Skeletal repair by in situ formation of the mineral phase of bone.
A process has been developed for the in situ formation of the mineral phase of bone. Inorganic calcium and phosphate sources are combined to form a paste that is surgically implanted by injection. Under physiological conditions, the material hardens in minutes concurrent with the formation of dahllite. After 12 hours, dahllite formation was nearly complete, and an ultimate compressive strength of 55 megapascals was achieved. The composition and crystal morphology of the dahllite formed are similar to those of bone. Animal studies provide evidence that the material is remodeled in vivo. A novel approach to skeletal repair is being tested in human trials for various applications; in one of the trials the new biomaterial is being percutaneously placed into acute fractures. After hardening, it serves as internal fixation to maintain proper alignment while healing occurs.


Solvent vapor annealing in the molecular regime drastically improves carrier transport in small-molecule thin-film transistors.
We demonstrate a new way to investigate and control the solvent vapor annealing of solution-cast organic semiconductor thin films. Solvent vapor annealing of spin-cast films of 6,13-bis(triisopropylsilylethynyl) pentacene (TIPS-Pn) is investigated in situ using quartz crystal microbalance with dissipation (QCM-D) capability, allowing us to monitor both solvent mass uptake and changes in the mechanical rigidity of the film. Using time-resolved grazing incidence wide angle X-ray scattering (GIWAXS) and complementary static atomic force microscopy (AFM), we demonstrate that solvent vapor annealing in the molecular regime can cause significant performance improvements in organic thin film transistors (OTFTs), whereas allowing the solvent to percolate and form a liquid phase results in catastrophic reorganization and dewetting of the film, making the process counterproductive. Using these lessons we devise processing conditions which prevent percolation of the adsorbed solvent vapor molecules for extended periods, thus extending the benefits of solvent vapor annealing and improving carrier mobility by nearly two orders of magnitude. Ultimately, it is demonstrated that QCM-D is a very powerful sensor of the state of the adsorbed solvent as well as the thin film, thus making it suitable for process development as well as in-line process monitoring both in laboratory and in future manufacturing settings.


An electrochemical and photophysical study of a covalently linked inorganic-organic dyad.
A molecular donor-acceptor dyad comprising a hexarhenium cluster core, [Re(6)(mu(3)-Se)(8)](2+), and a fullerene moiety which are covalently linked through a pyridine ligand was synthesized and fully characterized. The electrochemical and photophysical properties are reported. The detailed study includes cyclic voltammetry, steady-state absorption and fluorescence spectroscopy, radiation chemistry and transient absorption spectroscopy. A light-induced electron transfer between the inorganic cluster moiety and the fullerene can be excluded. However, a light-induced energy transfer from the rhenium cluster to the fullerene is proposed.


Differences in tolerability of pirfenidone between elderly and younger patients with idiopathic pulmonary fibrosis.
Although pirfenidone (PFD) is a key drug for the treatment of idiopathic pulmonary fibrosis (IPF), differences in tolerability between elderly and young patients remain unclear. This study aimed to investigate age-related differences in adverse drug reactions to PFD and to evaluate whether patient age influences the safety and tolerability of PFD in clinical practice. One hundred fifty-four patients with IPF were treated with PFD in our institution between May 2009 and April 2017; these patients were classified into 2 groups on the basis of age: ≥75 years of age (elderly patients) and <75 years of age (younger patients). In each group, the clinical course, laboratory data, radiographic findings, adverse events, and tolerability of PFD at 6 months and 1 year after administration were retrospectively analyzed. Among the 120 patients examined in this study, 31 patients (26%) were ≥75 years of age. The continuation rate of PFD at 1 year in the elderly patient group was significantly lower (n=11 [35%] vs 57 [64%], p=0.007) than in the younger patient group. Regarding adverse drug reactions to PFD, the incidence of gastrointestinal disorders including anorexia (n=24 [77%] vs 40 [45%], p=0.002) and the discontinuation caused by gastrointestinal disorders (n=11 [35%] vs 13 [15%], p=0.019) were significantly higher in elderly patients than those in younger patients. However, with the exception of gastrointestinal disorders, other adverse drug reactions did not significantly differ between elderly and younger patients. Compared with younger patients, elderly patients with IPF had a higher incidence of gastrointestinal disorders, along with an increased discontinuation rate of PFD. More careful management of gastrointestinal disorders may be required to ensure continuation of PFD in elderly patients.


Studies of dialysates from Dermatophagoides farinae: partial purification and haptenic properties of dialysates from Dermatophagoides farinae.
Dialysates from Dermatophagoides farinae were partially purified. Fractionation on HPLC and anion exchange chromatography revealed that the dialysates consisted of 5 major fractions of glycoprotein whose apparent molecular sizes were 5.1, 4.1, 3.2, and less than 1.35 kD on HPLC. The apparent molecular size of two fractions was 5.3 and 2.9 kD on SDS-PAGE. They were basic glycoproteins which had a pI ranging from 7.46 to 8.71 on PAG-IEF. These fractions were allergenic in the RAST and ELISA inhibition tests but not in the skin prick test (SPT). Our results suggest that the dialysates from D. farinae have haptenic properties. The dialysates from D. farinae (low molecular weight) and its 5 fractions bound noncovalently to human serum albumin (HSA) at the free tyrosine residues of HSA. They proved to bind noncovalently to serum proteins and collagens. Once they bound to proteins, the conjugates became allergenic not only in the RAST and ELISA inhibition test but also in the SPT. Our results provide evidence that the dialysates from D. farinae have haptenic properties.


Juvenile rheumatoid arthritis in Caribbean children: a clinical characterization.
The knowledge about rheumatic diseases in Latin American children is scanty. The features of Juvenile Rheumatoid Arthritis (J.R.A.) in the islands of the Caribbean had not been described. In this retrospective study, fifty cases of J.R.A. in a pediatric population of Puerto Rico and the Virgin Islands followed at a formal pediatric rheumatology service in San Juan, Puerto Rico, were demographically and clinically characterized. There was a female predominance of 1.4:1. The peak of occurrence was at 1-2 years of age. Pauciarticular onset was the most common through all ages. Antinuclear antibodies and rheumatoid factor were positive in 40% and 10% respectively. Although a larger and prospective study is necessary to determine if these clinical trends will prevail, the present findings suggest that the characteristics of J.R.A. among children in the Caribbean are similar to other previously published series in U.S.A. and Europe.


Loss of Shh signaling in the neocortex reveals heterogeneous cell recovery responses from distinct oligodendrocyte populations.
The majority of oligodendrocytes in the neocortex originate from neural progenitors that reside in the dorsal forebrain. We recently showed that Sonic Hedgehog (Shh) signaling in these dorsal progenitors is required to produce normal numbers of neocortical oligodendrocytes during embryonic development. Conditional deletion of the Shh signaling effector, Smo, in dorsal progenitors caused a dramatic reduction in oligodendrocyte numbers in the embryonic neocortex. In the current study, we show that the depleted oligodendrocyte lineage in Smo conditional mutants is able to recover to control numbers over time. This eventual recovery is achieved in part by expansion of the ventrally-derived wild-type lineage that normally makes up a minority of the total oligodendrocyte population. However, we find that the remaining dorsally-derived mutant cells also increase in numbers over time to contribute equally to the recovery of the total population. Additionally, we found that the ways in which the dorsal and ventral sources cooperate to achieve recovery is different for distinct populations of oligodendrocyte-lineage cells. Oligodendrocyte precursor cells (OPCs) in the neocortical white matter recover completely by expansion of the remaining dorsally-derived Smo mutant cells. On the other hand, mature oligodendrocytes in the white and gray matter recover through an equal contribution from dorsal mutant and ventral wild-type lineages. Interestingly, the only population that did not make a full recovery was OPCs in the gray matter. We find that gray matter OPCs are less proliferative in Smo cKO mutants compared to controls, which may explain their inability to fully recover. Our data indicate that certain populations of the dorsal oligodendrocyte lineage are more affected by loss of Shh signaling than others. Furthermore, these studies shed new light on the complex relationship between dorsal and ventral sources of oligodendrocytes in the developing neocortex.


Subacute intranasal administration of tissue plasminogen activator increases functional recovery and axonal remodeling after stroke in rats.
As a thrombolytic agent, application of recombinant tissue plasminogen activator (tPA) to ischemic stroke is limited by the narrow time window and side effects on brain edema and hemorrhage. This study examined whether tPA, administered by intranasal delivery directly targeting the brain and spinal cord, provides therapeutic benefit during the subacute phase after stroke. Adult male Wistar rats were subjected to permanent right middle cerebral artery occlusion (MCAo). Animals were treated intranasally with saline, 60 μg or 600 μg recombinant human tPA at 7 and 14days after MCAo (n=8/group), respectively. An adhesive-removal test and a foot-fault test were used to monitor functional recovery. Biotinylated dextran amine (BDA) was injected into the left motor cortex to anterogradely label the corticorubral tract (CRT) and the corticospinal tract (CST). Naive rats (n=6) were employed as normal control. Animals were euthanized 8 weeks after stroke. Compared with saline treated animals, significant functional improvements were evident in rats treated with 600 μg tPA (p<0.05), but not in 60 μg tPA treated rats. Furthermore, 600 μg tPA treatment significantly enhanced both CRT and CST sprouting originating from the contralesional cortex into the denervated side of the red nucleus and cervical gray matter compared with control group (p<0.01), respectively. The behavioral outcomes were highly correlated with CRT and CST axonal remodeling. Our data suggest that delayed tPA intranasal treatment provides therapeutic benefits for neurological recovery after stroke by, at least in part, promoting neuronal remodeling in the brain and spinal cord.


KCl-induced cortical spreading depression waves more heterogeneously propagate than optogenetically-induced waves in lissencephalic brain: an analysis with optical flow tools.
Although cortical spreading depolarizations (CSD) were originally assumed to be homogeneously and concentrically propagating waves, evidence obtained first in gyrencephalic brains and later in lissencephalic brains suggested a rather non-uniform propagation, shaped heterogeneously by factors like cortical region differences, vascular anatomy, wave recurrences and refractory periods. Understanding this heterogeneity is important to better evaluate the experimental models on the mechanistics of CSD and to make appropriate clinical estimations on neurological disorders like migraine, stroke, and traumatic brain injury. This study demonstrates the application of optical flow analysis tools for systematic and objective evaluation of spatiotemporal CSD propagation patterns in anesthetized mice and compares the propagation profile in different CSD induction models. Our findings confirm the asymmetric angular CSD propagation in lissencephalic brains and suggest a strong dependency on induction-method, such that continuous potassium chloride application leads to significantly higher angular propagation variability compared to optogenetically-induced CSDs.


