An empirical assessment of the focal species hypothesis.
Biodiversity surrogates and indicators are commonly used in conservation management. The focal species approach (FSA) is one method for identifying biodiversity surrogates, and it is underpinned by the hypothesis that management aimed at a particular focal species will confer protection on co-occurring species. This concept has been the subject of much debate, in part because the validity of the FSA has not been subject to detailed empirical assessment of the extent to which a given focal species actually co-occurs with other species in an assemblage. To address this knowledge gap, we used large-scale, long-term data sets of temperate woodland birds to select focal species associated with threatening processes such as habitat isolation and loss of key vegetation attributes. We quantified co-occurrence patterns among focal species, species in the wider bird assemblage, and species of conservation concern. Some, but not all, focal species were associated with high levels of species richness. One of our selected focal species was negatively associated with the occurrence of other species (i.e., it was an antisurrogate)-a previously undescribed property of nominated focal species. Furthermore, combinations of focal species were not associated with substantially elevated levels of bird species richness, relative to levels associated with individual species. Our results suggest that although there is some merit to the underpinning concept of the FSA, there is also a need to ensure that actions are sufficiently flexible because management tightly focused on a given focal species may not benefit some other species, including species of conservation concern, such of which might not occur in species-rich assemblages.


The molecular basis of variable phenotypic severity among common missense mutations causing Rett syndrome.
Rett syndrome is caused by mutations in the X-linked MECP2 gene, which encodes a chromosomal protein that binds to methylated DNA. Mouse models mirror the human disorder and therefore allow investigation of phenotypes at a molecular level. We describe an Mecp2 allelic series representing the three most common missense Rett syndrome (RTT) mutations, including first reports of Mecp2[R133C] and Mecp2[T158M] knock-in mice, in addition to Mecp2[R306C] mutant mice. Together these three alleles comprise ∼25% of all RTT mutations in humans, but they vary significantly in average severity. This spectrum is mimicked in the mouse models; R133C being least severe, T158M most severe and R306C of intermediate severity. Both R133C and T158M mutations cause compound phenotypes at the molecular level, combining compromised DNA binding with reduced stability, the destabilizing effect of T158M being more severe. Our findings contradict the hypothesis that the R133C mutation exclusively abolishes binding to hydroxymethylated DNA, as interactions with DNA containing methyl-CG, methyl-CA and hydroxymethyl-CA are all reduced in vivo. We find that MeCP2[T158M] is significantly less stable than MeCP2[R133C], which may account for the divergent clinical impact of the mutations. Overall, this allelic series recapitulates human RTT severity, reveals compound molecular aetiologies and provides a valuable resource in the search for personalized therapeutic interventions.


D120 and K152 within the PH Domain of T Cell Adapter SKAP55 Regulate Plasma Membrane Targeting of SKAP55 and LFA-1 Affinity Modulation in Human T Lymphocytes.
The β2-integrin lymphocyte function-associated antigen 1 (LFA-1) is needed for the T cell receptor (TCR)-induced activation of LFA-1 to promote T cell adhesion and interaction with antigen-presenting cells (APCs). LFA-1-mediated cell-cell interactions are critical for proper T cell differentiation and proliferation. The Src kinase-associated phosphoprotein of 55 kDa (SKAP55) is a key regulator of TCR-mediated LFA-1 signaling (inside-out/outside-in signaling). To gain an understanding of how SKAP55 controls TCR-mediated LFA-1 activation, we assessed the functional role of its pleckstrin homology (PH) domain. We identified two critical amino acid residues within the PH domain of SKAP55, aspartic acid 120 (D120) and lysine 152 (K152). D120 facilitates the retention of SKAP55 in the cytoplasm of nonstimulated T cells, while K152 promotes SKAP55 membrane recruitment via actin binding upon TCR triggering. Importantly, the K152-dependent interaction of the PH domain with actin promotes the binding of talin to LFA-1, thus facilitating LFA-1 activation. These data suggest that K152 and D120 within the PH domain of SKAP55 regulate plasma membrane targeting and TCR-mediated activation of LFA-1.


Phenotypic and genotypic typing of food and clinical isolates of Enterobacter sakazakii.
Enterobacter sakazakii, designated a unique species in 1980, has been implicated as the causative organism in a rare but severe form of neonatal meningitis. Dried infant formula milk has been identified as a potential source of the organism. E. sakazakii isolates from dried infant formula available in Canada and clinical isolates obtained from Canadian hospital culture collections were characterised by phenotypic (biotype and antibiograms) and genotypic (ribotyping, random amplification of polymorphic DNA and pulsed-field gel electrophoresis) methods. Three biotypes and four antibiogram patterns were observed in the 18 isolates examined. Ribotyping with the Dupont Riboprinter microbial identification system divided the 18 isolates into 10 ribotypes. Three isolates from the same hospital had indistinguishable ribotyping patterns although each was isolated in a different year, as did three food isolates from one company. Pulsed-field gel electrophoresis (PFGE) and random amplification of polymorphic DNA (RAPD) profiles indicated minor differences between the isolates that were indistinguishable by ribotyping. PFGE (with the restriction endonucleases Xba1 and Spe1) and RAPD gave discrete patterns that enabled easy comparison of E. sakazakii isolates, with a high degree of discrimination. The discriminatory index showed RAPD and PFGE were shown to be the most discriminatory typing schemes for E. sakazakii, followed by ribotyping, biotyping and antibiograms.


Preferential outgrowth of central nervous system neurites on astrocytes and Schwann cells as compared with nonglial cells in vitro.
I have compared central nervous system (CNS) neurite outgrowth on glial and nonglial cells. Monolayers of glial cells (astrocytes and Schwann cells) or nonglial cells (e.g., fibroblasts) were prepared and were shown to be greater than 95% pure as judged by cell type-specific markers. These monolayers were then tested for their ability to support neurite outgrowth from various CNS explants. While CNS neurites grew vigorously on the glial cells, most showed little growth on nonglial cell monolayers. Neurites grew singly or in fine fascicles on the glial cells at rates greater than 0.5 mm/d. The neurite outgrowth on astrocytes was investigated in detail. Scanning and transmission electron microscopy showed that the neurites were closely apposed to the astrocyte surface and that the growth cones were well spread with long filopodia. There was no evidence of significant numbers of explant-derived cells migrating onto the monolayers. Two types of experiments indicated that factors associated with the astrocyte surface were primarily responsible for the vigorous neurite outgrowth seen on these cells: (a) Conditioned media from either astrocytes or fibroblasts had no effect on the pattern of outgrowth on fibroblasts and astrocytes, and conditioned media factors from either cell type did not promote neurite outgrowth when bound to polylysine-coated dishes. (b) When growing CNS neurites encountered a boundary between astrocytes and fibroblasts, they stayed on the astrocytes and did not encroach onto the fibroblasts. These experiments strongly suggest that molecules specific to the surfaces of astrocytes make these cells particularly attractive substrates for CNS neurite outgrowth, and they raise the possibility that similar molecules on embryonic glial cells may play a role in guiding axonal growth during normal CNS development.


Quality of reporting internal and external validity data from randomized controlled trials evaluating stents for percutaneous coronary intervention.
Stents are commonly used to treat patients with coronary artery disease. However, the quality of reporting internal and external validity data in published reports of randomised controlled trials (RCTs) of stents has never been assessed.The objective of our study was to evaluate the quality of reporting internal and external validity data in published reports of RCTs assessing the stents for percutaneous coronary interventions. A systematic literature review was conducted. Reports of RCTs assessing stents for percutaneous coronary interventions indexed in MEDLINE and the Cochrane Central Register of Controlled Trials and published between January 2003 and September 2008 were selected. A standardized abstraction form was used to extract data. All analyses were adjusted for the effect of clustering articles by journal. 132 articles were analyzed. The generation of the allocation sequence was adequate in 58.3% of the reports; treatment allocation was concealed in 34.8%. Adequate blinding was reported in one-fifth of the reports. An intention-to-treat analysis was described in 79.5%. The main outcome was a surrogate angiographic endpoint in 47.0%. The volume of interventions per center was described in two reports. Operator expertise was described in five (3.8%) reports. The quality of reporting was better in journals with high impact factors and in journals endorsing the CONSORT statement. The current reporting of results of RCTs testing stents needs to be improved to allow readers to appraise the risk of bias and the applicability of the results.


Exploring the nitrogen ingestion of aphids--a new method using electrical penetration graph and (15)N labelling.
Studying plant-aphid interactions is challenging as aphid feeding is a complex process hidden in the plant tissue. Here we propose a combination of two well established methods to study nutrient acquisition by aphids focusing on the uptake of isotopically labelled nitrogen ((15)N). We combined the Electrical Penetration Graph (EPG) technique that allows detailed recording of aphid feeding behaviour and stable isotope ratio mass spectrometry (IRMS) to precisely measure the uptake of nitrogen. Bird cherry-oat aphids Rhopalosiphum padi L. (Hemiptera, Aphididae) fed for 24 h on barley plants (Hordeum vulgare L., cultivar Lina, Poaceae) that were cultivated with a (15)N enriched nutrient solution. The time aphids fed in the phloem was strongly positive correlated with their (15)N uptake. All other single behavioural phases were not correlated with (15)N enrichment in the aphids, which corroborates their classification as non-feeding EPG phases. In addition, phloem-feeding and (15)N enrichment of aphids was divided into two groups. One group spent only short time in the phloem phase and was unsuccessful in nitrogen acquisition, while the other group displayed longer phloem-feeding phases and was successful in nitrogen acquisition. This suggests that several factors such as the right feeding site, time span of feeding and individual conditions play a role for the aphids to acquire nutrients successfully. The power of this combination of methods for studying plant-aphid interactions is discussed.


