=18) received a direct injection of an adenoviral vector carrying the luciferase marker gene into the tendon on the left side, while on the right side the adenoviral vector was first injected into the bone trough and the tendon was later inserted into the trough. The analysis of this experiment showed that over a 4-week period a higher luciferase activity was achieved using the bone trough immersion technique. In the second group (n=8) we therefore used the qualitative marker virus (Ad/-LacZ) with the bone trough immersion technique in order to show the site of gene expression. The histological analysis of this experiment demonstrated the presence of beta-galactosidase positive cells within the tendon-bone interface over a 4-week period. Therefore we showed in the first part of this study that the bone canal provides a more efficient target for direct adenoviral gene delivery than the tendon. In the second part of the study we demonstrated the feasibility of the bone trough immersion technique since sustained gene expression within the tendon-bone interface was obtained for up to 4 weeks. This study has shown the feasibility of gene delivery to the tendon-bone interface and provides the basis for the application adenoviral delivery of growth factor genes to the tendon-bone insertion site.


Low Hemoglobin Levels Are Associated with Reduced Psychomotor and Language Abilities in Young Ugandan Children.
Children living in Sub-Saharan Africa are vulnerable to developmental delay, particularly in the critical first five years due to various adverse exposures including disease and nutritional deficiencies. Anemia and iron deficiency (ID) are highly prevalent in pregnant mothers and young children and are implicated in abnormal brain development. However, available evidence on the association between anemia, ID and neurodevelopment in sub-Saharan Africa is limited. Using data from the Entebbe Mother and Baby Study prospective birth cohort, we examined the effect of maternal and child hemoglobin (Hb) levels and child iron status on developmental scores in 933 and 530 pre-school Ugandan children respectively. Associations between Hb levels, iron status and developmental scores were assessed using regression analyses adjusting for potential confounders. Lower maternal and child Hb levels were associated with reduced psychomotor scores at 15 months, while only lower Hb levels in infancy were associated with reduced language scores. We found no evidence that anemia or ID was associated with cognitive or motor scores at five years. This study emphasizes the importance of managing anemia in pregnancy and infancy and highlights the need for further studies on the effects of anemia and ID in children living in Sub-Saharan Africa.


Silencing of GATA3 defines a novel stem cell-like subgroup of ETP-ALL.
GATA3 is pivotal for the development of T lymphocytes. While its effects in later stages of T cell differentiation are well recognized, the role of GATA3 in the generation of early T cell precursors (ETP) has only recently been explored. As aberrant GATA3 mRNA expression has been linked to cancerogenesis, we investigated the role of GATA3 in early T cell precursor acute lymphoblastic leukemia (ETP-ALL). We analyzed GATA3 mRNA expression by RT-PCR (n = 182) in adult patients with T-ALL. Of these, we identified 70 of 182 patients with ETP-ALL by immunophenotyping. DNA methylation was assessed genome wide (Illumina Infinium® HumanMethylation450 BeadChip platform) in 12 patients and GATA3-specifically by pyrosequencing in 70 patients with ETP-ALL. The mutational landscape of ETP-ALL with respect to GATA3 expression was investigated in 18 patients and validated by Sanger sequencing in 65 patients with ETP-ALL. Gene expression profiles (Affymetrix Human genome U133 Plus 2.0) of an independent cohort of adult T-ALL (n = 83) were used to identify ETP-ALL and investigate GATA3low and GATA3high expressing T-ALL patients. In addition, the ETP-ALL cell line PER-117 was investigated for cytotoxicity, apoptosis, GATA3 mRNA expression, DNA methylation, and global gene expression before and after treatment with decitabine. In our cohort of 70 ETP-ALL patients, 33 % (23/70) lacked GATA3 expression and were thus defined as GATA3low. DNA methylation analysis revealed a high degree of GATA3 CpG island methylation in GATA3low compared with GATA3high ETP-ALL patients (mean 46 vs. 21 %, p < 0.0001). Genome-wide expression profiling of GATA3low ETP-ALL exhibited enrichment of myeloid/lymphoid progenitor (MLP) and granulocyte/monocyte progenitor (GMP) genes, while T cell-specific signatures were downregulated compared to GATA3high ETP-ALL. Among others, FLT3 expression was upregulated and mutational analyses demonstrated a high rate (79 %) of FLT3 mutations. Hypomethylating agents induced reversal of GATA3 silencing, and gene expression profiling revealed downregulation of hematopoietic stem cell genes and upregulation of T cell differentiation. We propose GATA3low ETP-ALL as a novel stem cell-like leukemia with implications for the use of myeloid-derived therapies.