Yin Yang 1 overexpression in diffuse large B-cell lymphoma is associated with B-cell transformation and tumor progression.
Yin Yang 1 (YY1), a multifunctional transcription factor, has been shown to be involved in the pathogenesis of several cancer types. However, its role in hematological malignancies has not yet been fully investigated. In the present study, using computational methods, we showed that YY1 transcript levels were significantly increased in the high-grade lymphomas, including Burkitt's lymphoma and diffuse large B-cell lymphoma (DLBCL), compared with those of both low-grade lymphomas and normal B-cells. The significant increase in gene expression resulted in a significant increase also at protein level in three NHL cell lines. The association of YY1 expression with some clinical-pathological features in DLBCL showed a positive correlation between a high level of YY1 mRNA and high levels of BCL-6 protein. Moreover, by analyzing the large series of DLBCL in the Hummel dataset, we identified the transcription factor PAX-5 among the top 50 genes positively correlated with YY1. These findings are also supported by the biological network analysis in which the top network, with the highest score, associated with YY1 expression levels in DLBCL is cellular movement, hematological system development and function, and immune response. overall these data suggest that YY1 is involved in B cells transformation which gives rise to high-grade lymphomas through a dysregulation in the normal development of B cells affecting cell cycle and cellular motility.


Sexual and reproductive rights at the United Nations: frustration or fulfilment?
Over the past 20 years, advocates have gained formal recognition for some rights in sexuality and reproduction and established the application of human rights standards to sexual and reproductive health issues more generally. However, careful reflection on the state of norm development across sexuality and reproduction as a field reveals fractures and stagnation in the development of standards, and a lack of synergy among advocates and between frameworks for similar rights. This paper seeks to stimulate a more careful accounting for these realities. It examines the formal processes and rules guiding standard-setting, in light of the different intellectual and ideological genealogies of sexual and reproductive rights. We use (homo)sexual orientation and abortion as case studies of current high-profile human rights standard-setting, with specific attention to the contemporary state of human rights law-making in the United Nations today. By placing these two issues in conjunction, we seek to make visible relationships between the vicious political debates in the UN on abortion and sexual orientation, and the multiple and sometimes divergent statements of independent experts and expert bodies in the UN human rights system on these and other sexual and reproductive rights issues. We offer no answers but seek to highlight the need for more investigation and self-reflection by advocates and scholars on how these forces operate and how to work with them.


Neurofibromatosis type 1 (Recklinghausen's disease). Neurologic and cognitive assessment with sibling controls.
Neurologic and cognitive function in neurofibromatosis type 1 (NF1) were assessed in a controlled pilot study of 13 pairs of siblings aged 6 to 27 years. One subject in each pair was affected with NF1, and the other, the control subject, was unaffected. Subjects with evidence of focal central nervous system disease were excluded. The 13 subjects with NF1 had no excess of mental retardation, attention-deficit disorder, or specific learning disorders (using Wilcoxon's Signed Rank Test and McNemar's Test for Symmetry). These subjects, however, had significantly higher scores for subtle neurologic abnormalities (21 vs 6) and significantly lower full-scale IQ scores (94 vs 105) than their unaffected siblings. The IQ scores of the affected subjects were not clustered at the lower end of the scale but showed a slight downward shift in distribution compared with those of their siblings. In addition, a visual-spatial orientation deficit was present in eight of nine affected subjects so evaluated. The findings suggest that subjects with NF1 have a widespread alteration of the brain during development that manifests as one or more specific types of neuropsychologic deficits.


Liver fibrosis progression in HIV-HCV-coinfected patients treated with distinct antiretroviral drugs and impact of pegylated interferon/ribavirin therapy.
Advanced liver fibrosis frequently develops in patients with chronic hepatitis C coinfected with HIV. Non-invasive techniques for staging liver fibrosis, such as transient elastometry, may allow both periodic monitoring and examination of large patient populations. A programme of liver fibrosis assessment using transient elastometry has been ongoing at our institution since 2004. All HIV-HCV-coinfected patients having ≥2 examinations separated by >18 months were included. Liver fibrosis progression (LFP) was defined as an increase in liver stiffness from <9.5 kPa (Metavir F0-F2) to >9.5 kPa (Metavir F3-F4), or an increase >30% in patients with baseline Metavir F3-F4. A total of 545 HIV-HCV-coinfected patients were analysed (mean age 41 years, 71% male, 81% intravenous drug users, mean body mass index 23.3 kg/m(2), 4.2% hepatitis B surface antigen-positive, 8.4% alcohol abuse, mean CD4(+) T-cell count 519 cells/μl). At baseline, 527 patients were on antiretroviral therapy, with the most frequent third drug being atazanavir (19.7%), efavirenz (15.9%), lopinavir (13.1%) or nevirapine (7.2%). A total of 99 (18%) patients experienced LFP during a mean (sd) follow-up of 70.9 (15.7) months. Use of protease inhibitors (OR 4.93, 95% CI 1.73, 14.0; P=0.03) and male gender (OR 5.12, 95% CI 1.37, 19.1; P=0.01) were associated with LFP. By contrast, the achievement of HCV clearance following pegylated interferon/ribavirin (PEG-IFN/RBV) therapy (OR 0.27, 95% CI 0.1, 0.79; P=0.02) was protective. Lopinavir exposure was significantly associated with LFP (OR 1.02, 95% CI 1.0, 1.04; P=0.03), whereas nevirapine was protective (OR 0.94, 95% CI 0.9, 0.99; P=0.02). The use of protease inhibitors, mainly lopinavir, is associated with increased LFP in HIV-HCV-coinfected patients. By contrast, nevirapine therapy and, particularly, HCV clearance with PEG-IFN/RBV significantly reduce LFP.


Prospective Phase II Trial of Prognostication by 68Ga-NOTA-AE105 uPAR PET in Patients with Neuroendocrine Neoplasms: Implications for uPAR-Targeted Therapy.
The clinical course for patients with neuroendocrine neoplasms (NENs) ranges from indolent to highly aggressive. Noninvasive tools to improve prognostication and guide decisions on treatment are warranted. Expression of urokinase plasminogen activator receptor (uPAR) is present in many cancer types and associated with a poor outcome. Therefore, using an in-house-developed uPAR PET tracer [68Ga]Ga-NOTA-Asp-Cha-Phe-D-Ser-D-Arg-Tyr-Leu-Trp-Ser-OH (68Ga-NOTA-AE105), we aimed to assess uPAR expression in NENs. We hypothesized that uPAR expression was detectable in a significant proportion of patients and associated with a poorer outcome. In addition, as uPAR-targeted radionuclide therapy has previously proven effective in preclinical models, the study would also indicate the potential for uPAR-targeted radionuclide therapy in NEN patients. Methods: In a prospective clinical phase II trial, we included 116 patients with NENs of all grades, of whom 96 subsequently had uPAR PET/CT performed with evaluable lesions. PET/CT was performed 20 min after injection of approximately 200 MBq of 68Ga-NOTA-AE105. uPAR target-to-liver ratio was used to define lesions as uPAR-positive when lesion SUVmax-to-liver SUVmean ratio was at least 2. Patients were followed for at least 1 y to assess progression-free and overall survival. Results: Most patients had small intestinal NENs (n = 61) and metastatic disease (n = 86). uPAR-positive lesions were seen in 68% (n = 65) of all patients and in 75% (n = 18) of patients with high-grade (grade 3) NENs. During follow-up (median, 28 mo), 59 patients (62%) experienced progressive disease and 28 patients (30%) died. High uPAR expression, defined as a uPAR target-to-liver ratio above median, had a hazard ratio of 1.87 (95% CI, 1.11-3.17) and 2.64 (95% CI, 1.19-5.88) for progression-free and overall survival, respectively (P < 0.05 for both). Conclusion: When 68Ga-NOTA-AE105 PET was used to image uPAR in patients with NENs, uPAR-positive lesions were seen in most patients, notably in patients with both low-grade and high-grade NENs. Furthermore, uPAR expression was associated with a worse prognosis. We suggest that uPAR PET is relevant for risk stratification and that uPAR may be a promising target for therapy in patients with NENs.


Effects of mentally retarded children's behavioral competence on nonretarded peers' behaviors and attitudes: toward establishing ecological validity in attitude research.
This study was designed to begin establishing the ecological validity of laboratory research indicating that nonretarded children's responses toward mentally retarded peers are positively affected by the retarded peers' competence in different areas. Groups of 3 children (1 retarded and 2 nonretarded fourth through sixth-graders) played a bean bag toss game in which the retarded child was either the best or an average player. Then children selected one of their two partners as a partner for a subsequent game and indicated their attitudes toward each peer and their attribution for each peer's performance. Retarded children who were the best players were selected as partners significantly more often than retarded children who were average players. Nonretarded children's attitudes toward retarded peers, however, were the same regardless of the retarded peers' performance. A relationship was also found between children's attributions of retarded peers' success to ability and their selection of the same peers as partners.


Physical and biologic impermeability of intestinal sutures in the first twenty-four hours after operations on the gastrointestinal tract.
The author has substantiated a concept that in the general notion of the impermeability of intestinal sutures, it is necessary to distinguish between physical impermeability (imperviousness of sutures to liquids and gases) and biologic impermeability (imperviousness of sutures to microbes and toxins). It was established in experiments on 165 dogs that infection that invaded peritoneum during operation disappeared from the peritoneal cavity within an hour of small-bowel and stomach surgery and within 1 1/2 hours of colon surgery. Then a period of sterility of peritoneum was observed, which lasted for 8 to 9 hours after operations on the stomach and the small bowel (p = 0.01) and for 4 to 7 hours after operations on the colon (p = 0.01). In the following hours the peritoneal cavity was infected through physically hermetic intestinal sutures. At the end of 24 hours the peritoneum of operated organs showed the presence of 10(4) to 10(6) intestinal microbes. Greater omentum, bowel loops, and their mesentery adhered to the suture in response to the infection through the suture. Complete covering of the suture with adjacent viscera prevented infection of the peritoneal cavity through the suture. The author suggests a well-grounded concept of the leading role of infection through a physically hermetic intestinal suture in the development of postoperative peritonitis and peritoneal adhesions.