Prevention of acute graft-versus-host disease by magnetic nanoparticles of Fe₃O₄ combined with cyclosporin A in murine models.
To investigate the effect of magnetic nanoparticles (MNPs) of Fe(3)O(4) combined with cyclosporin A (CsA) on acute graft-versus-host disease (aGVHD) after allogeneic hematopoietic stem cell transplantation (allo-HSCT) in murine models. BALB/c mice preconditioned with total-body irradiation generated aGVHD and then were followed with allo-HSCT from allogeneic C57BL/6. Recipient mice were randomly divided into five groups and then given different supportive care and followed up. The physical signs and median survival time (MST) were recorded, peripheral blood cell counts were assessed, and histological changes of the main tissues were evaluated with hematoxylin-eosin staining. Furthermore, fluorescence polarization immunoassay was used to monitor the concentration of CsA. The irradiated-only mice developed typical aGVHD, and the typical signs of aGVHD in the skin, liver, and intestine were observed by histopathological examination. Both CsA alone and in combination with Fe(3)O(4) MNPs significantly prolonged the MST of recipient mice compared with both the control and the Fe(3)O(4) MNPs groups. Notably, a combination of CsA with Fe(3)O(4) MNPs can elevate the peripheral white blood cells and alleviate the symptoms of GVHD and the pathological damage after allo-HSCT. In addition, the concentration of CsA was higher in plasma, heart, liver, and spleen of recipient mice with supporting care of the combination of CsA with Fe(3)O(4) MNPs than with CsA alone. Taken together, Fe(3)O(4) MNPs may be used as a carrier of immunosuppressive agents to alleviate GVHD after allo-HSCT in murine models.


Magnetization transfer ratio does not correlate to myelin content in the brain in the MOG-EAE mouse model.
Magnetization transfer ratio (MTR) is a magnetic resonance imaging (MRI) method which may detect demyelination not detected by conventional MRI in the central nervous system of patients with multiple sclerosis (MS). A decrease in MTR value has previously been shown to correlate to myelin loss in the mouse cuprizone model for demyelination. In this study, we investigated the sensitivity of MTR for demyelination in the myelin oligodendrocyte (MOG) 1-125 induced experimental autoimmune encephalomyelitis (EAE) mouse model. A total of 24 female c57Bl/6 mice were randomized to a control group (N = 6) or EAE (N = 18). MTR images were obtained at a preclinical 7 Tesla Bruker MR-scanner before EAE induction (baseline), 17-19 days (midpoint) and 31-32 days (endpoint) after EAE induction. Mean MTR values were calculated in five regions of the brain and compared to weight, EAE severity score and myelin content assessed by immunostaining for proteolipid protein and luxol fast blue, lymphocyte and monocyte infiltration and iron deposition. Contrary to what was expected, MTR values in the EAE mice were higher than in the control mice at the midpoint and endpoint. No significant difference in myelin content was found according to histo- or immunohistochemistry. Changes in MTR values did not correlate to myelin content, iron content, lymphocyte or monocyte infiltration, weight or EAE severity scores. This suggest that MTR measures of brain tissue can give significant differences between control mice and EAE mice not caused by demyelination, inflammation or iron deposition, and may not be useful surrogate markers for demyelination in the MOG1-125 mouse model.


Palliative care; role of family physicians.
Palliative care is the holistic approach to provide relief to patients suffering from life threatening diseases and their families throughout the disease. This is mainly through the prevention and relief of suffering by means of early identification, comprehensive assessment and management of physical, psychosocial and spiritual problems. With the rise of elderly population in the world patients needing palliative care will also increase. Family physicians who are closest to the community and easily accessible has a major role to play in providing palliative care. Their broad knowledge, long standing relationship with patients and their families, ability to carry out home visits and communicate and coordinate with other health care resources place them in an ideal position to address complex issues faced by patients. Keeping up to date with knowledge, acquiring counseling skills, non availability of guide lines and medications, lack of support from team and time constraints are the challenges faced by family physicians in providing palliative care. With the aging population, demand on palliative care resources will increase markedly in the next few decades. Developing palliative care models, improving the skills and opportunities for doctors to learn sound palliative care principles have to be initiated without a delay in order to meet the challenges of the future.


MiR-146a negatively regulates TLR2-induced inflammatory responses in keratinocytes.
Keratinocytes represent the first line of defense against pathogens in the skin and have important roles in initiating and regulating inflammation during infection and autoimmunity. Here we investigated the role of miR-146a in the regulation of the innate immune response of keratinocytes. Toll-like receptor 2 (TLR2) stimulation of primary human keratinocytes resulted in an NF-κB- and mitogen-activated protein kinase-dependent upregulation of miR-146a expression, which was surprisingly long lasting, contrasting with the rapid and transient induction of inflammatory mediators. Overexpression of miR-146a significantly suppressed the production of IL-8, CCL20, and tumor necrosis factor-α, which functionally suppressed the chemotactic attraction of neutrophils by keratinocytes. Inhibition of endogenous miR-146a induced the production of inflammatory mediators even in nonstimulated keratinocytes, and potentiated the effect of TLR2 stimulation. Transcriptomic profiling revealed that miR-146a suppresses the expression of a large number of immune-related genes in keratinocytes. MiR-146a downregulated interleukin-1 receptor-associated kinase 1 and TNF receptor-associated factor 6, two key adapter molecules downstream of TLR signaling, and suppressed NF-κB promoter-binding activity as shown by promoter luciferase experiments. Together, these data identify miR-146a as a regulatory element in keratinocyte innate immunity, which prevents the production of inflammatory mediators under homeostatic conditions and serves as a potent negative feedback regulator after TLR2 stimulation.


Building characterization and aerosol infiltration into a naturally ventilated three-story apartment building.
Understanding infiltration of outdoor pollutants was an integral part of the Brooklyn Traffic Real-Time Ambient Pollutant Penetration and Environmental Dispersion (B-TRAPPED) study. For this reason, the structural and air exchange properties of the three-story row house in Brooklyn, NY, USA, that was used in the B-TRAPPED experiments were fully characterized. Factors investigated included representativeness of the construction and impact of building design features on the natural ventilation and infiltration of outdoor aerosol. Both blower door and perfluorocarbon tracer (PFT) air exchange rate (AER) experiments showed that the ventilation rates of the building were quite typical of similar structures in the New York City (NYC) metropolitan area. Indoor/outdoor (I/O) aerosol count ratios by particle size were comparable to a similar vintage naturally ventilated building in Boston, MA, USA. I/O ratio analyses were consistent with literature findings and showed I/O ratios ranging from 0.310 to 0.601, varying across particle sizes (from 0.3 to 5.0 [corrected] mum) and between first and second floor apartments. An effort to apply the rebound method of Thatcher et al. (Aerosol Sci. Technol., 2003, 37, 847-864) in determining aerosol infiltration rates proved unsuccessful due to unexpectedly long (>60 min) equilibration times after the filtration period. Uninsulated interior wall renovations in the study house created a cavity that resulted in a large intermediate dead volume (for infiltration) that apparently could not be accommodated by a simple infiltration model. Simple two-compartment models evidently have finite application limitations for even modestly complex settings.


The moderating effect of childhood disadvantage on the associations between smoking and occupational exposure and lung function; a cross sectional analysis of the UK Household Longitudinal Study (UKHLS).
Lung function is lower in people with disadvantaged socio-economic position (SEP) and is associated with hazardous health behaviours and exposures. The associations are likely to be interactive, for example, exposure to socially patterned environmental tobacco smoke (ETS) in childhood is associated with an increased effect of smoking in adulthood. We hypothesise that disadvantaged childhood SEP increases susceptibility to the effects of hazards in adulthood for lung function. We test whether disadvantaged childhood SEP moderates smoking, physical activity, obesity, occupational exposures, ETS and air pollution's associations with lung function. Data are from the Nurse Health Assessment (NHA) in waves two and three of the United Kingdom Household Longitudinal Study (UKHLS). Analysis is restricted to English residents aged at least 20 for women and 25 for men, producing a study population of 16,339. Lung function is measured with forced expiratory volume in the first second (FEV1) and standardised to the percentage of expected FEV1 for a healthy non-smoker of equivalent age, gender, height and ethnicity (FEV1%). Using STATA 14, a mixed linear model was fitted with interaction terms between childhood SEP and health behaviours and occupational exposures. Cross level interactions tested whether childhood SEP moderated household ETS and neighbourhood air pollution's associations with FEV1%. SEP, smoking, physical activity, obesity, occupational exposures and air pollution were associated with lung function. Interaction terms indicated a significantly stronger negative association between disadvantaged childhood SEP and currently smoking (coefficient -6.47 %, 95% confidence intervals (CI): 9.51 %, 3.42 %) as well as with formerly smoking and occupational exposures. Significant interactions were not found with physical activity, obesity, ETS and air pollution. The findings suggest that disadvantaged SEP in childhood may make people's lung function more susceptible to the negative effects of smoking and occupational exposures in adulthood. This is important as those most likely to encounter these exposures are at greater risk to their effects. Policy to alleviate this inequality requires intervention in health behaviours through public health campaigns and in occupational health via health and safety legislation.


Phase 2 Study of an Intravenous Busulfan and Melphalan Conditioning Regimen for Autologous Stem Cell Transplantation in Patients with Multiple Myeloma (KMM150).
This prospective study evaluated the efficacy and toxicity of intravenous busulfan and melphalan as a conditioning regimen for autologous stem cell transplantation (ASCT) in patients with multiple myeloma (MM). A total of 99 patients with MM, enrolled between January 2013 and March 2016, received intravenous busulfan (9.6 mg/kg) and melphalan (140 mg/m2) before ASCT. The median time to transplant was 6.2 months, and 90 (90.9%) patients underwent ASCT within 12 months of the diagnosis. The overall response rate after ASCT was 94.0%, including 43.5% with a stringent complete response/complete response, 27.3% with very good partial response, and 23.2% with partial response. The most common severe nonhematologic toxicity (grade 3 to 4) was infection (26.3%) and stomatitis (15.2%). Three (3.2%) patients developed veno-occlusive disease. No treatment-related mortality was observed. After a median follow-up of 26.1 months, the median progression-free survival was 27.2 months (range, 13.0 to 41.4 months) and median overall survival was not reached. In conclusion, a conditioning regimen of intravenous busulfan and melphalan was effective and tolerable. ClinicalTrials.gov. number: NCT01923935.


