Not always what closes best opens better: mesoporous nanoparticles capped with organic gates.
Four types of calcined MCM-41 silica nanoparticles, loaded with dyes and capped with different gating ensembles are prepared and characterized. N1 and N2 nanoparticles are loaded with rhodamine 6G and capped with bulky poly(ethylene glycol) derivatives bearing ester groups (1 and 2). N3-N4 nanoparticles are loaded with sulforhodamine B and capped with self-immolative derivatives bearing ester moieties. In the absence of esterase enzyme negligible cargo release from N1, N3 and N4 nanoparticles is observed whereas a remarkable release for N2 is obtained most likely due to the formation of an irregular coating on the outer surface of the nanoparticles. In contrast, a marked delivery is found in N1, N3, and N4 in the presence of esterase enzyme. The delivery rate is related to the hydrophilic/hydrophobic character of the coating shell. The use of hydrophilic poly(ethylene glycol) derivatives as gating ensembles on N1 and N2 enables an easy access of esterase to the ester moieties with subsequent fast cargo release. On the other hand, the presence of a hydrophobic monolayer on N3 and N4 partially hinders esterase enzyme access to the ester groups and the rate of cargo release was decreased.


A cell-type-specific abnormality of cell proliferation in mutant (curly tail) mouse embryos developing spinal neural tube defects.
The mouse mutant curly tail (ct) provides a model system for studies of neurulation mechanisms. 60% of ct/ct embryos develop spinal neural tube defects (NTD) as a result of delayed neurulation at the posterior neuropore whereas the remaining 40% of embryos develop normally. In order to investigate the role of cell proliferation during mouse neurulation, cell cycle parameters were studied in curly tail embryos developing spinal NTD and in their normally developing litter-mates. Measurements were made of mitotic index, median length of S-phase and percent reduction of labelling index during a [3H]thymidine pulse-chase experiment. These independent measures of cell proliferation rate indicate a reduced rate of proliferation of gut endoderm and notochord cells in the neuropore region of embryos developing spinal NTD compared with normally developing controls. The incidence of cell death and the relative frequency of mitotic spindle orientations does not differ consistently between normal and abnormal embryos. These results suggest a mechanism of spinal NTD pathogenesis in curly tail embryos based on failure of normal cell proliferation in gut endoderm and notochord.


Mouse pancreatic islet macrophages use locally released ATP to monitor beta cell activity.
Tissue-resident macrophages sense the microenvironment and respond by producing signals that act locally to maintain a stable tissue state. It is now known that pancreatic islets contain their own unique resident macrophages, which have been shown to promote proliferation of the insulin-secreting beta cell. However, it is unclear how beta cells communicate with islet-resident macrophages. Here we hypothesised that islet macrophages sense changes in islet activity by detecting signals derived from beta cells. To investigate how islet-resident macrophages respond to cues from the microenvironment, we generated mice expressing a genetically encoded Ca2+ indicator in myeloid cells. We produced living pancreatic slices from these mice and used them to monitor macrophage responses to stimulation of acinar, neural and endocrine cells. Islet-resident macrophages expressed functional purinergic receptors, making them exquisite sensors of interstitial ATP levels. Indeed, islet-resident macrophages responded selectively to ATP released locally from beta cells that were physiologically activated with high levels of glucose. Because ATP is co-released with insulin and is exclusively secreted by beta cells, the activation of purinergic receptors on resident macrophages facilitates their awareness of beta cell secretory activity. Our results indicate that islet macrophages detect ATP as a proxy signal for the activation state of beta cells. Sensing beta cell activity may allow macrophages to adjust the secretion of factors to promote a stable islet composition and size.


Chronic intermittent hypoxia increases sympathetic control of blood pressure: role of neuronal activity in the hypothalamic paraventricular nucleus.
Like humans with sleep apnea, rats exposed to chronic intermittent hypoxia (CIH) experience arterial hypoxemias and develop hypertension characterized by exaggerated sympathetic nerve activity (SNA). To gain insights into the poorly understood mechanisms that initiate sleep apnea/CIH-associated hypertension, experiments were performed in rats exposed to CIH for only 7 days. Compared with sham-treated normoxic control rats, CIH-exposed rats (n = 8 rats/group) had significantly increased hematocrit (P < 0.001) and mean arterial pressure (MAP; P < 0.05). Blockade of ganglionic transmission caused a significantly (P < 0.05) greater reduction of MAP in rats exposed to CIH than control rats (n = 8 rats/group), indicating a greater contribution of SNA in the support of MAP even at this early stage of CIH hypertension. Chemical inhibition of neuronal discharge in the hypothalamic paraventricular nucleus (PVN) (100 pmol muscimol) had no effect on renal SNA but reduced lumbar SNA (P < 0.005) and MAP (P < 0.05) more in CIH-exposed rats (n = 8) than control rats (n = 7), indicating that CIH increased the contribution of PVN neuronal activity in the support of lumbar SNA and MAP. Because CIH activates brain regions controlling body fluid homeostasis, the effects of internal carotid artery injection of hypertonic saline were tested and determined to increase lumbar SNA more (P < 0.05) in CIH-exposed rats than in control rats (n = 9 rats/group). We conclude that neurogenic mechanisms are activated early in the development of CIH hypertension such that elevated MAP relies on increased sympathetic tonus and ongoing PVN neuronal activity. The increased sensitivity of Na(+)/osmosensitive circuitry in CIH-exposed rats suggests that early neuroadaptive responses among body fluid regulatory neurons could contribute to the initiation of CIH hypertension.


Delta-toxin production deficiency in Staphylococcus aureus: a diagnostic marker of bone and joint infection chronicity linked with osteoblast invasion and biofilm formation.
Biofilm formation, intra-osteoblastic persistence, small-colony variants (SCVs) and the dysregulation of agr, the major virulence regulon, are possibly involved in staphylococcal bone and joint infection (BJI) pathogenesis. We aimed to investigate the contributions of these mechanisms among a collection of 95 Staphylococcus aureus clinical isolates from 64 acute (67.4%) and 31 chronic (32.6%) first episodes of BJI. The included isolates were compared for internalization rate, cell damage and SCV intracellular emergence using an ex vivo model of human osteoblast infection. Biofilm formation was assessed in a microbead immobilization assay (BioFilm Ring test). Virulence gene profiles were assessed by DNA microarray. Seventeen different clonal complexes were identified among the screened collection. The staphylococcal internalization rate in osteoblasts was significantly higher for chronic than acute BJI isolates, regardless of the genetic background. Conversely, no differences regarding cytotoxicity, SCV emergence, biofilm formation and virulence gene distribution were observed. Additionally, agr dysfunction, detected by the lack of delta-toxin production using whole-cell matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) analysis (n = 15; 15.8%), was significantly associated with BJI chronicity, osteoblast invasion and biofilm formation. These findings provide new insights into MSSA BJI pathogenesis, suggesting the correlation between chronicity and staphylococcal osteoblast invasion. This adaptive mechanism, along with biofilm formation, is associated with agr dysfunction, which can be routinely assessed by delta-toxin detection using MALDI-TOF spectrum analysis, possibly providing clinicians with a diagnostic marker of BJI chronicity at the time of diagnosis.


High EGFR mRNA expression is a prognostic factor for reduced survival in pancreatic cancer after gemcitabine-based adjuvant chemotherapy.
Pancreatic ductal adenocarcinoma (PDAC) still presents a major therapeutic challenge and a phase III clinical trial has revealed that the combination of gemcitabine and a human epidermal growth factor receptor type I (HER1/EGFR) targeting agent presented a significant benefit compared to treatment with gemcitabine alone. The aim of this study was to investigate EGFR mRNA expression in resected PDAC tissues and its correlation with patient prognosis. We obtained formalin-fixed paraffin-embedded (FFPE) tissue samples from 88 patients with PDAC who underwent pancreatectomy, and measured EGFR mRNA levels by quantitative real-time reverse transcription-polymerase chain reaction. The high-level EGFR group had significantly shorter disease-free-survival (p=0.029) and overall-survival (p=0.014) as shown by univariate analyses, although these did not reach statistical significance, as shown by multivariate analyses. However, we found that high EGFR expression was an independent prognostic factor in patients receiving gemcitabine-based adjuvant chemotherapy (p=0.023). Furthermore, we measured EGFR mRNA levels in 20 endoscopic ultrasound-guided fine needle aspiration (EUS-FNA) cytological specimens. Altered EGFR levels were distinguishable in microdissected neoplastic cells from EUS-FNA cytological specimens compared to those in whole cell pellets. In conclusion, quantitative analysis of EGFR mRNA expression using FFPE tissue samples and microdissected neoplastic cells from EUS-FNA cytological specimens could be useful in predicting prognosis and sensitivity to gemcitabine in PDAC patients.


Importance of P53, Ki-67 expression in the differential diagnosis of benign/malignant phyllodes tumors of the breast.
Conventionally growth pattern, stromal overgrowth, stromal cellularity and stromal mitotic activity are the main parameters in the grading of phyllodes tumors (PTs). Recent studies revealed that both p53 and Ki-67 expressions are correlated with grade of PTs of the breast. Expression of hormone receptors and overexpression/amplification of HER2 has been studied in PTs to discover the roles of these markers as new treatment modalities. We studied 26 PT cases. Seventeen benign and nine malignant PTs were re-evaluated as regards stromal cellularity mitotic activity, p53/Ki-67 expression rates and the relation between these parameters. Estrogen receptor and progesterone receptor (ER, PR) positivity were determined by counting nuclear staining in five high-power fields. Also, the presence of any HER2 staining and staining patterns were documanted. Stromal cellularity, mitotic rate, p53 and Ki-67 expression rates were all correlated with benign and malignant histologic subgroups (P = 0.000-0.001). Ki-67 and p53 expressions were statistically significantly correlated with histologic subgroups, stromal cellularity and mitotic rate (P < 0.005). ER and PR expressions in the epithelial component were not statistically significant between the two groups. HER2 showed different staining patterns in the epithelial component, and there was no staining in the stromal component. Ki-67 and p53 expression rates were statistically significantly correlated with grade of mammary PTs; therefore, they can be used in the determination of tumor grade, especially for the differential diagnosis of benign and malignant tumors. Malignant and benign tumors did not differ significantly in terms of hormone receptor and HER2 expression. HER2 expression showed different patterns in the epithelial component of the PTs.


Employment and self-management: a meta-evaluation of seven literature reviews.
Efforts focused on teaching individuals with intellectual disabilities to manage their own affairs have evolved over the past 30 years. Self-management strategies, in particular, hold much promise when the goal is to promote self-determination. In this article, the authors describe trends in the evolution of self-management strategies by analyzing seven literature reviews. The authors conclude with thoughts related to jump-starting an intervention that has appeared to lose momentum, namely, self-management.