GWAS-identified genetic variants associated with medication-assisted treatment outcomes in patients with opioid use disorder: a systematic review and meta-analysis protocol.
The burden of opioid use disorder (OUD) has been increasing in North America. Administration of medication-assisted treatments (MATs) for OUD on an individual-dose basis has been shown to affect patient responses to treatment, proving to be, on occasion, dangerous. A genetic basis has been identified for some MAT responses in a candidate gene context, but consensus has not been reached for any genome-wide significant associations. This systematic review aims to identify and assess any genetic variants associated with MAT patient outcomes at genome-wide significance. The databases searched by the authors will be: MEDLINE, Web of Science, EMBASE, CINAHL and Pre-CINAHL, GWAS Catalog, GWAS Central, and NIH Database of Genotypes and Phenotypes. A title and abstract screening, full-text screening, data extraction, and quality assessment will be completed in duplicate for each study via Covidence. Treatment outcomes of interest include continued opioid use or abstinence during treatment or at follow-up, time to relapse, treatment retention rates, opioid overdose, other substance use, comorbid psychiatric disorders, risk taking behaviors, MAT plasma concentrations, and mortality rates. Analysis methods applied, if appropriate, will include random effects meta-analysis with pooled odds ratios for all outcomes. Subgroup analyses will also be implemented, when possible. This systematic review can hopefully inform the direction of future research, aiding in the development of a safer and more patient-centered treatment. It will be able to highlight genome-wide significant variants that are replicable and associated with MAT patient outcomes. This systematic review protocol has been registered with the International Prospective Register of Systematic Reviews (PROSPERO) (registration ID CRD42020169121).


Technical and ethical problems associated with patch testing.
When relevant allergens are identified by patch testing, and patients can avoid them in their environment, improvement of dermatitis is the rule. Some forms of chronic dermatitis may not clear completely, but patients are usually satisfied with modification of their previously more severe problem. In 1981 when asked what he felt were the five most important advances in clinical dermatology during the 20th century, Dr. Marion B. Sulzburger, an icon in American dermatology, said that, "The increased use and usefulness of the patch test and the international standardization of test concentrations and methods" was number one. Those of us who are enthusiastic patch testers and fascinated by the evaluation of patients with irritant and allergic contact dermatitis would agree. It is a thrilling clinical experience to be able to tell a machinist that he need not stop his lifelong occupation, but instead will do fine if he will simply avoid the waterless hand cleanser he has been using which is preserved with glutaraldehyde. The woman whose facial dermatitis has embarrassed her for years and clears when she stops using the Quaternium-15 preserved moisturizer that you have identified in your patch testing is grateful to you forever, and again happy in her own life. In 1991, my research assistants, Patricia Norris and Mary Lou Belozer, and I studied 30 university hospital workers who answered our advertisement asking for individuals who believed they were troubled by their rubber gloves (unreported study). By evaluating these people through history, physical examination, and patch testing, we were able to prove glove relatedness in 14 of them. Nine of the 14 had contact urticaria to latex, and only 5 had allergic contact dermatitis to rubber glove ingredients. Fifteen of our patients had irritant dermatitis. In this study, none of the patients with allergic contact dermatitis to glove ingredients had contact urticaria. However, since that time, we have observed a number of patients who had both forms of allergic reaction. Three of our patients who presented with nummular (patchy) hand dermatitis also had contact urticaria to latex, but no positive patch tests. With latex glove avoidance, their dermatitis resolved; an example of how scratching urticaria can eventuate in longer lasting dermatitis in some people ("the itch that rashes"). The patients presented to their dermatologist with dermatitis, but their true initiating event was urticaria which lasted only hours. The gratifying part of this study was that patch testing and contact urticaria testing allowed us to discover the 5 patients with allergic contact dermatitis and the 9 patients with contact urticaria who could benefit from glove alternatives. We were also able to assure patients in the remainder of the group that their hand eczema was not glove induced, but rather was related to their wet work. In most instances, therapeutic intervention helped, but in several cases job changes were required. Patch testing, when done properly, produces exciting results. When done improperly, it confuses and misleads patients and results in embarrassment to physicians who cannot properly interpret their results. Should a physician choose to include patch testing in his or her evaluation of patients with contact dermatitis it is essential, in my view, that he or she have highly developed skills in the diagnosis and treatment of skin diseases, and that these physicians be elaborately trained in the techniques of application and the methods of interpretation of patch tests.


Excited-state properties of heteroleptic iridium(III) complexes bearing aromatic hydrocarbons with extended cores.
The synthesis, complete structural characterization, electrochemistry, and excited-state dynamics of a series of four bis-heteroleptic iridium(III) charge-transfer complexes composed of a single acac-functionalized and two ortho-metalated 2-phenylpyridine ligands. The formed iodophenyl complex (2) was used as a metallosynthon to introduce extended-core ethynyltolyl (3), ethynylpyrene (4), and ethynylperylene (5) residues into these structures projecting from the acac ancillary ligand. Static and dynamic photoluminescence along with ultrafast and conventional transient absorption measurements in conjunction with cyclic voltammetry were employed to elucidate the nature of the intramolecular energy-transfer processes occurring in the excited states of polychromophores 4 and 5 and are directly compared with those of model complexes 2 and 3. Upon charge-transfer excitation of these molecules, the long-lived triplet-state metal-to-ligand charge-transfer ((3)MLCT)-based photoluminescence readily observed in 2 and 3 (τ = 1 μs) is nearly quantitatively quenched, resulting from production of the associated triplet intraligand ((3)IL) excited states in 4 and 5 through intramolecular triplet-triplet energy transfer. The respective formation of the extended-core (3)*pyrenyl and (3)*perylenyl-localized excited states in 4 and 5 is confirmed by their ultrafast excited-state evolution, which ultimately generates features associated with these (3)IL excited states and their greatly extended excited-state lifetimes with respect to the parent complexes 2 and 3.


Effects of sodium valproate and diazepam on beta-endorphin, beta-lipotropin and cortisol secretion induced by hypoglycemic stress in humans.
Evidence that gamma-aminobutyric acid (GABA) and benzodiazepine receptors play a role in the inhibition of ACTH-cortisol secretion in humans has until now been drawn only from data indicating that sodium valproate, a GABA mimetic, and diazepam, a benzodiazepine, decrease hypothalamus-pituitary-adrenal (HPA) axis secretion in patients affected by pathological hypersecretion of the axis. Therefore, the present study investigated the effects, in the same healthy subjects, of sodium valproate or diazepam, on both basal and stress-stimulated concentrations of beta-endorphin (beta-EP), beta-lipotropin (beta-LPH) and cortisol. A single maximal dose of sodium valproate (400 mg) or diazepam (10 mg) did not significantly modify basal concentrations of beta-EP, beta-LPH and cortisol. On the other hand, in the same subjects, pretreatment with sodium valproate (20 mg X 3) or diazepam (10 mg X 2) blocked the increases in these hormones produced by hypoglycemic stress in all patients tested (p less than 0.01 vs. placebo at 45, 60 and 90 min after insulin injection), without affecting the decrease in blood glucose levels. The present data show that sodium valproate and diazepam inhibit stress-induced beta-EP, beta-LPH and cortisol secretion in humans, suggesting that endogenous GABA and benzodiazepine receptors participate in physiological mechanisms regulating the activity of the HPA axis.


Neurological trypanosomiasis in quinapyramine sulfate-treated horses--a breach of the blood-brain barrier?
Trypanosoma evansi infection typically produces wasting disease, but it can also develop into a neurological or meningoencephalitis form in equids. Trypanosomiasis in horses was treated with quinapyramine sulfate, and all the 14 infected animals were recovered clinically. After clinical recovery, four animals developed a neurological form of the disease at various intervals. Two of these animals treated with diminazene aceturate recovered temporarily. Repeated attempts failed to find the parasite in the blood or the cerebrospinal fluid (CSF), but all of the animals were positive in enzyme-linked immunosorbent assay. The calculation of the antibody index (AI) in the serum and the CSF and polymerase chain reaction (PCR) analysis of the CSF and brain tissue were carried out to confirm the neuro-infection. We found PCR and AI analyses of the CSF to be useful tools in the diagnosis of the neurological form of trypanosomiasis when the organism cannot be found in the blood or CSF. The increased albumin quotient is indicative of barrier leakage due to neuroinflammation. The biochemical changes in the CSF due to nervous system trypanosomiasis include increases in the albumin quotient, total protein, and urea nitrogen. It seems to be the first report on relapse of the nervous form of trypanosomiasis in equids even after quinapyramine treatment in endemic areas.


Anxiolytic and anti-inflammatory role of thymoquinone in arsenic-induced hippocampal toxicity in Wistar rats.
Arsenic (As) is a widely existing metalloid in the biosphere. Drinking water contamination by arsenic is a major route of human exposure, either by natural means or through industrial pollution. Numerous evidence form earlier reports suggest that arsenic exposure causes cerebral neurodegeneration which initiates behavioral disturbances concomitant to psychiatric disorders. Also, mood disorders in humans as well as in animals correlate with arsenic exposure; the present study is carried out to implore the neuroprotective potential of thymoquinone (TQ) in arsenic-stressed rats. TQ is an active component of Nigella sativa (Kalonji) seed oil. Arsenic exposure in the form of sodium arsenate (10 mg/kg/day; p.o) caused neurobehavioral deficits as evidenced by changes in locomotion and exploratory behavior in open-field and elevated plus maze tasks. Alongside this, arsenate also elevated hippocampal oxidative stress parameters like lipid peroxidation (TBARS) and protein carbonyl formation with a decrease in superoxide dismutase (SOD) and reduced glutathione (GSH) content. Genotoxicity assessment by Comet assay also showed prominent levels of DNA damage. Furthermore, arsenic also elevated hippocampal cytokine levels, TNF-α and INF-γ. However, TQ supplementation (2.5 and 5 mg/kg/day, p.o) preceded three days before arsenic administration, significantly attenuated arsenic-associated anxiogenic changes which majorly attributed to its antioxidant and anxiolytic potential. Also, TQ pre-treated rats expressed positive shifts in the hippocampal oxidative stress and cytokine levels with decreased DNA fragmentation. Thus, this study concludes that TQ might serve as a strong therapeutic agent for management of anxiety and depressive outcomes of arsenic intoxication.