Inositol 1,2,3-trisphosphate and inositol 1,2- and/or 2,3-bisphosphate are normal constituents of mammalian cells.
1. An inositol trisphosphate (InsP3) distinct from Ins(1,4,5)P3 and Ins(1,3,4)P3, which we previously observed in myeloid and lymphoid cells [French, Bunce, Stephens, Lord, McConnell, Brown, Creba and Michell (1991) Proc R. Soc. London B 245, 193-201; Bunce, French, Allen, Mountford, Moore, Greaves, Michell and Brown (1993) Biochem. J. 289, 667-673], is present in WRK1 rat mammary tumour cells and pancreatic endocrine beta-cells. 2. It has been identified as Ins(1,2,3)P3 by a combination of oxidation to ribitol, a structurally diagnostic polyol, and ammoniacal hydrolysis to identified inositol monophosphates. 3. Ins(1,2,3)P3 concentration in HL60 cells changed little during stimulation by ATP or fMetLeuPhe or during neutrophilic or monocytic differentiation, and Ins(1,2,3)P3 was unresponsive to vasopressin in WRK1 cells. 4. Ins(1,2,3)P3 was usually more abundant than Ins(1,4,5)P3, often being present at concentrations between approximately 1 microM and approximately 10 microM. 5. HL60, WRK-1 and lymphoid cells also contain Ins(1,2)P2 or Ins(2,3)P2, or a mixture of these two enantiomers, as a major InsP2 species. 6. Ins(1,2,3)P3 and Ins(1,2)P2/Ins(2,3)P2 are readily detected in cells labelled for long periods, but not in acutely labelled cells. This behaviour resembles that of InsP6, the most abundant cellular inositol polyphosphate that includes the 1,2,3-trisphosphate motif, which also achieves isotopic equilibrium with inositol only slowly. 7. Ins(1,2,3)P3 is the major InsP3 that accumulates during metabolism of InsP6 by WRK-1 cell homogenates. 8. Possible metabolic relationships between Ins(1,2,3)P3, Ins(1,2)P2/Ins(2,3)P2 and other inositol polyphosphates in cells, and a possible role for Ins(1,2,3)P3 in cellular iron handling, are considered.


The distribution of cytochrome P-450 monooxygenase in cells of the rabbit lung: an ultrastructural immunocytochemical characterization.
The cytochrome P-450 monooxygenase system of the mammalian lung is known to be associated with the microsomal subcellular fraction and has been demonstrated in two pulmonary cell types rich in endoplasmic reticulum: Clara cells and type II pneumocytes. However, analysis of ultracellular fractions, isolated cell preparations, or light microscopic immunohistochemical studies of tissue sections has permitted only limited resolution of the distribution of this enzyme system within the 40 or more cell types of the lung. Therefore, we have used the greater resolving power of transmission electron microscopy and immunogold labeling to characterize the cellular and subcellular distribution of the cytochrome P-450 system in the lung. In Lowicryl-embedded sections of lung from adult rabbits, antisera (1:10,000) against the constitutive pulmonary microsomal cytochrome P-450 monooxygenase isozymes 2 and 5 and NADPH-cytochrome P-450 reductase (anti-2, anti-5 and anti-R) bound specifically to regions known to be rich in agranular endoplasmic reticulum (AER) in the cytoplasm of Clara cells. The plasma membranes of bronchiolar Clara cells, the tips of microvillae of ciliated cells, secretory granules of goblet cells, and the cell membrane and pinocytotic vesicles of endothelial cells were all intensely labeled with anti-2 and anti-5 but not with anti-R, even at a 10-fold higher concentration. The intensity of labeling of AER in Clara cells with anti-R and anti-2, but not anti-5, appeared to correlate positively with the cellular content of secretory granules. The Golgi membranes of ciliated cells were labeled intensely with anti-5 only. The plasma membrane of type II pneumocytes was not labeled by any of the antisera, but with anti-2 or anti-5 there was labeling of AER-associated vacuoles, the membranous residue of lamellar bodies, and, to some extent, mitochondria; at 1:5,000 but not 1:10,000 dilution, staining with anti-R was qualitatively similar. Type I pneumocytes, ciliated cell cytoplasm, and nuclei were essentially unlabeled. Immunoblots (Western) of tracheal homogenates yielded no evidence for epitopes other than those in microsomal fractions from whole lung. Contact blots of fresh whole trachea, before but not after lavage, bound anti-2 and anti-R. Thus, we have demonstrated for the first time that components of the pulmonary cytochrome P-450 monooxygenase, although localized in the AER-rich regions of the Clara cells and type II pneumocytes, are not restricted to these cell types or to the endoplasmic reticulum.(ABSTRACT TRUNCATED AT 400 WORDS)


CT scan range estimation using multiple body parts detection: let PACS learn the CT image content.
The aim of this study was to develop an efficient CT scan range estimation method that is based on the analysis of image data itself instead of metadata analysis. This makes it possible to quantitatively compare the scan range of two studies. In our study, 3D stacks are first projected to 2D coronal images via a ray casting-like process. Trained 2D body part classifiers are then used to recognize different body parts in the projected image. The detected candidate regions go into a structure grouping process to eliminate false-positive detections. Finally, the scale and position of the patient relative to the projected figure are estimated based on the detected body parts via a structural voting. The start and end lines of the CT scan are projected to a standard human figure. The position readout is normalized so that the bottom of the feet represents 0.0, and the top of the head is 1.0. Classifiers for 18 body parts were trained using 184 CT scans. The final application was tested on 136 randomly selected heterogeneous CT scans. Ground truth was generated by asking two human observers to mark the start and end positions of each scan on the standard human figure. When compared with the human observers, the mean absolute error of the proposed method is 1.2% (max: 3.5%) and 1.6% (max: 5.4%) for the start and end positions, respectively. We proposed a scan range estimation method using multiple body parts detection and relative structure position analysis. In our preliminary tests, the proposed method delivered promising results.


Increased CD86 but Not CD80 and PD-L1 Expression on Liver CD68+ Cells during Chronic HBV Infection.
The failure to establish potent anti-HBV T cell responses suggests the absence of an effective innate immune activation. Kupffer cells and liver-infiltrating monocytes/macrophages have an essential role in establishing anti-HBV responses. These cells express the costimulatory molecules CD80 and CD86. CD80 expression on antigen-presenting cells (APCs) induces Th1 cell differentiation, whereas CD86 expression drives the differentiation towards a Th2 profile. The relative expression of CD80, CD86 and PD-L1 on APCs, regulates T cell activation. Few studies investigated CD80 and CD86 expression on KCs and infiltrating monocytes/macrophages in HBV-infected liver and knowledge about the expression of PD-L1 on these cells is controversial. The expression of these molecules together in CD68+ cells has not been explored in HBV-infected livers. Double staining immunohistochemistry was applied to liver biopsies of HBV-infected and control donors to explore CD80, CD86 and PD-L1 expression in the lobular and portal areas. Chronic HBV infection was associated with increased CD68+CD86+ cell count and percentage in the lobular areas, and no changes in the count and percentage of CD68+CD80+ and CD68+PD-L1+ cells, compared to the control group. While CD68+CD80+ cell count in portal areas correlated with the fibrosis score, CD68+CD80+ cell percentage in lobular areas correlated with the inflammation grade. The upregulation of CD86 but not CD80 and PD-L1 on CD68+ cells in HBV-infected livers, suggests that these cells do not support the induction of potent Th1. Moreover, the expression of CD80 on CD68+ cells correlates with liver inflammation and fibrosis.


Sensitivity of coded aperture Raman spectroscopy to analytes beneath turbid biological tissue and tissue-simulating phantoms.
Traditional slit-based spectrometers have an inherent trade-off between spectral resolution and throughput that can limit their performance when measuring diffuse sources such as light returned from highly scattering biological tissue. Recently, multielement fiber bundles have been used to effectively measure diffuse sources, e.g., in the field of spatially offset Raman spectroscopy, by remapping the source (or some region of the source) into a slit shape for delivery to the spectrometer. Another approach is to change the nature of the instrument by using a coded entrance aperture, which can increase throughput without sacrificing spectral resolution.In this study, two spectrometers, one with a slit-based entrance aperture and the other with a coded aperture, were used to measure Raman spectra of an analyte as a function of the optical properties of an overlying scattering medium. Power-law fits reveal that the analyte signal is approximately proportional to the number of transport mean free paths of the scattering medium raised to a power of -0.47 (coded aperture instrument) or -1.09 (slit-based instrument). These results demonstrate that the attenuation in signal intensity is more pronounced for the slit-based instrument and highlight the scattering regimes where coded aperture instruments can provide an advantage over traditional slit-based spectrometers.


Sex/Gender-Based Pharmacology and Statin Utilization Amongst Elderly Patients with Diabetes.
The purpose of this study was to explore the ten-year trends in utilization of bioequivalent doses of statin amongst elderly patients with diabetes according to sex/gender in Ontario, Canada. A cohort of patients with diabetes (>65 years) was constructed using the Ontario Diabetes Database Statin utilization data (2003-2012) was obtained from the Ontario Drug Benefit Program for both women and men. Bioequivalent doses for statins were calculated according to the dosing conversion factor in therapeutic interchange programs in clinical practice. Utilization pattern of high potency (Atorvastatin and Rosuvastatin) vs. low potency statins (Simvastatin, Lovastatin, Fluvastatin, Pravastatin) were also analyzed. The average bioequivalent Simvastatin utilization in 2003 was 29.22 mg/day for women and 30.35 mg/day for men. By 2008, this gap in dosing was higher for both women and men and by 2013 it had increased to 47.75 mg/day for women and 52.98 mg/day for men. For average number of day supply per year, there was no significant trend of changes over the 10-year period, although the use of high potency statins increased significantly (P<0.001) for both women and men. No differences were seen for sex/gender; either for the 10-year period or for each year. There has been significant increase in bioequivalent statin utilization amongst elderly patients with diabetes in Ontario; for both men and women. In a publicly-funded healthcare system such as Ontario, there were no sex/gender differences in the utilization of high potency statin (Atorvastatin and Rosuvastatin) amongst elderly patient with diabetes.