Pulsed photothermal radiometry in turbid media: internal reflection of backscattered radiation strongly influences optical dosimetry.
The integrated irradiance (energy fluence rate) within tissue can exceed the incident irradiance due to backscattered and multiply reflected light near the sample surface. This was studied quantitatively using pulsed photothermal radiometry, which measures blackbody radiation emitted by a sample during and after absorption of an optical pulse. Aqueous gels containing absorbing dye with or without various scattering materials were studied using a fast sensitive IR detector system and 1-micros tunable pulsed dye laser. For nonscattering samples, the temperature transient (T-jump) due to absorption of a laser pulse was consistent with Beer's law for homogeneous absorbing media. When scattering was present, increases of up to almost an order of magnitude in the T-jump were observed. For a given absorption coefficient, there was a proportional relationship between the increase in the T-jump and the sample's diffuse reflectance. A model describing the reflectance of diffuse radiation at the sample boundary was derived to explain this result. To test the model, the refractive index was varied with air as the external medium and was also matched to that of BaF(2) as the externalmedium. The subsurface fluence is, to areasonable approximation, given by E congruent with E(0)(l + 2bR),where E(0) is the incident fluence of an infinitely wide collimated beam, b is a coefficient strongly dependent on only the refractive index, and R is the measured diffuse reflectance of the sample. This study shows that irradiance within tissues can greatly exceed the irradiance of incident collimated light, an effect that should be accounted for in photomedical dosimetry or research.


Diagnosing depression: there is no blood test.
To explore and describe primary care physicians' experiences in providing care to depressed patients and to increase understanding of the possibilities and constraints around diagnosing and treating depression in primary care. Qualitative study using personal interviews. A hospital region in eastern Canada. A purposely diverse sample of 20 physicians chosen from among all 100 practising family physicians in the region. Invitations were mailed to all physicians practising in the region. Twenty physicians were chosen from among the 39 physicians responding positively to the invitation. Location of practice, sex, and year of graduation from medical school were used as sampling criteria. The 20 physicians were then interviewed, and the interviews were audiotaped and transcribed verbatim. Data were analyzed using a constant comparative approach involving handwritten notes on transcripts and themes created using qualitative data analysis software. Three themes related to diagnosis emerged. The first concerns use of checklists. Physicians said they needed an efficient but effective means of diagnosing depression and often used diagnostic aids, such as checklists. Some physicians, however, were reluctant to use such aids. The second theme, interpersonal processes, involved the investment of time needed for diagnosing depression and the importance of establishing rapport. The final theme, intuition, revealed how some physicians relied on "gut sense" and years of experience to make a diagnosis. Diagnosis of depression by primary care physicians involves a series of often complicated negotiations with patients. Such negotiations require expertise gained through experience, yet prior research has not recognized the intricacies of this diagnostic process. Our findings suggest that future research must recognize the complex and multidisciplinary nature of physicians' approaches to diagnosis of depression in order to better reflect how they practise.


Aripiprazole Lauroxil Long-Acting Injectable: The Latest Addition to Second-Generation Long-Acting Agents.
Antipsychotics have long been the mainstay for the treatment of schizophrenia and other psychotic disorders. Long-acting injectables (LAI) of antipsychotics-provided once every two weeks to once every three months-promise to reduce the incidence of nonadherence. ARISTADA(™) (aripiprazole lauroxil; ALLAI) extended-release injectable suspension was approved by the U.S. Food and Drug Administration in October 2015 for the treatment of schizophrenia, and is the newest entrant in the LAI market. ALLAI is available as a single-use, pre-filled syringe, can be started in three different dosages, and also has the option of every six-week dosing. Treatment with oral aripiprazole is recommended for the first twenty-one days after the first ALLAI injection, which is a potential disadvantage. Adverse effects include sensitivity to extrapyramidal symptoms, especially akathisia, which is well documented in other aripiprazole preparations. There is no available data comparing ALLAI to other antipsychotics, and more head-to-head trials comparing different LAI formulations are needed. Based on the available data, ALLAI is an effective and safe option for treatment of schizophrenia. Further studies and post-marketing data will provide better understanding of this formulation.


Quantification of Contractile Dynamic Complexities Exhibited by Human Stem Cell-Derived Cardiomyocytes Using Nonlinear Dimensional Analysis.
Understanding the complexity of biological signals has been gaining widespread attention due to increasing knowledge on the nonlinearity that exists in these systems. Cardiac signals are known to exhibit highly complex dynamics, consisting of high degrees of interdependency that regulate the cardiac contractile functions. These regulatory mechanisms are important to understand for the development of novel in vitro cardiac systems, especially with the exponential growth in deriving cardiac tissue directly from human induced pluripotent stem cells (hiPSCs). This work describes a unique analytical approach that integrates linear amplitude and frequency analysis of physical cardiac contraction, with nonlinear analysis of the contraction signals to measure the signals' complexity. We generated contraction motion waveforms reflecting the physical contraction of hiPSC-derived cardiomyocytes (hiPSC-CMs) and implemented these signals to nonlinear analysis to compute the capacity and correlation dimensions. These parameters allowed us to characterize the dynamics of the cardiac signals when reconstructed into a phase space and provided a measure of signal complexity to supplement contractile physiology data. Thus, we applied this approach to evaluate drug response and observed that relationships between contractile physiology and dynamic complexity were unique to each tested drug. This illustrated the applicability of this approach in not only characterization of cardiac signals, but also monitoring and diagnostics of cardiac health in response to external stress.


Prevalence of toxoplasmosis, HIV, syphilis and rubella in a population of puerperal women using Whatman 903 filter paper.
to determine the seroprevalence rate of toxoplasmosis, HIV, syphilis and rubella in a population of puerperal women. a prospective, cross-sectional study was performed from February 2007 to April 2008 at Hospital Geral, Universidade de Caxias do Sul in a population of 1,510 puerperal women. Women that gave birth to live born or stillborn infants were included in the study; maternal and perinatal variables were analyzed. Descriptive statistics and Pearson's chi-square with occasional Fisher's correction were used for comparisons. Alpha was set in 5%. a total of 148 cases of congenital infection (9.8%) were identified: 66 cases of syphilis (4.4%), 40 cases of HIV (2.7%), 27 cases of toxoplasmosis (1.8%) and 15 cases of rubella (1.0%). In ten cases there was co-infection (four cases of HIV and syphilis, two cases of HIV and rubella, one case of HIV and toxoplasmosis, two cases of rubella and syphilis, and one case of toxoplasmosis and rubella). In a comparison between puerperal women with and without infection there was no statistical significance in relation to incidence of abortions, small for gestational age, prematurity, live births and stillbirths, and prenatal care. Need of neonatal intensive care unit (NICU), maternal schooling, maternal age higher than 35 years and drug use (alcohol, cocaine and crack) had statistical significance. the prevalence rate of infections was 9.8%. Need of NICU, maternal schooling lower than eight years, maternal age higher than 35 years and drug use were significantly associated with occurrence of congenital infection.


Repetitive sequences (RPSs) in the chromosomes of Candida albicans are sandwiched between two novel stretches, HOK and RB2, common to each chromosome.
A novel sequence designated HOK, which is next to the RPS, a repetitive sequence specific to Candida albicans, was cloned and sequenced. HOK hybridized with all of the chromosomes on which the RPSs were located, but did not hybridize with chromosome 3, which does not harbour any RPSs. Sequence determination revealed that a portion of HOK has significant homology with the B and C1 fragments of Ca3, which is used as a molecular epidemiological probe. A homology search of the deduced amino acids of HOK against the protein database showed partial homology with an isocitrate dehydrogenase of Saccharomyces cerevisiae, although an ORF large enough to encode the enzyme was not detected. To verify the existence of other sequences homologous with HOK, a portion of the HOK sequence was amplified using PCR. Sequence determination of the 41 clones from the PCR products resulted in at least six HOK-homologous clones. Another RPS-containing clone, RB2, was isolated from the Pstl-digested chromosome R or 1. It was determined that RB2a, one of the subclones from RB2, hybridized with all of the chromosomes, including chromosome 3, with which neither HOK nor RPS hybridized. The hybridization profile also showed that RPS is located between HOK and RB2a on chromosomes other than chromosome 3.


Prospective analysis of waiting times for emergency plastic surgery in four units.
Trauma is a significant part of the workload in plastic surgery. There are currently wide variations in the available resource for dealing with these patients. Delays to treatment currently exist and may result in poorer clinical outcomes. Data was collected prospectively in 4 centres (Cork University Hospital, John Radcliffe Hospital, Stoke Mandeville Hospital and Salisbury District Hospital) assessing delays to theatre. We assessed time to theatre, both from injury and from review, cancellation rate, starvation time and patient satisfaction. 424 patients were audited over an 8-week period. The average time from review to theatre was 15.7 h and the average injury-to-theatre time was 58.6 h. The average starvation time was 10 h; the mean cancellation rate was 25%. Patients are satisfied overall with the service provided with 83% rating the service as excellent or good, and 63% feeling that there wait was not too long. Despite an increase in provision of emergency plastic surgery trauma lists, the average wait for emergency plastic surgery is increasing. Despite this patients remain, on the whole, satisfied with the service that they are receiving.


Nardostachys jatamansi improves learning and memory in mice.
Cure of cognitive disorders such as amnesia, attention deficit, and Alzheimer's disease is still far from being realized in the field of medicine. Nootropic agents such as piracetam, aniracetam, and choline esterase inhibitors like donepezil are being used for improving memory, mood, and behavior, but the resulting side effects associated with these agents have made their applicability limited. In Ayurveda, the roots of Nardostachys jatamansi have been clinically employed for their anti-ischemic, antioxidant, anticonvulsant, and neuroprotective activities. The present study was undertaken to assess the potential of N. jatmansi as a memory enhancer. The elevated plus maze and the passive avoidance paradigm were employed to evaluate learning and memory parameters. Three doses (50, 100, and 200 mg/kg, p.o.) of an ethanolic extract of N. jatamansi were administered for 8 successive days to both young and aged mice. The 200 mg/kg dose of N. jatmansi ethanolic extract significantly improved learning and memory in young mice and also reversed the amnesia induced by diazepam (1 mg/kg, i.p.) and scopolamine (0.4 mg/kg, i.p.). Furthermore, it also reversed aging-induced amnesia due to natural aging of mice. As scopolamine-induced amnesia was reversed, it is possible that the memory improvement may be because of facilitation of cholinergic transmission in the brain. Hence, N. jatmansi might prove to be a useful memory restorative agent in the treatment of dementia seen in elderly persons. The underlying mechanism of action can be attributed to its antioxidant property.