Dispersal and reformation of acetylcholine receptor clusters of cultured rat myotubes treated with inhibitors of energy metabolism.
The effects of energy metabolism inhibitors on the distribution of acetylcholine receptors (AChRs) in the surface membranes of non-innervated, cultured rat myotubes were studied by visualizing the AChRs with monotetramethylrhodamine-alpha-bungarotoxin. Incubation of myotubes with inhibitors of energy metabolism causes a large decrease in the fraction of myotubes displaying clusters of AChR. This decrease is reversible, and is dependent on temperature, the concentration of inhibitor, and the duration of treatment. Cluster dispersal is probably not the result of secondary effects on Ca++ or cyclic nucleotide metabolism, membrane potential, cytoskeletal elements, or protein synthesis. Sequential observations of identified cells treated with sodium azide showed that clusters appear to disperse by movements of receptors within the sarcolemma without accompanying changes in cell shape. AChR clusters dispersed by pretreating cells with sodium azide rapidly reform upon removal of the inhibitor. Reclustering involves the formation of small aggregates of AChR, which act as foci for further aggregation and which appear to be precursors of large AChR clusters. Small AChR aggregates also appear to be precursors of clusters which form on myotubes never exposed to azide. Reclustering after azide treatment does not necessarily occur at the same sites occupied by clusters before dispersal, nor does it employ only receptors which had previously been in clusters. Cluster reformation can be blocked by cycloheximide, colchicine, and drugs which alter the intracellular cation composition.


Multiplexed Phosphoproteomic Study of Brain in Patients with Alzheimer's Disease and Age-Matched Cognitively Healthy Controls.
Alzheimer's disease (AD) is the most common neurodegenerative disorder caused by neuronal loss that results in cognitive and functional impairment. Formation of neurofibrillary tangles composed of abnormal hyperphosphorylation of tau protein is one of the major pathological hallmarks of AD. Importantly, several neurodegenerative disorders, including AD, are associated with abnormal protein phosphorylation events. However, little is known thus far on global protein phosphorylation changes in AD. We report a phosphoproteomics study examining the frontal gyrus of people with AD and age-matched cognitively normal subjects, using tandem mass tag (TMT) multiplexing technology along with immobilized metal affinity chromatography to enrich phosphopeptides. We identified 4631 phosphopeptides corresponding to 1821 proteins with liquid chromatography-mass spectrometry (MS)/MS analysis on an Orbitrap Fusion Lumos Tribrid mass spectrometer. Of these, 504 phosphopeptides corresponding to 350 proteins were significantly altered in the AD brain: 389 phosphopeptides increased whereas 115 phosphopeptides decreased phosphorylation. We observed significant changes in phosphorylation of known as well as novel molecules. Using targeted parallel reaction monitoring experiments, we validated the phosphorylation of microtubule-associated protein tau and myristoylated alanine-rich protein kinase C substrate (MARCKS) in control and AD (Control = 6, AD = 11) brain samples. In conclusion, our study provides new evidence on alteration of RNA processing and splicing, neurogenesis and neuronal development, and metabotropic glutamate receptor 5 (GRM5) calcium signaling pathways in the AD brain, and it thus offers new insights to accelerate diagnostics and therapeutics innovation in AD.


Cardiac transplantation.
Cardiac transplantation is now an accepted therapeutic option for patients with end-stage myocardial failure. Provided donor and recipient are appropriately selected and adequately matched, expected survival rates at one and five years are 85% and 65%, respectively. Two major challenges are encountered in clinical heart transplantation. The first is monitoring immunosuppression for adequate prevention of acute rejection and surveillance for side effects. The endomyocardial biopsy remains the gold standard for rejection surveillance, but since it is an invasive procedure which can only be performed at arbitrary time intervals, the search for non-invasive methods continues. The approach to immunosuppression currently practised by most centers is that of combination drug therapy, which allows low doses with decreased potential for side effects. At Stanford, immunosuppression is usually initiated with OKT3, corticosteroids, and cyclosporine, and maintained with a combination of steroids, cyclosporine, and azathioprine. The most frequently encountered complications include bacterial and opportunistic infections, cyclosporine nephrotoxicity, and malignancy. The second challenge is accelerated coronary disease, which has emerged as the major factor limiting long-term survival. It is usually clinically silent and often presents with sudden death, acute myocardial infarction, or progressive unexplained graft failure. Coronary arteriography is currently the only method for premorbid diagnosis, and retransplantation the only effective therapy.


Stimulation of hepatic RNA synthesis by phenobarbital.
Administration of phenobarbital in rats is followed by an in vitro stimulation of hepatic ribosomal protein synthesis. The increase of polyribosomes which is also observed is paralleled by a phenobarbital-dependent enhancement of uridine incorporation into ribonucleic acids. Sedimentation of either ribosomes or ribosomal RNA extracted from ribosomal subunits through sucrose gradients reveals stimulation by phenobarbital of 3H-uridine incorporation into messenger RNA as well as into ribosomal RNA. These experiments together with the findings of stimulation of RNA polymerase activity and blockade of the effects of phenobarbital by the administration of Actinomycin D indicate a primary action of phenobarbital on RNA synthesis.


The load of amyloid-β oligomers is decreased in the cerebrospinal fluid of Alzheimer's disease patients.
Amyloid-β (Aβ) oligomers are heterogeneous and instable compounds of variable molecular weight. Flow cytometry and fluorescence resonance energy transfer (FRET)-based methods allow the simultaneous detection of Aβ oligomers with low and high molecular weight in their native form. We evaluated whether an estimate of different species of Aβ oligomers in the cerebrospinal fluid (CSF) with or without dilution with RIPA buffer could be more useful in the diagnosis of Alzheimer's disease (AD) than the measurement of Aβ42 monomers, total tau (t-tau), and phosphorylated tau (p-tau). Increased t-tau (p < 0.01) and p-tau (p < 0.01), and decreased Aβ42 (p < 0.01), were detected in the CSF of patients with AD (n = 46), compared to patients with other dementia (OD) (n = 35) or with other neurological disorders (OND) (n = 56). In native CSF (n = 137), the levels of Aβ oligomers were lower (p < 0.05) in AD than in OD and OND patients; in addition, the ratio Aβ oligomers/p-tau was lower in AD than in OD (p < 0.01) and OND (p < 0.05) patients, yielding a sensitivity of 75% and a specificity of 64%. However, in CSF diluted with RIPA (n = 30), Aβ oligomers appeared higher (p < 0.05) in AD than in OND patients, suggesting they become partially disaggregated and more easily detectable after RIPA. In conclusion, FRET analysis in native CSF is essential to correctly determine the composition of Aβ oligomers. In this experimental setting, the simultaneous estimate of low and high molecular weight Aβ oligomers is as useful as the other biomarkers in the diagnosis of AD. The low amount of Aβ oligomers detected in native CSF of AD may be inversely related to their levels in the brain, as occurs for Aβ monomers, representing a biomarker for the amyloid pathogenic cascade.


Development and evaluation of 4 loop-mediated isothermal amplification assays to detect mastitis-causing bacteria in bovine milk samples.
Farmers prefer fast, sensitive, and on-site tests for treatment decisions on mastitis. Due to the time to results of the currently available diagnostic tools, these are rarely used for that purpose. Genotypic tests that do not require a growth step may be suitable for on-site testing, for example loop-mediated isothermal amplification (LAMP), which has been described as a sensitive test that can be used on-site. Therefore, this study aimed to develop and evaluate LAMP assays for the detection of a subset of mastitis-causing pathogens, Escherichia coli, Klebsiella pneumoniae, Staphylococcus aureus, and Streptococcus spp., in milk from cows with clinical mastitis. Furthermore, a generic nucleic acid lateral flow immunoassay (NALFIA) was evaluated as a potential on-site readout of the LAMP assays. For each assay of LAMP and NALFIA, the limit of detection and analytical specificity were determined using isolates, and the diagnostic specificity was determined using selected samples with known etiology. In addition, the diagnostic specificity of LAMP was determined using field samples with unknown etiology at testing. Bacteriological culture with identification by mass spectrometry was used as a reference method. The 4 assays had a kappa ≥0.73 with the reference method when testing the selected samples, but ≥0.47 when testing field samples. After correcting for prevalence, kappa was ≥0.80 for the E. coli, K. pneumoniae, and Staph. aureus assays. The Streptococcus spp. assay had a kappa of 0.47 (0.48 after correction) with the reference method, probably caused by the assay broadly targeting a genus instead of a particular species. The NALFIA readout was found to have kappa ≥0.81 for the E. coli, Staph. aureus, and Streptococcus spp. assays at a generic runtime, but for the K. pneumoniae assay a shorter runtime could be used. In conclusion, LAMP is a promising method for fast on-site tests for mastitis-causing pathogens if the current elaborate method for sample preparation is replaced by a simplified protocol. The NALFIA is an easy and reliable readout for on-site use, with the observation that for the current assay designs a generic runtime is not yet possible for the chosen set of pathogens. If associated with a simple and fast sample preparation protocol, the combination of LAMP and NALFIA has the potential to enable fast and reliable on-site testing of clinical mastitis milk samples.


QUANTITATIVE ANALYSIS OF RESPONSES FROM LATERAL-LINE NERVES OF FISHES. II.
1. The lateral-line nerves of trout as well as those of catfish are found to discharge impulses spontaneously at a high frequency. 2. The frequency of nerve impulse discharge is measured as a function of the number of participating receptor groups (lateral-line sense organs). A quantitative analysis is made of the contribution to the total response made by each group of sense organs. 3. An analysis of the variability of the response is presented which makes it possible to estimate quantitatively the longitudinal extent of damage to the neuromasts due to surgical manipulation. 4. A method is described for recording the response of a single nerve fiber in the lateral-line trunk. 5. The frequency of the spontaneous discharge from the lateral-line nerve trunk when plotted as a function of temperature according to the Arrhenius equation yields a temperature characteristic of approximately 5000 calories. 6. The variability of the frequency of response as a function of temperature indicates the existence of temperature thresholds for the spontaneous activity of the neuromasts. 7. A possible basis for the spontaneous activity is considered. It is pointed out that the lateral-line system may serve as a model of the Purkinje cells of the cerebellum.