Sub-nanometer Resolution Imaging with Amplitude-modulation Atomic Force Microscopy in Liquid.
Atomic force microscopy (AFM) has become a well-established technique for nanoscale imaging of samples in air and in liquid. Recent studies have shown that when operated in amplitude-modulation (tapping) mode, atomic or molecular-level resolution images can be achieved over a wide range of soft and hard samples in liquid. In these situations, small oscillation amplitudes (SAM-AFM) enhance the resolution by exploiting the solvated liquid at the surface of the sample. Although the technique has been successfully applied across fields as diverse as materials science, biology and biophysics and surface chemistry, obtaining high-resolution images in liquid can still remain challenging for novice users. This is partly due to the large number of variables to control and optimize such as the choice of cantilever, the sample preparation, and the correct manipulation of the imaging parameters. Here, we present a protocol for achieving high-resolution images of hard and soft samples in fluid using SAM-AFM on a commercial instrument. Our goal is to provide a step-by-step practical guide to achieving high-resolution images, including the cleaning and preparation of the apparatus and the sample, the choice of cantilever and optimization of the imaging parameters. For each step, we explain the scientific rationale behind our choices to facilitate the adaptation of the methodology to every user's specific system.


Traveling Time of a translating ribosome along messenger RNA monitored directly on a quartz crystal microbalance.
During translation, the biosynthesis of polypeptides is dynamically regulated. The translation rate along messenger RNA (mRNA), which is dependent on the codon, structure, and sequence, is not always constant. However, methods for measuring the duration required for polypeptide elongation on an mRNA of interest have not been developed. In this work, we used a quartz crystal microbalance (QCM) technique to monitor mRNA translation in an Escherichia coli cell-free translation system in real time. This method permitted us to evaluate the translation of proteins of interest fused upstream of a streptavidin-binding peptide (SBP) fusion protein. The translation of mRNA encoding the SBP fusion protein alone was observed as a mass increase on a streptavidin-modified QCM plate. Addition of the protein of interest resulted in a delay in the mass change corresponding to the traveling time of the ribosome along the coding region of the protein of interest. With this technique, the lengths of coding sequences, codon usages, influences of unique sequences, and various protein-coding sequences were evaluated. The results showed that the traveling time of the translating ribosome depends on the length of the coding region translated but is also affected by the sequence itself. Differences in the time lags for various proteins imply that mRNA coding sequences may regulate gene expression.


Equine arteritis virus is not a togavirus but belongs to the coronaviruslike superfamily.
The nucleotide sequence of the genome of equine arteritis virus (EAV) was determined from a set of overlapping cDNA clones and was found to contain eight open reading frames (ORFs). ORFs 2 through 7 are expressed from six 3'-coterminal subgenomic mRNAs, which are transcribed from the 3'-terminal quarter of the viral genome. A number of these ORFs are predicted to encode structural EAV proteins. The organization and expression of the 3' part of the EAV genome are remarkably similar to those of coronaviruses and toroviruses. The 5'-terminal three-quarters of the genome contain the putative EAV polymerase gene, which also shares a number of features with the corresponding gene of corona- and toroviruses. The gene contains two large ORFs, ORF1a and ORF1b, with an overlap region of 19 nucleotides. The presence of a "shifty" heptanucleotide sequence in this region and a downstream RNA pseudoknot structure indicate that ORF1b is probably expressed by ribosomal frameshifting. The frameshift-directing potential of the ORF1a/ORF1b overlap region was demonstrated by using a reporter gene. Moreover, the predicted ORF1b product was found to contain four domains which have been identified in the same relative positions in coronavirus and torovirus ORF1b products. The sequences of the EAV and coronavirus ORF1a proteins were found to be much more diverged. The EAV ORF1a product contains a putative trypsinlike serine protease motif. Our data indicate that EAV, presently considered a togavirus, is evolutionarily related to viruses from the coronaviruslike superfamily.


The antibacterial properties of Malaysian tualang honey against wound and enteric microorganisms in comparison to manuka honey.
Antibiotic resistance of bacteria is on the rise, thus the discovery of alternative therapeutic agents is urgently needed. Honey possesses therapeutic potential, including wound healing properties and antimicrobial activity. Although the antimicrobial activity of honey has been effectively established against an extensive spectrum of microorganisms, it differs depending on the type of honey. To date, no extensive studies of the antibacterial properties of tualang (Koompassia excelsa) honey on wound and enteric microorganisms have been conducted. The objectives of this study were to conduct such studies and to compare the antibacterial activity of tualang honey with that of manuka honey. Using a broth dilution method, the antibacterial activity of tualang honey against 13 wound and enteric microorganisms was determined; manuka honey was used as the control. Different concentrations of honey [6.25-25% (w/v)] were tested against each type of microorganism. Briefly, two-fold dilutions of honey solutions were tested to determine the minimum inhibitory concentration (MIC) against each type of microorganism, followed by more assays within a narrower dilution range to obtain more precise MIC values. MICs were determined by both visual inspection and spectrophotometric assay at 620 nm. Minimum bactericidal concentration (MBC) also was determined by culturing on blood agar plates. By visual inspection, the MICs of tualang honey ranged from 8.75% to 25% compared to manuka honey (8.75-20%). Spectrophotometric readings of at least 95% inhibition yielded MIC values ranging between 10% and 25% for both types of honey. The lowest MBC for tualang honey was 20%, whereas that for manuka honey was 11.25% for the microorganisms tested. The lowest MIC value (8.75%) for both types of honey was against Stenotrophomonas maltophilia. Tualang honey had a lower MIC (11.25%) against Acinetobacter baumannii compared to manuka honey (12.5%). Tualang honey exhibited variable activities against different microorganisms, but they were within the same range as those for manuka honey. This result suggests that tualang honey could potentially be used as an alternative therapeutic agent against certain microorganisms, particularly A. baumannii and S. maltophilia.


Comparative descriptions of eggs from three species of Rhodnius (Hemiptera: Reduviidae: Triatominae).
The authors describe and compare the morphological and ultrastructural characteristics of eggs from the three most recent described species of the genus Rhodnius Stål, 1859, which have not previously been studied. These species are Rhodnius colombiensis (Mejia, Galvão & Jurberg 1999), Rhodnius milesi (Carcavallo, Rocha, Galvão & Jurberg 2001) and Rhodnius stali (Lent, Jurberg & Galvão 1993). The results revealed that there are similarities in the exochorial architecture of optical microscopy and scanning electron microscopy; these include the predominance of hexagonal cells that are common to all Rhodnius species and variable degrees of lateral flattening, which is common not only to species of this genus, but also to the Rhodniini tribe. Differences in overall colour, the presence of a collar in R. milesi, a longitudinal bevel in R. stali and the precise length of R. colombiensis can be useful distinguishing features. As a result of this study, the key for egg identification proposed by Barata in 1981 can be updated.


Long-term effects of ileal bypass on lipoproteins in patients with familial hypercholesterolemia.
Partial ileal bypass is effective in reducing circulating cholesterol levels. In our study the 10 year effects of the procedure on serum lipids and lipoproteins were studied in 27 patients with heterozygous familial hypercholesterolemia. The ileal bypass patients were compared with conservatively treated case controls matched for age, sex, serum cholesterol level, relative body weight, blood pressure, and smoking habits and also for the existence of diabetes and coronary heart disease. Serum triglycerides were initially slightly higher in the patients undergoing ileal bypass. During the 10 year follow-up eight surgically treated and seven control patients suffered fatal or nonfatal myocardial infarctions. Of these all but one male subject who underwent surgery had had manifest coronary heart disease at entry. Male sex, smoking, triglyceride levels, and angina were significant predictors of new coronary events. The fall in serum cholesterol in patients who underwent ileal bypass and had fatal myocardial infarctions was smaller than in the corresponding subjects without events. The serum lipid levels of the survivors at the end of the 10 year follow-up showed that ileal bypass, as compared with the conservative treatment, had led to a larger decrease in total serum cholesterol (-33% vs -11% in the control patients; p less than .001); lower total serum cholesterol (360 vs 468 mg/dl; p less than .001), low-density lipoprotein (LDL) cholesterol (236 vs 324 mg/dl, p less than .001), and LDL apoprotein B levels (186 vs 231 mg/dl; p less than .001); and higher serum high-density lipoprotein (HDL) (46 vs 38 mg/dl; p less than .05) and HDL2 cholesterol levels (25 vs 16 mg/dl; p less than .05).(ABSTRACT TRUNCATED AT 250 WORDS)


Nucleus accumbens core and pathogenesis of compulsive checking.
To investigate the role of the nucleus accumbens core (NAc) in the development of quinpirole-induced compulsive checking, rats received an excitotoxic lesion of NAc or sham lesion and were injected with quinpirole (0.5 mg/kg) or saline; development of checking behavior was monitored for 10 biweekly tests. The results showed that even after the NAc lesion, quinpirole still induced compulsive checking, suggesting that the pathogenic effects produced by quinpirole lie outside the NAc. Although the NAc lesion did not prevent the induction of compulsive checking, it altered how quickly it develops, suggesting that the NAc normally contributes toward the induction of compulsive checking. Saline-treated rats with an NAc lesion were hyperactive, but did not develop compulsive checking, indicating that hyperactivity by itself is not sufficient for the pathogenesis of compulsive checking. It is proposed that compulsive checking is the exaggerated output of a security motivation system and that the NAc serves as a neural hub for coordinating the orderly activity of neural modules of this motivational system. Evidence is considered suggesting that the neurobiological condition for the pathogenesis of compulsive checking is two-fold: activation of dopamine D2/D3 receptors without concurrent stimulation of D1-like receptors and long-term plastic changes related to quinpirole-induced sensitization.


Optimizing person-centered transitions in the dementia journey: a comparison of national dementia strategies.
The journey for people with Alzheimer's disease or another dementia involves the need for increasing levels of support, with transitions across care settings. Although transitional care has received increasing attention in the health care arena, no widely accepted transitions typology exists for the dementia journey. At the same time, national dementia strategies are proliferating. We developed a typology containing six transitions that cover the dementia journey from symptom recognition to end-of-life care. We then critically evaluated whether and how the national dementia strategies of Australia, England, France, the Netherlands, Norway, Scotland, and the United States addressed each transition. Adopting a person-centered perspective, we found that most or all of the national strategies adequately address earlier transitions in the journey, but fewer strategies address the later transitions. We recommend that next-generation national dementia strategies focus on later transitions, specify how care coordination and workforce training should make transitions more person centered, and use person-centered outcomes in evaluating the success of the strategies' implementation and dissemination.