EXUDATIVE LESIONS IN DIABETES MELLITUS.
Exudative lesions and capsular drops have been examined in a renal biopsy series of 25 diabetics and a post-mortem series of 100 consecutive diabetics. Electron microscopy of a capsular drop and an exudative lesion is described. Exudative lesions were found in 48% of the biopsies and 47% of the necropsy material. Capsular drops were found in 60% of the biopsies and 29% of the necropsy material. No correlation with age, proteinuria, hypertension, or uraemia was noted. The incidence of both types of lesion increased with the known duration of the diabetes.


Development of the pseudothumb in frogs.
Frogs have highly conserved hand and foot morphology, possessing four fingers and five toes. As an exception, two Japanese ranid frog species, the Otton frog Babina subaspera and the dagger frog Babina holsti, possess a unique thumb-like structure (the pseudothumb) in the forelimb, giving an appearance of a total of five fingers on the hand. To obtain insights into the developmental mechanisms that generate this novel character, we investigated the hand morphogenesis of the Otton frog. The unique morphological pattern of the pseudothumb was already established in juveniles. Surprisingly, the bud-like structure, which is similar to the area of inductive activity (e.g. feather buds in birds and the carapacial ridge in turtles), was detected over the site where the future prepollex develops in larvae. By contrast, this bud-like structure was not found in larvae of other ranid species. We discuss possible scenarios that would favour the evolution of this very unusual trait in frogs.


Understanding reasons for and outcomes of patients lost to follow-up in antiretroviral therapy programs in Africa through a sampling-based approach.
Losses to follow-up after initiation of antiretroviral therapy (ART) are common in Africa and are a considerable obstacle to understanding the effectiveness of nascent treatment programs. We sought to characterize, through a sampling-based approach, reasons for and outcomes of patients who become lost to follow-up. Cohort study. We searched for and interviewed a representative sample of lost patients or close informants in the community to determine reasons for and outcomes among lost patients. Three thousand six hundred twenty-eight HIV-infected adults initiated ART between January 1, 2004 and September 30, 2007 in Mbarara, Uganda. Eight hundred twenty-nine became lost to follow-up (cumulative incidence at 1, 2, and 3 years of 16%, 30%, and 39%). We sought a representative sample of 128 lost patients in the community and ascertained vital status in 111 (87%). Top reasons for loss included lack of transportation or money and work/child care responsibilities. Among the 111 lost patients who had their vital status ascertained through tracking, 32 deaths occurred (cumulative 1-year incidence 36%); mortality was highest shortly after the last clinic visit. Lower pre-ART CD4 T-cell count, older age, low blood pressure, and a central nervous system syndrome at the last clinic visit predicted deaths. Of patients directly interviewed, 83% were in care at another clinic and 71% were still using ART. Sociostructural factors are the primary reasons for loss to follow-up. Outcomes among the lost are heterogeneous: both deaths and transfers to other clinics were common. Tracking a sample of lost patients is an efficient means for programs to understand site-specific reasons for and outcomes among patients lost to follow-up.


Pectinesterase Inhibitor from Jelly Fig (Ficus awkeotsang Makino) Achene Inhibits Surface Antigen Expression by Human Hepatitis B Virus.
Pectinesterase inhibitor (PEI) isolated from jelly fig (Ficus awkeotsang Makino) is an edible component of a popular drink consumed in Asia. Hepatitis B virus (HBV) infection is prevalent in Asia, and current treatments for HBV infection need improvement. This study aimed to evaluate the effect of PEI on the surface antigen expression by HBV (HBsAg). Human hepatoma cell lines Hep3B and Huh7 served as in vitro models for assessing the cytotoxicity and HBsAg expression. A culture of primary hepatocytes cultured from mice served as the normal counterpart. Cell viability was measured by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) colorimetric assay. HBsAg expression was evaluated by measuring HBsAg secretion into the culture medium using an enzyme-linked immunosorbent assay. The results showed that PEI did not affect the viability of the human hepatoma cell lines or primary mouse hepatocytes. PEI inhibited the expression of HBsAg in hepatoma cell lines harboring endogenous (Hep3B) and integrated (Huh7) HBV genomes in a concentration- and time-dependent manner, thus implicating a universal activity against HBV gene expression. In conclusion, it suggests that PEI from jelly fig inhibits the expression of human HBsAg in host cells without toxic effects on normal primary hepatocytes.


The role of adiponectin in human vascular physiology.
Adiponectin (ApN) is an adipose tissue-derived hormone which is involved in a wide variety of physiological processes including energy metabolism, inflammation, and vascular physiology via actions on a broad spectrum of target organs including liver, skeletal muscle, and vascular endothelium. Besides possessing insulin sensitizing and anti-inflammatory properties ApN also exerts a pivotal role in vascular protection through activation of multiple intracellular signaling cascades. Enhancement of nitric oxide generation and attenuation of reactive oxygen species production in endothelial cells along with reduced vascular smooth muscle cell proliferation and migration constitute some of ApN's vasoprotective actions. Additionally, recent data indicate that ApN has direct myocardio-protective effects. Decreased plasma ApN levels are implicated in the pathogenesis of the metabolic syndrome and atherosclerosis and may serve as a diagnostic and prognostic biomarker as well as a rational pharmaco-therapeutic target to treat these disorders. This review article summarizes recent work on the cardiovascular actions of ApN.


Precursor cells from Atlantic salmon (Salmo salar) visceral fat holds the plasticity to differentiate into the osteogenic lineage.
In order to study the potential plasticity of Atlantic salmon (Salmo salar) precursor cells (aSPCs) from the adipogenic mesenchyme cell lineage to differentiate to the osteogenic lineage, aSPCs were isolated and cultivated under either osteogenic or adipogenic promoting conditions. The results strengthen the hypothesis that aSPCs most likely are predestined to the adipogenic lineage, but they also hold the flexibility to turn into other lineages given the right stimuli. This assumption is supported by the fact that the transcription factor pparγ , important for regulation of adiopogenesis, was silent in aSPCs grown in osteogenic media, while runx2, important for osteogenic differentiation, was not expressed in aSPCs cultivated in adipogenic media. After 2 weeks in osteogenic promoting conditions the cells started to deposit extracellular matrix and after 4 weeks, the cells started mineralizing secreted matrix. Microarray analyses revealed large-scale transcriptome responses to osteogenic medium after 2 days, changes remained stable at day 15 and decreased by magnitude at day 30. Induction was observed in many genes involved in osteogenic differentiation, growth factors, regulators of development, transporters and production of extracellular matrix. Transcriptome profile in differentiating adipocytes was markedly different from differentiating osteoblasts with far fewer genes changing activity. The number of regulated genes slowly increased at the mature stage, when adipocytes increased in size and accumulated lipids. This is the first report on in vitro differentiation of aSPCs from Atlantic salmon to mineralizing osteogenic cells. This cell model system provides a new valuable tool for studying osteoblastogenesis in fish.


Chronobiology of interspecific interactions in a changing world.
Animals should time activities, such as foraging, migration and reproduction, as well as seasonal physiological adaptation, in a way that maximizes fitness. The fitness outcome of such activities depends largely on their interspecific interactions; the temporal overlap with other species determines when they should be active in order to maximize their encounters with food and to minimize their encounters with predators, competitors and parasites. To cope with the constantly changing, but predictable structure of the environment, organisms have evolved internal biological clocks, which are synchronized mainly by light, the most predictable and reliable environmental cue (but which can be masked by other variables), which enable them to anticipate and prepare for predicted changes in the timing of the species they interact with, on top of responding to them directly. Here, we review examples where the internal timing system is used to predict interspecific interactions, and how these interactions affect the internal timing system and activity patterns. We then ask how plastic these mechanisms are, how this plasticity differs between and within species and how this variability in plasticity affects interspecific interactions in a changing world, in which light, the major synchronizer of the biological clock, is no longer a reliable cue owing to the rapidly changing climate, the use of artificial light and urbanization.This article is part of the themed issue 'Wild clocks: integrating chronobiology and ecology to understand timekeeping in free-living animals'.


Role of Sortilin in Models of Autoimmune Neuroinflammation.
The proneurotrophin receptor sortilin is a protein with dual functions, being involved in intracellular protein transport, as well as cellular signal transduction. The relevance of the receptor for various neuronal disorders, such as dementia, seizures, and brain injury, is well established. In contrast, little is known about the role of sortilin in immune cells and inflammatory diseases. The aim of our study was to elucidate the distribution of sortilin in different immune cell types in mice and humans and to analyze its function in autoimmune CNS inflammation. Sortilin was expressed most profoundly in murine and human macrophages and dendritic cells and to a much lesser extent in B and T cells. In dendritic cells, sortilin had an impact on Ag processing. Accordingly, sortilin was highly expressed by infiltrated perivascular myeloid cells, mainly in vessel cuffs, in the CNS of patients suffering from multiple sclerosis, the most common inflammatory autoimmune disease of the CNS. Yet, sortilin gene-targeted mice (Sort1(-/-)) and chimeras deficient in sortilin in the immune system were as susceptible as wild-type littermates to T cell-dependent experimental autoimmune encephalomyelitis. Considering our results and recent data from other investigators, we conclude that the proneurotrophin receptor sortilin plays a role in innate, rather than in adaptive, immune processes and, thus, not in autoimmune neuroinflammation.


Identification of fodrin as a major calmodulin-binding protein in postsynaptic density preparations.
A major protein of postsynaptic densities (PSDs), a doublet of 230,000 and 235,000 Mr that becomes enriched in PSDs after treatment of synaptic membranes with 0.5% Triton X-100, has been found to be identical to fodrin (Levine, J., and M. Willard, 1981, J. Cell Biol. 90:631) by the following criteria. The upper bands of the PSD doublet and purified fodrin (alpha-fodrin) were found to be identical since both bands (a) co-migrated on SDS gels, (b) reacted with antifodrin, (c) bound calmodulin, and (d) had identical peptide maps after Staphylococcus aureus protease digestion. The lower bands of the PSD doublet and of purified fodrin (beta-fodrin) were found to be identical since both bands co-migrated on SDS gels and both had identical peptide maps after S. aureus protease digestion. The binding of calmodulin to alpha-fodrin was confirmed by cross-linking azido-125I-calmodulin to fodrin before running the protein on SDS gels. No binding of calmodulin to beta-fodrin was observed with either the gel overlay or azido-calmodulin techniques. A second calmodulin binding protein in the PSD has been found to be the proteolytic product of alpha-fodrin. This band (140,000 Mr), which can be created by treating fodrin with chymotrypsin, both binds calmodulin and reacts with antifodrin.