A high observed substitution rate in the human mitochondrial DNA control region.
The rate and pattern of sequence substitutions in the mitochondrial DNA (mtDNA) control region (CR) is of central importance to studies of human evolution and to forensic identity testing. Here, we report a direct measurement of the intergenerational substitution rate in the human CR. We compared DNA sequences of two CR hypervariable segments from close maternal relatives, from 134 independent mtDNA lineages spanning 327 generational events. Ten substitutions were observed, resulting in an empirical rate of 1/33 generations, or 2.5/site/Myr. This is roughly twenty-fold higher than estimates derived from phylogenetic analyses. This disparity cannot be accounted for simply by substitutions at mutational hot spots, suggesting additional factors that produce the discrepancy between very near-term and long-term apparent rates of sequence divergence. The data also indicate that extremely rapid segregation of CR sequence variants between generations is common in humans, with a very small mtDNA bottleneck. These results have implications for forensic applications and studies of human evolution.


Envelope glycoproteins of HIV-1 interfere with T-cell-dependent B cell differentiation: role of CD4-MHC class II interaction in the effector phase of T cell help.
T-cell-dependent B cell differentiation involves two phases: an inductive phase of T cell activation followed by an effector phase, which involves stimulation of B cells by activated T cells. We have previously demonstrated that anti-CD3 mAb and antigen-induced T-cell-dependent B cell functions are inhibited by HIV-1 envelope glycoprotein, gp120, at the inductive phase of T-cell-dependent B cell response. In this study we have investigated whether gp120 also inhibits the effector phase of interactions involved in T-cell-dependent-B cell differentiation response. For these studies, CD4+ T cells were first activated with antigen or pokeweed mitogen, cultured with soluble HIV-gp120 or medium for 2 hr, and washed. Coculture of gp120-treated preactivated T cells with autologous B cells resulted in impairment of IgG secretion, but did not affect IgM secretion significantly. The IgG secretion was restored by the addition of PMA (activator of protein kinase C) or forskolin (activator of adenylate cyclase), but not by the addition of ionomycin (inducer of intracellular calcium) to the T plus B cell cultures. A similar pattern of Ig secretion (IgM, no IgG) was observed with B cells of a patient with bare lymphocyte syndrome, indicating a requirement for MHC class II molecule interaction with T cells. These studies suggest that the effector phase of T-B cell interactions are impaired by gp120, and that the mechanism involves a signal transducing event(s), which is dependent upon cyclic AMP and/or protein kinase C. Furthermore, these latter reactions occur subsequent to T-B cell contact-dependent interactions at the effector phase, which involve MHC class II molecules on B cells and CD4 molecules on T cells.


A critical examination of home care: end of life care as an illustrative case.
Drawing on end of life care as an illustrative case, this paper critically examines the provision of care in the home, identifying a number of inherent tensions. For 60 years the hospital has been the preferred site of care. However, the UK caring division of labour is currently undergoing a process of (re)domestication and the provision of home care is increasingly regarded as a 'gold standard' for the organisation of care, in institutional and domestic contexts. In this paper we argue that while 'home care' policies serve a range of professional and political agendas, they contain unacknowledged contradictions and strains, creating challenges for both family and professional carers. The realities of home care are examined through reconceptualising qualitative data generated from three research projects concerned with dying in the community. We argue that, whilst previous work has highlighted the burdens the redomestication of care places upon carers, home care philosophies and policies have led to over-romanticised notions of care which privilege the value of caring relationships without acknowledging the dynamic interaction of such social relationships with the actual work of caring. Moreover, such policy trends have created a nexus of social expectations and obligations for which modern society is unprepared. With reference to both end of life care, and home care more widely, we argue that health care planners and professionals need to think more critically about the way care is delivered. Home is not merely about a physical space, but the social and emotional relationships therein. Good 'home care,' characterised by attention to patient-centred needs and flexible in design and scope, does not have to be located within the private sphere; relationships may actually be maintained and nurtured by enabling people to have a realistic choice of care in an institution.


[Anodic stripping voltammetry of promethazine hydrochloride with Nafion coated glassy carbon electrode].
This paper reports the determination of promethazine hydrochloride with Nafion coated glassy carbon electrode by anodic stripping voltammetry. There is a linear relationship between the concentration and peak height in the range of 4 x 10(-8)-5 x 10(-5) mol/L and 8 x 10(-5)-1 x 10(-3) mol/L for promethazine hydrochloride. The raw material and tablets of promethazine hydrochloride have been analysed and good results were obtained.


Blood pressure tracking as an indicator of hypertension risk.
The significance of high blood pressure to cardio- and cerebrovascular disease is well recognized. The associated mortality and morbidity risks call for multiple approaches to control the development of high blood pressure. Recent studies have shown that precursors of cardiovascular disorders may be apparent from a young age, and that these can persist and contribute to disease in later life. With regard to blood pressure, the phenomenon of persisting at the same rank has been referred to as tracking. This allows for the possible identification of high-risk groups, for whom early intervention can be initiated. In this paper, some of the findings of studies on blood pressure tracking are reviewed.


Chromosomal abnormalities associated with cyclopia and synophthalmia.
At the present time, essentially all known facts concerning cyclopia are consistent with some chromosomal disease, including clinical features of the pregnancy (fetal wastage, prematurity, intrauterine growth retardation, maternal age factor, complications of pregnancy), the generalized developmental abnormalities, specific ocular dysgenesis, by the high incidence of chromosomal abnormality already demonstrated, and the possibility of error in those cases of cyclopia with normal chromosomes. Even if chromosomal aberrations represent only one group of several different etiologic factors leading to cyclopia, at the present time chromosomal errors would seem to be the most common cause of cyclopia now recognized. Further studies will establish or disprove a chromosomal error in those instances which are now considered to be the result of an environmental factor alone or those with apparent familial patterns of inheritance. This apparent diverse origin of cyclopia can be clarified if future cyclopic specimens are carefully investigated. The evaluation should include a careful gross and microscopic examination of all organs, including the eye, and chromosome banding studies of all organs, including the eye, and chromosome banding studies of at least two cyclopic tissues. Then the presence or absence of multiple causative factors can be better evaluated.


Dipolar response of hydrated proteins.
The paper presents an analytical theory and numerical simulations of the dipolar response of hydrated proteins in solution. We calculate the effective dielectric constant representing the average dipole moment induced at the protein by a uniform external field. The dielectric constant shows a remarkable variation among the proteins, changing from 0.5 for ubiquitin to 640 for cytochrome c. The former value implies a negative dipolar susceptibility, that is a dia-electric dipolar response and negative dielectrophoresis. It means that ubiquitin, carrying an average dipole of ≃240 D, is expected to repel from the region of a stronger electric field. This outcome is the result of a negative cross-correlation between the protein and water dipoles, compensating for the positive variance of the intrinsic protein dipole in the overall dipolar susceptibility. In contrast to the neutral ubiquitin, charged proteins studied here show para-electric dipolar response and positive dielectrophoresis. The study suggests that the dipolar response of proteins in solution is strongly affected by the coupling of the protein surface charge to the hydration water. The protein-water dipolar cross-correlations are long-ranged, extending ~2 nm from the protein surface into the bulk. A similar correlation length of about 1 nm is seen for the electrostatic potential produced by the hydration water inside the protein. The analysis of numerical simulations suggests that the polarization of the protein-water interface is highly heterogeneous and does not follow the standard dielectric results for cavities carved in dielectrics. The polarization of the water shell gains in importance, relative to the intrinsic protein dipole, at high frequencies, above the protein Debye peak. The induced interfacial dipole can be either parallel or antiparallel to the protein dipole, depending on the distribution of the protein surface charge. As a result, the high-frequency absorption of the protein solution can be either higher or lower than the absorption of water. Both scenarios have been experimentally observed in the THz window of radiation.


TranscriptomeBrowser: a powerful and flexible toolbox to explore productively the transcriptional landscape of the Gene Expression Omnibus database.
As public microarray repositories are constantly growing, we are facing the challenge of designing strategies to provide productive access to the available data. We used a modified version of the Markov clustering algorithm to systematically extract clusters of co-regulated genes from hundreds of microarray datasets stored in the Gene Expression Omnibus database (n = 1,484). This approach led to the definition of 18,250 transcriptional signatures (TS) that were tested for functional enrichment using the DAVID knowledgebase. Over-representation of functional terms was found in a large proportion of these TS (84%). We developed a JAVA application, TBrowser that comes with an open plug-in architecture and whose interface implements a highly sophisticated search engine supporting several Boolean operators (http://tagc.univ-mrs.fr/tbrowser/). User can search and analyze TS containing a list of identifiers (gene symbols or AffyIDs) or associated with a set of functional terms. As proof of principle, TBrowser was used to define breast cancer cell specific genes and to detect chromosomal abnormalities in tumors. Finally, taking advantage of our large collection of transcriptional signatures, we constructed a comprehensive map that summarizes gene-gene co-regulations observed through all the experiments performed on HGU133A Affymetrix platform. We provide evidences that this map can extend our knowledge of cellular signaling pathways.


A mathematical model of diurnal variations in human plasma melatonin levels.
Studies in animals and humans suggest that the diurnal pattern in plasma melatonin levels is due to the hormone's rates of synthesis, circulatory infusion and clearance, circadian control of synthesis onset and offset, environmental lighting conditions, and error in the melatonin immunoassay. A two-dimensional linear differential equation model of the hormone is formulated and is used to analyze plasma melatonin levels in 18 normal healthy male subjects during a constant routine. Recently developed Bayesian statistical procedures are used to incorporate correctly the magnitude of the immunoassay error into the analysis. The estimated parameters [median (range)] were clearance half-life of 23.67 (14.79-59.93) min, synthesis onset time of 2206 (1940-0029), synthesis offset time of 0621 (0246-0817), and maximum N-acetyltransferase activity of 7.17(2.34-17.93) pmol x l(-1) x min(-1). All were in good agreement with values from previous reports. The difference between synthesis offset time and the phase of the core temperature minimum was 1 h 15 min (-4 h 38 min-2 h 43 min). The correlation between synthesis onset and the dim light melatonin onset was 0.93. Our model provides a more physiologically plausible estimate of the melatonin synthesis onset time than that given by the dim light melatonin onset and the first reliable means of estimating the phase of synthesis offset. Our analysis shows that the circadian and pharmacokinetics parameters of melatonin can be reliably estimated from a single model.