Posttransplant diabetes mellitus in pediatric renal transplant recipients: a report of the North American Pediatric Renal Transplant Cooperative Study (NAPRTCS).
The incidence of renal post transplant diabetes mellitus (PTDM) in adults varies from 3-46%. We did a retrospective analysis of 1365 children in The North American Pediatric Renal Transplant Cooperative Study with renal transplant (Tx) reported between January 92 and July 1997. PTDM, defined as >2 weeks of insulin therapy after Tx, developed in 36 patients. A control group of 153/1329 non-PTDM patients was selected and matched for age at Tx and primary diagnosis. African-Americans were overrepresented (36.1 vs. 17.6%, P=0.017) and Hispanics were underrepresented (5.6 vs. 26.1%, P=0.019) among cases. Although prednisone dose 30 days post-Tx was higher among cases (0.89 mg/kg/day) versus controls (0.71 mg/kg/day), P=0.019, cyclosporine dose was similar. No differences in prednisone or cyclosporine doses were observed at 6, 12, or 24 months post-Tx. Tacrolimus use in PTDM group was high (45%). The estimated incidence of first acute rejection at 1, 3, and 12 months was higher among cases, 0.41+/-0.08, 0.52+/-0.08, 0.61+/-0.08, compared to controls, 0.23+/-0.02, 0.37+/-0.02, and 47+/-0.02 (P=0.058). Crude graft failure rates of 13.5% (5/36) and 12.4% (19/153) were similar between the two groups, so was the calculated creatinine clearance at 12 and 24 months and post-Tx hospitalization days. PTDM occurs in <3% of children. African-Americans are at higher risk and Hispanics at lower risk for PTDM. Tacrolimus is a significant risk factor for PTDM. Children with PTDM had a higher incidence of acute rejection, but graft survival, kidney function, and hospitalization rates were similar to selected controls.


Recurrent Neural Networks for Early Detection of Heart Failure From Longitudinal Electronic Health Record Data: Implications for Temporal Modeling With Respect to Time Before Diagnosis, Data Density, Data Quantity, and Data Type.
We determined the impact of data volume and diversity and training conditions on recurrent neural network methods compared with traditional machine learning methods. Using longitudinal electronic health record data, we assessed the relative performance of machine learning models trained to detect a future diagnosis of heart failure in primary care patients. Model performance was assessed in relation to data parameters defined by the combination of different data domains (data diversity), the number of patient records in the training data set (data quantity), the number of encounters per patient (data density), the prediction window length, and the observation window length (ie, the time period before the prediction window that is the source of features for prediction). Data on 4370 incident heart failure cases and 30 132 group-matched controls were used. Recurrent neural network model performance was superior under a variety of conditions that included (1) when data were less diverse (eg, a single data domain like medication or vital signs) given the same training size; (2) as data quantity increased; (3) as density increased; (4) as the observation window length increased; and (5) as the prediction window length decreased. When all data domains were used, the performance of recurrent neural network models increased in relation to the quantity of data used (ie, up to 100% of the data). When data are sparse (ie, fewer features or low dimension), model performance is lower, but a much smaller training set size is required to achieve optimal performance compared with conditions where data are more diverse and includes more features. Recurrent neural networks are effective for predicting a future diagnosis of heart failure given sufficient training set size. Model performance appears to continue to improve in direct relation to training set size.