Genetic and Non-Genetic Factor-Adjusted Association between Coffee Drinking and High-Density Lipoprotein Cholesterol in Taiwanese Adults: Stratification by Sex.
Low high-density lipoprotein cholesterol (HDL-C) is a major risk factor for cardiovascular diseases (CVDs), the leading cause of global mortality. We aimed to determine the effect of coffee drinking and sex and their interaction, as well as rs1800588 and rs1800775 polymorphisms on HDL-C levels in Taiwanese adults. Data of 4262 men and 4813 women, aged 30-70 years, were retrieved from Taiwan Biobank. The interaction between sex and coffee drinking on HDL-C was significant (p = 0.0452). Coffee consumption was significantly associated with higher HDL-C levels in only women (β = 0.81679; p = 0.0246). However, rs1800588 and rs1800775 variants were significantly associated with HDL-C in both sexes. In women, β-values were 0.99080; p = 0.0059 and 3.16277; p < 0.0001 for rs1800588 CT and TT genotypes, respectively and -1.80954; p < 0.0001 and -2.81512; p < 0.0001 for rs1800775 AC and CC genotypes, respectively. In men, β-values were 1.32430; p < 0.0001 and 3.24976; p < 0.0001 for rs1800775 CT and TT genotypes, respectively and -1.96232; p < 0.0001 and -2.71536, p < 0.0001 for the AC and CC genotypes, respectively. In conclusion, coffee drinking was significantly associated with higher high-density lipoprotein (HDL) levels in women but not men after adjusting for confounders including rs1800588 (LIPC) and rs1800775 (CETP) variants.


Managing Lafora body disease with vagal nerve stimulation.
A 17-year-old female, of consanguineous parents, presented with a history of seizures and cognitive decline since the age of 12 years. She had absence, focal dyscognitive, generalized myoclonic, and generalized tonic-clonic seizures, all of which were drug resistant. The diagnosis of Lafora body disease was made based on a compatible clinical, EEG, seizure semiology picture and a disease-causing homozygous mutation in the EPM2A gene. A vagus nerve stimulator (VNS) was inserted and well tolerated with a steady decrease and then stabilization in seizure frequency during the six months following insertion (months 1-6). At follow-up, at 12 months after VNS insertion, there was a persistent improvement. Seizure frequency during months 7-12, compared to pre-VNS, was documented as follows: the absence seizures observed by the family had decreased from four episodes per month to 0 per month, the focal dyscognitive seizures from 300 episodes per month to 90 per month, the generalized myoclonic seizures from 90 clusters per month to eight per month, and the generalized tonic-clonic seizures from 30 episodes per month to 1.5 per month on average. To our knowledge, this is the second case reported in the literature showing efficacy of VNS in the management of seizures in Lafora body disease.


Predictive gene signatures: molecular markers distinguishing colon adenomatous polyp and carcinoma.
Cancers exhibit abnormal molecular signatures associated with disease initiation and progression. Molecular signatures could improve cancer screening, detection, drug development and selection of appropriate drug therapies for individual patients. Typically only very small amounts of tissue are available from patients for analysis and biopsy samples exhibit broad heterogeneity that cannot be captured using a single marker. This report details application of an in-house custom designed GenomeLab System multiplex gene expression assay, the hCellMarkerPlex, to assess predictive gene signatures of normal, adenomatous polyp and carcinoma colon tissue using archived tissue bank material. The hCellMarkerPlex incorporates twenty-one gene markers: epithelial (EZR, KRT18, NOX1, SLC9A2), proliferation (PCNA, CCND1, MS4A12), differentiation (B4GANLT2, CDX1, CDX2), apoptotic (CASP3, NOX1, NTN1), fibroblast (FSP1, COL1A1), structural (ACTG2, CNN1, DES), gene transcription (HDAC1), stem cell (LGR5), endothelial (VWF) and mucin production (MUC2). Gene signatures distinguished normal, adenomatous polyp and carcinoma. Individual gene targets significantly contributing to molecular tissue types, classifier genes, were further characterised using real-time PCR, in-situ hybridisation and immunohistochemistry revealing aberrant epithelial expression of MS4A12, LGR5 CDX2, NOX1 and SLC9A2 prior to development of carcinoma. Identified gene signatures identify aberrant epithelial expression of genes prior to cancer development using in-house custom designed gene expression multiplex assays. This approach may be used to assist in objective classification of disease initiation, staging, progression and therapeutic responses using biopsy material.


The chemokine CXCL12 and its receptor CXCR4 are implicated in human seminoma metastasis.
Seminoma and non-seminoma tumours increasingly occur within the western population. These tumours originate from carcinoma in situ (CIS) cells, which arise from dysfunctional gonocytes. CXCL12 and its receptors, CXCR4 and CXCR7, have been implicated in migration, proliferation and survival of gonocytes and their precursors and progeny, primordial germ cells and spermatogonial stem cells respectively. We previously found evidence that several miRNA molecules predicted to modulate CXCR4 signalling are differentially expressed during the differentiation of gonocytes into spermatogonia in mice. Bioinformatic analysis predicted these miRNA to modulate CXCR4 signalling, leading us to hypothesize that CXCL12-mediated CXCR4 signalling is involved in the disrupted differentiation of gonocytes that underpins CIS formation. Indeed, we detected CXCL12 in Sertoli cells of normal human testis, and relatively high expression in tumour stroma with concomitant weak staining in dispersed tumour cells. In contrast, CXCR4 was expressed in spermatogonial and meiotic germ cells of normal testis and in the majority of tumour cells. Quantitative RT-PCR identified elevated CXCR4 transcript levels in seminoma compared with normal testis and to non-seminoma, potentially reflecting the higher proportion of dysfunctional germ cells within seminomas. In the normal testis, expression of CXCR4 downstream signalling molecules phospho-MEK1/2 and phospho-ERK1/2 correlated with CXCR4/CXCL12 expression. Strikingly, this correlation was absent in seminoma and non-seminoma samples, suggesting that CXCL12 signalling is disrupted. Proliferation rate and cell survival were not altered by CXCL12 in either seminoma (TCam-2) or non-seminoma (833ke) cell lines. However, CXCL12 exposure induced TCam-2 cell invasion though simulated basement membrane, while in contrast, we provide the novel evidence that CXCR4-expressing non-seminoma cell lines 833ke and NTera2/D1 do not invade in response to CXCL12. These findings indicate that CXCL12 expression in the human testis may selectively influence seminoma migration and metastasis, correlating with its importance in gonocyte and spermatogonial stem cell biology.


CD8alpha- CD11b+ dendritic cells present exogenous virus-like particles to CD8+ T cells and subsequently express CD8alpha and CD205 molecules.
Recombinant porcine parvovirus virus-like particles (PPV-VLPs) are particulate exogenous antigens that induce a strong, specific cytotoxic T lymphocyte (CTL) response in the absence of adjuvant. In the present report, we demonstrate in vivo that dendritic cells (DCs) present PPV-VLPs to CD8+ T cells after intracellular processing. PPV-VLPs are captured by DCs with a high efficacy, which results in the delivery of these exogenous antigens to 50% of the whole spleen DC population. In vivo, a few hours after injection, PPV-VLPs are presented exclusively to CD8+ T cells by CD8alpha- DCs, whereas 15 hours later they are presented mainly by CD8alpha+ DCs. After PPV-VLPs processing, a fraction of CD11b+ DCs undergo phenotypic changes, i.e., the up-regulation of CD8alpha and CD205 and the loss of CD4 molecules on their surface. The failure to detect mRNA coding for CD8alpha in CD11b+ DCs suggests that CD8alpha expression by these cells is not due to de novo synthesis. In recombination-activating gene knockout mice (Rag-/-), CD11b+ DCs did not express CD8alpha and PPV-VLPs presentation by CD8alpha+ DCs was severely diminished. These results indicate that both CD8alpha- and CD8alpha+ DCs play an important role in the induction of CTL responses by exogenous antigens, such as VLP.


Predicting unilateral prostate cancer based on biopsy features: implications for focal ablative therapy--results from the SEARCH database.
For men with low risk prostate cancer it was recently proposed that ablative treatment to the affected side may decrease morbidity, while maintaining good oncological outcomes. However, few studies have assessed the correlation between biopsy parameters and pathological outcome (unilateral vs bilateral disease). Using the Shared Equal Access Regional Cancer Hospital Database of men treated with radical prostatectomy at multiple equal access medical centers we retrospectively examined the records of 261 men with clinical stage T1c or T2a prostate cancer, prostate specific antigen less than 10 ng/ml, Gleason sum 6 or less and only 1 or 2 ipsilateral positive cores on at least sextant biopsy. We compared clinical characteristics between men with pathologically unilateral disease or less (pT2b or less) and men with pathologically bilateral disease or extraprostatic extension (pT2c or greater). To determine the significant predictors of pT2c or greater disease we used a multivariate logistic regression model. Of the cohort of 261 men with low risk prostate cancer only 93 (35.1%) had unilateral or no evidence of disease following examination of radical prostatectomy specimens. Men with pathologically unilateral or less disease did not differ from those with bilateral or more advanced disease by age, prostate specific antigen, clinical stage, body mass index or number of positive biopsy cores (1 vs 2). On multivariate analysis no clinical feature was significantly related to pathologically unilateral or less vs bilateral or greater disease. The majority of men with low risk prostate cancer and 1 or 2 ipsilateral positive biopsy cores have pathologically bilateral disease. Therefore, strategies for unilateral treatment of prostate cancer are unlikely to be curative for these men.


In Crystallo Selection to Establish New RNA Crystal Contacts.
Crystallography is a major technique for determining large RNA structures. Obtaining diffraction-quality crystals has been the bottleneck. Although several RNA crystallization methods have been developed, the field strongly needs additional approaches. Here we invented an in crystallo selection strategy for identifying mutations that enhance a target RNA's crystallizability. The strategy includes constructing an RNA pool containing random mutations, obtaining crystals, and amplifying the sequences enriched by crystallization. We demonstrated a proof-of-principle application to the P4-P6 domain from the Tetrahymena ribozyme. We further determined the structures of four selected mutants. All four establish new crystal lattice contacts while maintaining the native structure. Three mutants achieve this by relocating bulges and one by making a helix more flexible. In crystallo selection provides opportunities to improve crystals of RNAs or RNA-ligand complexes. Our results also suggest that mutants may be rationally designed for crystallization by "walking" a bulge along the RNA chain.