Feasibility Study of a Newly Developed Hybrid Energy Device Used During Laparoscopic Liver Resection in a Porcine Model.
Although various devices have been clinically used for laparoscopic liver resection (LLR), the best device for liver parenchymal transection remains unknown. Olympus Corp (Tokyo, Japan) developed a laparoscopic hybrid pencil (LHP) device, which is the first electric knife to combine ultrasound and electric energy with a monopolar output. We aimed to evaluate the feasibility of using the LHP device and to compare it with the laparoscopic monopolar pencil (LMP) and laparoscopic ultrasonic shears (LUS) devices for LLR in a porcine model. Nine male piglets underwent laparoscopic liver lobe transections using each device. The operative parameters were evaluated in the 3 groups (n = 24 lobes) during the acute study period. The imaging findings from contrast-enhanced computed tomography and histopathological findings of autopsy on postoperative day 7 were compared among groups (n = 6 piglets) during the long-term study. The transection time was shorter ( P = .001); there was less blood loss ( P = .018); and tip cleaning ( P < .001) and instrument changes were less often required ( P < .001) in the LHP group than in the LMP group. The LHP group had fewer instances of bleeding ( P < .001) and coagulator usage ( P < .001) than did the LUS group. In the long-term study, no postoperative adverse events occurred in the 3 groups. The thermal spread and depth of the LHP device were equivalent to those of the LMP and LUS devices (vs LMP: P = .226 and .159; vs LUS: P = 1.000 and .574). The LHP device may be an efficient device for LLR if it can be applied to human surgery.


Curcumin Can Decrease Tissue Inflammation and the Severity of HSV-2 Infection in the Female Reproductive Mucosa.
Herpes Simplex Virus Type 2 (HSV-2) is one of the most prevalent sexually transmitted viruses and is a known risk factor for HIV acquisition in the Female Genital Tract (FGT). Previously, we found that curcumin can block HSV-2 infection and abrogate the production of inflammatory cytokines and chemokines by genital epithelial cells in vitro. In this study, we investigated whether curcumin, encapsulated in nanoparticles and delivered by various in vivo routes, could minimize inflammation and prevent or reduce HSV-2 infection in the FGT. Female mice were pre-treated with curcumin nanoparticles through oral, intraperitoneal and intravaginal routes, and then exposed intravaginally to the tissue inflammation stimulant CpG-oligodeoxynucleotide (ODN). Local intravaginal delivery of curcumin nanoparticles, but not intraperitoneal or oral delivery, reduced CpG-mediated inflammatory histopathology and decreased production of pro-inflammatory cytokines Interleukin (IL)-6, Tumor Necrosis Factor Alpha (TNF-α) and Monocyte Chemoattractant Protein-1 (MCP-1) in the FGT. However, curcumin nanoparticles did not demonstrate anti-viral activity nor reduce tissue pathology when administered prior to intravaginal HSV-2 infection. In an alternative approach, intravaginal pre-treatment with crude curcumin or solid dispersion formulations of curcumin demonstrated increased survival and delayed pathology following HSV-2 infection. Our results suggest that curcumin nanoparticle delivery in the vaginal tract could reduce local tissue inflammation. The anti-inflammatory properties of curcumin delivered to the vaginal tract could potentially reduce the severity of HSV-2 infection and decrease the risk of HIV acquisition in the FGT of women.


Homozygous deletions at chromosome 9p21 and mutation analysis of p16 and p15 in microdissected primary non-small cell lung cancers.
Loss of heterozygosity on chromosome 9p has been detected in many primary human tumors and cell lines, suggesting that this chromosomal arm harbors one or more tumor suppressor genes. The recently cloned p16 and p15 genes, mapped to 9p21, are likely candidates for such tumor suppressors. To map the deletion at chromosome 9p21 in non-small cell lung tumors, we analyzed DNA from 25 tumors and matching normal DNAs at six microsatellite markers that flank the region occupied by the p16 and p15 genes. Loss of heterozygosity of at least one microsatellite marker on chromosome 9p21 was detected in 13 (52%) of 25 tumors, including one tumor that exhibited homozygous deletion of both human IFNalpha and D9S171. Six tumors analyzed by a comparative multiplex PCR technique showed homozygous deletions of the sequence tag site marker c5.1 (within p16). Screening for mutations in p16 and p15 revealed one tumor with a non-sense mutation in exon 2 of p16, but no mutations were detected in p15 in any of the tumors. Thus, in these analyses approximately one-half of the non-small cell lung tumors had loss of heterozygosity at chromosome 9p21, and of these tumors, one-half had homozygous deletions of the region that includes p16. This appears to confirm the importance of a locus in this region critical to growth control in lung. The apparent lack of other mutations in p16 and p15 in the tumors with loss of heterozygosity leaves open the possibility of an unidentified gene in this region that may function as a tumor suppressor.


Challenges and opportunities for smallholder livestock production in post-conflict South Kivu, eastern DR Congo.
A survey on smallholder livestock production with emphasis on monogastric animals was conducted in 20 villages of seven so-called 'groupements' of South Kivu province in DR Congo, situated along a north to south-west axis with the town of Bukavu in the center. This land adjacent to Lake Kivu is located at elevations ranging around 900- 1900 m asl, experiencing tropical highland climate. A diagnostic survey helped to rapidly obtain in-depth knowledge of constraints and opportunities in this environment. Correspondence analysis and multiple regression analysis were used to investigate the association of production constraints with particular livestock species and to understand the factors that govern the number of livestock that people owned (converted to tropical livestock units [TLU]), respectively. Responses of 112 informants demonstrated that livestock is an integral part of the region's mixed farming systems. Low livestock numbers per household at present reflect the poverty as a consequence of recent violent conflicts. Currently, farmers focus on small livestock, like poultry, swine, cavies (i.e., Guinea pigs) and rabbits. Families keep livestock to accumulate household reserves that are strongly invested in children's education. Major issues of animal husbandry were related to animal diseases and lack of feed resources, particularly in the dry season. Lack of feed or forages were unrelated to a particular livestock species. Livestock holdings depended on animal diversity, location, land size available and respondents' education level. The potential introduction of improved forages is challenged by their dry-season tolerance, compatibility with cropping on small farms; and people's readiness to cultivate forages.


First Report of Rose yellow vein virus in Rosa sp. in New Zealand.
Rose is the top selling cut flower in New Zealand and is the most popular garden plant in the world. Several virus-like diseases have been described in roses, but the causal agents for many remain unknown. Most of the described viruses infecting rose belong to the genera Ilarvirus and Nepovirus. Only recently, a number of new viruses have been or are in the process of being characterized (1,2,3,4). In January 2011, 10 rose samples showing virus-like symptoms were collected from the Wanganui region on the North Island of New Zealand. Total nucleic acid was extracted from these samples using an InviMag Plant DNA Mini Kit (Invitek GmbH, Berlin, Germany) and a KingFisher mL workstation (Thermo Scientific, Waltham, MA). PCR and reverse transcription (RT)-PCR was conducted using specific primers for Arabis mosaic virus (ArMV), Cherry leaf roll virus, Prunus necrotic ringspot virus (PNRSV), Rosa rugosa leaf distortion virus, Rose spring dwarf associated virus, Rose yellow leaf virus, Rose yellow mosaic virus, Rose yellow vein virus (RYVV), and Strawberry latent ringspot virus. Samples were also tested using generic primers for carlavirus, potexvirus, potyvirus, tombusvirus, and phytoplasmas. Two samples (cvs. Pauls Himalayan Musk and Bloomfield) were positive for ArMV, four samples (cvs. Leda, Rosa Mundi, Charles de Mills, and Indica Major) were positive for PNRSV, and two samples (cvs. Leda and Zephirine Drouhin) were positive for RYVV. Samples were negative for all other tested viruses and phytoplasmas. RYVV was detected using two sets of primers (D. Mollov, personal communication) designed to amplify fragments of estimated sizes of 797 bp and 684 bp of the movement protein (MP) and coat protein (CP) genes of RYVV, respectively. RYVV amplicons were sequenced directly (GenBank Accession Nos. JX887423 to JX887426). A BLASTn search of the MP and CP fragments showed the highest nucleotide identity of 98% and 96 to 97%, respectively, with the type isolate of RYVV (JX028536). RYVV has been reported as the causal agent of a vein yellowing disease in rose (2). Symptoms observed in the 'Leda' sample infected with PNRSV and RYVV (vein yellowing and chlorotic mottle in the apex of leaves) were not typical of PNRSV, so they may be caused by RYVV. Symptoms in samples of cv. Zephirine Drouhin (curling of leaves and mottle), observed in both RYVV-positive and -negative samples, may not be associated with RYVV infection. This suggests that vein yellowing may be influenced by cultivar. RYVV has been reported in several rose cultivars, but only in the United States (2). To the best of our knowledge, this is the first report of RYVV infecting rose in New Zealand, where it is likely that the virus has been present for some time. The virus may have a much wider geographical distribution than that reported as the virus was only recently characterized (3). References: (1) B. Lockhart et al. Page 31 in: Program and Abstracts of The 12th International Symposium on Virus Diseases of Ornamental Plants, 2008. (2) D. Mollov et al. Phytopathology 99:S87, 2009. (3) D. Mollov et al. Arch Virol. 158:877, 2012. (4) N. Salem et al. Plant Dis. 92:508, 2008.


Bunk bed injuries.
Bunk beds are commonly used in American households, yet to our knowledge, no studies have been done to determine if they are safe. We prospectively studied the incidence, epidemiology, and outcome of injuries related to bunk beds. We interviewed all patients with such injuries who presented to the emergency department between February 1987 and February 1988. A control group of children who use bunk beds but who came to the emergency department for another reason were also interviewed. There were 68 injured children and 54 controls during the 1-year study period. There were 47 injured children (70% of this group) and 26 control children (48% of this group) younger than 6 years, which is below the age recommended by the Consumer Product Safety Commission for bunk bed use. Carpeted floors were significantly more common in the control group, 67% (36 children) vs 42% (26 children). Injuries occurred most often when the child fell from the top bed (38 children [58%]), fell off the ladder (7 children [11%]), or fell off the bottom bed (8 children [12%]). Injuries occurred during sleep (19 children [29%]), getting in or out of the bunk bed (13 children [20%]), or playing in or near the beds (28 children [43%]). Of those injured while asleep, 13 of 19 children were younger than 6 years. Head injuries accounted for half the trauma (35 children [52%]), and extremities were involved in 16 patients [24%]. The most common injuries were lacerations (27 children [40%]) and contusions (19 children [28%]), but 8 children (12%) had concussions and 7 children (10%) had fractures. Six children (9%) required admission to the hospital. Head and face injuries were significantly more likely if the top bed had no side rails. These data suggest injuries could be prevented if side rails were mandatory for all top beds, young children were not permitted to sleep in bunk beds, and all children were encouraged not to use the beds for play.