P-glycoprotein mediates resistance to histidine kinase inhibitors.
Histidine kinase inhibitors are being developed as a new class of antimicrobial drugs. We recently demonstrated the activity of a class of histidine kinase inhibitors against a mammalian enzyme, elongation factor-2 kinase (eEF-2K), and the effect of these compounds on cancer cell viability (Arora et al., 2003). To further characterize these compounds, we studied their interaction with ATP-binding cassette transporters, which are known to mediate resistance to a variety of chemotherapeutic agents. The 24 compounds studied belong to three structural series of derivatives of 2-methylimidazolium iodide. We focused this work on a representative compound (NH125) because we found it to be most potent against both histidine kinase and eEF-2K among the series. Cell lines that expressed P-glycoprotein (P-gp) were 2- to 5-fold resistant to NH125. NH125 increased accumulation of P-gp substrates such as paclitaxel and doxorubicin but had no effect on the accumulation of non-P-gp substrates. P-gp modulators verapamil and trans-flupenthixol and MDR1-targeted siRNA increased sensitivity of multidrug-resistant cell lines to NH125. The presence of a benzyl group on the N-3 position of the 2-methylimidazolium iodide was important for the interaction with P-gp. C6-NH, an NH125-resistant cell line, markedly overexpressed P-gp compared with the parental cell line. In animal models, we found that NH125 increased by 129% the survival of sensitive P388 cells bearing mice but had no effect on mice harboring the resistant cell line. These observations indicate that certain histidine kinase inhibitors are substrates for P-gp and hence an important consideration in development of these agents as potential antimicrobial and anticancer agents.


Unraveling the Catalytic Pathway of Metalloenzyme Farnesyltransferase through QM/MM Computation.
The protein farnesyltransferase (FTase) is a Zn(2+)-metalloenzyme that catalyzes the farnesylation reaction, i.e., the transfer of the 15-carbon atom farnesyl group from farnesyl diphosphate (FPP) to a specific cysteine of protein substrates. Oncogenic Ras proteins, which are among the FTase substrates, are observed in about 20-30% of human cancer cells. Thus, FTase represents a target for anticancer drug design. Herein, we present a classical force-field-based and quantum mechanics/molecular mechanics (QM/MM) computational study of the FTase reaction mechanism. Our findings offer a detailed picture of the FTase catalytic pathway, describing structural features and the energetics of its saddle points. A moderate dissociation of the diphosphate group from the FPP is observed during the nucleophilic attack of the zinc-bound thiolate. At the transition state, a resonance structure is observed, which indicates the formation of a metastable carbocation. However, no stable intermediate is found along the reaction pathway. Thus, the reaction occurs via an associative mechanism with dissociative character, in agreement with the mechanism proposed by Fierke et al. ( Biochemistry 2000, 39, 2593-2602 and Biochemistry 2003, 42, 9741-9748 ). Moreover, a fluorine-substituted FPP analogue (CF3-FPP) is used to investigate the inhibitory effect of fluorine, which in turn provides additional agreement with experimental data.


Tai chi for upper limb rehabilitation in stroke patients: the patient's perspective.
This study aimed at exploring the perceived benefits and drawbacks of practicing tai chi, an alternative therapy that can be implemented in the community, as part of upper-limb rehabilitation following stroke. Semistructured interviews were carried out with participants with chronic stroke (>6 months). The participants took part in 16 tai chi sessions over 8 weeks. Interviews were conducted in person using an interview guide based on the theory of planned behavior (TPB), and a thematic analysis was conducted. Eight interviews were carried out with participants at various stages of motor recovery. Participants perceived a number of physical, functional, and psychological benefits. They found tai chi to be a global exercise, including both physical and mental aspects, and suggested that it can be included as part of rehabilitation for stroke patients. Many participants expressed a desire to continue practicing tai chi after completion of the study because it exceeded their expectations, among other reasons. This study can serve to guide future tai chi interventions and research on tai chi for rehabilitation in terms of the characteristics of the intervention and the various areas to assess in order to measure the overall benefits. IMPLICATIONS FOR REHABILITATION Tai chi was perceived as a good way of integrating various skills learned during rehabilitation. Despite having different functional abilities, all the participants noted various physical, functional, and psychological benefits from participating in the tai chi sessions. Tai chi seems to be a form of exercise that stroke patients would perform more long-term since all the participants in this study expressed the desire to continue practicing tai chi.