The impact of burial period on compact bone microstructure: Histological analysis of matrix loss and cell integrity in human bones exhumed from tropical soil.
Human bone histological analysis is a useful tool to assess post mortem diagenesis and to predict successful nuclear DNA typing of forensic material. This study is part of a series of studies developed by the authors intended to improve the understanding of post mortem diagenesis and to develop applications for DNA analysis of skeletal species from tropical soils, in order to optimize genetic and anthropological protocols. The aim of this study was to analyze the impact of burial period on the integrity of exhumed compact bone microstructure from tropical climate. In fragments of exhumed human femora from 39 individuals from the same cemetery (exhumed group) and 5 fresh femora from routine autopsies (control group), sections stained by hematoxylin-eosin were analyzed in order to measure bone microstructural integrity. We found that bone integrity index in exhumed group was negatively influenced by the period of burial (r = -0.37, p < 0.05) and highly significantly decreased (p < 0.0001) in comparison to control group. The period of burial and nitric acid decalcification time was positively correlated (r = 0.51; p < 0.01), leading to imply a bone petrification process during inhumation. Exhumed group showed higher level of matrix bone loss (p < 0.001), as expected, and 87% of cases analyzed were "tunneled" as described by Hackett. Bone integrity index and bone matrix tend to decrease in bones buried in tropical soil between 8-14 years of inhumation. This period is short if we consider cases in which there are preserved bones interred for longer periods in other environments. These data must be considered in cases where genetic identification of exhumed skeletons from tropical environment is required. The diagenesis in these bones and the variations of results found are discussed, clarifying some challenges for forensic laboratories, especially in DNA analysis.


Efficacy of irrigation systems on penetration of sodium hypochlorite to working length and to simulated uninstrumented areas in oval shaped root canals.
To assess the ability of sodium hypochlorite (NaOCl) to penetrate simulated lateral canals and to reach working length (WL) when using the self-adjusting file (SAF). Seventy single-rooted teeth with oval-shaped canals were used. Upon access, presence of a single canal was confirmed by direct visualization under a dental-operating microscope. Canal length and patency were obtained using a size 10 K-file and root length standardized to 18 mm. Pre-enlargement was restricted to the coronal one-third. The apical size of each canal was gauged at WL and samples larger than size 30 were excluded. Canals were instrumented for 5 min using the SAF system while delivering a total of 20 mL of 5.25% NaOCl and 5 mL of 17% EDTA. Then, the apical diameters were standardized to size 35 using hand files. Four hundred and twenty simulated lateral canals were then created during the clearing process and roots coated with wax to create a closed system. All samples were then cleared and randomly assigned to four experimental groups: 1 (n = 15) positive pressure; 2 (n = 15) SAF without pecking motion; 3 (n = 15) SAF with pecking motion; 4 (n = 15) apical negative pressure (ANP) irrigation and (n = 10) control groups. Samples were scored on the basis of the ability of the contrast solution to reach WL and permeate into the simulated lateral canals to at least 50% of the total length. The Kruskal-Wallis test was used to analyse irrigant penetration and the Tukey test to determine statistical differences between groups (P < 0.05). All samples irrigated with ANP were associated with irrigant penetration to WL (Table 1). The differences between group 4 (ANP) and all other groups were significant in penetration to WL (P < 0.05). The pecking motion allowed for further penetration of the irrigant when using the SAF system but failed to irrigate at WL. None of the experimental groups demonstrated predictable irrigation of simulated lateral canals. In this laboratory model, ANP was the only delivery system capable of irrigating consistently to full WL. None of the systems tested produced complete irrigation in artificial lateral canals.


Retirement: What will you do? A narrative inquiry of occupation-based planning for retirement: Implications for practice.
To examine, using a retrospective narrative study, the factors influencing four older people's decision to plan for the activities they would undertake once retired, the planning process undertaken and their subsequent experience of retirement. Semi-structured interviews were conducted to examine the factors influencing older people's decision to commence pre-retirement planning, the planning process undertaken and their experience of retirement. In keeping with narrative inquiry, paradigmatic-type narrative analysis led to the development of categories and subsequent themes to reveal the participants' experiences of these issues. Three themes: environmental influences, the planning process and retirement experiences: the outcome of planning, were derived from the participants' narratives. Overall, retirement activities that were continued or initiated led to a positive experience even when these planned activities were temporarily interrupted or altered. Planning for future years in retirement and older age also continued. Occupational therapists have a unique understanding of the centrality of occupation to health and wellbeing. Thus occupational therapists are well-positioned to assist people identify, plan and engage in meaningful occupations outside work in retirement.


A comparison of volume growth during bud and mycelium formation in Candida albicans: a single cell analysis.
When stationary phase cells of the dimorphic yeast Candida albicans are diluted into fresh medium at pH 4.5 (low pH), they synchronously form ellipsoidal buds, but when diluted into the same medium at pH 6.7 (high pH), they synchronously form elongate mycelia. Using a perfusion chamber to monitor single cells, we have compared the rates of volume growth between budding and mycelium-forming cells. Results are presented which demonstrate that: (1) after release from stationary phase into medium of low or high pH, each original sphere grows in volume to the time of initial evagination, but does not grow subsequently; (2) successive budding on the original mother cell occurs without interruption resulting in continuous volume growth; however, an interruption in volume growth of the initial bud (B1) occurs before it in turn evaginates; and (3) the rate of volume growth of the first bud at low pH is identical to the rate of volume growth of the mycelium at high pH even though the surface to volume ratios are quite different. The last result is unexpected and is therefore considered in relation to cell wall deposition.


The relationship between postural control and fine manual dexterity.
The influence of the development of postural control of the trunk and center of the body on the development of fine manual dexterity ability is a common assumption. The objectives of this study were: (1) to examine the relationship between the adult version of the Nine-Hole Peg Test and the "children's version" in order to establish construct validity for the latter, and (2) to investigate the relationship between postural control and fine motor performance of children developing typically aged five to six. The study sample included 47 children, 25 males and 22 females, with a mean age of 5 years and 8 months and a standard deviation of 3.8 months. Two tests were administered: (1) The balance subtest of the Bruininks-Oseretsky Test of Motor Proficiency (BOTMP; Bruininks, 1978); and (2) the Nine-Hole Peg Test (NHPT; Kellor, Frost, Silberg Iversen & Cumming, 1971), as well as the "children's version" of rotation and translation tasks (Case-Smith, 1993, 1995). The adult version of the NHPT revealed significant moderate correlations with most of the BOTMP tasks for both hands. Secondary results of the study support the concurrent validity of the two "children's version" tasks. Results show low to moderate correlations between performance on the NHPT subtests and the BOTMP subtest tasks, with significant correlations ranging from -.31 to -.47. Given the negative and low correlations obtained in this study, it is suggested that more extensive longitudinal research examine the relationship between postural control and fine motor manual dexterity performance.


[What's new in clinical dermatology?].
Dermatoses have been better characterized as hypochromic vitiligo in subjects with dark skin or the acquired racquet nails associated with hyperparathyroidism. The innate immunity has a central role in the new classification of auto inflammatory diseases, neutrophilic dermatoses, neutrophilic urticaria or pseudo-neutrophilic urticaria. Comorbidities have been studied e.g. cardiovascular co morbidities for psoriasis, IBD associated with hidradenitis suppurativa or neurological disorders associated with pemphigoid. Bullous pemphigoid could be renamed as cutaneous pemphigoid, it can be induced by drugs especially gliptins. Genetic predispositions are analyzed in auto inflammatory diseases, psoriasis (HLA), drug eruptions (HLA or cytokines). Telaprevir often induces rashes, which can be severe but other treatments against hepatitis C as interferon can also induce debilitating rashes, some eligible for drug tolerance induction. European guidelines for the definition, classification, diagnosis of chronic spontaneous urticaria have been published. Severe cutaneous adverse drug reactions may be associated with severe systemic symptoms such as organ involvement in AGEP or the occurrence of a shock in DRESS. Allover Europe, there is now an epidemic of contact allergies to methylisothiazolinone (MIT), contained in cosmetics, wet wipes but also in paints. MIT should be tested at 2.000 ppm in water, included in the standard series for patch tests. Its role has to be evoked in any case localized on the face or hand, airborne or generalized eczema. Among infectious diseases, skin manifestations due to Parvovirus have been specified, a febrile rash can be related to dengue infection or to Zika virus.


Tissue-specific expression and cDNA cloning of small nuclear ribonucleoprotein-associated polypeptide N.
Sera from some patients with systemic lupus erythematosus and other autoimmune diseases have antibodies against nuclear antigens. An example is anti-Sm sera, which recognize proteins associated with small nuclear RNA molecules [small nuclear ribonucleoprotein (snRNP) particles]. In this paper anti-Sm sera were used to probe immunoblots of various rat tissues. A previously unidentified Mr 28,000 polypeptide was recognized by these anti-Sm sera. This polypeptide, referred to as "N," is expressed in a tissue-specific manner, being most abundant in rat brain, less so in heart, and undetectable in the other tissues examined. Immunoprecipitation experiments using antibodies directed against the cap structure of small nuclear RNAs have demonstrated that N is a snRNP-associated polypeptide. Anti-Sm serum was also used to isolate a partial cDNA clone (lambda rb91) from a rat brain phage lambda gt11 cDNA expression library. On RNA blots, the 450-base-pair cDNA insert of this clone hybridized to a 1600-nucleotide mRNA species with an identical tissue distribution to N, suggesting that lambda rb91 encodes at least part of N. A longer cDNA clone was obtained by rescreening the library with lambda rb91. In vitro transcription and subsequent translation of this subcloned, longer insert (pGMA2) resulted in a protein product with the same electrophoretic and immunological properties as N, confirming that pGMA2 encodes N. The tissue distribution of N and the involvement of snRNP particles in nuclear pre-mRNA processing may imply a role for N in tissue-specific pre-mRNA splicing.