In vitro antiprotozoal activity of extract and compounds from the stem bark of Combretum molle.
The antiprotozoal activity of the Ethiopian medicinal plant Combretum molle (R. Br. ex G. Don.) Engl & Diels (Combretaceae) was evaluated by in vitro testing against Plasmodium falciparum, Trypanosoma brucei rhodesiense, Trypanosoma cruzi and Leishmania donovani. The acetone fraction of the stem bark of this plant prepared by soxhlet extraction was inactive against the intracellular amastigotes of L. donovani and T. cruzi in murine peritoneal macrophages but showed significant activity against extracellular T. b. rhodesiense blood stream form trypomastigotes and trophozoites of P. falciparum with IC(50) values of 2.19 and 8.17 microg/mL, respectively. Phytochemical examination of the bioactive fraction resulted in the isolation of two tannins and two oleanane-type pentacyclic triterpene glycosides. One of the tannins was identified as the ellagitannin, punicalagin, whilst the structure of the other (CM-A) has not yet been fully elucidated. The saponins that were characterized as arjunglucoside (also called 4-epi-sericoside) and sericoside displayed no activity against any of the four species of protozoa tested. On the other hand, punicalagin and CM-A had IC(50) values of 1.75 and 1.50 microM, respectively, against T. b. rhodesiense and were relatively less toxic to KB cells (cytotoxic/antiprotozoal ratios of 70 and 48, respectively). The tannins also showed intermediate activity against P. falciparum, although their selectivity against these parasites was less favourable than the above. It appears that our findings are the first report of hydrolysable tannins exhibiting antitrypanosomal and antiplasmodial activities.


Changes in the trauma narratives of youth receiving trauma-focused cognitive behavioral therapy in relation to posttraumatic stress symptoms.
To understand the meaning of trauma narration, we examined changes in the trauma narratives of youth receiving trauma-focused cognitive behavioral therapy (TF-CBT) and explored the relationship between changes in narratives and in posttraumatic stress. The sample consisted of 12 non-responders and 12 maximum-responders to treatment (M = 14.3, SD = 2.35, range = 10-17; 75% girls). The youth were assessed with the Clinical-Administered PTSD-Scale for Children and Adolescents both pre- and post-treatment. Their first and last narratives were coded according to a standardized coding manual. For the group as a whole there was an increase in organized thoughts and reports of internal events (e.g., descriptions of thoughts and feelings), while narrative fragmentation decreased. There were no significant narrative changes in external events (e.g., descriptions of actions and dialogues). Max-responders differed significantly from non-responders in developing more organized thoughts. We did not find a significant relationship between changes in narratives and changes in posttraumatic stress symptoms (PTSS). Youth receiving TF-CBT develop narratives that contain more organized thoughts and a greater internal focus, which are both thought to be helpful for traumatized youth. However, more coherent and organized trauma narratives were not related to reductions in PTSS. Clinical or methodological significance of this article: This study suggests that trauma-focused cognitive behavioral therapy contributes to more organized and coherent trauma narratives for traumatized youth. Although, this may be important and contribute to meaning making, therapist should be aware that this may not be sufficient in reducing posttraumatic stress symptoms in youth.


The effect of acupuncture needle combination on central pain processing--an fMRI study.
Empirical acupuncture treatment paradigm for acute pain utilizing Tendinomuscular Meridians (TMM) calls for the stimulation of Ting Points (TPs) and Gathering point(GP). This study aims to compare the supraspinal neuronal mechanisms associated with both TPs and GP needling (EA3), and TPs needling alone (EA2) with fMRI. A significant (P < 0.01) difference between pre-scan (heat Pain) HP, and post-EA HP VAS scores in both paradigms was noted (n = 11). The post-EA HP VAS score was significantly (P < 0.05) lower with EA3 comparing to EA2 Within-group random effect analysis indicated that EA3+HP>EA3 (condition EA3+HP subtracted by condition EA3) appeared to exert a significant degree of activity suppression in the affective supraspinal regions including the IPL, anterior cingulate cortex (ACC) and the insular cortex (IN). This level of suppression was not observed in the EA2+HP>EA2 (condition EA2+HP subtracted by condition EA2) within-group random effect analysis Between-group random effect analysis indicated that EA3 induced a significantly (P < 0.01, cluster size threshold 150) higher degree of deactivation than EA2 in several pain related supraspinal regions including the right prefrontal cortex, rostral anterior cingulate (rACC), medial cingulate cortex, left inferior frontal lobe and posterior cerebellum. The 2-factor ANOVA in those regions indicated both rACC and posterior cerebellum had a significant (P < 0.01) needle effect, and the right prefrontal area showed a significant (P < 0.01) HP effect. However, a significant interaction between the two factors was only found in the right prefrontal lobe. Granger causality analysis showed EA3 induced a much higher degree of inference among HP related supraspinal somatosensory, affective and modulatory components than EA2. Deactivation pattern at the medullary-pontine area casted a direct inference on the deactivation pattern of secondary somatosensory cortices which also affected the deactivation of the IN. While both EA2 and EA3 induced a significant degree of deactivation in the human brain regions related to pain processing, the addition of GP stimulation further exerts an inhibitory effect on the ascending spinoreticular pain pathway. Therefore, different needling position as mandated in different empirical acupuncture treatment paradigms may play a different role in modulating pain related neuronal functions.


Antiamnesic effect of metoprine and of selective histamine H(1) receptor agonists in a modified mouse passive avoidance test.
The aim of this study was to elucidate the effect caused by the inhibition of histamine catabolism by means of metoprine and the activation of histamine H(1) receptors by selective agonists on learning and memory processes, using a modified method of the mouse passive avoidance test. The administration of scopolamine 1 mg/kg (i. p.) immediately after the training session caused statistically-significant amnesia during the retention trial performed 24 h later. Piracetam (30 mg/kg (i.p.)), used as a positive control, and administered 20 min before the training session, prevented scopolamine-induced memory impairment. The histamine-N-methyltransferase inhibitor, metoprine, (2 and 5 mg/kg (s.c.)) had effects similar to those of this nootropic drug. The highly-selective H(1) receptor agonist, 2-(3-trifluoromethylphenyl)histamine (FMPH) (2.65 and 6.5 microg/mouse (i.c.v.)) and the less selective agonist, 2-thiazolylethylamine (2-TEA) (0.1 and 0.3 microg/mouse (i.c.v.)) both antagonized the scopolamine-induced amnesia significantly and in a dose-related manner. The selective H(1) receptor antagonist, pyrilamine (20 mg/kg (i.p.)), revealed no effect by itself, but significantly prevented the antiamnesic action both that of the H(1) receptor agonists, and that of endogenous histamine, released by metoprine, thus suggesting a cognitive improvement via the activation of H(1) receptors.


Systemic arterial blood pressure and urine protein/creatinine ratio in dogs with hyperadrenocorticism.
To determine prevalence and severity of systemic arterial hypertension and proteinuria in dogs with naturally developing hyperadrenocorticism and to determine whether these abnormalities resolve with adequate management of the disease. Case series and cohort study. 77 dogs with naturally developing hyper-adrenocorticism examined once; 15 dogs examined before and after treatment. Among dogs examined only once, hypertension was diagnosed in 21 of 26 dogs with untreated pituitary-dependent hyperadrenocorticism (PDH), 17 of 21 with inadequately controlled PDH, 8 of 16 with well-controlled PDH, 10 of 10 with an untreated adrenocortical tumor, and 0 of 4 that had undergone adrenalectomy because of an adrenocortical tumor. Untreated dogs and dogs with inadequately controlled PDH had significantly higher blood pressures than did other dogs. Proteinuria was documented in 12 of 26 dogs with untreated PDH, 5 of 16 with inadequately controlled PDH, 3 of 14 with well-controlled PDH, 5 of 8 with an untreated adrenocortical tumor, and 1 of 3 that had undergone adrenalectomy. Dogs with untreated PDH and dogs with an untreated adrenocortical tumor had higher urine protein/creatinine ratios than did dogs with well-controlled PDH. Among dogs evaluated before and after treatment, blood pressure and urine protein/creatinine ratio did not change in 8 dogs with inadequately controlled hyperadrenocorticism, but decreased in 7 dogs with well-controlled disease. Results suggest that systemic hypertension and proteinuria are common in dogs with untreated hyperadrenocorticism and that successful treatment of hyperadrenocorticism will result in resolution of these abnormalities in many, but not all, dogs.


Average Weight of Seminal Vesicles: An Adjustment Factor for Radical Prostatectomy Specimens Weighed With Seminal Vesicles.
The International Society of Urological Pathology in 2010 recommended weighing prostates without seminal vesicles (SV) to include only prostate weight in prostate-specific antigen (PSA) density (PSAD) calculation, because SV do not produce PSA. Large retrospective cohorts exist with combined weight recorded that needs to be modified for retrospective analysis. Weights of prostates and SV were separately recorded in 172 consecutive prostatectomies. The average weight of SV and proportion of prostate weight from combined weight were calculated. The adjustment factors were then validated on databases of 2 other institutions. The average weight of bilateral SV was 6.4 g (range = 1-17.3 g). The prostate constituted on average 87% (range = 66% to 98%) of the total specimen weight. There was no correlation between patient age and prostate weight with SV weight. The best performing correction method was to subtract 6.4 g from total radical prostatectomy weight and to use this weight for PSAD calculation. The average weights of retrospective specimens weighed with SV were not significantly different between the 3 institutions. Using our data allowed calibration of the weights and PSAD between the cohorts weighed with and without SV. Thus, prostate weight in specimens including SV weight can be adjusted by subtracting 6.4 g, resulting in significant change of PSAD. Some institution-specific variations may exist, which could further increase the precision of retrospective analysis involving prostate weight and PSAD. However, unless institution-specific adjustment parameters are developed, we recommend that this correction factor be used for retrospective cohorts or in institutions where combined weight is still recorded.