Management of severe hypertriglyceridemia due to lipoprotein lipase deficiency in children
Severe hypertriglyceridemia is an endocrine emergency and is associated with acute pancreatitis and hyperviscosity syndrome. We describe an infant with lipoprotein lipase deficiency with severe hypertriglyceridemia who presented with acute pancreatitis. She was managed acutely with fasting and intravenous insulin infusion, followed by low-fat diet with no pharmacological agent. Subsequent follow-up until the age of 5 years showed satisfactory lipid profile and she has normal growth and development. Hypertriglyceridemia-induced acute pancreatitis has significant morbidity and mortality, and prompt treatment is imperative. When no secondary causes are readily identified, genetic evaluation should be pursued in hypertriglyceridemia in children. Intravenous insulin is a safe and effective acute treatment for hypertriglyceridemia in children, even in infants. Long-term management with dietary modifications alone could be effective for primary hypertriglyceridemia due to lipoprotein lipase deficiency, at least in early childhood phase.


Hepatitis C virus increases the risk of kidney disease among HIV-positive patients: Systematic review and meta-analysis.
Kidney disease has become an important co-morbidity among human immunodeficiency virus-infected patients as they live longer in the era of highly effective antiretroviral therapy. It remains unclear how co-infection with hepatitis C virus impacts on the trajectory of kidney disease among HIV-infected patients. To evaluate the effect of co-infection with HCV on the risk of kidney disease in HIV-infected populations. We conducted a systematic review of the published medical literature to determine if hepatitis C co-infection is associated with increased likelihood of chronic kidney disease in HIV-positive adults. We used the random effects model of DerSimonian and Laird to generate a summary estimate of the relative risk for chronic kidney disease (defined by reduced glomerular filtration rate and/or detectable proteinuria) with hepatitis C virus across the published studies. Meta-regression and stratified analysis were also conducted. We identified 19 studies (146,151 unique patients with HIV) and separate meta-analyses were performed according to the outcome. Aggregation of longitudinal studies (n = 8, 105,462 unique patients) showed a relationship between HCV infection and increased risk of reduced glomerular filtration rate among HIV-infected individuals, the summary estimate for adjusted hazard ratio was 1.64 (95%CI, 1.28; 2.0, P < 0.001) in HIV-HCV co-infected individuals compared with those having HIV mono-infection. No between-studies heterogeneity was noted (P-value by Q test = 0.08). HCV positive serology was an independent risk factor for proteinuria; adjusted effect estimate, 1.23 (95% confidence interval, 1.18; 1.28, P = 0.001) (n = 6 studies; 26,835 unique patients). In meta-regression, we noted the impact of ageing (P = 0.0001) upon the adjusted hazard ratio of incidence of reduced glomerular filtration rate among HCV-HIV co-infected patients; a negative association between frequency of males (P = 0.001) and the adjusted hazard ratio of prevalence of low glomerular filtration rate was found. Hepatitis C co-infection is associated with a significant increase in the risk of reduced glomerular filtration rate and/or detectable proteinuria among HIV-infected individuals.


[Glucose tolerance alterations and frequency of metabolic syndrome among patients with non alcoholic fatty liver disease].
Non alcoholic fatty liver disease (NAFLD) is associated to diabetes mellitus, obesity, disturbances in serum lipid levels, insulin resistance and metabolic syndrome. To assess glucose tolerance and the presence of metabolic syndrome among patients with biopsy proven NAFLD. Serum lipid levels, hepatic function tests were measured and an oral glucose tolerance test was performed in 46 patients (mean age 45+/-12 years, 36 females) without history of diabetes mellitus and with steatosis in a liver biopsy. Mean body mass index of the sample was 37+/-12 kg/m2. Seventeen percent had pure steatosis, 78% had steatohepatitis with or without fibrosis and 50% had fibrosis in the liver biopsy. Glucose intolerance and diabetes was found in 57% and 15% of cases, respectively. The presence of steatohepatitis was higher in diabetics, compared with subjects with glucose intolerance or a normal glucose response (43, 38 and 8%, respectively, p < 0.0001). Ninety three percent had a metabolic syndrome and the proportion of biopsies with fibrosis was higher among subjects with more than three diagnostic criteria for metabolic syndrome compared with those with three or less criteria (59 and 46%, respectively, p < 0.05). Glucose intolerance, diabetes and metabolic syndrome are common among patients with NAFLD, even when they are not obese.