The relationships among consumption of fruits, tooth loss and obesity.
to explore whether consumption of fruits attenuates the relationship between tooth loss and obesity among adult Americans. data from the Behavioral Risk Factor Surveillance System 2012, a cross-sectional survey. 20,103 adults aged 18 and over who participated in the Behavioral Risk Factor Surveillance System 2012, a nationally representative survey of non-institutionalized adult American. Obesity (Body Mass Index ≥ 30 kg/m²). Logistic regression analysis was used to assess the association between obesity and number of missing teeth adjusting for consumption of fruits socio-demographic factors, physical activity, diabetes and self-rated general health. The prevalence of obesity was higher among adults with ⟩ 6 missing teeth and among those with fruit intakes less than once a day. Individuals with ⟩ 6 missing teeth were at higher risk of obesity with odds ratios 1.25 (95% CI: 1.04, 1.50), the relationship attenuated after adjusting for fruit consumption. Frequent consumption of fruits was inversely associated with each of obesity and number of missing teeth. The findings imply a mediating role of fruit consumption in the relationship between tooth loss and obesity.


New technology to assess sleep apnea: wearables, smartphones, and accessories.
Sleep medicine has been an expanding discipline during the last few decades. The prevalence of sleep disorders is increasing, and sleep centers are expanding in hospitals and in the private care environment to meet the demands. Sleep medicine has evidence-based guidelines for the diagnosis and treatment of sleep disorders. However, the number of sleep centers and caregivers in this area is not sufficient. Many new methods for recording sleep and diagnosing sleep disorders have been developed. Many sleep disorders are chronic conditions and require continuous treatment and monitoring of therapy success. Cost-efficient technologies for the initial diagnosis and for follow-up monitoring of treatment are important. It is precisely here that telemedicine technologies can meet the demands of diagnosis and therapy follow-up studies. Wireless recording of sleep and related biosignals allows diagnostic tools and therapy follow-up to be widely and remotely available. Moreover, sleep research requires new technologies to investigate underlying mechanisms in the regulation of sleep in order to better understand the pathophysiology of sleep disorders. Home recording and non-obtrusive recording over extended periods of time with telemedicine methods support this research. Telemedicine allows recording with little subject interference under normal and experimental life conditions.


Are demographic and socioeconomic factors predictive for perceived tinnitus impairment?
Patients suffering from chronic tinnitus were analyzed to examine whether tinnitus impairments can be predicted by demographic and socioeconomic factors. For this purpose, subjective tinnitus complaints were measured in 153 patients using the tinnitus impairment questionnaire (THI-12) that distinguishes between emotional-cognitive and functional-communicative factors. Age, gender, marital status, and education levels were assessed and treated as predictive variables. In computing canonical correlations, only the level of education served as a significant emotional-cognitive predictor for tinnitus impairment. Patients with lower education demonstrated impairments in the emotional-cognitive domain that were more marked than those in higher-educated people. The results indicated that demographic and socioeconomic variables have no predictive value for tinnitus impairments, with the exception of a person's level of education.


Vaccination Hesitancy and Conspiracy Beliefs in the UK During the SARS-COV-2 (COVID-19) Pandemic.
Conspiracy beliefs about vaccination along with vaccination hesitancy are threats to achieving population immunity during the SARS-COV-2 (COVID-19) pandemic. This longitudinal study aimed to clarify the association between these and non-monetary incentives to vaccination in the UK. Data were collected at three points: (1) before and (2) after the development of a vaccine and (3) after the vaccination programme was underway. At Time 1, participants completed measures of general and COVID-19-specific concerns about vaccination and belief in conspiracy theories. At times 2 and 3, participants reported their intentions whether or not to have the SARS-CoV-2 vaccine. Those who were hesitant provided qualitative comments about what might change their decision. Vaccination hesitancy decreased between times 1 (54%) and 3 (13%). There were small effects of conspiracy beliefs on vaccine hesitancy, but only at time 1. Most concerns and reported incentives were related to safety, although at time 2, incentives included endorsement by trusted public figures. By time 3, only a minority of participants (N = 18) were adamantly against vaccination, stating that nothing would change their minds. Vaccination hesitancy declined in the UK during the course of the study. However, concerns about vaccine safety remained and could jeopardise the vaccination programme should any adverse effects be reported. Conspiracy beliefs seem to play only a minor role in hesitancy and may continue to decrease in importance with a successful vaccination programme. Understanding motivations behind vaccination hesitancy is vital if we are to achieve population immunity.


[Treatment of idiopathic pulmonary arterial hypertension].
In the past 5-10 years, drug treatment of idiopathic pulmonary arterial hypertension has evolved considerably. Experience and results from use of such updated treatment in Norway has not been reported. 32 patients newly diagnosed with idiopathic pulmonary arterial hypertension, were consecutively assessed with respect to hemodynamics and physical capacity. The results after three months were compared with those after 12 months. Observed survival was compared with estimated survival from the time when only conventional treatment was available. The patients (78% women) were 42 ± 14 years, had dyspnea in NYHA class 2.9 ± 0.4 and a maximal oxygen uptake of 12.0 ± 3.9 ml/kg/min (37 ± 13% of the expected). Updated treatment led to significantly improved hemodynamics and physical capacity, which persisted during follow-up. During 43 ± 31 months follow-up, seven patients died while two underwent bilateral lung transplantation. Observed transplantation-free survival was 81% after one, two and three years, while that for estimated transplantation-free survival was 70%, 58% and 49% respectively. Treatment of idiopathic pulmonary arterial hypertension with updated treatment improves hemodynamics and thereby symptoms. Mortality remains high, but is probably lower than it was when only conventional treatment was available.


Energy and coincidence time resolution measurements of CdTe detectors for PET.
We report on the characterization of 2 mm thick CdTe diode detector with Schottky contacts to be employed in a novel conceptual design of PET scanner. Results at -8°C with an applied bias voltage of -1000 V/mm show a 1.2% FWHM energy resolution at 511 keV. Coincidence time resolution has been measured by triggering on the preamplifier output signal to improve the timing resolution of the detector. Results at the same bias and temperature conditions show a FWHM of 6 ns with a minimum acceptance energy of 500 keV. These results show that pixelated CdTe Schottky diode is an excellent candidate for the development of next generation nuclear medical imaging devices such as PET, Compton gamma cameras, and especially PET-MRI hybrid systems when used in a magnetic field immune configuration.


Comparing the costs of alternative models of end-of-life care.
This study explores the financial consequences of decreased acute care utilization and expanded community-based care for patients at the end of life in England. A Markov model based on cost and utilization data was used to estimate the costs of care for cancer and organ failure in the last year of life and to simulate reduced acute care utilization. We estimated at pounds 1.8 billion the cost to the taxpayer of care for the 127,000 patients dying from cancer in 2006. The equivalent cost for the 30,000 people dying from organ failure was pounds 553 million. Resources of pounds 16 to pounds 171 million could be released for cancer. People generally prefer to die outside hospital. Our results suggest that reducing reliance on acute care could release resources and better meet peoples' preferences. Better data on the cost-effectiveness of interventions are required. Similar models would be useful to decision-makers evaluating changes in service provision.


Hepatitis Delta Virus Genome RNA Synthesis Initiates at Position 1646 with a Nontemplated Guanosine.
Hepatitis delta virus (HDV) is a significant human pathogen that causes acute and chronic liver disease; there is no licensed therapy. HDV is a circular negative-sense single-stranded RNA (ssRNA) virus that produces three RNAs in infected cells, genome, antigenome, and mRNA; the latter encodes hepatitis delta antigen (HDAg), the viral protein. These RNAs are synthesized by host DNA-dependent RNA polymerase acting as an RNA-dependent RNA polymerase. Although HDV genome RNA accumulates to high levels in infected cells, the mechanism by which this process occurs remains poorly understood. For example, the nature of the 5' end of the genome, including the synthesis start site and its chemical composition, is not known. Analysis of this process has been challenging because the initiation site is part of an unstable precursor in the rolling-circle mechanism by which HDV genome RNA is synthesized. In this study, circular HDV antigenome RNAs synthesized in vitro were used to directly initiate HDV genome RNA synthesis in transfected cells, thus enabling the detection of the 5' end of the genome RNA. The 5' end of this RNA is capped, as expected for a polymerase II product. Initiation begins at position 1646 on the genome, which is located near the loop end proximal to the start site for HDAg mRNA synthesis. Unexpectedly, synthesis begins with a guanosine that is not conventionally templated by the HDV RNA. IMPORTANCE Hepatitis delta virus (HDV) is a unique virus that causes severe liver disease. It uses host RNA polymerase II to copy its circular RNA genome in a unique and poorly understood process. Although the virus RNA accumulates to high levels within infected cells, it is not known how the synthesis of the viral RNA begins or even where on the genome synthesis starts. Here, we identify the start site for the initiation of HDV genome RNA synthesis as position 1646, which is at one end of the closed-hairpin-like structure of the viral RNA. The 5' end of the RNA is capped, as expected for polymerase II products. However, RNA synthesis begins with a guanosine that is not present in the genome. Thus, although HDV uses polymerase II to synthesize the viral genome, some details of the initiation process are different. These differences could be important for successfully targeting virus replication.


Self-organized periodic structures on Ge-S based chalcogenide glass induced by femtosecond laser irradiation.
Self-organized periodic structures have been observed on the surface of the ablation craters of Ge-S based chalcogenide glass produced after irradiation by a focused beam of a femtosecond Ti:sapphire laser (1 kHz, 34 fs, 806 nm). Scanning electron microscopy and atomic force microscopy images of irradiated spots show a periodic structure of ripples with a spatial period of 720 nm (close to the wavelength of fs laser pulses) and an alignment parallel to the electric field of light. With an increasing number of pulses, from 5 to 50 pulses, a characteristic evolution of ripples was observed from a random structure to a series of generally aligned peaks-and-valleys self-organized periodic structures. Additionally, at the center of the ablated spot, micro-domains appear where the ripples are still regular but are assembled in a more complex fashion. The experimental observations are interpreted in terms of strong temperature gradients combined with interference of the incident laser irradiation and a scattered surface electromagnetic wave.