Cost-Effectiveness of HIV Pre-exposure Prophylaxis Among Heterosexual Men in South Africa: A Cost-Utility Modeling Analysis.
Heterosexual men are not considered a key population in the HIV response and are mostly absent from pre-exposure prophylaxis (PrEP) studies to date. Yet, South African men face considerable HIV risk. We estimate the incremental cost-effectiveness of providing oral PrEP, injectable PrEP, or a combination of both to heterosexual South African men to assess whether providing PrEP would efficiently use resources. Epidemiological and costing models estimated the one-year costs and outcomes associated with PrEP use in 3 scenarios. PrEP uptake was estimated for younger (aged 18-24) and older (aged 25-49) men using a discrete choice experiment. Scenarios were compared with a baseline scenario of male condom use, while a health system perspective was used to estimate discounted lifetime costs averted per HIV infection. PrEP benefit was estimated in disability-adjusted life years (DALYs) averted. Uncertainty around the estimated incremental cost-effectiveness ratios (ICERs) was assessed using deterministic and probabilistic sensitivity analyses. No PrEP intervention scenarios were cost-effective for both age groups at a willingness-to-pay threshold of $1175/DALY averted. The lowest ICER ($2873/DALY averted) was for the provision of oral PrEP to older men, although probability of cost-effectiveness was just 0.26%. Results found that ICERs were sensitive to HIV incidence and antiretroviral coverage. This study estimates that providing PrEP to heterosexual South African men is not cost-effective at current cost-effectiveness thresholds. Given the ICERs' sensitivity to several variables, alongside the heterogeneity of HIV infection among South African men, PrEP may be cost-effective for older men with high incidence and other subgroups based on locality and race. We recommend further investigation to better identify and target these groups.


[Community composition and diversity of endophytic fungi from roots of Sinopodophyllum hexandrum in forest of Upper-north mountain of Qinghai province].
High throughput sequencing technology is also called Next Generation Sequencing (NGS), which can sequence hundreds and thousands sequences in different samples at the same time. In the present study, the culture-independent high throughput sequencing technology was applied to sequence the fungi metagenomic DNA of the fungal internal transcribed spacer 1(ITS 1) in the root of Sinopodophyllum hexandrum. Sequencing data suggested that after the quality control, 22 565 reads were remained. Cluster similarity analysis was done based on 97% sequence similarity, which obtained 517 OTUs for the three samples (LD1, LD2 and LD3). All the fungi which identified from all the reads of OTUs based on 0.8 classification thresholds using the software of RDP classifier were classified as 13 classes, 35 orders, 44 family, 55 genera. Among these genera, the genus of Tetracladium was the dominant genera in all samples(35.49%, 68.55% and 12.96%).The Shannon's diversity indices and the Simpson indices of the endophytic fungi in the samples ranged from 1.75-2.92, 0.11-0.32, respectively.This is the first time for applying high through put sequencing technol-ogyto analyze the community composition and diversity of endophytic fungi in the medicinal plant, and the results showed that there were hyper diver sity and high community composition complexity of endophytic fungi in the root of S. hexandrum. It is also proved that the high through put sequencing technology has great advantage for analyzing ecommunity composition and diversity of endophtye in the plant.


Therapy with efavirenz plus indinavir in patients with extensive prior nucleoside reverse-transcriptase inhibitor experience: a randomized, double-blind, placebo-controlled trial.
A randomized, double-blind, placebo-controlled trial compared efavirenz (600 mg every 24 h) plus indinavir (1000 mg every 8 h) with placebo (every 24 h) plus indinavir (800 mg every 8 h) among 327 nucleoside analogue reverse-transcriptase inhibitor (NRTI)-experienced human immunodeficiency virus (HIV)-infected adults. Patients received </=2 concomitant NRTIs. Eligible patients had CD4 cell counts >50 cells/mm(3), >10,000 plasma HIV-1 RNA copies/mL, and no prior protease inhibitor or non-NRTI therapy. Patients had a mean of 2.8 years of prior NRTI therapy. At 24 weeks, plasma HIV-1 RNA level was <400 copies/mL in 68.2% of efavirenz versus 52.4% of placebo recipients (P=.004). CD4 cell count increases were 104+/-9 cells/mm(3) and 77+/-10 cells/mm(3) in efavirenz and placebo recipients, respectively (P=.023). Responses in efavirenz recipients were sustained at 48 weeks. Thus, efavirenz plus indinavir with concomitant NRTIs is effective therapy for NRTI-experienced patients.


Tunable optical parametric oscillators pumped by Ti:sapphire lasers.
We report on the application of tunable, pulsed Ti:sapphire lasers as efficient pump sources for optical parametric oscillators. Using a 90-deg phase-matched KTP crystal, we have generated output from 1030 to 1280 nm (signal) and 2180 to 3030 nm (idler) by tuning the pump laser from 700 to 900 nm, with 49 mJ of combined signal and idler output energy at a pump level of 110 mJ. In addition, we have demonstrated degenerate optical parametric oscillator operation with a KNbO(3) crystal over the pump range of 720-818 nm and observed a 44% conversion efficiency.


Assessing the impact of differential genotyping errors on rare variant tests of association.
Genotyping errors are well-known to impact the power and type I error rate in single marker tests of association. Genotyping errors that happen according to the same process in cases and controls are known as non-differential genotyping errors, whereas genotyping errors that occur with different processes in the cases and controls are known as differential genotype errors. For single marker tests, non-differential genotyping errors reduce power, while differential genotyping errors increase the type I error rate. However, little is known about the behavior of the new generation of rare variant tests of association in the presence of genotyping errors. In this manuscript we use a comprehensive simulation study to explore the effects of numerous factors on the type I error rate of rare variant tests of association in the presence of differential genotyping error. We find that increased sample size, decreased minor allele frequency, and an increased number of single nucleotide variants (SNVs) included in the test all increase the type I error rate in the presence of differential genotyping errors. We also find that the greater the relative difference in case-control genotyping error rates the larger the type I error rate. Lastly, as is the case for single marker tests, genotyping errors classifying the common homozygote as the heterozygote inflate the type I error rate significantly more than errors classifying the heterozygote as the common homozygote. In general, our findings are in line with results from single marker tests. To ensure that type I error inflation does not occur when analyzing next-generation sequencing data careful consideration of study design (e.g. use of randomization), caution in meta-analysis and using publicly available controls, and the use of standard quality control metrics is critical.


Viruses disrupt functions of human lymphocytes. Effects of measles virus and influenza virus on lymphocyte-mediated killing and antibody production.
We present experimental data that offer, in part, a better understanding of the immunosuppression that accompanies measles virus infection. We note that measles virus "silently" infects human lymphocytes and that the infection does not alter lymphocyte survival in vitro. Yet such infected lymphocytes fail to generate natural killer (NK) cell activity or synthesize immunoglobulins (Ig). Thus, the presence of virus within lymphocytes impairs their specific immune functions in the absence of cytolysis. Influenza virus also infects human lymphocytes. In contrast to measles virus infection of resting lymphocytes in which viral antigen is rarely expressed, influenza virus infection of these cells yields viral antigens expressed in the cytoplasm and on the cell surface. Influenza virus-infected lymphocytes have normal NK cell activity but fail to synthesize IgG or IgM.


Gene-based testing of interactions in association studies of quantitative traits.
Various methods have been developed for identifying gene-gene interactions in genome-wide association studies (GWAS). However, most methods focus on individual markers as the testing unit, and the large number of such tests drastically erodes statistical power. In this study, we propose novel interaction tests of quantitative traits that are gene-based and that confer advantage in both statistical power and biological interpretation. The framework of gene-based gene-gene interaction (GGG) tests combine marker-based interaction tests between all pairs of markers in two genes to produce a gene-level test for interaction between the two. The tests are based on an analytical formula we derive for the correlation between marker-based interaction tests due to linkage disequilibrium. We propose four GGG tests that extend the following P value combining methods: minimum P value, extended Simes procedure, truncated tail strength, and truncated P value product. Extensive simulations point to correct type I error rates of all tests and show that the two truncated tests are more powerful than the other tests in cases of markers involved in the underlying interaction not being directly genotyped and in cases of multiple underlying interactions. We applied our tests to pairs of genes that exhibit a protein-protein interaction to test for gene-level interactions underlying lipid levels using genotype data from the Atherosclerosis Risk in Communities study. We identified five novel interactions that are not evident from marker-based interaction testing and successfully replicated one of these interactions, between SMAD3 and NEDD9, in an independent sample from the Multi-Ethnic Study of Atherosclerosis. We conclude that our GGG tests show improved power to identify gene-level interactions in existing, as well as emerging, association studies.


Primary and secondary allergies to laboratory animals.
Although laboratory animal allergy (LAA) is a significant occupational hazard among workers exposed to laboratory animals, few studies have evaluated long-term risks to workers who remain in the workplace. This short-term focus has obscured the evaluation of subsequent animal allergies (secondary LAA). We analyzed surveillance data from a 10-year LAA prevention program to estimate incidence rates of primary and secondary LAA and to evaluate the effectiveness of the prevention program in reducing the development of primary LAA. The 10-year incidence rates of primary and secondary LAA were 1.34 (95% CI, 0.78-1.90) and 11 (95% CI, 7.4-14.6) cases per 100 person-years, respectively. The annual incidence of primary LAA was reduced from 3.6% to 0 in the first 5 years and did not rise above 1.2% over the remaining years, whereas the incidence of secondary LAA was greater than 8% in most years. These findings suggest that programs effective at preventing primary LAA may need to be evaluated for their effectiveness at protecting against further risk.


Heterospecies partition analysis reveals binding curve and stoichiometry of protein interactions in living cells.
Measuring the binding curve and stoichiometry of protein complexes in living cells is a prerequisite for quantitative modeling of cellular processes. Dual-color fluorescence fluctuation spectroscopy provides a general framework for detecting protein interactions, but lacks suitable methods for quantifying protein heterointeractions in the cell. We address this challenge by introducing heterospecies partition (HSP) analysis for protein heterointeractions of the type D + nA<-->DA(n). HSP directly identifies the heterointeracting species from the sample mixture and determines the binding curve and stoichiometry of the protein complex. The HSP method is applied to provide the first direct characterization of the ligand-dependent binding of the retinoic X receptor to the coactivator transcription intermediate factor 2. A previous study based on protein fragments observed a higher binding stoichiometry than biologically expected. We address this difference in stoichiometry by measuring the binding curves of the full-length proteins in living cells. This study provides proof-of-principle experiments that illustrate the potential of HSP as a general and robust analysis tool for the quantitative characterization of protein heterointeractions by dual-color fluorescence fluctuation spectroscopy in living cells.