Characterization of SNARE cleavage products generated by formulated botulinum neurotoxin type-a drug products.
The study evaluated substrate cleavage product(s) generated by three botulinum neurotoxin serotype A (BoNT/A) medicinal drug products utilizing a novel and highly specific, light-chain activity, high-performance liquid chromatography (LCA-HPLC) method. Samples were reacted with a commercially available BoNT/A fluorescent substrate derived from the SNAP-25 sequence. Reaction products were separated by reversed-phase HPLC. The method detected an atypical cleavage pattern by one of the formulated drug products. IncobotulinumtoxinA produced two cleavage fragments rather than the single fragment typically generated by BoNT/A. Identification confirmed the secondary cleavage at a position corresponding to SNAP-25 Arg198-Ala199 (normal BoNT/A cleavage is Gln197-Arg198). Arg198-Ala199 is also the cleavage site for trypsin and serotype C toxin. Normal cleavage was observed for all other BoNT/A drug product samples, as well as 900-kD and 150-kD bulk toxin BoNT/A. The reason for this unexpected secondary cleavage pattern by one formulated BoNT/A drug product is unknown. Possible explanations include a contaminating protease and/or damage to the 150-kD type-A toxin causing nonspecific substrate recognition and subsequent cleavage uncharacteristic of type-A toxin. The BoNT/A drug products were also analyzed via the LCA-HPLC assay using a commercial BoNT/C fluorescent substrate derived from the syntaxin sequence. Cleavage of the serotype C substrate by incobotulinumtoxinA was also confirmed whilst neither of the other drug products cleaved the syntaxin substrate.


Localization of tektin filaments in microtubules of sea urchin sperm flagella by immunoelectron microscopy.
Extraction of doublet microtubules from the sperm flagella of the sea urchin Strongylocentrotus purpuratus with sarkosyl (0.5%)-urea (2.5 M) yields a highly pure preparation of "tektin" filaments that we have previously shown to resemble intermediate filament proteins. They form filaments 2-3 nm in diameter as seen by negative stain electron microscopy and are composed of approximately equal amounts of three polypeptide bands with apparent molecular weights of 47,000, 51,000, and 55,000, as determined by SDS PAGE. We prepared antibodies to this set of proteins to localize them in the doublet microtubules of S. purpuratus and other species. Tektins and tubulin were antigenically distinct when tested by immunoblotting with affinity-purified antitektin and antitubulin antibodies. Fixed sperm or axonemes from several different species of sea urchin showed immunofluorescent staining with antitektin antibodies. We also used antibodies coupled to gold spheres to localize the proteins by electron microscopy. Whereas a monoclonal antitubulin (Kilmartin, J.V., B. Wright, and C. Milstein, 1982, J. Cell Biol. 93:576-582) decorates intact microtubules along their lengths, antitektins labeled only the ends of intact microtubules and sarkosyl-insoluble ribbons. However, if microtubules and ribbons attached to electron microscope grids were first extracted with sarkosyl-urea, the tektin filaments that remain were decorated by antitektin antibodies throughout their length. These results suggest that tektins form integral filaments of flagellar microtubule walls, whose antigenic sites are normally masked, perhaps by the presence of tubulin around them.