CD2F-10: a new member of the CD2 subset of the immunoglobulin superfamily.
The CD2 subset of the immunoglobulin superfamily consists of a rapidly expanding family of leukocyte cell surface receptors, at least five of which (CD2, CD48, CD58, CD150, and CD244) are involved in lymphocyte activation as either receptors or ligands. Completion of the draft sequence of the human genome offers the possibility of systematically identifying the full set of proteins and interactions of this important family. Here we describe the identification and characterization of the first new member of the subset, CD2F-10, found exclusively by genome searching.


Is structural sensitivity a problem of oversimplified biological models? Insights from nested Dynamic Energy Budget models.
Many current issues in ecology require predictions made by mathematical models, which are built on somewhat arbitrary choices. Their consequences are quantified by sensitivity analysis to quantify how changes in model parameters propagate into an uncertainty in model predictions. An extension called structural sensitivity analysis deals with changes in the mathematical description of complex processes like predation. Such processes are described at the population scale by a specific mathematical function taken among similar ones, a choice that can strongly drive model predictions. However, it has only been studied in simple theoretical models. Here, we ask whether structural sensitivity is a problem of oversimplified models. We found in predator-prey models describing chemostat experiments that these models are less structurally sensitive to the choice of a specific functional response if they include mass balance resource dynamics and individual maintenance. Neglecting these processes in an ecological model (for instance by using the well-known logistic growth equation) is not only an inappropriate description of the ecological system, but also a source of more uncertain predictions.


Biological evaluation of neopeptins isolated from a Streptomyces strain.
Microbial secondary metabolites are a rich source of antifungal agents and have merit as alternatives to synthetic fungicides. To develop disease control agents against powdery mildew, the lipopeptide antibiotic neopeptins were identified from the culture broth of a Streptomyces sp., and in vivo control efficacy of the compounds was evaluated on cucumber plants under glasshouse conditions. The Streptomyces sp. KNF2047 antagonistic against powdery mildew development in cucumber plants was isolated from a soil sample. Antifungal compounds were purified from the culture broth and identified as neopeptin A and B. In vitro microtitre assays revealed the inhibitory activities of the compounds in the range 128-512 microg mL(-1) against the mycelial growth of Alternaria mali, Botrytis cinerea, Cladosporium cucumerinum, Colletotrichum lagenarium, Didimella bryoniae and Magnaporthe grisea. Although neither compound showed remarkable in vitro antifungal activity against other plant pathogenic fungi, a mixture of neopeptins (484 mg of neopeptin A and 290 mg of neopeptin B per gram of partially purified powder) showed potent protective and curative activity against cucumber powdery mildew in vivo. The disease control activity of the neopeptins at a concentration of 2.4 mg L(-1) was 92.1%, which was similar to that of the commercial fungicide fenarimol (89.3% at 63 mg L(-1)) and that of the commercial biocontrol agent Actinovate (67.4% at 2 x 10(7) cfu L(-1)). Neopeptin mixtures isolated from Streptomyces sp. KNF2047 showed potent disease control activity against powdery mildew development on cucumber plants. .


Type II Supernova Matter in a Silicon Carbide Grain from the Murchison Meteorite
The circumstellar silicon carbide (SiC) grain X57 from the Murchison meteorite contains large amounts of radiogenic calcium-44 (20 times its solar system abundance) and has an anomalous silicon isotopic composition, different from other circumstellar SiC grains. Its inferred initial 44Ti/Si and 44Ti/48Ti ratios are 1.6 x 10(-4) and 0.37. In addition, it contains radiogenic magnesium-26; the inferred initial 26Al/27Al ratio is 0.11. The isotopic and elemental data of X57 can be explained by selective mixing of matter from different zones of a typical type II supernova of 25 solar masses during its explosion. The high 44Ti/Si ratio requires contributions from the innermost nickel zone of the supernova to the SiC condensation site, as similarly suggested by astronomical observations.


Microbiota-induced active translocation of peptidoglycan across the intestinal barrier dictates its within-host dissemination.
Peptidoglycan, the major structural polymer forming the cell wall of bacteria, is an important mediator of physiological and behavioral effects in mammalian hosts. These effects are frequently linked to its translocation from the intestinal lumen to host tissues. However, the modality and regulation of this translocation across the gut barrier has not been precisely addressed. In this study, we characterized the absorption of peptidoglycan across the intestine and its systemic dissemination. We report that peptidoglycan has a distinct tropism for host organs when absorbed via the gut, most notably by favoring access to the brain. We demonstrate that intestinal translocation of peptidoglycan occurs through a microbiota-induced active process. This process is regulated by the parasympathetic pathway via the muscarinic acetylcholine receptors. Together, this study reveals fundamental parameters concerning the uptake of a major microbiota molecular signal from the steady-state gut.


Management of uncomplicated malaria in children under 13 years of age at a district hospital in Senegal: from official guidelines to usual practices.
To be effective, national malaria guidelines must be properly followed. This study evaluated nurses' practices in the management of uncomplicated malaria cases at a District Hospital. Its objective was to identify the reasons for discrepancies between official guidelines and usual practices. This study took place at Oussouye hospital, south-western Senegal. Blood smears were available for biological diagnosis in patients aged more than five years while the Integrated Management of Childhood Illness recommended treating fevers presumptively in children under five. First line anti-malarial was Amodiaquine plus sulphadoxine-pyrimethamine (AQ+SP) bi-therapy. Hospital records of children under 13 years of age seen between 2004 and 2005 were reviewed. Among children treated with anti-malarials, 74% (2, 063/2, 789) received AQ+SP. However, only 22% (406/1, 879) of febrile children and 19% (429/2, 198) of children diagnosed with malaria got a blood smear. Moreover, an anti-malarial was prescribed for 80% (377/474) of children with a negative blood smear. The transition from chloroquine to AQ+SP was well followed. Nonetheless, blood smear use was very low and many over-prescriptions were reported. Reasons for discrepancies between guidelines and practices can be classified in three main categories: ambiguous guidelines, health system's dysfunctions and nurses' own considerations. Aside from the strengthening of the public health system, in order to guarantee practices complying with guidelines, training content should be more adapted to nurses' own considerations.


Impact of early-life stress and resilience on patients with major depressive disorder.
Early-life stress (ELS) has a long-lasting effect on affective function and may entail an increased risk for major depressive disorder (MDD). However, resilience can play a protective role against developing psychopathology. In this study, we investigated the relationships of depressive symptoms with ELS and resilience in MDD. Twenty-six patients with MDD as well as age- and gender-matched healthy controls were included in this study. Each subject was assessed concerning ELS, resilience, and depressive symptom severity with self-report questionnaires. Independent samples t-test and Mann-Whitney test were performed to compare ELS and resilience between the patient and control groups. Spearman correlation analyses and linear regression analysis were conducted to investigate significant ELS and resilience factors associated with depressive symptoms. In the MDD patient group, subjects reported greater exposure to inter- parental violence, and five factor scores on the resilience scale were significantly lower in comparison to the control group. In linear regression analysis, in regards to resilience, depressive symptom score was significantly associated with self-confidence and self-control factors; however, ELS demonstrated no significant association with depressive symptoms. Among resilience factors, self-confidence and self-control may ameliorate depressive symptoms in MDD. ELS, including inter-parental violence, physical abuse and emotional abuse, might be a risk factor for developing depression. Assessment of early-life stress and intervention programs for increasing resilience capacity would be helpful in treating MDD.


Electrophoretically mediated microanalysis for characterization of the enantioselective CYP3A4 catalyzed N-demethylation of ketamine.
Execution of an enzymatic reaction performed in a capillary with subsequent electrophoretic analysis of the formed products is referred to as electrophoretically mediated microanalysis (EMMA). An EMMA method was developed to investigate the stereoselectivity of the CYP3A4-mediated N-demethylation of ketamine. Ketamine was incubated in a 50 μm id bare fused-silica capillary together with human CYP3A4 Supersomes using a 100 mM phosphate buffer (pH 7.4) at 37°C. A plug containing racemic ketamine and the NADPH regenerating system including all required cofactors for the enzymatic reaction was injected, followed by a plug of the metabolizing enzyme CYP3A4 (500 nM). These two plugs were bracketed by plugs of incubation buffer to ensure proper conditions for the enzymatic reaction. The rest of the capillary was filled with a pH 2.5 running buffer comprising 50 mM Tris, phosphoric acid, and 2% w/v of highly sulfated γ-cyclodextrin. Mixing of reaction plugs was enhanced via application of -10 kV for 10 s. After an incubation of 8 min at 37°C without power application (zero-potential amplification), the capillary was cooled to 25°C within 3 min followed by application of -10 kV for the separation and detection of the formed enantiomers of norketamine. Norketamine formation rates were fitted to the Michaelis-Menten model and the elucidated values for V(max) and K(m) were found to be comparable to those obtained from the off-line assay of a previous study.


A phosphotyrosine-imprinted polymer receptor for the recognition of tyrosine phosphorylated peptides.
Hyperphosphorylation at tyrosine is commonly observed in tumor proteomes and, hence, specific phosphoproteins or phosphopeptides could serve as markers useful for cancer diagnostics and therapeutics. The analysis of such targets is, however, a challenging task, because of their commonly low abundance and the lack of robust and effective preconcentration techniques. As a robust alternative to the commonly used immunoaffinity techniques that rely on phosphotyrosine(pTyr)-specific antibodies, we have developed an epitope-imprinting strategy that leads to a synthetic pTyr-selective imprinted polymer receptor. The binding site incorporates two monourea ligands placed by preorganization around a pTyr dianion template. The tight binding site displayed good binding affinities for the pTyr template, in the range of that observed for corresponding antibodies, and a clear preference for pTyr over phosphoserine (pSer). In further analogy to the antibodies, the imprinted polymer was capable of capturing short tyrosine phosphorylated peptides in the presence of an excess of their non-phosphorylated counterparts or peptides phosphorylated at serine.