Neurodegeneration and unfolded-protein response in mice expressing a membrane-tethered flexible tail of PrP.
The cellular prion protein (PrPC) consists of a flexible N-terminal tail (FT, aa 23-128) hinged to a membrane-anchored globular domain (GD, aa 129-231). Ligation of the GD with antibodies induces rapid neurodegeneration, which is prevented by deletion or functional inactivation of the FT. Therefore, the FT is an allosteric effector of neurotoxicity. To explore its mechanism of action, we generated transgenic mice expressing the FT fused to a GPI anchor, but lacking the GD (PrPΔ141-225, or "FTgpi"). Here we report that FTgpi mice develop a progressive, inexorably lethal neurodegeneration morphologically and biochemically similar to that triggered by anti-GD antibodies. FTgpi was mostly retained in the endoplasmic reticulum, where it triggered a conspicuous unfolded protein response specifically activating the PERK pathway leading to phosphorylation of eIF2α and upregulation of CHOP ultimately leading to neurodegeration similar to what was observed in prion infection.


Mutations in the gene encoding the serine protease inhibitor, Kazal type 1 are associated with chronic pancreatitis.
Chronic pancreatitis (CP) is a continuing or relapsing inflammatory disease of the pancreas. In approximately one-third of all cases, no aetiological factor can be found, and these patients are classified as having idiopathic disease. Pathophysiologically, autodigestion and inflammation may be caused by either increased proteolytic activity or decreased protease inhibition. Several studies have demonstrated mutations in the cationic trypsinogen gene (PRSS1) in patients with hereditary or idiopathic CP. It is thought that these mutations result in increased trypsin activity within the pancreatic parenchyma. Most patients with idiopathic or hereditary CP, however, do not have mutations in PRSS1 (ref. 4). Here we analysed 96 unrelated children and adolescents with CP for mutations in the gene encoding the serine protease inhibitor, Kazal type 1 (SPINK1), a pancreatic trypsin inhibitor. We found mutations in 23% of the patients. In 18 patients, 6 of whom were homozygous, we detected a missense mutation of codon 34 (N34S). We also found four other sequence variants. Our results indicate that mutations in SPINK1 are associated with chronic pancreatitis.


Parent-Adolescent Communication and Adolescent Delinquency: Unraveling Within-Family Processes from Between-Family Differences.
Understanding the factors that predict adolescent delinquency is a key topic in parenting research. An open question is whether prior results indicating relative differences between families reflect the dynamic processes occurring within families. Therefore, this study investigated concurrent and lagged associations among parental behavioral control, parental solicitation, adolescent disclosure, and adolescent delinquency by separating between-family and within-family effects in three-wave annual data (N = 1515; Mage = 13.01 years at T1; 50.6% girls). At the within-family level, parental behavioral control negatively predicted adolescent delinquency. Adolescent disclosure and delinquency, and adolescent disclosure and parental solicitation, reciprocally predicted each other. Parental solicitation negatively predicted parental behavioral control. The findings indicate a prominent role of adolescent disclosure in within-family processes concerning parental-adolescent communication and adolescent delinquency.


Melatonin as an endogenous regulator of diseases: The role of autophagy.
Melatonin has long been known for its apparent effects on sleep and circadian rhythm. It may participate as a possible therapeutic agent in neurodegenerative, cardiovascular, and endocrine disorders as well. Autophagy is a lysosomal degradation process that occurs in response to starvation and other stresses. Recent studies have reported that melatonin may modulate the autophagy process. We reviewed the current literature that has reported on how different diseases and/or experimental models change autophagy parameters. We also discussed the effect of melatonin on autophagy parameters in the central nervous, cardiovascular, gastrointestinal and endocrine systems as reported in nonclinical studies. Moreover, the molecular targets for melatonin are discussed in details. In summary, melatonin has been reported to enhance significant protective effects in different in vitro and in vivo studies either through enhancement or inhibition of the autophagy process. Melatonin holds promise in the treatment of several major diseases. Regulation of autophagy by melatonin is a determinant parameter that should be considered in the future studies.


Potent and specific genetic interference by double-stranded RNA in Caenorhabditis elegans.
Experimental introduction of RNA into cells can be used in certain biological systems to interfere with the function of an endogenous gene. Such effects have been proposed to result from a simple antisense mechanism that depends on hybridization between the injected RNA and endogenous messenger RNA transcripts. RNA interference has been used in the nematode Caenorhabditis elegans to manipulate gene expression. Here we investigate the requirements for structure and delivery of the interfering RNA. To our surprise, we found that double-stranded RNA was substantially more effective at producing interference than was either strand individually. After injection into adult animals, purified single strands had at most a modest effect, whereas double-stranded mixtures caused potent and specific interference. The effects of this interference were evident in both the injected animals and their progeny. Only a few molecules of injected double-stranded RNA were required per affected cell, arguing against stochiometric interference with endogenous mRNA and suggesting that there could be a catalytic or amplification component in the interference process.


Variability of respiration and sleep during polysomnography in individuals with TBI.
Obstructive sleep apnea (OSA) is commonly found in individuals with traumatic brain injury (TBI) and may exacerbate TBI-related symptoms. Nocturnal polysomnography (NPSG) is considered the gold standard for detecting the presence of sleep apnea. However, there is a limitation with its use known as the "first-night effect" (aberrant polysomnography findings on the first night in a sleep lab). The primary objectives were to investigate the night-to-night consistency of diagnosing and classifying obstructive sleep apnea in individuals with TBI, and ascertain if individuals with TBI are prone to a first-night effect. 47 community-dwelling adults with self-reported mild-to-severe TBI underwent two nights of in-laboratory NPSG to examine variability between the first and second night with regards to OSA diagnosis and severity as well as sleep architecture. OSA detection and severity were consistent from night-to-night in 89% of participants with TBI. Participants with TBI demonstrated longer REM latency on the first night compared to the second night of sleep study. These findings indicate that two nights of in-laboratory NPSG are generally consistent in reliably diagnosing OSA in individuals with TBI and that first-night effects are minimal. One night of NPSG has diagnostic utility in the evaluation of sleep disorders in individuals with TBI.


Placental mesenchymal stromal cells derived from blood vessels or avascular tissues: what is the better choice to support endothelial cell function?
Mesenchymal stromal cells (MSCs) are promising tools for therapeutic revascularization of ischemic tissues and for support of vessel formation in engineered tissue constructs. Recently, we could show that avascular-derived MSCs from placental amnion release soluble factors that exhibit survival-enhancing effects on endothelial cells (ECs). We hypothesize that MSCs derived from placental blood vessels might have even more potent angiogenic effects. Therefore, we isolated and characterized MSCs from placental chorionic blood vessels (bv-MSCs) and tested their angiogenic potential in comparison to amnion-derived avascular MSCs (av-MSCs). bv-MSCs express a very similar surface marker profile compared with av-MSCs and could be differentiated toward the adipogenic and osteogenic lineages. bv-MSCs exert immunosuppressive properties on peripheral blood mononuclear cells, suggesting that they are suitable for cell transplantation settings. Conditioned medium (Cdm) from av-MSCs and bv-MSCs significantly enhanced EC viability, whereas only Cdm from bv-MSCs significantly increased EC migration and network formation (Matrigel assay). Angiogenesis array analysis of av- and bv-MSC-Cdm revealed a similar secretion pattern of angiogenic factors, including angiogenin, interleukins-6 and -8, and tissue inhibitors of matrix metalloproteinase-1 and 2. Enzyme-linked immunosorbent assay analysis showed that, in contrast to av-MSCs, bv-MSCs secreted vascular endothelial growth factor. In direct coculture with bv-MSCs, ECs showed a significantly increased formation of vessel-like structures compared with av-MSCs. With regard to therapeutic treatment, bv-MSCs and particularly their Cdm might be valuable to stimulate angiogenesis especially in ischemic tissues. av-MSCs and their Cdm could be beneficial in conditions when it is required to promote the survival and stabilization of blood vessels without the risk of unmeant angiogenesis.


Management of otitis media-related diseases in children with a cochlear implant.
In all, 4 of 19 children with a cochlear implant (CI) with otitis media developed cholesteatoma. The insertion of a CI may interfere with normal mastoid pneumatization, stimulate mucosa as foreign body, or act as a nidus of infection. Regular follow-up is necessary for children with a CI at the prevalent ages of otitis media. Antibiotic treatment is suggested whenever acute otitis media is suspected, to prevent chronic progress of infection, cholesteatoma, or even meningitis. To evaluate treatment modality and outcomes of otitis media-related diseases in children with a CI. This was a retrospective case review in a tertiary referral center of Taipei Medical University, Chi Mei Medical Center. All patients had a history of otitis media or related disease in the implanted ear. Treatments included antibiotics, tympanotomy, and tympanomastoidectomy. In all, 19 of 186 children with a CI were identified as having otitis media, and 4 of them as having cholesteatoma. Among the others, 10 were identified as having acute otitis media, 4 as having middle ear effusion, and 1 as having mastoiditis in the implanted ear. Surgery was performed in children with cholesteatoma and mastoiditis. The CIs of three children were eventually explanted to eradicate cholesteatoma.


Evidence of plasma CoQ10-lowering effect by HMG-CoA reductase inhibitors: a double-blind, placebo-controlled study.
Inhibitors of HMG-CoA reductase are new safe and effective cholesterol-lowering agents. Elevation of alanine-amino transferase (ALT) and aspartate-amino transferase (AST) has been described in a few cases and a myopathy with elevation of creatinine kinase (CK) has been reported rarely. The inhibition of HMG-CoA reductase affects also the biosynthesis of ubiquinone (CoQ10). We studied two groups of five healthy volunteers treated with 20 mg/day of pravastatin (Squibb, Italy) or simvastatin (MSD) for a month. Then we treated 30 hypercholesterolemic patients in a double-blind controlled study with pravastatin, simvastatin (20 mg/day), or placebo for 3 months. At the beginning, and 3 months thereafter we measured plasma total cholesterol, CoQ10, ALT, AST, CK, and other parameters (urea, creatinine, uric acid, total bilirubin, gamma GT, total protein). Significant changes in the healthy volunteer group were detected for total cholesterol and CoQ10 levels, which underwent about a 40% reduction after the treatment. The same extent of reduction, compared with placebo was measured in hypercholesterolemic patients treated with pravastatin or simvastatin. Our data show that the treatment with HMG-CoA reductase inhibitors lowers both total cholesterol and CoQ10 plasma levels in normal volunteers and in hypercholesterolemic patients. CoQ10 is essential for the production of energy and also has antioxidative properties. A diminution of CoQ10 availability may be the cause of membrane alteration with consequent cellular damage.