Biological or psychological? Effects of eating disorder psychoeducation on self-blame and recovery expectations among symptomatic individuals.
Recent years have witnessed increasing popularity and promotion of biological influences (e.g., genetics) in eating disorder (ED) development. Although research suggests biological models of EDs reduce blame-oriented stigma in the general public, their effect on symptomatic individuals' attitudes toward themselves, treatment, and their prognosis has not been studied. Additionally, little is known about how other credible forms of conceptualizing ED development (e.g., cognitive-behavioral) affect individuals with disordered eating. Accordingly, the present study assessed the effects of three different forms of psychoeducation about ED development (biology-only, malleability of biology, cognitive-behavioral) among a sample high in ED symptoms. Participants (N = 216) viewed an audiovisual presentation describing ED development from one of the three perspectives before completing measures of self-blame for symptoms, prognostic expectations, self-efficacy in recovering, and attitudes toward a description of cognitive-behavioral therapy. There were no significant differences between conditions in self-blame. Relative to biology-only, the psychoeducational messages emphasizing malleable biology and cognitive-behavioral factors produced more prognostic optimism and self-efficacy in recovering. Perceived credibility of cognitive-behavioral therapy and expectations for its efficacy were highest in the cognitive-behavioral psychoeducation condition. Implications for efforts to educate the public and treatment-seeking individuals about the nature of EDs are discussed.


Does the Meeting Centre Support Programme decrease the experience of stigmatisation among people with cognitive deficits?
This is the first study to focus on the role and impact of a psychosocial intervention, the Meeting Centre Support Programme (MCSP), for people living with dementia and mild cognitive impairment (MCI) on the experience of stigmatisation across three different European countries. A pre/post-test control group study design compared outcomes for 114 people with dementia (n=74) and MCI (n=40) in Italy, Poland and the UK who received either the MCSP or usual care (UC). The 'Stigma Impact Scale, neurological disease' (SIS) was administered at two points in time, 6 months apart. The Global Deterioration Scale (GDS) was used to assess the level of cognitive impairment. Although statistical analysis did not show any significant differences between MCSP and UC at pre/post-test for the 3 countries combined, there were significant results for individual countries. In Italy, the level of SIS was significantly lower (p=0.02) in the MCSP group following the intervention. The level of Social Isolation increased significantly (p=0.05) in the UC group at follow-up in Poland. The level of Social Rejection was significantly higher (p=0.03) over time for UK participants receiving MCSP compared to UC. The experience of stigma by people living with dementia and MCI is complex and there may be different country specific contexts and mechanisms. The results do not enable us to confirm or disconfirm the impact of a social support programme, such as MCSP, on this experience. Difficulties in directly measuring the level of stigma in this group also requires further research.


Community-directed interventions for priority health problems in Africa: results of a multicountry study.
To determine the extent to which the community-directed approach used in onchocerciasis control in Africa could effectively and efficiently provide integrated delivery of other health interventions. A three-year experimental study was undertaken in 35 health districts from 2005 to 2007 in seven research sites in Cameroon, Nigeria and Uganda. Four trial districts and one comparison district were randomly selected in each site. All districts had established ivermectin treatment programmes, and in the trial districts four other established interventions - vitamin A supplementation, use of insecticide-treated nets, home management of malaria and short-course, directly-observed treatment for tuberculosis patients - were progressively incorporated into a community-directed intervention (CDI) process. At the end of each of the three study years, we performed quantitative evaluations of intervention coverage and provider costs, as well as qualitative assessments of the CDI process. With the CDI strategy, significantly higher coverage was achieved than with other delivery approaches for all interventions except for short-course, directly-observed treatment. The coverage of malaria interventions more than doubled. The district-level costs of delivering all five interventions were lower in the CDI districts, but no cost difference was found at the first-line health facility level. Process evaluation showed that: (i) participatory processes were important; (ii) recurrent problems with the supply of intervention materials were a major constraint to implementation; (iii) the communities and community implementers were deeply committed to the CDI process; (iv) community implementers were more motivated by intangible incentives than by external financial incentives. The CDI strategy, which builds upon the core principles of primary health care, is an effective and efficient model for integrated delivery of appropriate health interventions at the community level in Africa.