Changes in Malaria Epidemiology in Africa and New Challenges for Elimination.
Although the burden of Plasmodium falciparum malaria is gradually declining in many parts of Africa, it is characterized by spatial and temporal variability that presents new and evolving challenges for malaria control programs. Reductions in the malaria burden need to be sustained in the face of changing epidemiology whilst simultaneously tackling significant pockets of sustained or increasing transmission. Large-scale, robust surveillance mechanisms that measure rather than estimate the actual burden of malaria over time from large areas of the continent where such data are lacking need to be prioritized. We review these fascinating developments, caution against complacency, and make the case that improving the extent and quality of malaria surveillance is vital for Africa as she marches on towards elimination.


The uncertainty of predicting intact anterior cruciate ligament degeneration in terms of structural properties using T(2)(*) relaxometry in a human cadaveric model.
The combination of healing anterior cruciate ligament (ACL) volume and the distributions of T2(*) relaxation times within it have been shown to predict the biomechanical failure properties in a porcine model. This MR-based prediction model has not yet been used to assess ligament degeneration in the aging human knee. Using a set of 15 human cadaveric knees of varying ages, we obtained in situ MR measures of volume and T2(*) of the intact ACL and then related these MR variables to biomechanical outcomes (maximum and yield loads, linear stiffness) obtained via ex vivo failure testing. Using volume in conjunction with the median T2(*) value, the multiple linear regression model did not predict maximum failure load for the intact human ACL; R(2)=0.23, p=0.200. Similar insignificant results were found for yield load and linear stiffness. Naturally restricted distributions of the intact ligament volume and T2(*) (demonstrated by the respective Z-scores) in an older cadaveric population were the likely reason for the insignificant results. These restricted distributions may negatively affect the ability to detect a correlation when one exists. Further research is necessary to understand the relationship of MRI variables and ligament degeneration. While this study failed to find a significant prediction of human biomechanical outcome using these MR variables, with further research, an MR-based approach may offer a tool to longitudinally assess changes in cruciate ligament degradation.


Multicenter evaluation of infant lung function tests as cystic fibrosis clinical trial endpoints.
The conducting of clinical trials in infants with cystic fibrosis (CF) has been hindered by lack of sensitive outcome measures. To evaluate safety, feasibility, and ability to detect abnormalities in lung function of serial pulmonary function tests (PFTs) in infants with CF. Multicenter observational study using a commercial device, rigorous training, ongoing quality control, and over-reading of data by an independent panel. Raised volume rapid thoracoabdominal compression technique and plethysmography were performed at enrollment and at 6 and 12 months, with an additional 1-month reproducibility visit. A total of 342 procedures were performed in 100 infants with CF at 10 centers. FRC measurements were acceptable at a higher proportion of study visits (89%) than raised volume (72%) or fractional lung volume (68%) measurements. Average Z scores for many parameters differed significantly from historical control values. Mean (95% confidence interval) Z scores were: -0.52 (-0.78 to -0.25) for forced expiratory flow at 75% (FEF₇₅) for FVC; 1.92 (1.39-2.45) for FRC; 1.22 (0.68-1.76) for residual volume; 0.87 (0.60-1.13) for FRC/total lung capacity; and 0.66 (0.27-1.06) for residual volume/total lung capacity. For future multicenter clinical trials using infant PFTs as primary endpoints, minimum detectable treatment effects are presented for several sample sizes. In this 10-center study, key PFT measures were significantly different in infants with CF than in historical control subjects. However, infant PFTs do not yet appear ready as primary efficacy endpoints for multicenter clinical trials, particularly at inexperienced sites, based on acceptability rates, variability, and potentially large sample sizes required to detect reasonable treatment effects.


Use of NIR light and upconversion phosphors in light-curable polymers.
Light-curable polymers are commonly used in restorative surgery, prosthodontics and surgical procedures. Despite the fact of wide application, there are clinical problems due to limitations of blue light penetration: application is restricted to defects exposed to the light source, layered filling of defect is required. Combining photo-activation and up conversion allows efficient polymer hardening by deep penetrating near-infrared (NIR) light. The prerequisite 450 nm blue light to polymerize dental resins could be achieved by filler particles, which absorb the incident NIR irradiation and convert it into visible light. The on spot generated blue light results in uniform polymer hardening. Composite samples of 5mm thickness were cured two times faster than pure polymer cured by blue light (30 and 60 s, respectively). Overall degree of monomer conversion resulted in higher values of more than 40%. The enhanced transmission of NIR light was confirmed by optical analysis of dentin and enamel. The NIR transmittance surge in the 800-1200 nm window could improve sealing of complex and deep caries lesions. We demonstrate faster curing and an improved degree of polymerization by using upconversion filler particles as multiple light emission centers. This study represents an alternative approach in curing dental resins by NIR source.


Pathophysiological mechanisms underlying the adverse effects of calcium channel-blocking drugs in patients with chronic heart failure.
Although the vasodilator and anti-ischemic actions of calcium channel blocking drugs might be expected to produce hemodynamic benefits in chronic heart failure, none of these agents has been shown to exert favorable clinical effects in these patients. Furthermore, when left ventricular function is markedly impaired, short- and long-term therapy with calcium channel blockers can increase cardiovascular morbidity and mortality. Previous investigators have attributed these unfavorable results to the established negative inotropic effects of calcium channel blockers on the failing heart. Yet, this cardiodepressant action seems to account for only the deleterious responses seen during short-term therapy with these drugs, whereas the detrimental effects of long-term treatment can be more readily explained by the capacity of calcium channel blockers to activate endogenous neurohormonal systems, especially the renin-angiotensin system. According to this hypothesis, a number of cardioactive drugs (including calcium channel blockers, beta-blockers, and converting enzyme inhibitors) can depress myocardial contractility, but only calcium channel blockers, which activate neurohormonal systems, adversely affect patients with left ventricular dysfunction, whereas beta-blockers and converting enzyme inhibitors, which interfere with neurohormonal activity, can favorably modify the long-term outcome of these high-risk patients. If confirmed by future studies, these observations suggest that the effect of antianginal drugs on neurohormonal systems, as well as on cardiac contractility, should be an important consideration in the selection of a therapeutic agent in patients with coronary artery disease and chronic heart failure.


Predicting concentrations of organic chemicals in fish by using toxicokinetic models.
Quantification of chemical toxicity continues to be generally based on measured external concentrations. Yet, internal chemical concentrations have been suggested to be a more suitable parameter. To better understand the relationship between the external and internal concentrations of chemicals in fish, and to quantify internal concentrations, we compared three toxicokinetic (TK) models with each other and with literature data of measured concentrations of 39 chemicals. Two one-compartment models, together with the physiologically based toxicokinetic (PBTK) model, in which we improved the treatment of lipids, were used to predict concentrations of organic chemicals in two fish species: rainbow trout (Oncorhynchus mykiss) and fathead minnow (Pimephales promelas). All models predicted the measured internal concentrations in fish within 1 order of magnitude for at least 68% of the chemicals. Furthermore, the PBTK model outperformed the one-compartment models with respect to simulating chemical concentrations in the whole body (at least 88% of internal concentrations were predicted within 1 order of magnitude using the PBTK model). All the models can be used to predict concentrations in different fish species without additional experiments. However, further development of TK models is required for polar, ionizable, and easily biotransformed compounds.


Fatherhood and suffering: a qualitative exploration of Swedish men's experiences of care after the death of a baby.
This study was designed to evaluate fathers' experiences of stillbirth and psychosocial care. Data were collected between 27 March 2008 and 1 April 2010 via a questionnaire posted on the homepage of the Swedish National Infant Foundation. The responses to the following open-ended questions were analyzed using content analysis: "Are you grateful today for anything that health care professionals did in connection with the birth of your child?" and "Are you sad, hurt or angry today about something personnel did in connection with the birth of your baby?". 113/131 (86%) fathers reported feelings of being grateful. Only 22/131 (16%) fathers reported feeling sad, hurt, or angry. Fathers expressed gratitude when health care professionals treated their newborn "with respect and without fear", "with extraordinary reverence", and when their fatherhood was validated by providers. They were also grateful when providers helped them to create memories of their baby. Fathers also reported feeling sad, hurt, or angry when providers were nonchalant and indifferent and when they perceived providers to be uncaring and disrespectful toward their baby. Bereaved fathers experience overall gratitude for person-centered psychosocial care in the aftermath of stillbirth, particularly when they feel validated as a grieving father and their child is acknowledged with reverence. Health care professionals should support fathers by treating the baby who died with respect and dignity and by validating and acknowledging both his grief experiences and his fatherhood just as they would for a grieving mother.


Mitochondrial form, function and signalling in aging.
Aging is often accompanied by a decline in mitochondrial mass and function in different tissues. Additionally, cell resistance to stress is frequently found to be prevented by higher mitochondrial respiratory capacity. These correlations strongly suggest mitochondria are key players in aging and senescence, acting by regulating energy homeostasis, redox balance and signalling pathways central in these processes. However, mitochondria display a wide array of functions and signalling properties, and the roles of these different characteristics are still widely unexplored. Furthermore, differences in mitochondrial properties and responses between tissues and cell types, and how these affect whole body metabolism are also still poorly understood. This review uncovers aspects of mitochondrial biology that have an impact upon aging in model organisms and selected mammalian cells and tissues.