Electrochemical behavior and electrodeposition of gallium in 1,2-dimethoxyethane-based electrolytes.
The electrochemical behavior and electrodeposition of gallium was studied in a non-aqueous electrolyte comprising of gallium(iii) chloride and 1,2-dimethoxyethane (DME). Electrochemical quartz crystal microbalance (EQCM) and rotating ring disk electrode (RRDE) measurements indicate that reduction of gallium(iii) is a two-step process: first from gallium(iii) to gallium(i), and then from gallium(i) to gallium(0). The morphology and elemental composition of the electrodeposited layer were examined using scanning electron microscopy (SEM) and energy-dispersive X-ray spectroscopy (EDX). Metallic gallium was deposited as spheres with diameters of several hundred nanometers that were stacked on top of each other. X-ray photoelectron spectroscopy (XPS) revealed that each gallium sphere was covered by a thin gallium oxide shell. Electrochemical experiments indicated that these oxide layers are electrically conductive, as gallium can be electrodeposited and partially stripped on or from the layer of spheres below. This was further evidenced by simultaneous electrodeposition of gallium and indium, using indium as a tracer. Electrodeposition of gallium from an O2-containing electrolyte resulted in spheres with smaller diameters. This was due to the formation thicker oxide shells, through which diffusion of gallium atoms that were electrodeposited on the surface, was slower. The concentration of gallium adatoms on top of the gallium spheres to form a new sphere therefore reaches the critical concentration for nucleating a new gallium sphere sooner, leading to smaller spheres.


Leukocyte entry into the CNS of Leishmania chagasi naturally infected dogs.
In dogs, there is an association of chronic visceral leishmaniasis with neurological symptoms, and very few publications have investigated whether these neurological manifestations correlate with specific alterations in brain. A total of 42 mixed-breed adult dogs were selected from the Veterinary Hospital of UNESP-Araçatuba and the Control Zoonosis Center in Araçatuba, São Paulo State, Brazil, which is an endemic area for visceral leishmaniasis. Animals presenting positive ELISA and/or positive parasitological diagnosis of Leishmania were enrolled in the group of infected dogs (n=32). Animals with negative ELISA results and parasitological tests for Leishmania, including a negative immunofluorescence test for toxoplasmosis and neosporosis, were included as the control group (n=10). Brain samples were collected, stored in 10% buffered formalin and subjected to routine histological procedures, following by staining with haematoxylin-eosin (HE) and immunohistochemical examination for T and B lymphocytes and phagocytic cells. Cerebrospinal fluid was collected to determine the anti-Leishmania antibody titers. Histological examination of HE stains demonstrated intense inflammatory infiltrate, primarily in the choroid plexus, which was composed of mononuclear cells with no detectable parasites. Immunohistochemistry revealed that CD3(+) T lymphocytes were the major components of the inflammatory infiltrate at the choroid plexus and in the brain. Infected dogs had more CD3(+) T cells than uninfected animals (P=0.0002). Cerebrospinal fluid from infected dogs contained high titers of anti-Leishmania antibodies in comparison with control animals (P<0.0001), which suggests a compromise of the blood-cerebrospinal fluid barrier. Leukocyte entry into the brain suggests the participation of these cells in the pathogenesis of neurological disorders during the advanced stages of leishmaniasis and confirms that the choroid plexus is an important structure for T cell influx.


Getting to the core: Internal body temperatures help reveal the ecological function and thermal implications of the lions' mane.
It has been proposed that there is a thermal cost of the mane to male lions, potentially leading to increased body surface temperatures (Ts), increased sperm abnormalities, and to lower food intake during hot summer months. To test whether a mane imposes thermal costs on males, we measured core body temperature (Tb) continuously for approximately 1 year in 18 free-living lions. There was no difference in the 24-hr maximum Tb of males (n = 12) and females (n = 6), and males had a 24-hr mean Tb that was 0.2 ± 0.1°C lower than females after correcting for seasonal effects. Although feeding on a particular day increased 24-hr mean and 24-hr maximum Tb, this phenomenon was true of both male and female lions, and females had higher 24-hr mean and 24-hr maximum Tb than males, on both days when lions did not feed, and on days when lions did feed. Twenty-four-hour Tb was not influenced by mane length or color, and 24-hr mean Tb was negatively correlated with mane length. These data contradict the suggestion that there exists a thermal cost to male lions in possessing a long dark mane, but do not preclude the possibility that males compensate for a mane with increased heat loss. The increased insulation caused by a mane does not necessarily have to impair heat loss by males, which in hot environments is primarily through respiratory evaporative cooling, nor does in necessarily lead to increased heat gain, as lions are nocturnal and seek shade during the day. The mane may even act as a heat shield by increasing insulation. However, dominant male lions frequent water points more than twice as often as females, raising the possibility that male lions are increasing water uptake to facilitate increased evaporative cooling. The question of whether male lions with manes compensate for a thermal cost to the mane remains unresolved, but male lions with access to water do not have higher Tb than females or males with smaller manes.


Rehabilitating mangrove ecosystem services: A case study on the relative benefits of abandoned pond reversion from Panay Island, Philippines.
Mangroves provide vital climate change mitigation and adaptation (CCMA) ecosystem services (ES), yet have suffered extensive tropics-wide declines. To mitigate losses, rehabilitation is high on the conservation agenda. However, the relative functionality and ES delivery of rehabilitated mangroves in different intertidal locations is rarely assessed. In a case study from Panay Island, Philippines, using field- and satellite-derived methods, we assess carbon stocks and coastal protection potential of rehabilitated low-intertidal seafront and mid- to upper-intertidal abandoned (leased) fishpond areas, against reference natural mangroves. Due to large sizes and appropriate site conditions, targeted abandoned fishpond reversion to former mangrove was found to be favourable for enhancing CCMA in the coastal zone. In a municipality-specific case study, 96.7% of abandoned fishponds with high potential for effective greenbelt rehabilitation had favourable tenure status for reversion. These findings have implications for coastal zone management in Asia in the face of climate change.


Validation of the Endopep-MS method for qualitative detection of active botulinum neurotoxins in human and chicken serum.
Botulinum neurotoxins (BoNTs) are highly toxic proteases produced by anaerobic bacteria. Traditionally, a mouse bioassay (MBA) has been used for detection of BoNTs, but for a long time, laboratories have worked with alternative methods for their detection. One of the most promising in vitro methods is a combination of an enzymatic and mass spectrometric assay called Endopep-MS. However, no comprehensive validation of the method has been presented. The main purpose of this work was to perform a validation for the qualitative analysis of BoNT-A, B, C, C/D, D, D/C, and F in serum. The limit of detection (LOD), selectivity, precision, stability in matrix and solution, and correlation with the MBA were evaluated. The LOD was equal to or even better than that of the MBA for BoNT-A, B, D/C, E, and F. Furthermore, Endopep-MS was for the first time successfully used to differentiate between BoNT-C and D and their mosaics C/D and D/C by different combinations of antibodies and target peptides. In addition, sequential antibody capture was presented as a new way to multiplex the method when only a small sample volume is available. In the comparison with the MBA, all the samples analyzed were positive for BoNT-C/D with both methods. These results indicate that the Endopep-MS method is a valid alternative to the MBA as the gold standard for BoNT detection based on its sensitivity, selectivity, and speed and that it does not require experimental animals.


Studies on the activator 1 protein complex, an accessory factor for proliferating cell nuclear antigen-dependent DNA polymerase delta.
Activator 1 (A1) is a multiprotein complex which is essential for proliferating cell nuclear antigen (PCNA)-dependent DNA polymerase delta (pol delta) activity and efficient in vitro DNA synthesis in the SV40 dipolymerase replication system. In this report, we describe the isolation of A1 from HeLa cytosolic extracts. A1 stimulated pol delta activity in singly primed phi X174 DNA or (dA)4500.oligo(dT)12-18 in reactions containing PCNA, single-stranded DNA binding protein (SSB), and ATP. Using this assay, A1 has been extensively purified. Purified preparations contained five discrete subunits of 145, 40, 38, 37, and 36.5 kDa. ATP hydrolysis to ADP and Pi is essential for A1-dependent pol delta activity, and we have shown that A1 contains an intrinsic ATPase which is stimulated by DNA. The DNA-dependent hydrolysis of ATP can be stimulated by PCNA and further activated by PCNA plus the human single-stranded DNA binding protein. These stimulatory effects were observed with (dA)4500.oligo(dT)12-18, but were not detected with each poly-deoxynucleotide alone. Furthermore, A1 formed a complex with (dA)4500.oligo(dT)12-18 which could be measured by nitrocellulose binding. No complex with (dA)4500 or oligo(dT)12-18 alone was detected by this procedure. Data are also presented which indicate that A1, in conjunction with PCNA, functions as a primer-recognition factor for pol delta, increasing its ability to utilize low levels of primer ends, but it does not increase the size of the DNA products. A1 also markedly reduced the amount of PCNA required for pol delta activity on a multiply primed DNA suggesting that PCNA interacts with A1 at the primer end. These multiple effects of A1 closely resemble the properties of the multisubunit protein RF-C described by Tsurimoto and Stillman (Tsurimoto, T., and Stillman, B. (1990) Proc. Natl. Acad. Sci. U.S.A. 87, 1023-1027).


Dda helicase tightly couples translocation on single-stranded DNA to unwinding of duplex DNA: Dda is an optimally active helicase.
Helicases utilize the energy of ATP hydrolysis to unwind double-stranded DNA while translocating on the DNA. Mechanisms for melting the duplex have been characterized as active or passive, depending on whether the enzyme actively separates the base pairs or simply sequesters single-stranded DNA (ssDNA) that forms due to thermal fraying. Here, we show that Dda translocates unidirectionally on ssDNA at the same rate at which it unwinds double-stranded DNA in both ensemble and single-molecule experiments. Further, the unwinding rate is largely insensitive to the duplex stability and to the applied force. Thus, Dda transduces all of its translocase activity into DNA unwinding activity so that the rate of unwinding is limited by the rate of translocation and that the enzyme actively separates the duplex. Active and passive helicases have been characterized by dividing the velocity of DNA unwinding in base pairs per second (V(un)) by the velocity of translocation on ssDNA in nucleotides per second (V(trans)). If the resulting fraction is 0.25, then a helicase is considered to be at the lower end of the "active" range. In the case of Dda, the average DNA unwinding velocity was 257±42 bp/s, and the average translocation velocity was 267±15 nt/s. The V(un)/V(trans) value of 0.96 places Dda in a unique category of being an essentially "perfectly" active helicase.