Activation of murine natural killer cells and macrophages by 8-bromoguanosine.
It has been shown that 8-bromoguanosine (8-BrGuo) activates T and B cells, but the underlying mechanism of this effect is not clear. We found that it also activates NK cells and macrophages by induction of IFN production. Culturing spleen cells with 8-BrGuo for 16 to 18 h induced cytotoxic activity to YAC cells. The cytotoxic cells expressed Qa-5, asialo-Gm-1, and NK-1.1 Ag. Similarly, preincubation of peptone-elicited peritoneal macrophages with 8-BrGuo induced macrophage cytolytic activity to P815 cells in an 18-h assay. Anti-IFN antibodies to IFN-alpha -beta and -gamma abolished these inductive events. Furthermore, elimination of asialo-Gm-1+ cells reduced the responses of NK and macrophages to 8-BrGuo, indicating that these cells possibly produce the IFN. We conclude from these observations that one biologic effect of 8-BrGuo is due to IFN production.


Nod-like receptor protein 3 (NLRP3) inflammasome activation and podocyte injury via thioredoxin-interacting protein (TXNIP) during hyperhomocysteinemia.
NADPH oxidase-derived reactive oxygen species (ROS) have been reported to activate NLRP3 inflammasomes resulting in podocyte and glomerular injury during hyperhomocysteinemia (hHcys). However, the mechanism by which the inflammasome senses ROS is still unknown in podocytes upon hHcys stimulation. The current study explored whether thioredoxin-interacting protein (TXNIP), an endogenous inhibitor of the antioxidant thioredoxin and ROS sensor, mediates hHcys-induced NLRP3 inflammasome activation and consequent glomerular injury. In cultured podocytes, size exclusion chromatography and confocal microscopy showed that inhibition of TXNIP by siRNA or verapamil prevented Hcys-induced TXNIP protein recruitment to form NLRP3 inflammasomes and abolished Hcys-induced increases in caspase-1 activity and IL-1β production. TXNIP inhibition protected podocytes from injury as shown by normal expression levels of podocyte markers, podocin and desmin. In vivo, adult C57BL/6J male mice were fed a folate-free diet for 4 weeks to induce hHcys, and TXNIP was inhibited by verapamil (1 mg/ml in drinking water) or by local microbubble-ultrasound TXNIP shRNA transfection. Evidenced by immunofluorescence and co-immunoprecipitation studies, glomerular inflammasome formation and TXNIP binding to NLRP3 were markedly increased in mice with hHcys but not in TXNIP shRNA-transfected mice or those receiving verapamil. Furthermore, TXNIP inhibition significantly reduced caspase-1 activity and IL-1β production in glomeruli of mice with hHcys. Correspondingly, TXNIP shRNA transfection and verapamil attenuated hHcys-induced proteinuria, albuminuria, glomerular damage, and podocyte injury. In conclusion, our results demonstrate that TXNIP binding to NLRP3 is a key signaling mechanism necessary for hHcys-induced NLRP3 inflammasome formation and activation and subsequent glomerular injury.


Intramedullary spinal cord lipomas.
Spinal cord lipomas are rare lesions, accounting for approximately 1% of all spinal cord tumors. True intramedullary spinal cord lipomas are extremely rare and are represented in the literature as scattered, single case reports. The authors present a series of six patients with intramedullary spinal cord lipomas managed at our institution from July, 1985 to July, 1993. The patients' ages ranged from 8 to 45 years. Four patients presented with newly diagnosed tumors and two had undergone previous surgery. Patients usually presented with long histories of disability followed by rapid progression of their symptoms. Most patients were in poor neurological condition on presentation. Presenting symptoms included spinal pain, dysesthetic sensory changes, gait difficulties, weakness, and incontinence. Three patients had cervical tumors, two had cervicothoracic tumors, and one patient had a thoracic tumor. Diagnostic studies, including magnetic resonance imaging, were obtained in all patients. No patient exhibited any form of spinal dysraphism or had a dural defect. All patients underwent decompressive, subtotal resections of 40% to 70% of their lesions. Follow-up times ranged from 12 to 96 months. All patients had resolution of their pain, but they generally showed no neurological improvement. As of their most recent follow-up visit, none of the patients was neurologically normal; three can function independently, although with neurological deficits. The other three patients cannot function independently and have severe neurological deficits. The authors conclude that patients with intramedullary spinal cord lipoma who present with significant neurological compromise have a very poor prognosis with regard to neurological function and generally show no improvement with surgical resection.