Analysis of the Binding of Expansin Exl1, from Pectobacterium carotovorum, to Plant Xylem and Comparison to EXLX1 from Bacillus subtilis.
The plant xylem is a preferred niche for some important bacterial phytopathogens, some of them encoding expansin proteins, which bind plant cell walls. Yet, the identity of the substrate for bacterial expansins within the plant cell wall and the nature of its interaction with it are poorly known. Here, we determined the localization of two bacterial expansins with differing isoelectric points (and with differing binding patterns to cell wall extracts) on plant tissue through in vitro fluorophore labeling and confocal imaging. Differential localization was observed, in which Exl1 from Pectobacterium carotovorum located into the intercellular spaces between xylem vessels and adjacent cells of the plant xylem, whereas EXLX1 from Bacillus subtilis bound cell walls of most cell types. In isolated vascular tissue, however, both PcExl1 and BsEXLX1 preferentially bound to tracheary elements over the xylem fibers, even though both are composed of secondary cell walls. Fluorescence correlation spectroscopy, employed to analyze the interaction of expansins with isolated xylem, indicates that binding is governed by more than one factor, which could include interaction with more than one type of polymer in the fibers, such as cellulose and hemicellulose or pectin. Binding to different polysaccharides could explain the observed reduction of cellulolytic and xylanolytic activities in the presence of expansin, possibly because of competition for the substrate. Our findings are relevant for the comprehensive understanding of the pathogenesis by P. carotovorum during xylem invasion, a process in which Exl1 might be involved.


Evaluation of a commercially-available block for spatially fractionated radiation therapy.
In this paper, we present the dosimetric characteristics of a commercially-produced universal GRID block for spatially fractioned radiation therapy. The dosimetric properties of the GRID block were evaluated. Ionization chamber and film measurements using both Kodak EDR2 and Gafchromic EBT film were performed in a solid water phantom to determine the relative output of the GRID block as well as its spatial dosimetric characteristics. The surface dose under the block and at the openings was measured using ultra thin TLDs. After introducing the GRID block into the treatment planning system, a treatment plan was created using the GRID block and also by creating a GRID pattern using the multi-leaf collimator. The percent depth doses measured with film showed that there is a shift of the dmax towards shallower depths for both energies (6 MV and 18 MV) under investigation. It was observed that the skin dose at the GRID openings was higher than the corresponding open field by a factor as high as 50% for both photon energies. The profiles showed the transmission under the block was in the order of 15-20% for 6 MV and 30% for 18 MV. The MUs calculated for a real patient using the block were about 80% less than the corresponding MUs for the same plan using the multileaf collimator to define the GRID. Based on this investigation, this brass GRID compensator is a viable alternative to other solid compensators or MLC-based fields currently in use. Its ease of creation and use give it decided advantages. Its ability to be created once and used for multiple patients (by varying the collimation of the linear accelerator jaws) makes it attractive from a cost perspective. We believe this compensator can be put to clinical use, and will allow more centers to offer GRID therapy to their patients.


Uncertainty estimation using fuzzy measures for multiclass classification.
Uncertainty arises in classification problems when the input pattern is not perfect or measurement error is unavoidable. In many applications, it would be beneficial to obtain an estimate of the uncertainty associated with a new observation and its membership within a particular class. Although statistical classification techniques base decision boundaries according to the probability distributions of the patterns belonging to each class, they are poor at supplying uncertainty information for new observations. Previous research has documented a multiarchitecture, monotonic function neural network model for the representation of uncertainty associated with a new observation for two-class classification. This paper proposes a modification to the monotonic function model to estimate the uncertainty associated with a new observation for multiclass classification. The model, therefore, overcomes a limitation of traditional classifiers that base decisions on sharp classification boundaries. As such, it is believed that this method will have advantages for applications such as biometric recognition in which the estimation of classification uncertainty is an important issue. This approach is based on the transformation of the input pattern vector relative to each classification class. Separate, monotonic, single-output neural networks are then used to represent the "degree-of-similarity" between each input pattern vector and each class. An algorithm for the implementation of this approach is proposed and tested with publicly available face-recognition data sets. The results indicate that the suggested approach provides similar classification performance to conventional principle component analysis (PCA) and linear discriminant analysis (LDA) techniques for multiclass pattern recognition problems as well as providing uncertainty information caused by misclassification.


Wave propagation retrieval method for chiral metamaterials.
In this paper we present the wave propagation method for the retrieving of effective properties of media with circularly polarized eigenwaves, in particularly for chiral metamaterials. The method is applied for thick slabs and provides bulk effective parameters. Its strong sides are the absence of artificial branches of the refractive index and simplicity in implementation. We prove the validity of the method on three case studies of homogeneous magnetized plasma, bi-cross and U-shaped metamaterials.


αvβ3, αvβ5 and αvβ6 integrins in brain metastases of lung cancer.
Integrins are transmembranous adhesion molecules postulated to be involved in the brain metastatic cascade. We investigated the correlation of alpha v beta 3 (αvβ3), alpha v beta 5 (αvβ5) and alpha v beta 6 (αvβ6) integrin isoform expression with clinical characteristics including survival times in lung cancer patients with brain metastases (BM). All BM from lung cancer operated at our institution between 1990 and 2011, were identified; where available, primary tumors were retrieved as well. Immunohistochemical analysis for αvβ3, αvβ5 and αvβ6 integrin subunits was performed and correlated with Ki67 and hypoxia-inducible factor (HIF)-1α indexes. Clinical data including survival data were obtained by chart review. 191 BM specimens of 191 patients with histologically confirmed lung cancer (172 non-small cell lung cancer and 19 small cell lung cancer) were included. In 18 patients matched primary tumor samples were available. αvβ6 expression was commonly found on BM tumor cells (103/191; 53.9 %) and showed a significant association with low Ki67 proliferation indices (46 vs. 36 %, p = 0.001, Mann-Whitney U test) and favorable survival times (p = 0.020; log rank test) in patients with non-squamous NSCLC BM. αvβ5 expression was highly expressed on vascular structures (167/191; 87.4 %) and tumor stroma in BM (151/191; 79.1 %) and associated with high HIF-1α indices (60 vs. 90, p = 0.007, Mann-Whitney U test). αvβ3 expression was more frequently found on vascular structures in BM than in primary tumors (68.1 vs. 5.6 %; p = 0.645; Chi square test) and its expression in BM tumor cells correlated with low Ki67 indices (41 vs. 28 %; p = 0.046, Mann-Whitney U test). Expression of αv integrin subunits seem to be of pathobiological and clinical relevance in patients with NSCLC BM. Further investigations of their involvement in the brain metastatic cascade and their role as biomarkers are warranted.


Transfer of yeast artificial chromosomes into mammalian cells and comparative study of their integrity.
Yeast artificial chromosomes (YACs) from the CEPH MegaYAC library (Paris, France) ranging in size from 350 to 1600 kb and mapping to the q22.1 and q22.2 regions of human chromosome 21 were transferred into mammalian cells by spheroplast fusion. The integrity of the YACs from two adjacent parts of the region was compared after retrofitting and stable transfer into mammalian cells. We found that large YACs could easily be manipulated to allow transfer of the YAC material into mammalian cells and that the size of the YAC did not appear to be limiting for fusion. However, we show that there was great variability in the integrity of the YACs from the two regions, which was not related to the size of the YACs. Four YACs in region I from sequence-tagged site (STS) G51E05 up to STS LL103 showed, in general, no loss of material and correct gene transfer into mammalian cells. In contrast, the three YACs in the more centromeric region II (from STS G51B09 up to G51E05) frequently showed a loss of human material during handling, retrofitting and transfer. As a YAC from another library covering region II was also found to be unstable, we propose that the integrity of the YACs is highly dependent on the incorporated human chromosomal DNA.


Inhibition of Japanese encephalitis virus infection by flavivirus recombinant E protein domain III.
Japanese encephalitis virus (JEV) is a mosquito-borne flavivirus closely related to the human pathogens including yellow fever virus, dengue virus and West Nile virus. There are currently no effective antiviral therapies for all of the flavivirus and only a few highly effective vaccines are licensed for human use. In this paper, the E protein domain III (DIII) of six heterologous flaviviruses (DENV1-4, WNV and JEV) was expressed in Escherichia coli successfully. The proteins were purified after a solubilization and refolding procedure, characterized by SDS-PAGE and Western blotting. Competitive inhibition showed that all recombinant flavivirus DIII proteins blocked the entry of JEV into BHK-21 cells. Further studies indicated that antibodies induced by the soluble recombinant flavivirus DIII partially protected mice against lethal JEV challenge. These results demonstrated that recombinant flavivirus DIII proteins could inhibit JEV infection competitively, and immunization with proper folding flavivirus DIII induced cross-protection against JEV infection in mice, implying a possible role of DIII for the cross-protection among flavivirus as well as its use in antigens for immunization in animal models.


Titanium nanostructures for biomedical applications.
Titanium and titanium alloys exhibit a unique combination of strength and biocompatibility, which enables their use in medical applications and accounts for their extensive use as implant materials in the last 50 years. Currently, a large amount of research is being carried out in order to determine the optimal surface topography for use in bioapplications, and thus the emphasis is on nanotechnology for biomedical applications. It was recently shown that titanium implants with rough surface topography and free energy increase osteoblast adhesion, maturation and subsequent bone formation. Furthermore, the adhesion of different cell lines to the surface of titanium implants is influenced by the surface characteristics of titanium; namely topography, charge distribution and chemistry. The present review article focuses on the specific nanotopography of titanium, i.e. titanium dioxide (TiO2) nanotubes, using a simple electrochemical anodisation method of the metallic substrate and other processes such as the hydrothermal or sol-gel template. One key advantage of using TiO2 nanotubes in cell interactions is based on the fact that TiO2 nanotube morphology is correlated with cell adhesion, spreading, growth and differentiation of mesenchymal stem cells, which were shown to be maximally induced on smaller diameter nanotubes (15 nm), but hindered on larger diameter (100 nm) tubes, leading to cell death and apoptosis. Research has supported the significance of nanotopography (TiO2 nanotube diameter) in cell adhesion and cell growth, and suggests that the mechanics of focal adhesion formation are similar among different cell types. As such, the present review will focus on perhaps the most spectacular and surprising one-dimensional structures and their unique biomedical applications for increased osseointegration, protein interaction and antibacterial properties.


Irish general practice study of acetylcysteine (Fabrol) in chronic bronchitis.
The results of this general practice study in 248 patients suggest that acetylcysteine (Fabrol) administered orally for 2 months to patients with chronic bronchitis effectively changes the viscosity and character of sputum with resultant ease of expectoration and cough severity. There was a notable improvement in associated abnormal physical signs such as the presence of rhonchi, crepitations and symptoms including dyspnoea at rest. Tolerability was good, with 77% of patients experiencing no side-effects.