Nuclear medicine staff exposure to ionising radiation in 18F-FDG PET/CT practice: a preliminary retrospective study.
This retrospective study provides an insight into the levels of radiation exposure of six nuclear medicine (NM) staff (four technologists and two nurses) performing routine diagnostic 18F-fluorodeoxyglucose (18F-FDG) positron emission tomography-computed tomography (PET/CT) at the University Clinical Centre of the Republic of Srpska, Department of Nuclear Medicine and Thyroid Disorders, Banja Luka, Bosnia and Herzegovina. Data analysis included monthly staff exposure measured with personal thermoluminescent dosimeters (TLD) between June and December 2018, quantified in terms of normalised dose for the whole body [Hp(10)] and dominant hand [Hp(0.07)] and their comparison between each staff member and between the two groups (technologists and nurses). The study goal was to establish how our Department compared with reports from other PET/CT centres worldwide in terms of annual number of procedures and exposure limits and whether there could be room for further improvements in radiation protection. The number of procedures rose considerably from 208 in 2016 to 876 in 2019 and was 423 in the observed seven-month period. Mean individual whole-body exposure dose per GBq of injected 18F-FDG activity, [Hp(10)/A] was 18.55 μSv/GBq for the four technologists and 15.61 μSv/GBq for the two nurses. Mean dominant-hand exposure dose per GBq of injected 18F-FDG activity [Hp(0.07)/A] was 16.99 μSv/GBq and 25.44 μSv/GBq for the two groups, respectively. The average annual cumulative dose for all staff was (1.06±0.29) mSv for Hp(10) and (1.15±0.32) mSv for Hp(0.07). These results are comparable with those of similar studies. Staff doses were well below the annual limits. Nurses received slightly higher extremity doses than technologists. In view of the increasing trends in the number of PET/CT procedures, dose monitoring should be continued to identify exposure hotspots and maintain doses as low as possible.


Immunogenicity and safety of a recombinant vaccinia virus vaccine expressing the carcinoembryonic antigen gene in a nonhuman primate.
We have previously reported the development of a recombinant vaccinia virus vaccine expressing the human carcinoembryonic antigen (CEA) gene, designated rV(NYC)-CEA. This construct has been shown to elicit specific anti-CEA immune responses and an antitumor effect in a murine tumor model. In the studies reported here, the safety and immunogenicity of this recombinant vaccinia virus were evaluated in a rhesus monkey model. Human CEA is a M(r) 180,000 glycoprotein expressed in approximately 90% of gastrointestinal carcinomas and in some breast and non-small cell lung carcinomas. This family also includes normal cross-reacting antigen (NCA). Rhesus monkeys, like humans, have some NCA on the surface of their granulocytes. Eight monkeys were immunized 3 or 4 times by skin scarification with the recombinant CEA vaccine and four monkeys received wild-type vaccinia virus as control. After three vaccinations, all rV(NYC)-CEA-vaccinated animals exhibited a strong anti-CEA antibody response as measured by enzyme-linked immunosorbent assay. The functional ability of these antibodies to mediate lysis of a CEA-bearing tumor cell was demonstrated using human effector cells. This response could be enhanced by interleukin 2. Cellular immunity to CEA was measured by delayed-type hypersensitivity upon intradermal challenge with purified CEA. Only those animals receiving the recombinant vaccine displayed significant anti-CEA responses. Furthermore, peripheral blood mononuclear cells from immunized monkeys were found to proliferate in response to CEA stimulation. All vaccinated monkeys developed local skin irritation at the site of the vaccination, regional lymphadenopathy, and low-grade fevers after immunization. Following immunization with rV(NYC)-CEA, the response was consistent with the usual constitutional symptoms seen with human smallpox virus immunization. Blood counts, differentials, and hepatic and renal chemistries remained normal in all animals throughout the study and for up to 1 year following the primary vaccination. No evidence of immunological cross-reactivity to NCA was found by either a fall in the granulocyte count or analyses for anti-NCA antibodies. Thus, the rV(NYC)-CEA vaccine appears to be safe in rhesus monkeys. The administration of a CEA recombinant vaccine to rhesus monkeys induces both a humoral and a cell-mediated immune response directed against human CEA.


Combined prophylactic and therapeutic immune responses against human papillomaviruses induced by a thioredoxin-based L2-E7 nanoparticle vaccine.
Global burden of cervical cancer, the most common cause of mortality caused by human papillomavirus (HPV), is expected to increase during the next decade, mainly because current alternatives for HPV vaccination and cervical cancer screening programs are costly to be established in low-and-middle income countries. Recently, we described the development of the broadly protective, thermostable vaccine antigen Trx-8mer-OVX313 based on the insertion of eight different minor capsid protein L2 neutralization epitopes into a thioredoxin scaffold from the hyperthermophilic archaeon Pyrococcus furiosus and conversion of the resulting antigen into a nanoparticle format (median radius ~9 nm) upon fusion with the heptamerizing OVX313 module. Here we evaluated whether the engineered thioredoxin scaffold, in addition to humoral immune responses, can induce CD8+ T-cell responses upon incorporation of MHC-I-restricted epitopes. By systematically examining the contribution of individual antigen modules, we demonstrated that B-cell and T-cell epitopes can be combined into a single antigen construct without compromising either immunogenicity. While CD8+ T-cell epitopes had no influence on B-cell responses, the L2 polytope (8mer) and OVX313-mediated heptamerization of the final antigen significantly increased CD8+ T-cell responses. In a proof-of-concept experiment, we found that vaccinated mice remained tumor-free even after two consecutive tumor challenges, while unvaccinated mice developed tumors. A cost-effective, broadly protective vaccine with both prophylactic and therapeutic properties represents a promising option to overcome the challenges associated with prevention and treatment of HPV-caused diseases.


[Development of the urogenital tract in double-bodied pleurodele chimeras. Effect of position on interactions between the genital glands of different sex].
Chimeric grafts of the embryos of the urodele amphibian (Pleurodeles waltlii Michah.) produced viable, double-bodied animals having two pairs of genital glands of the same sex [homosexual chimeras: anterior testes--posterior testes (male----male) and anterior ovaries--posterior ovaries (female----female)] or of different sex [(heterosexual chimeras: anterior ovaries--posterior testes (female----male) and anterior testes--posterior ovaries (male----female)]. 115 chimeras aged 5 months to 15 years were analysed. Gonadal development in a same animal was observed at different times by exploratory laparotomy. The testes of heterosexual chimeras were removed in order to prevent their inhibitory effect on the ovaries (freemartin effect). The 115 experimental animals were classified according to gonadal development as follows: 47 male----male chimeras (41%), 26 female----female chimeras (23%), 13 female----male chimeras (11%), 29 male----female chimeras (25%). As a rule, male----male chimeras only developed three normal testes. The fourth testis, always situated in the anterior part of the chimera, remained vestigial; nevertheless, it could attain a normal size if the other three testes were removed. female----female chimeras usually had four ovaries; the times of vitellogenesis and of sexual maturity were synchronous in all four ovaries. The anterior ovaries of female----male chimeras were always inhibited and the posterior testes were always normal. male----female chimeras always had normal anterior testes but their posterior ovaries were often weakly inhibited. The high number of male----male chimeras and the low number of female----female ones could be due to a phenotypic reversal of the anterior ovaries in some of the female----male chimeras. The number of male----female chimeras (corresponding to the theoretical number) suggests that the phenotypic sex of the posterior ovaries was not reversed. The freemartin effect in double-bodied heterosexual chimeras depended on the location of the ovaries in relation to the testes. The effect was more significant on the anterior ovaries when posterior testes were present. No or weak freemartin effect was observed when the posterior ovaries developed facing the anterior testes; in about 60% of cases, a reciprocal inhibition or an equilibrium between gonads of different sex occurred.(ABSTRACT TRUNCATED AT 400 WORDS)


Topoisomerase II, scaffold component, promotes chromatin compaction in vitro in a linker-histone H1-dependent manner.
TopoisomeraseII (Topo II) is a major component of chromosomal scaffolds and essential for mitotic chromosome condensation, but the mechanism of this action remains unknown. Here, we used an in vitro chromatin reconstitution system in combination with atomic force and fluorescence microscopic analyses to determine how Topo II affects chromosomal structure. Topo II bound to bare DNA and clamped the two DNA strands together, even in the absence of ATP. In addition, Topo II promoted chromatin compaction in a manner dependent on histone H1 but independent of ATP. Histone H1-induced 30-nm chromatin fibers were converted into a large complex by Topo II. Fluorescence microscopic analysis of the Brownian motion of chromatin stained with 4',6-diamidino-2-phenylindole showed that the reconstituted chromatin became larger following the addition of Topo II in the presence but not the absence of histone H1. Based on these findings, we propose that chromatin packing is triggered by histone H1-dependent, Topo II-mediated clamping of DNA strands.


Expression of a heat stable phytase from Aspergillus fumigatus in tobacco (Nicotiana tabacum L. cv. NC89).
Aspergillus fumigatus contains a heat-stable phytase of great potential. To determine whether this phytase could be expressed in plants as a functional enzyme, we introduced the phytase gene from A. fumigatus (fphyA) in tobacco (Nicotiana tabacum L. cv. NC89) by Agrobacterium-mediated transformation. Phytase expression was controlled by the cauliflower mosaic virus (CaMV) 35S promoter. Secretion of recombinant phytase (tfphyA) to the extracellular fluid was established by use of the signal sequence from tobacco calreticulin. Forty-one independent transgenic plants were generated. Single-copy line A was selected based on segregation of T1 seeds for kanamycin resistance, phytase expression and Southern blotting analysis for use in further study. After 4-weeks of plant growth, the phytase was accumulated in leaves up to 2.3% of total soluble protein. tfphyA was functional and shared similar profiles of pH, temperature and thermal stability to the same enzyme expressed in Pichia pastoris (pfphyA). The expressed enzyme had an apparent molecular mass of 63 kDa and showed maximum activity at pH 5.5, and temperature, 55 degrees C. It had a high thermostability and retained 28.7% of the initial activity even after incubation at 90 degrees C for 15 min. The above results showed that the thermostable A. fumigatus phytase could be expressed in tobacco as a functional enzyme and thus has the potential of overexpressing it in other crop plants also.